Systems, methods, and apparatuses for embedded printing of thick structured meat constructs

Abstract

Systems, apparatuses, methods, and computer program products are described herein for performing embedded printing of thick structured meat constructs. Bioinks can be formed from mixing myoblasts and a hydrogel precursor with a porous microgel to form a myoblast-microgel composite ink and from mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink. A cellular yield-stress support matrix material can be formed by mixing adipose-derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel. A thick, structured, meat construct can be formed by iteratively printing, into the cellular yield-stress matrix material, the myoblast-microgel composite ink to form muscle portions, the adipocyte progenitor cell-microgel composite ink to form lipid portions, and a sacrificial ink to form vascular network portions. Sacrificial ink is then removed to define a perfusable vascular network through the thick structured meat construct.

Description

Attorney Docket No.: 049648 / 636743SYSTEMS, METHODS, AND APPARATUSES FOR EMBEDDED PRINTING OF THICK STRUCTURED MEAT CONSTRUCTSCross-Reference to Related Applications

[0001] This application claims the benefit of priority to U.S. Provisional Application Serial No. 63 / 689,446, filed August 30, 2024 and entitled “Systems, Methods, and Apparatuses for Embedded Printing of Thick Structured Meat Constructs,” the entire disclosure of which is hereby incorporated herein by reference in its entirety' for all purposes.Statement of Government Support

[0002] This invention was made with government support under grant number 2233814 awarded by the National Science Foundation. The government has certain rights in the invention.Technical Field

[0003] This disclosure relates to thick structured meat constructs and additive manufacturing processes and devices for printing the same.Background

[0004] Additive manufacturing, also commonly known as three-dimensional (3D) printing, encompasses a range of technologies used to fabricate parts by adding material to build up the part rather than by subtracting unwanted material away from a bulk starting workpiece. For freeform 3D printing of functional structures, liquid extrusion, sometimes known as direct ink writing, can be used due to its ease of implementation, high efficiency, and wide range of printable materials. However, conventional direct ink writing methods are typically not appropriate for cell-laden or cellularly viable structures, such as meat constructs and the like.Brief Summary

[0005] Described herein are systems, methods, apparatuses, compositions of matter, materials, devices, processes, approaches, and computer program products, such as storage media (e.g, a non-transitory computer-readable storage medium) for performing in full or in part embedded printing of thick structured meat constructs and thick structured meat constructs printed (or bioprinted) accordingly. Thick structured meat constructs can be bioprinted using one or more bioinks. Bioinks can be formed from mixing myoblasts and a hydrogel precursor with a porous microgel to form a my oblast-mi crogel composite ink and from mixing adipocyte1LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink. A cellular yield-stress support matrix material can be formed by mixing adipose-derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel. A thick, structured, meat construct can be formed by iteratively printing, into the cellular yield-stress matrix material, the myoblast-microgel composite ink to form muscle portions, the adipocyte progenitor cell-microgel composite ink to form lipid portions, and a sacrificial ink to form vascular network portions. Sacrificial ink is then removed to define a perfusable vascular network through the thick structured meat construct.

[0006] According to an embodiment, a method of preparing a thick structured meat construct can be carried out, the method comprising: mixing myoblasts and a hydrogel precursor with a porous microgel to form a myoblast-microgel composite ink. In some embodiments, the method can further comprise: mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink. In some embodiments, the method can further comprise: mixing adipose- derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel to form a cellular yield-stress matrix material. In some embodiments, the method can further comprise: iteratively printing one or more portions of the myoblast-microgel composite ink into the cellular yield-stress matrix material according to first spatial portions of a meat construct print design, the first spatial portions of the meat construct print design being associated with one or more muscle portions of the thick structured meat construct, one or more portions of the adipocyte progenitor cell-microgel composite ink into the cellular yield-stress matrix material according to second spatial portions of the meat construct print design, the second spatial portions of the meat construct print design being associated with one or more lipid portions of the thick structured meat construct, and one or more portions of a sacrificial ink into the cellular yield-stress matrix material according to third spatial portions of the meat construct print design, the third spatial portions of the meat construct print design being associated with a perfusable vascular network within the thick structured meat construct. In some embodiments, the method can further comprise: at least partially solidifying one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, or the cellular yield-stress matrix material. In some embodiments, the method can further comprise: removing the one or more portions of the sacrificial ink printed into the cellular yield-stress matrix material to form the perfusable vascular network within the thick structured meat construct.2LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743

[0007] In some embodiments, the method can further comprise: disposing a cross-linking agent into one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cellmicrogel composite ink, or the cellular yield-stress matrix material. In some embodiments, the cross-linking agent comprises transglutaminase. In some embodiments, the hydrogel precursor comprises a hydrogel precursor-based cellular suspension. In some embodiments, the hydrogel precursor-based cellular suspension comprises gelatin. In some embodiments, the support cells in the cellular yield-stress matrix material comprises one or more from among: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, the support cells facilitate the endothelial cells in the cellular yield-stress matrix material to form one or more capillary structures for microcirculation across the muscle portions of the thick structured meat construct.

[0008] In some embodiments, the porous microgel in the myoblast-microgel composite ink comprises gelatin. In some embodiments, the method further comprises: loading one or more of differentiation media or culturing media into the porous microgel. In some embodiments, the porous microgel in the adipocyte progenitor microgel composite ink comprises gelatin. In some embodiments, the method further comprises: loading one or more of differentiation media or culturing media into the porous microgel. In some embodiments, the cellular yield-stress matrix material is retained within a yield-stress support bath. In some embodiments, the printing of the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink. and the sacrificial ink into the cellular yield-stress matrix material comprises: adjacently printing the myoblast-microgel composite ink, the adipocyte progenitor cellmicrogel composite ink, and the sacrificial ink into the cellular yield-stress matrix material from a first side of the yield-stress support bath to a second side of the yield-stress support bath based on the meat construct print design associated with the thick structured meat construct.

[0009] In some embodiments, the sacrificial ink comprises one or more water-soluble polymers. In some embodiments, the thick structured meat construct comprises multiple layers of myoblasts, multiple layers of adipocytes, and multiple sacrificial ink filaments, and wherein a spatial arrangement of the multiple layers of myoblasts, adipocytes, and sacrificial ink filaments is controlled during the panting based upon the meat construct print design associated with the thick structured meat construct. In some embodiments, the printing of the myoblast- microgel composite ink into the cellular yield-stress support material to form the one or more muscle portions of the thick structured meat construct is carried out using one or more extrusion printing nozzles. In some embodiments, the one or more extrusion printing nozzles are3LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 dimensioned and configured to exert extrusion printing-induced shear forces on the myoblastmicrogel composite ink, thereby facilitating mature myofiber formation therefrom.

[0010] In some embodiments, the method further comprises: tensioning the myoblastmicrogel composite ink to further facilitate myofiber formation therefrom. In some embodiments, the tensioning comprises: performing mechanical tensioning of the myoblasts in the myoblast-microgel composite ink to facilitate one or more of: cell alignment, cell fusion, myotubule formation, an increased number of cell nuclei, increased myogenic protein expression, or increased cell differentiation. In some embodiments, the mechanical tensioning of the myoblasts comprises exerting on the muscle tissues one or more of: shear strain, volumetric strain, longitudinal strain, tensile strain, compressive strain, bending strain, stretching strain, multiaxial strain, uniaxial strain, randomized iterative strain, periodic strain, cyclic strain, or continuous strain.

[0011] According to another embodiment, an apparatus can be provided that comprises at least one processor and at least one memory' storing instructions thereon that, when executed by the at least one processor, cause the apparatus to perform preparation (e.g., embedded bioprinting) of a thick structured meat construct. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, cause the apparatus to perform: mixing myoblasts and a hydrogel precursor with a porous microgel to form a myoblast-microgel composite ink. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, can further cause the apparatus to perform: mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink. In some embodiments, the instructions stored on the at least one memory', when executed by the at least one processor, can further cause the apparatus to perform: mixing adipose-derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel to form a cellular yieldstress matrix material. In some embodiments, the instructions stored on the at least one memory7, when executed by the at least one processor, can further cause the apparatus to perform: iteratively printing one or more portions of the myoblast-microgel composite ink into the cellular yield-stress matrix material according to first spatial portions of a meat construct print design, the first spatial portions of the meat construct print design being associated with one or more muscle portions of the thick structured meat construct, one or more portions of the adipocyte progenitor cell-microgel composite ink into the cellular yield-stress matrix material according to second spatial portions of the meat construct print design, the second spatial portions of the meat construct print design being associated with one or more lipid portions of4LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 the thick structured meat construct, and one or more portions of a sacrificial ink into the cellular yield-stress matrix material according to third spatial portions of the meat construct print design, the third spatial portions of the meat construct print design being associated with a perfusable vascular network within the thick structured meat construct. In some embodiments, the instructions stored on the at least one memory. when executed by the at least one processor, can further cause the apparatus to perform: at least partially solidifying one or more of: the my oblast-mi crogel composite ink. the adipocyte progenitor cell-microgel composite ink, or the cellular yield-stress matrix material. In some embodiments, the instructions stored on the at least one memory. when executed by the at least one processor, can further cause the apparatus to perform: removing the one or more portions of the sacrificial ink printed into the cellular yield-stress matrix material to form the perfusable vascular network within the thick structured meat construct.

[0012] In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, further cause the apparatus to perform: disposing a cross-linking agent into one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink. or the cellular yield-stress matrix material. In some embodiments, the cross-linking agent comprises transglutaminase. In some embodiments, the hydrogel precursor comprises a hydrogel precursor-based cellular suspension. In some embodiments, the hydrogel precursor-based cellular suspension comprises gelatin. In some embodiments, the support cells in the cellular yield-stress matrix material comprises one or more from among: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, the support cells facilitate the endothelial cells in the cellular yieldstress matrix material to form one or more capillary structures for microcirculation across the muscle portions of the thick structured meat construct.

[0013] In some embodiments, the porous microgel in the myoblast-microgel composite ink comprises gelatin. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, further cause the apparatus to perform: loading one or more of differentiation media or culturing media into the porous microgel. In some embodiments, the porous microgel in the adipocyte progenitor microgel composite ink comprises gelatin. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, further cause the apparatus to perform: loading one or more of differentiation media or culturing media into the porous microgel. In some embodiments, the cellular yield-stress matrix material is retained within a yield-stress support bath. In some embodiments, the printing of the myoblast-microgel composite ink, the5LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 adipocyte progenitor cell-microgel composite ink, and the sacrificial ink into the cellular yieldstress matrix material comprises: adjacently printing the my oblast-mi crogel composite ink. the adipocyte progenitor cell-microgel composite ink, and the sacrificial ink into the cellular yieldstress matrix material from a first side of the yield-stress support bath to a second side of the yield-stress support bath based on the meat construct print design associated with the thick structured meat construct.

[0014] In some embodiments, the sacrificial ink comprises one or more water-soluble polymers. In some embodiments, the thick structured meat construct comprises multiple layers of myoblasts, multiple layers of adipocytes, and multiple sacrificial ink filaments, and wherein a spatial arrangement of the multiple layers of myoblasts, adipocytes, and sacrificial ink filaments is controlled during the printing based upon the meat construct print design associated with the thick structured meat construct. In some embodiments, the printing of the myoblastmicrogel composite ink into the cellular yield-stress support material to form the one or more muscle portions of the thick structured meat construct is carried out using one or more extrusion printing nozzles. In some embodiments, the one or more extrusion printing nozzles are dimensioned and configured to exert extrusion printing-induced shear forces on the myoblastmicrogel composite ink, thereby facilitating mature myofiber formation therefrom.

[0015] In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, further cause the apparatus to perform: tensioning the myoblast-microgel composite ink to further facilitate myofiber formation therefrom. In some embodiments, the tensioning comprises: performing mechanical tensioning of the myoblasts in the myoblast-microgel composite ink to facilitate one or more of: cell alignment, cell fusion, myotubule formation, an increased number of cell nuclei, increased myogenic protein expression, or increased cell differentiation. In some embodiments, the mechanical tensioning of the myoblasts comprises exerting on the muscle tissues one or more of: shear strain, volumetric strain, longitudinal strain, tensile strain, compressive strain, bending strain, stretching strain, multiaxial strain, uniaxial strain, randomized iterative strain, periodic strain, cyclic strain, or continuous strain.

[0016] According to another embodiment, a computer program product can be provided that comprises a non-transitory computer-readable storage medium storing instructions thereon that, when executed by at least one processor of an apparatus, cause the apparatus to perform at least a portion of preparing (e.g. , embedded bioprinting) of a thick structured meat construct. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, cause the apparatus to perform: mixing6LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 myoblasts and a hydrogel precursor with a porous microgel to form a myoblast-microgel composite ink. In some embodiments, the instructions stored on the non-transitory computer- readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: mixing adipose-derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel to form a cellular yield-stress matrix material. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: iteratively printing one or more portions of the myoblast-microgel composite ink into the cellular yield-stress matrix material according to first spatial portions of a meat construct print design, the first spatial portions of the meat construct print design being associated with one or more muscle portions of the thick structured meat construct, one or more portions of the adipocyte progenitor cell-microgel composite ink into the cellular yield-stress matrix material according to second spatial portions of the meat construct print design, the second spatial portions of the meat construct print design being associated with one or more lipid portions of the thick structured meat construct, and one or more portions of a sacrificial ink into the cellular yield-stress matrix material according to third spatial portions of the meat construct print design, the third spatial portions of the meat construct print design being associated with a perfusable vascular network within the thick structured meat construct; at least partially solidifying one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, or the cellular yield-stress matrix material. In some embodiments, the instructions stored on the non- transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: removing the one or more portions of the sacrificial ink printed into the cellular yield-stress matrix material to form the perfusable vascular network within the thick structured meat construct.

[0017] In some embodiments, the instructions stored on the non-transitory computer- readable storage medium, when executed by the at least one processor, further cause the apparatus to perform: disposing a cross-linking agent into one or more of: the myoblast- microgel composite ink, the adipocyte progenitor cell-microgel composite ink, or the cellular yield-stress matrix material. In some embodiments, the cross-linking agent comprises transglutaminase. In some embodiments, the hydrogel precursor comprises a hydrogel7LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 precursor-based cellular suspension. In some embodiments, the hydrogel precursor-based cellular suspension comprises gelatin. In some embodiments, the support cells in the cellular yield-stress matrix material comprises one or more from among: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, the support cells facilitate the endothelial cells in the cellular yield-stress matrix material to form one or more capillary structures for microcirculation across the muscle portions of the thick structured meat construct.

[0018] In some embodiments, the porous microgel in the myoblast-microgel composite ink comprises gelatin. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, further cause the apparatus to perform: loading one or more of differentiation media or culturing media into the porous microgel. In some embodiments, the porous microgel in the adipocyte progenitor microgel composite ink comprises gelatin. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, further cause the apparatus to perform: loading one or more of differentiation media or culturing media into the porous microgel. In some embodiments, the cellular yield-stress matrix material is retained within a yield-stress support bath. In some embodiments, the printing of the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, and the sacrificial ink into the cellular yield-stress matrix material comprises: adjacently printing the myoblast-microgel composite ink, the adipocyte progenitor cellmicrogel composite ink, and the sacrificial ink into the cellular yield-stress matrix material from a first side of the yield-stress support bath to a second side of the yield-stress support bath based on the meat construct print design associated with the thick structured meat construct.

[0019] In some embodiments, the sacrificial ink comprises one or more water-soluble polymers. In some embodiments, the thick structured meat construct comprises multiple layers of myoblasts, multiple layers of adipocytes, and multiple sacrificial ink filaments, and wherein a spatial arrangement of the multiple layers of myoblasts, adipocytes, and sacrificial ink filaments is controlled during the printing based upon the meat construct print design associated wi th the thick structured meat construct. In some embodiments, the printing of the myoblast- microgel composite ink into the cellular yield-stress support material to form the one or more muscle portions of the thick structured meat construct is carried out using one or more extrusion printing nozzles. In some embodiments, the one or more extrusion printing nozzles are dimensioned and configured to exert extrusion printing-induced shear forces on the myoblast- microgel composite ink, thereby facilitating mature myofiber formation therefrom.8LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743

[0020] In some embodiments, the instructions stored on the non-transitory computer- readable storage medium, when executed by the at least one processor, further cause the apparatus to perform: tensioning the myoblast-mi crogel composite ink to further facilitate myofiber formation therefrom. In some embodiments, the tensioning comprises: performing mechanical tensioning of the myoblasts in the myoblast-microgel composite ink to facilitate one or more of: cell alignment, cell fusion, myotubule formation, an increased number of cell nuclei, increased myogenic protein expression, or increased cell differentiation. In some embodiments, the mechanical tensioning of the myoblasts comprises exerting on the muscle tissues one or more of: shear strain, volumetric strain, longitudinal strain, tensile strain, compressive strain, bending strain, stretching strain, multiaxial strain, uniaxial strain, randomized iterative strain, periodic strain, cyclic strain, or continuous strain.

[0021] According to another embodiment, a system or device can be provided that is configured to perform at least a portion of preparing (e.g., embedded bioprinting) of a thick structured meat construct. In some embodiments, the system or device comprises: means for mixing myoblasts and a hydrogel precursor with a porous microgel to form a myoblast- microgel composite ink. In some embodiments, the system or device further comprises: means for mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink. In some embodiments, the system or device further comprises: means for mixing adipose-derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel to form a cellular yield-stress matrix material. In some embodiments, the system or device further comprises: means for iteratively printing one or more portions of the myoblast-microgel composite ink into the cellular yield-stress matrix material according to first spatial portions of a meat construct print design, the first spatial portions of the meat construct print design being associated with one or more muscle portions of the thick structured meat construct, one or more portions of the adipocyte progenitor cell-microgel composite ink into the cellular yield-stress matrix material according to second spatial portions of the meat construct print design, the second spatial portions of the meat construct print design being associated with one or more lipid portions of the thick structured meat construct, and one or more portions of a sacrificial ink into the cellular yield-stress matrix material according to third spatial portions of the meat construct print design, the third spatial portions of the meat construct print design being associated with a perfusable vascular network within the thick structured meat construct. In some embodiments, the system or device further comprises: means for at least partially solidifying one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, or9LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 the cellular yield-stress matrix material. In some embodiments, the system or device further comprises: means for removing the one or more portions of the sacrificial ink printed into the cellular yield-stress matrix material to form the perfusable vascular network within the thick structured meat construct.

[0022] In some embodiments, the system or device can further comprise: means for disposing a cross-linking agent into one or more of: the my oblast-mi crogel composite ink, the adipocyte progenitor cell-microgel composite ink, or the cellular yield-stress matrix material. In some embodiments, the cross-linking agent comprises transglutaminase. In some embodiments, the hydrogel precursor comprises a hydrogel precursor-based cellular suspension. In some embodiments, the hydrogel precursor-based cellular suspension comprises gelatin. In some embodiments, the support cells in the cellular yield-stress matrix material comprises one or more from among: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, the support cells facilitate the endothelial cells in the cellular yield-stress matrix material to form one or more capillary structures for microcirculation across the muscle portions of the thick structured meat construct.

[0023] In some embodiments, the porous microgel in the myoblast-microgel composite ink comprises gelatin. In some embodiments, the system or device can further comprise: means for loading one or more of differentiation media or culturing media into the porous microgel. In some embodiments, the porous microgel in the adipocyte progenitor microgel composite ink comprises gelatin. In some embodiments, the system or device can further comprise: means for loading one or more of differentiation media or culturing media into the porous microgel. In some embodiments, the cellular yield-stress matrix material is retained within a yield-stress support bath. In some embodiments, the printing of the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, and the sacrificial ink into the cellular yieldstress matrix material comprises: adjacently printing the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, and the sacrificial ink into the cellular yieldstress matrix material from a first side of the yield-stress support bath to a second side of the yield-stress support bath based on the meat construct print design associated with the thick structured meat construct.

[0024] In some embodiments, the sacrificial ink comprises one or more water-soluble polymers. In some embodiments, the thick structured meat construct comprises multiple layers of myoblasts, multiple layers of adipocytes, and multiple sacrificial ink filaments, and wherein a spatial arrangement of the multiple layers of myoblasts, adipocytes, and sacrificial ink filaments is controlled during the printing based upon the meat construct print design associated10LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 with the thick structured meat construct. In some embodiments, the printing of the myoblastmicrogel composite ink into the cellular yield-stress support material to form the one or more muscle portions of the thick structured meat construct is carried out using one or more extrusion printing nozzles. In some embodiments, the one or more extrusion printing nozzles are dimensioned and configured to exert extrusion printing-induced shear forces on the myoblastmicrogel composite ink, thereby facilitating mature myofiber formation therefrom.

[0025] In some embodiments, the system or device can further comprise: means for tensioning the myobl ast-microgel composite ink to further facilitate myofiber formation therefrom. In some embodiments, the means for tensioning comprise means for performing mechanical tensioning of the myoblasts in the myoblast-microgel composite ink to facilitate one or more of: cell alignment, cell fusion, myotubule formation, an increased number of cell nuclei, increased myogenic protein expression, or increased cell differentiation. In some embodiments, the means for mechanical tensioning of the myoblasts comprises means for exerting, on the muscle tissues, one or more of: shear strain, volumetric strain, longitudinal strain, tensile strain, compressive strain, bending strain, stretching strain, multiaxial strain, uniaxial strain, randomized iterative strain, penodic strain, cyclic strain, or continuous strain.

[0026] According to another embodiment, a method of preparing a thick structured meat construct can be carried out, the method comprising: mixing a plurality of first living cells with one or more porous microgels and a hydrogel precursor to form a first ink. In some embodiments, the method can further comprise: mixing a plurality of second living cells with one or more porous microgels and a hydrogel precursor to form a second ink. In some embodiments, the method can further comprise: mixing one or more porous microgels, a plurality of endothelial cells, and a plurality of support cells with a hydrogel precursor to form a cellular yield-stress matrix material. In some embodiments, the method can further comprise: adding a crosslinking agent to at least one of the first ink, the second ink, or the cellular yieldstress matrix material; forming a third ink comprising a sacrificial material. In some embodiments, the method can further comprise: selectively and iteratively dispensing one or more volumes of the first ink, one or more volumes of the second ink, and one or more volumes of the third ink into the cellular yield-stress matrix material to form an uncross-linked printed tissue construct. In some embodiments, the method can further comprise: exposing the uncrosslinked printed tissue construct to ultraviolet radiation or a to cross-link the cellular yield-stress matrix material and the cellular microgel composite ink, thereby forming a cross-linked printed tissue construct. In some embodiments, the method can further comprise: immersing the crosslinked printed tissue construct in phosphate-buffered saline to at least partially dilute or11LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 dissolve the third ink. In some embodiments, the method can further comprise: flushing the third ink and phosphate-buffered saline from the cross-linked printed tissue construct to form one or more channels of a perfusable network within the cross-linked printed tissue construct. In some embodiments, the method can further comprise: after flushing the sacrificial ink and phosphate-buffered saline from the cross-linked printed tissue construct, connecting the perfusable network of the cross-linked printed tissue construct to a peristaltic pump-based perfusion system together with a post-printing tensile-force-based tensioning system applied to the tissue construct. In some embodiments, the method can further comprise: perfusing a cellular media through the one or more channels of the perfusable network within the crosslinked printed tissue construct.

[0027] In some embodiments, the cellular yield-stress matrix comprises one or more solid porous microgels, a liquid hydrogel precursor, a plurality of endothelial cells, and a plurality of support cells. In some embodiments, the plurality of support cells comprises one or more of: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, the crosslinking agent comprises transglutaminase. In some embodiments, the first ink is a first cellular microgel composite ink. In some embodiments, the method further comprises: mixing a plurality of living cells with one or more solid porous microgels and a liquid hydrogel precursor to form the second ink, wherein the second ink is a second cellular microgel composite ink.

[0028] In some embodiments, the first cellular microgel composite ink is a myoblast-laden microgel composite ink comprised of at least a plurality of living cells, solid porous microgels, and a liquid hydrogel precursor. In some embodiments, the plurality of living cells comprises animal stem cells. In some embodiments, the solid porous microgels comprise one or more of: polyethylene glycol (PEG), hyaluronic acid, chitosan, or gelatin. In some embodiments, the liquid hydrogel precursor is comprised of at least collagen, gelatin, gellan, alginate, fibrin, silk, chitosan, or hyaluronic acid.

[0029] In some embodiments, the second cellular microgel composite in is an adipocyte progenitor cell-laden microgel composite ink comprised of at least a plurality of living cells, solid porous microgels, and a liquid hydrogel precursor. In some embodiments, the third ink comprises one or more of: a hydrophilic non-ionic surfactant, a poloxamer, a material comprising a central hydrophobic block of polypropylene glycol flanked by two hydrophilic blocks of polyethylene glycol, C572H1146O259, methylcellulose, polyvinyl alcohol, xanthangum, or carbohydrates.12LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743

[0030] In some embodiments, the first ink, second ink, or cellular matrix are not exposed to a chemical or enzymatic crosslinking agent. In some embodiments, the method further comprises: exposing the thick structured meat construct to ultraviolet radiation to cause at least partial crosslinking of one or more of the first ink, the second, the third ink, or the cellular yield-stress matrix material. In some embodiments, the phosphate buffered saline solution has a pH level of between about 5.8 and about 7.4. In some embodiments, the phosphate buffered saline comprises one or more of: sodium chloride, potassium chloride, potassium dihydrogen phosphate, or disodium hydrogen phosphate.

[0031] In some embodiments, the method further comprises: culturing the thick structured meat construct under physiologically relevant medium perfusion conditions and necessary tensioning conditions to mature the myoblasts and adipocytes while endothelial cells selfassemble into capillary structures around the main perfusable channel. In some embodiments, the method further comprises: when selectively and iteratively dispensing the one or more volumes of the first ink and the one or more volumes of the second ink through the printing nozzle into the cellular yield-stress matrix material to form the uncross-linked printed tissue construct, selectively and iteratively dispensing one or more volumes of the second cellular microgel composite ink through the printing nozzle into the cellular yield-stress matrix material. In some embodiments, the method further comprises: printing myoblast filaments into the cellular yield-stress matrix material; and enculturing the myoblast filaments to form a muscle tissue layer.

[0032] According to another embodiment, an apparatus can be provided that comprises at least one processor and at least one memory storing instructions thereon that, when executed by the at least one processor, cause the apparatus to perform at least in part preparing (e.g., embedded bioprinting) of a thick structured meat construct. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, cause the apparatus to perform: mixing a plurality of first living cells with one or more porous microgels and a hydrogel precursor to form a first ink. In some embodiments, the instructions stored on the at least one memory', when executed by the at least one processor, can further cause the apparatus to perform: mixing a plurality of second living cells with one or more porous microgels and a hydrogel precursor to form a second ink. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, can further cause the apparatus to perform: mixing one or more porous microgels, a plurality of endothelial cells, and a plurality of support cells with a hydrogel precursor to form a cellular yield-stress matrix material. In some embodiments, the instructions stored on the at least one13LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 memory', when executed by the at least one processor, can further cause the apparatus to perform: adding a crosslinking agent to at least one of the first ink, the second ink, or the cellular yield-stress matrix material. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, can further cause the apparatus to perform: forming a third ink comprising a sacrificial material. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, can further cause the apparatus to perform: selectively and iteratively dispensing one or more volumes of the first ink, one or more volumes of the second ink, and one or more volumes of the third ink into the cellular yield-stress matrix material to form an uncross-linked printed tissue construct. In some embodiments, the instructions stored on the at least one memory', when executed by the at least one processor, can further cause the apparatus to perform: exposing the uncross-linked printed tissue construct to ultraviolet radiation or a to cross-link the cellular yield-stress matrix material and the cellular microgel composite ink, thereby forming a cross-linked printed tissue construct. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, can further cause the apparatus to perform: immersing the cross-linked printed tissue construct in phosphate- buffered saline to at least partially7dilute or dissolve the third ink; flushing the third ink and phosphate-buffered saline from the cross-linked printed tissue construct to form one or more channels of a perfusable network within the cross-linked printed tissue construct. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, can further cause the apparatus to perform: after flushing the sacrificial ink and phosphate-buffered saline from the cross-linked printed tissue construct, connecting the perfusable network of the cross-linked printed tissue construct to a peristaltic pump-based perfusion system together with a post-printing tensile-force-based tensioning system applied to the tissue construct. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, can further cause the apparatus to perform: perfusing a cellular media through the one or more channels of the perfusable netw ork within the cross-linked printed tissue construct.

[0033] In some embodiments, the cellular yield-stress matrix comprises one or more solid porous microgels, a liquid hydrogel precursor, a plurality' of endothelial cells, and a plurality of support cells. In some embodiments, the plurality' of support cells comprises one or more of: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, the crosslinking agent comprises transglutaminase. In some embodiments, the first ink is a first cellular microgel composite ink. In some embodiments, the instructions stored14LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 on the at least one memory, when executed by the at least one processor, further cause the apparatus to perform: mixing a plurality of living cells with one or more solid porous microgels and a liquid hydrogel precursor to form the second ink, wherein the second ink is a second cellular microgel composite ink.

[0034] In some embodiments, the first cellular microgel composite ink is a myoblast-laden microgel composite ink comprised of at least a plurality of living cells, solid porous microgels, and a liquid hydrogel precursor. In some embodiments, the plurality of living cells comprises animal stem cells. In some embodiments, the solid porous microgels comprise one or more of: polyethylene glycol (PEG), hyaluronic acid, chitosan, or gelatin. In some embodiments, the liquid hydrogel precursor is comprised of at least collagen, gelatin, gellan, alginate, fibrin, silk, chitosan, or hyaluronic acid.

[0035] In some embodiments, the second cellular microgel composite in is an adipocyte progenitor cell-laden microgel composite ink comprised of at least a plurality of living cells, solid porous microgels, and a liquid hydrogel precursor. In some embodiments, the third ink comprises one or more of: a hydrophilic non-ionic surfactant, a poloxamer, a material comprising a central hydrophobic block of polypropylene glycol flanked by two hydrophilic blocks of polyethylene glycol, C572H1146O259, methylcellulose, polyvinyl alcohol, xanthan gum, or carbohydrates.

[0036] In some embodiments, the first ink, second ink, or cellular matrix are not exposed to a chemical or enzymatic crosslinking agent. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, further cause the apparatus to perform: exposing the thick structured meat construct to ultraviolet radiation to cause at least partial crosslinking of one or more of the first ink, the second, the third ink, or the cellular yield-stress matrix material. In some embodiments, the phosphate buffered saline solution has a pH level of between about 5.8 and about 7.4. In some embodiments, the phosphate buffered saline comprises one or more of: sodium chloride, potassium chloride, potassium dihydrogen phosphate, or disodium hydrogen phosphate.

[0037] In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, further cause the apparatus to perform: culturing the thick structured meat construct under physiologically relevant medium perfusion conditions and necessary tensioning conditions to mature the myoblasts and adipocytes while endothelial cells self-assemble into capillary structures around the main perfusable channel. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, further cause the apparatus to perform: when selectively and iteratively15LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 dispensing the one or more volumes of the first ink and the one or more volumes of the second ink through the printing nozzle into the cellular yield-stress matrix material to form the uncrosslinked printed tissue construct, selectively and iteratively dispensing one or more volumes of the second cellular microgel composite ink through the printing nozzle into the cellular yieldstress matrix material. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, further cause the apparatus to perform: printing myoblast filaments into the cellular yield-stress matrix material; and enculturing the myoblast filaments to form a muscle tissue layer.

[0038] According to another embodiment, a computer program product can be provided that comprises anon-transitory computer-readable storage medium storing instructions thereon that, when executed by’ at least one processor of an apparatus, cause the apparatus to perform at least in part preparing (, embedded bioprinting) of a thick structured meat construct. In some embodiments, the instructions stored on the non- transitory' computer-readable storage medium, when executed by the at least one processor, cause the apparatus to perform: mixing a plurality of first living cells with one or more porous microgels and a hydrogel precursor to form a first ink. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: mixing a plurality’ of second living cells with one or more porous microgels and a hydrogel precursor to form a second ink. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: mixing one or more porous microgels, a plurality’ of endothelial cells, and a plurality of support cells with a hydrogel precursor to form a cellular yield-stress matrix material. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: adding a crosslinking agent to at least one of the first ink, the second ink, or the cellular yield-stress matrix material. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: forming a third ink comprising a sacrificial material; selectively and iteratively dispensing one or more volumes of the first ink, one or more volumes of the second ink, and one or more volumes of the third ink into the cellular yield-stress matrix material to form an uncross-linked printed tissue construct. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: exposing the uncross-linked printed tissue construct to16LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 ultraviolet radiation or a to cross-link the cellular yield-stress matrix material and the cellular microgel composite ink, thereby forming a cross-linked printed tissue construct. In some embodiments, the instructions stored on the non- transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: immersing the cross-linked printed tissue construct in phosphate-buffered saline to at least partially dilute or dissolve the third ink; flushing the third ink and phosphate-buffered saline from the cross-linked printed tissue construct to form one or more channels of a perfusable network within the cross-linked printed tissue construct. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: after flushing the sacrificial ink and phosphate-buffered saline from the cross-linked printed tissue construct, connecting the perfusable network of the cross-linked printed tissue construct to a peristaltic pump-based perfusion system together with a post-printing tensile-force-based tensioning system applied to the tissue construct. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: perfusing a cellular media through the one or more channels of the perfusable network within the cross-linked printed tissue construct.

[0039] In some embodiments, the cellular yield-stress matrix comprises one or more solid porous microgels, a liquid hydrogel precursor, a plurality of endothelial cells, and a plurality of support cells. In some embodiments, the plurality of support cells comprises one or more of: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, the crosslinking agent comprises transglutaminase. In some embodiments, the first ink is a first cellular microgel composite ink. In some embodiments, the instructions stored on the non-transitory’ computer-readable storage medium, when executed by the at least one processor of the apparatus, further cause the apparatus to perform: mixing a plurality of living cells with one or more solid porous microgels and a liquid hydrogel precursor to form the second ink, wherein the second ink is a second cellular microgel composite ink.

[0040] In some embodiments, the first cellular microgel composite ink is a myoblast-laden microgel composite ink comprised of at least a plurality of living cells, solid porous microgels, and a liquid hydrogel precursor. In some embodiments, the plurality of living cells comprises animal stem cells. In some embodiments, the solid porous microgels comprise one or more of: polyethylene glycol (PEG), hyaluronic acid, chitosan, or gelatin. In some embodiments, the liquid hydrogel precursor is comprised of at least collagen, gelatin, gellan, alginate, fibrin, silk, chitosan, or hyaluronic acid.17LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743

[0041] In some embodiments, the second cellular microgel composite in is an adipocyte progenitor cell-laden microgel composite ink comprised of at least a plurality of living cells, solid porous microgels, and a liquid hydrogel precursor. In some embodiments, the third ink comprises one or more of: a hydrophilic non-ionic surfactant, a poloxamer, a material comprising a central hydrophobic block of polypropylene glycol flanked by two hydrophilic blocks of polyethylene glycol, C572H1146O259, methylcellulose, polyvinyl alcohol, xanthangum, or carbohydrates.

[0042] In some embodiments, the first ink, second ink, or cellular matrix are not exposed to a chemical or enzymatic crosslinking agent. In some embodiments, the instructions stored on the non- transitory computer-readable storage medium, when executed by the at least one processor of the apparatus, further cause the apparatus to perform: exposing the thick structured meat construct to ultraviolet radiation to cause at least partial crosslinking of one or more of the first ink, the second, the third ink, or the cellular yield-stress matrix material. In some embodiments, the phosphate buffered saline solution has a pH level of between about 5.8 and about 7.4. In some embodiments, the phosphate buffered saline comprises one or more of: sodium chloride, potassium chloride, potassium dihydrogen phosphate, or disodium hydrogen phosphate.

[0043] In some embodiments, the instructions stored on the non-transitory computer- readable storage medium, when executed by the at least one processor of the apparatus, further cause the apparatus to perform: culturing the thick structured meat construct under physiologically relevant medium perfusion conditions and necessary tensioning conditions to mature the myoblasts and adipocytes while endothelial cells self-assemble into capillary structures around the main perfusable channel. In some embodiments, the instructions stored on the non-transitory’ computer-readable storage medium, when executed by the at least one processor of the apparatus, further cause the apparatus to perform: when selectively and iteratively dispensing the one or more volumes of the first ink and the one or more volumes of the second ink through the printing nozzle into the cellular yield-stress matrix material to form the uncross-linked printed tissue construct, selectively and iteratively dispensing one or more volumes of the second cellular microgel composite ink through the printing nozzle into the cellular yield-stress matrix material. In some embodiments, the instructions stored on the non- transitory computer-readable storage medium, when executed by the at least one processor of the apparatus, further cause the apparatus to perform: printing myoblast filaments into the cellular yield-stress matrix material; and enculturing the myoblast filaments to form a muscle tissue layer.18LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743

[0044] According to another embodiment, a system or device can be provided that is configured for carrying out at least in part preparing (e.g.. embedded bioprinting) of a thick structured meat construct. In some embodiments, the system or device can comprise: means for mixing a plurality of first living cells with one or more porous microgels and a hydrogel precursor to form a first ink; mixing a plurality' of second living cells with one or more porous microgels and a hydrogel precursor to form a second ink. In some embodiments, the system or device can further comprise: means for mixing one or more porous microgels, a plurality' of endothelial cells, and a plurality of support cells with a hydrogel precursor to form a cellular yield-stress matrix material. In some embodiments, the system or device can further comprise: means for adding a crosslinking agent to at least one of the first ink, the second ink, or the cellular yield-stress matrix material. In some embodiments, the system or device can further comprise: means for forming a third ink comprising a sacrificial material. In some embodiments, the system or device can further comprise: means for selectively and iteratively dispensing one or more volumes of the first ink, one or more volumes of the second ink, and one or more volumes of the third ink into the cellular yield-stress matrix material to form an uncross-linked printed tissue construct. In some embodiments, the system or device can further comprise: means for exposing the uncross-linked printed tissue construct to ultraviolet radiation or a to cross-link the cellular yield-stress matrix material and the cellular microgel composite ink, thereby forming a cross-linked printed tissue construct. In some embodiments, the system or device can further comprise: means for immersing the cross-linked printed tissue construct in phosphate-buffered saline to at least partially dilute or dissolve the third ink. In some embodiments, the system or device can further comprise: means for flushing the third ink and phosphate-buffered saline from the cross-linked printed tissue construct to form one or more channels of a perfusable network within the cross-linked printed tissue construct. In some embodiments, the system or device can further comprise: means for after flushing the sacrificial ink and phosphate-buffered saline from the cross-linked printed tissue construct, connecting the perfusable network of the cross-linked printed tissue construct to a peristaltic pump-based perfusion system together with a post-printing tensile-force-based tensioning system applied to the tissue construct. In some embodiments, the system or device can further comprise: means for perfusing a cellular media through the one or more channels of the perfusable network yvithin the cross-linked printed tissue construct.

[0045] In some embodiments, the cellular yield-stress matrix comprises one or more solid porous microgels, a liquid hydrogel precursor, a plurality of endothelial cells, and a plurality of support cells. In some embodiments, the plurality of support cells comprises one or more19LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 of: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, the crosslinking agent comprises transglutaminase. In some embodiments, the first ink is a first cellular microgel composite ink. In some embodiments, the system or device can further comprise: means for mixing a plurality of living cells with one or more solid porous microgels and a liquid hydrogel precursor to form the second ink, wherein the second ink is a second cellular microgel composite ink.

[0046] In some embodiments, the first cellular microgel composite ink is a myoblast-laden microgel composite ink comprised of at least a plurality of living cells, solid porous microgels, and a liquid hydrogel precursor. In some embodiments, the plurality of living cells comprises animal stem cells. In some embodiments, the solid porous microgels comprise one or more of: polyethylene glycol (PEG), hyaluronic acid, chitosan, or gelatin. In some embodiments, the liquid hydrogel precursor is comprised of at least collagen, gelatin, gellan, alginate, fibrin, silk, chitosan, or hyaluronic acid.

[0047] In some embodiments, the second cellular microgel composite in is an adipocyte progenitor cell-laden microgel composite ink comprised of at least a plurality of living cells, solid porous microgels, and a liquid hydrogel precursor. In some embodiments, the third ink comprises one or more of: a hydrophilic non-ionic surfactant, a poloxamer, a material comprising a central hydrophobic block of polypropylene glycol flanked by two hydrophilic blocks of polyethylene glycol, C572H1146O259, methylcellulose, polyvinyl alcohol, xanthangum, or carbohydrates.

[0048] In some embodiments, the first ink, second ink, or cellular matrix are not exposed to a chemical or enzymatic crosslinking agent. In some embodiments, the system or device can further comprise: means for exposing the thick structured meat construct to ultraviolet radiation to cause at least partial crosslinking of one or more of the first ink, the second, the third ink, or the cellular yield-stress matrix material. In some embodiments, the phosphate buffered saline solution has a pH level of between about 5.8 and about 7.4. In some embodiments, the phosphate buffered saline comprises one or more of: sodium chloride, potassium chloride, potassium dihydrogen phosphate, or disodium hydrogen phosphate.

[0049] In some embodiments, the system or device can further comprise: means for culturing the thick structured meat construct under physiologically relevant medium perfusion conditions and necessary’ tensioning conditions to mature the myoblasts and adipocytes while endothelial cells self-assemble into capillary structures around the main perfusable channel. In some embodiments, the system or device can further comprise: means for when selectively and iteratively dispensing the one or more volumes of the first ink and the one or more volumes of20LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 the second ink through the printing nozzle into the cellular yield-stress matrix material to form the uncross-linked printed tissue construct, selectively and iteratively dispensing one or more volumes of the second cellular microgel composite ink through the printing nozzle into the cellular yield-stress matrix material. In some embodiments, the system or device can further comprise: means for printing myoblast fdaments into the cellular yield-stress matrix material; and enculturing the myoblast fdaments to form a muscle tissue layer.

[0050] According to another embodiment, a thick structured meat construct can be printed, formed, or provided. In some embodiments, the thick structured meat construct comprises: a first ink comprising a plurality of living cells, one or more solid porous microgels, and a liquid hydrogel precursor; and a cellular yield-stress matrix material comprising one or more solid porous microgels, a liquid hydrogel precursor, a plurality of endothelial cells, and a plurality of support cells. In some embodiments, the plurality of support cells comprise one or more of: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, at least one of the first ink or the cellular yield-stress matrix material further comprises a crosslinking agent. In some embodiments, the first ink and the second ink are disposed within the cellular yield-stress matrix material according to a 3D printing design / printing pathway and cured using ultraviolet radiation to form the thick structured meat / tissue construct.

[0051] In some embodiments, the crosslinking agent comprising transglutaminase. In some embodiments, the first ink comprises a cellular microgel composite. In some embodiments, the second ink comprises a sacrificial material. In some embodiments, the third ink is configured to be diluted or dissolved via phosphate-buffered saline to form a perfusable network within the thick structured meat / tissue construct. In some embodiments, the perfusable network is configured to be placed in fluidic communication with a perfusion system or a post-printing tensioning system to facilitate perfusion of the thick structured meat / tissue construct. In some embodiments, the perfusable network is configured to fluidically receive a cellular media from the perfusion system or post-printing tensioning system to enhance cellular survival within the thick structured meat / tissue construct.

[0052] According to another embodiment, an apparatus can be provided that comprises: at least one processor; and at least one memory storing instructions thereon that, when executed by the at least one processor, cause the apparatus to perform at least: mixing myoblasts and a hydrogel precursor with a porous microgel to form a my oblast-mi crogel composite ink; mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink; mixing adipose-derived stem cells, endothelial cells,21LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 support cells, and a hydrogel precursor with a porous microgel to form a cellular yield-stress matrix material; iteratively printing one or more portions of the myoblast-microgel composite ink into the cellular yield-stress matrix material according to first spatial portions of a meat construct print design, the first spatial portions of the meat construct print design being associated with one or more muscle portions of the thick structured meat construct, one or more portions of the adipocyte progenitor cell-microgel composite ink into the cellular yield-stress matrix material according to second spatial portions of the meat construct print design, the second spatial portions of the meat construct print design being associated with one or more lipid portions of the thick structured meat construct, and one or more portions of a sacrificial ink into the cellular yield-stress matrix material according to third spatial portions of the meat construct print design, the third spatial portions of the meat construct print design being associated with a perfusable vascular network within the thick structured meat construct; at least partially solidifying one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, or the cellular yield-stress matrix material; and removing the one or more portions of the sacrificial ink printed into the cellular yield-stress matrix material to form the perfusable vascular network within the thick structured meat construct.

[0053] According to another embodiment, a computer program product can be provided that comprises a non-transitory computer-readable storage medium storing computerexecutable instructions thereon that, when executed by at least one processor, cause an apparatus to perform at least: mixing myoblasts and a hydrogel precursor with a porous microgel to form a myoblast-microgel composite ink; mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink; mixing adipose-derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel to form a cellular yield-stress matrix material; iteratively printing one or more portions of the myoblast-microgel composite ink into the cellular yield-stress matrix material according to first spatial portions of a meat construct print design, the first spatial portions of the meat construct print design being associated with one or more muscle portions of the thick structured meat construct, one or more portions of the adipocyte progenitor cell-microgel composite ink into the cellular yield-stress matrix material according to second spatial portions of the meat construct print design, the second spatial portions of the meat construct print design being associated with one or more lipid portions of the thick structured meat construct, and one or more portions of a sacrificial ink into the cellular yield-stress matrix material according to third spatial portions of the meat construct print22LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 design, the third spatial portions of the meat construct print design being associated with a perfusable vascular network within the thick structured meat construct; at least partially solidifying one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cellmicrogel composite ink, or the cellular yield-stress matrix material; and removing the one or more portions of the sacrificial ink printed into the cellular yield-stress matrix material to form the perfusable vascular network within the thick structured meat construct.Brief Description of the Drawings

[0054] Having thus described the invention in general terms, reference will now be made to the accompanying drawings. The skilled artisan will understand that the drawings primarily are for illustrative purposes and are not intended to limit the scope of the inventive subject matter described herein. The drawings are not necessarily to scale; in some instances, various aspects of the inventive subject matter disclosed herein may be shown exaggerated or enlarged in the drawings to facilitate an understanding of different features. In the drawings, like reference characters generally refer to like features (e.g, functionally similar and / or structurally similar elements).

[0055] FIG. 1 provides a block flow diagram of an example system for carrying out embedded printing of thick structured meat constructs, according to embodiments of the present disclosure.

[0056] FIG. 2 provides a block flow diagram of an example processes for carrying out embedded printing of thick structured meat constructs, according to embodiments of the present disclosure.

[0057] FIG. 3 provides a process flow diagram for an example process for embedded printing of thick structured meat constructs, according to embodiments of the present disclosure.

[0058] FIG. 4 provides a schematic of an example computing device configured to perform some or all aspects of embedded printing of thick structured meat constructs, according to embodiments of the present disclosure.

[0059] FIG. 5 provides a schematic of an example external computing device configured to perform some or all aspects of embedded printing of thick structured meat constructs, according to embodiments of the present disclosure.

[0060] FIG. 6 illustrates a process flow diagram of a method for forming cellular inks and a cellular yield-stress support matrix material for embedded printing of thick structured meat constructs, according to an embodiment of the present disclosure.23LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743

[0061] FIG. 7 illustrates a process flow diagram of a method for earn ing out embedded printing of thick structured meat constructs, according to an embodiment of the present disclosure.

[0062] FIG. 8 illustrates a process flow diagram of a method for carrying out embedded printing of thick structured meat constructs, according to an embodiment of the present disclosure.

[0063] FIG. 9 illustrates a process flow diagram of a method for carrying out embedded printing of thick structured meat constructs, according to an embodiment of the present disclosure.

[0064] FIG. 10 illustrates a process flow diagram of a method for carrying out embedded printing of thick structured meat constructs, according to an embodiment of the present disclosure.

[0065] FIG. 11 illustrates a process flow diagram of a method for carrying out embedded printing of thick structured meat constructs, according to an embodiment of the present disclosure.Detailed Description

[0066] The present disclosure more fully describes various embodiments with reference to the accompanying drawings. It should be understood that some, but not all embodiments are shown and described herein. Indeed, the embodiments may take many different forms, and accordingly this disclosure should not be construed as limited to the embodiments set forth herein. Rather, these embodiments are provided so that this disclosure will satisfy applicable legal requirements. Like numbers refer to like elements throughout.

[0067] Various embodiments of the present disclosure will now be described more fully hereinafter with reference to the accompanying drawings, in which some, but not all embodiments of the inventions are shown. Indeed, these inventions may be embodied in many different forms and should not be construed as limited to the embodiments set forth herein; rather, these embodiments are provided so that this disclosure will satisfy applicable legal requirements. The term “or” is used herein in both the alternative and conjunctive sense, unless otherwise indicated. The terms “illustrative” and “exemplary” are used to be examples with no indication of quality level. Like numbers refer to like elements throughout.

[0068] As used herein, the terms “about,” “substantially,” and “approximately” generally mean plus or minus 10% of the value stated, e.g, about 250 pm would include 225 pm to 275 pm, about 1,000 pm would include 900 pm to 1,100 pm. Any provided value, whether or24LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 not it is modified by terms such as “about,"’ “substantially,” or “approximately ,” all refer to and hereby disclose associated values or ranges of values thereabout, as described above.

[0069] Continued global population growth has imposed a significant challenge for traditional meat production as demand for animal products grows. To meet the dramatically increased human needs for food and nutrition, cultured meat, produced using a bioreactor, has emerged as a promising solution. Cultured meat (CM), also known as cultivated meat, is a promising alternative to animal meat products by avoiding animal meat-associated environmental, health, and ethical challenges.

[0070] Described herein are systems, methods, apparatuses, and computer program products for scalable three-dimensional (3D) printing and fabrication of 3D thick structured meat constructs with textures similar to those of traditional meat using embedded printing in a yield-stress cellular matrix bath. The meat texture or structure is made possible by using embedded printing, and the meat volumetric size or thickness is enabled by incorporating a perfusable network in the meat construct to support the growth and maturation of the whole construct. According to some embodiments, both myoblasts and adipocyte progenitor cells are individually printed into specific layouts in a yield-stress cellular matrix bath to mimic the meat structure, and the cellular matrix bath is a bulky microgel-based matrix with endothelial cells and other support cells such as the adipose-derived stem cells, fibroblast cells, and / or stromal vascular fraction cells. In addition, main perfusable channels can be created in the bulky meat construct using a sacrificial ink-assisted embedded printing process, which support the coculture of endothelial cells and support cells in the matrix to self-assemble into capillary structures. Made of the main perfusable channels and capillary’ structures, a perfusable network is formed in the meat construct, serving as a vessel system.

[0071] Among different biofabrication approaches for cultured meat construction, the leading technologies generally involve creating acellular scaffolds then seeding with cells and allowing cellular assembly. Such resulting constructs are partially or completely fabricated using external force-assisted assembly to form the structure. Recently, the development of cultured meat has been promoted by advances in biofabrication, in particular, 3D printing in addition to cell cultivation, stem cell biology, biological process engineering, nutritional science, etc. 3D printing, also known as additive manufacturing AM, builds objects layer by layer, forming 3D structures from different materials including living cells. Compared to traditional manufacturing approaches, 3D printing enables mass customization, more freedom of designs to achieve complex structures, and reduced waste. By adopting 3D printing for25LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 cultured meat production, structured scaffolds and constructs can be printed, supporting better cell growth and maturation.

[0072] Generally, 3D meat printing can be classified into two approaches: printed acellular scaffolds with cell seeding and cellular construct printing. During the approach of acellular scaffolds with cell seeding, a scaffold suitable for cell growth is first printed, and the shape of the scaffold is the shape of the final cultured meat. The acellular inks for printing scaffolds may contain the necessary materials for good printabihty. structural stability, cytocompatibility, and non-cytotoxicity. Once a scaffold is printed, cells are seeded onto it and proliferate, maturate, and eventually form a meat construct together with the scaffold. During the approach of cellular structure printing, a cell-laden ink is printed directly into the final shape of the meat construct. Muscle, adipose, and vascular tissues are printed respectively using different kinds of bioink. It is noted that some meat constructs may be assembled from printed patterns such as cellular fibers.

[0073] Unfortunately, the extra assembly process may make this method inefficient, less robust, and difficult to scale up. While 3D printing has made a great contribution to making cultured meat more structured, challenges still exist in making cultured meat thicker. To create full-thickness cultured meat, a thickness of 1 cm or more is preferred — this scale is far beyond the diffusion limit of oxygen and nutrients, which is generally less than 200 pm. To maintain full thickness printed meat constructs, a perfusion system that allows stable and sufficient delivery of oxygen and nutrients and adequate effusion of metabolic waste is required. Unfortunately, there is currently no scalable 3D printing method for producing cultured meat that can be structured as well as thick.

[0074] Herein, we describe a scalable 3D printing method for fabricating thick structured meat constructs by utilizing embedded extrusion printing, which prints three different composite inks (myoblast-laden microgel composite ink, adipocyte progenitor cell-laden microgel composite ink, and sacrificial ink) in a yield-stress cellular matrix bath. This method can make the printed meat construct structured by the embedded printing of muscle and fat tissues as well as thick by forming a perfusable network via embedded sacrificial printing and cell self-assembly.

[0075] Briefly, two different cellular microgel composite inks and one sacrificial ink are first printed into a designed pattern in a yield-stress cellular matrix bath, which is made from endothelial cells and other support cells such as the adipose-derived stem cells, fibroblast cells, and / or stromal vascular fraction cells. The two cellular microgel composite inks are myoblastladen microgel composite ink and adipocyte progenitor cell-laden microgel composite ink. The26LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 function of the myoblast and adipocyte progenitor cell microgel composite inks is to construct muscle and fat structures in the final meat product, respectively. Then the sacrificial ink is removed after all the printing processes are completed, forming main perfusable channels in the meat construct that can be connected with an external perfusion system for dynamic cell culturing. It is worth noting that the printing order of these three inks can be determined based on the design of the meat construct with a ratio of muscle tissue, fat tissue, and perfusable vessels in a specific construct. Finally, the printed meat construct is connected to a perfusion system and cultured under dynamic medium perfusion and necessary tensioning conditions to mature the myoblasts and adipocytes while endothelial cells self-assemble into capillary structures around the main perfusable channel. The resulting capillary structures together with the main perfusable channels support the growth and maturation of bulky meat constructs for meat conversion.

[0076] Systems, apparatuses, methods, and computer program products are described herein for embedded printing of thick structured meat constructs using cellular inks and a cellular yield-stress support matrix material. An initial construct of the cellular inks in the build material can be achieved by choosing a particular dispensing nozzle that exerts suitable shear forces on the build material, causing the development of mature myofibers. Once the build material is dispensed in the yield-stress support bath, an intermediate article is formed. A crosslinking agent, such as transglutaminase or ultraviolet radiation, can be dispensed in the cellular inks or after the intermediate article is formed, causing subsequent crosslinking in initial meat construct. The yield-stress support bath can be rotated relative to the printing nozzle such that any number of directions can be achieved for the subsequent alignment of the cellular inks to create the meat construct.

[0077] According to some embodiments of the present disclosure, two different cellular microgel composite inks and one sacrificial ink can be printed into a designed pattern in a yieldstress cellular matrix bath, made from endothelial cells and other support cells such as the adipose-derived stem cells, fibroblast cells, and / or stromal vascular fraction cells. The two cellular microgel composite inks are myoblast-laden microgel composite ink and adipocyte progenitor cell-laden microgel composite ink. The function of the myoblast and adipocyte progenitor cell microgel composite inks is to construct muscle and fat structures in the final meat product, respectively. Then the sacrificial ink is removed after all the printing processes are completed, forming main perfusable channels in the meat construct that can be connected with an external perfusion system for dynamic cell culturing.27LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743

[0078] According to some embodiments, the printing order of these three inks can be determined based on the design of the meat construct with a ratio of muscle tissue, fat tissue, and perfusable vessels in a specific construct. The printed meat construct is connected to a perfusion system and cultured under dynamic medium perfusion and necessary tensioning conditions to mature the myoblasts and adipocytes while endothelial cells self-assemble into capillary structures around the main perfusable channel. The resulting capillary structures together with the main perfusable channels support the growth and maturation of thick structured meat constructs. It is important to note that it is only possible to print a thick meat construct that is cellularly viable if the meat construct comprises a perfusable vascular network formed throughout the meat construct, otherwise the thick muscle tissue is not cellularly viable. In addition to that, the relevancy of structured is also important to the present disclosure. It does not just need to be a “complex” tissue / meat construct, meaning that the construct includes muscle tissue, fatty tissue, and a vascular network. Instead, the meat construct needs to be physiologically relevant, or “structured” as a physiologically relevant / accurate facsimile of the human tissue(s) being modeled.

[0079] Embodiments of the present invention are described below with reference to block diagrams and flowchart illustrations. Thus, it should be understood that each block of the block diagrams and flowchart illustrations may be implemented in the form of a computer program product, an entirely hardware embodiment, a combination of hardware and computer program products, and / or apparatus, systems, computing devices, computing entities, and / or the like carrying out instructions, operations, steps, and similar words used interchangeably (e.g., the executable instructions, instructions for execution, program code, and / or the like) on a computer-readable storage medium for execution. For example, retrieval, loading, and execution of code may be performed sequentially such that one instruction is retrieved, loaded, and executed at a time. In some exemplary embodiments, retneval, loading, and / or execution may be performed in parallel such that multiple instructions are retrieved, loaded, and / or executed together. Thus, such embodiments can produce specifically configured machines performing the steps or operations specified in the block diagrams and flowchart illustrations. Accordingly, the block diagrams and flowchart illustrations support various combinations of embodiments for performing the specified instructions, operations, or steps.Exemplary Systems, Apparatuses, and Approaches

[0080] FIG. 1 provides, according to one or more embodiments of the present disclosure, an exemplary apparatus 100 for carrying out embedded printing of thick structured meat constructs. The apparatus 100 comprises a cellular yield-stress support bath 101 comprising a28LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 yield-stress support matrix material. The apparatus 100 comprises build material reservoirs, including the myoblast-microgel composite ink reservoir 104, the adipocyte progenitor cellmicrogel composite ink reservoir 105, and the sacrificial ink reservoir 106, configured to store a supply of a build material, which is also referred to herein as a printing material, a composite ink, a cross-linkable material, or the like. The build material reservoir 101 can also be configured to communicate volume(s) of the build material to or towards a printing environment. The apparatus 100 can further comprise a printing nozzle 102 in fluidic communication with the cellular yield-stress support bath 101. The printing nozzle 102 can be configured to be moved in three dimensions (in the x, y, and z directions) within the cellular yield-stress support bath 101 and to dispose discrete volumes or continuous flows of one or more build materials / inks to particular locations within the cellular yield-stress support bath 101. The cellular yield-stress matrix material is retained within a yield-stress support bath.

[0081] In some embodiments, the printing nozzle 102 can be configured to exert shear forces on the cellular yield-stress support bath 101 and / or the build material / ink during printing. The shear forces can cause the non-covalent and reversible bonds of the yield-stress support material to be interrupted and then reestablish after the printing nozzle 102 has moved in one of three dimensions to a different part of the medium. In some embodiments, the printing environment can be or comprise a yield-stress support bath 101 comprising a yield-stress support material. Said otherwise, the printing nozzle 102 can be configured to dispose volumes or a flow of the build material into the yield-stress support bath 101. The yield-stress support material in the yield-stress support bath 101 can be configured to support (e.g, against deformation) the portions or volumes of build material that are disposed discretely or continuously into the yield-stress support material. Said otherwise, the yield-stress support material can be configured such that the build material can be disposed within a volume of the yield-stress support bath 101 during a first time to achieve a particular size and form factor, and the yield-stress support material can be configured to maintain the size and the form factor of the build material during a second time subsequent to the first time without the need for curing, cross-linking, binding, gelling, solidifying, or otherwise changing the chemical or physical state of the portion of build material being supported in the yield-stress support material.

[0082] More details about yield-stress support materials and freeform additive manufacturing techniques are provided in U.S. Patent No. 10,974.441, U.S. Patent No. 11,426.945, U.S. Patent No. 11.731,343, U.S. Patent No. 11,413,808. U.S. Patent No. 11,724,460, U.S. Patent No. 11,207,841, U.S. Patent No. 11,759,999, U.S. Patent No.29LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 63674311,724.440, U.S. Patent No. 11,745,412, U.S. Patent Publication No. 2023-0226772, U.S. Patent Publication No. 2021-0236386. U.S. Patent Publication No. 2021-0236697, U.S. Patent Publication No. 2020-0276761, U.S. Patent Publication No. 2023-0256668, U.S. Patent Application No. 18 / 211,827, U.S. Patent Application No. 18 / 223,844, and International Patent Application No. PCT / US2023 / 067838, the entire disclosures of each of which are hereby incorporated herein by reference in their entireties for all purposes.

[0083] Once the build material is communicated into the yield-stress support bath 101, and since the yield-stress support material can maintain the size and form factor of the volume(s) of build material being disposed within the yield-stress support material, an intermediate article can be formed in the yield-stress support bath 101 without curing the curable liquid matrix of the build material. This can be helpful if a further / subsequent alignment or partial alignment of the 2D material (platelets) is desired before curing the curable liquid matrix of the build material in the intermediate article to form a finished article.

[0084] The apparatus 100 can further comprise a rotating element 103 that is operably- coupled to the yield-stress support bath 101. In some embodiments, the rotating element 103 can be disposed beneath or about the yield-stress support bath 101 and configured to rotate the entire yield-stress support bath 101 about a center point of the yield-stress support bath 101. In some embodiments, the rotating element 103 can be disposed beneath or about the yieldstress support bath 101 and configured to rotate the entire yield-stress support bath 101 relative to the printing nozzle 102.

[0085] In some embodiments, the apparatus 100 can further comprise a myoblast-microgel composite ink reservoir 104 as a component of the build material. The myoblast-microgel composite ink comprises at least a porous gelatin microgel which includes differentiation and culturing media. In some embodiments, the apparatus 100 can further comprise an adipocyte progenitor cell-microgel composite ink reservoir 105 as a component of the build material. The adipocyte progenitor cell-microgel comprises at least porous gelatin microgels which includes differentiation and culturing media.

[0086] In some embodiments, the apparatus 100 can further comprise a sacrificial ink 106. The sacrificial ink comprises one or more water-soluble polymers. The adipocyte progenitor cell-microgel may comprise of solid gelatin microgels. In some embodiments, one or more portions of the myoblast-microgel composite ink 104 are iteratively printed through the printing nozzle 102 into the cellular yield-stress matrix material 101 on the rotating element 103 according to first spatial portions of a meat construct print design. The first spatial portions of the meat construct print design being associated with one or more muscle portions30LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 of the thick structured meat construct. One or more portions of the adipocyte progenitor cellmicrogel composite ink 105 are iteratively printed through the printing nozzle 102 into the cellular yield-stress matrix material 101 on the rotating element 103 according to second spatial portions of the meat construct print design. The second spatial portions of the meat construct print design being associated with one or more lipid portions of the thick structured meat construct. One or more portions of a sacrificial ink 106 are iteratively printed through the printing nozzle 102 into the cellular yield-stress matrix material 101 on the rotating element 103 according to third spatial portions of the meat construct print design. The third spatial portions of the meat construct print design being associated with a perfusable vascular network within the thick structured meat construct; at least partially solidifying one or more of: the myoblast-microgel composite ink 104, the adipocyte progenitor cell-microgel composite ink 105, or the cellular yield-stress matrix material 101; and removing the one or more portions of the sacrificial ink 106 printed into the cellular yield-stress matrix material to form the perfusable vascular network within the thick structured meat construct.

[0087] Referring now to FIG. 2. a block flow diagram is provided that illustrates a process 200 for embedded printing of thick structured meat constructs. The process 200 can comprise three-dimensional (3D) printing using two or more cellular inks. For example, the process 200 can comprise printing a myoblast-microgel composite ink (e.g, from the myoblast-microgel composite ink reservoir 104) and adipocyte progenitor cell-microgel composite ink (e.g., from the adipocyte progenitor cell-microgel composite ink reservoir 105), and a sacrificial ink (e.g, from the sacrificial ink reservoir 106) into a yield-stress support bath (e.g., 101). The process 200 can comprise mixing myoblasts and a hydrogel precursor with a porous microgel to form the myoblast-microgel composite ink. The process 200 can comprise mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink. The process 200 can comprise mixing adipose-derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel to form a cellular yield-stress matrix material. The support cells in the cellular yield-stress matrix material comprises one or more from among: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. The support cells facilitate the endothelial cells in the cellular yield-stress matrix material to form one or more capillary structures for microcirculation across the muscle portions of the thick structured meat construct. The process 200 can further comprise disposing a cross-linking agent into one or more of the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, or the cellular yield-stress matrix material.31LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743

[0088] Further, as illustrated in FIG. 2, the process 200 can comprise a step of iteratively printing the myoblast-microgel composite ink iteratively through a printing nozzle (e.g.. 102) into the cellular yield-stress matrix material. The cellular yield-stress matrix material can be disposed within a yield-stress support bath, which may be mounted on a rotating element (e.g. , 103). Printing of the myoblast-microgel composite ink into the cellular yield-stress matrix material can be carried out according to one or more first spatial portions of a meat construct print design. The first spatial portions of the meat construct print design can be associated with one or more muscle portions of a thick structured meat construct being printed using the process 200.

[0089] Additionally or alternatively, the process 200 can comprise iteratively printing one or more portions of the adipocyte progenitor cell-microgel composite ink through the printing nozzle into the cellular yield-stress matrix material can be carried out according to one or more second spatial portions of the meat construct print design. The one or more second spatial portions of the meat construct print design can be associated with one or more lipid portions of the thick structured meat construct.

[0090] Additionally or alternatively, the process 200 can comprise iteratively printing one or more portions of a sacrificial ink through the printing nozzle into the cellular yield-stress matrix material. Printing of the one or more portions of the sacrificial ink into the cellular yield-stress matrix material can be carried out according to one or more third spatial portions of the meat construct print design. The one or more third spatial portions of the meat construct print design can be associated with a perfusable vascular network within the thick structured meat construct being printed using the process 200.

[0091] In some embodiments, at least some portions of the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, and / or the sacrificial ink can be iteratively and adjacently printed into the cellular yield-stress matrix material from a first side of the yield-stress support bath to a second side of the yield-stress support bath based on the meat construct print design associated with the thick structured meat construct.

[0092] In some embodiments, the process 200 can further comprise at least partially solidifying one or more of the myoblast-microgel composite ink, the adipocyte progenitor cell- microgel composite ink, or the cellular yield-stress matrix material.

[0093] In some embodiments, the process 200 can comprise defining one or more portions of the sacrificial ink 106 printed into the cellular yield-stress matrix material being removed to form the perfusable vascular network within the thick structured meat construct.32LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743

[0094] Furthermore, FIG. 2 illustrates a third step of the block flow diagram of an example processes 200 for carrying out embedded printing of thick structured meat constructs. The third step defines the perfusion for endothelial cell self-assembly and post-printing tensioning for myofiber formation to enable the formation of the thick structured meat construct. According to other embodiments, the process 200 for printing (e.g., embedded bioprinting) a thick structured meat construct illustrated in FIG. 2 involves extrusion printing of two cellular inks and a sacrificial ink in a yield-stress cellular matrix bath.

[0095] Referring now to FIG. 3, a process flow diagram is provided for a process 300 for embedded printing of thick structured meat constructs. The process 300 can be carried out in order to print thick structured meat construct that comprise multiple layers of myoblasts, multiple layers of adipocytes, and multiple sacrificial ink filaments. In some embodiments, a spatial arrangement of the multiple layers of myoblasts, adipocytes, and sacrificial ink filaments is controlled during the process 300 by printing the thick structured meat construct based upon a meat construct print design associated with the thick structured meat construct.

[0096] One or more portions of the myoblast-microgel composite ink can be iteratively printed through the printing nozzle into the cellular yield-stress matrix material on the rotating element according to first spatial portions of a meat construct print design. The first spatial portions of the meat construct print design being associated with one or more muscle portions of the thick structured meat construct. The process 300 of printing of the myoblast-microgel composite ink into the cellular yield-stress support material to form the one or more muscle portions of the thick structured meat construct is carried out using one or more extrusion printing nozzles. According to some embodiments, the process 300 for printing can be carried out using one or more extrusion printing nozzles that are dimensioned and configured to exert extrusion printing-induced shear forces on the myoblast-microgel composite ink, thereby facilitating mature myofiber formation therefrom.

[0097] One or more portions of the adipocyte progenitor cell-microgel composite ink are iteratively printed through the printing nozzle into the cellular yield-stress matrix material on the rotating element according to second spatial portions of the meat construct print design. The second spatial portions of the meat construct print design being associated with one or more lipid portions of the thick structured meat construct. One or more portions of a sacrificial ink are iteratively printed through the printing nozzle into the cellular yield-stress matrix material on a rotating element according to third spatial portions of the meat construct print design. The third spatial portions of the meat construct print design being associated with a33LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 perfusable vascular network within the thick structured meat construct. The three iteratively printed inks compose a printed meat construct with sacrificial patterns.

[0098] According to some embodiments, the process can comprise printing the muscle, fat, and vascular tissues in a cellular yield-stress matrix bath using two cellular inks and one sacrificial ink, as illustrated in FIG. 3. The two cellular microgel composite inks are myoblastladen microgel composite ink and adipocyte progenitor cell-laden microgel composite ink. Generally, both the cellular inks and matrix bath consist of a discrete microgel phase (either porous or solid microgel) and a continuous hydrogel precursor phase (hydrogel precursor-based cellular suspension). Such microgel-based materials have yield-stress rheological properties providing good printability for embedded extrusion printing. The hydrogel precursor that makes up the continuous phase can be made of cross-linkable hydrogels, such as gelatin. The cellular inks, including myoblasts-laden and adipocyte progenitor cells-laden inks, are prepared by mixing living cells with porous or solid microgels and a liquid hydrogel precursor. The yield-stress cellular matrix bath consists of porous or solid microgels, a liquid hydrogel precursor, endothelial cells, and support cells such as adipose-derived stem cells, fibroblasts, and / or stromal vascular fraction cells, which facilitate endothelial cells to form capillary structures for microcirculation across the whole volume of the meat construct being printed during the post-printing perfusion step. Right before printing, a cross-linking agent such as transglutaminase (TG) may be added to the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, and the cellular yield-stress matrix bath for in-situ cross-linking. Alternatively, the meat construct may be cross-linked using a different mechanism such as ultraviolet radiation after the printing process is completed. The sacrificial ink should be easily removable after printing and is usually made of a water-soluble polymer and rheology modifiers.

[0099] Printed myoblast filaments are cultured to form a muscle tissue layer, where extrusion printing-induced shear force and / or post-printing tensioning are crucial for the development of mature myofibers. A meat construct may consist of multiple layers of myoblasts, multiple layers of adipocytes, and several sacrificial ink filaments, and their spatial arrangement should be controlled per the texture and perfusable network as designed.

[0100] The second step is to have sacrificial patterns removed after the printing process is completed and the meat construct is sufficiently cross-linked. Depending on the type of sacrificial materials used, some sacrificial patterns may be simply removed by immersing the printed meat construct in phosphate-buffered saline until the sacrificial material becomes diluted and much less viscous and then followed by flushing gently using a syringe. Once the34LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 sacrificial material is completely removed, main perfusable channels are formed for postprinting perfusion.

[0101] The third step is to connect the printed meat construct to a perfusion system and post-printing tensioning setup for cell growth and maturation as well as capillary structure formation by perfusing cell media through the created main perfusable channels. The main perfusable channels and capillary structures form a perfusable network in the meat construct, enabling it to be thick.

[0102] The meat texture or structure is made possible by using embedded printing, and the meat volumetric size or thickness is enabled by incorporating a perfusable network in the meat construct to support the growth and maturation of the whole construct. The described thick structured meat printing technology may be applied to print various meat types such as fatless meat constructs and meat constructs with controllable muscle-to-fat ratios or customized nutrition as well as other non-meat printing-related tissue constructs such as:

[0103] Thick muscular tissues such as heart tissues to model muscular diseases or test related pharmaceuticals;

[0104] Muscular tissues for repairing or replacing damaged muscles in patients with injuries or degenerative diseases; and

[0105] Fat transplants, that can be used for cosmetic and reconstructive surgeries such as for breast reconstruction or facial contouring.Example Computer-Comprising Entities and Devices

[0106] According to some embodiments, some or all elements, aspects, or steps of the approaches and methods described herein can be carried out by one or more apparatuses, such as computing devices. An apparatus can comprise one or more processors and one or more memory devices having instructions, such as computer-executable program codes or the like, stored thereon that, when executed by the one or more processors, cause the apparatus to perform the one or more elements, aspects, or steps of the approaches and methods described herein.

[0107] As used herein, the terms “instructions.” “file,” “designs,” “data,” “content,” “information,” and similar terms may be used interchangeably, according to some example embodiments of the present invention, to refer to data capable of being transmitted, received, operated on, displayed, and / or stored. Thus, use of any such terms should not be taken to limit the spirit and scope of the disclosure. Further, where a computing device is described herein to receive data from another computing device, it will be appreciated that the data may be received directly from the other computing device or may be received indirectly via one or more35LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 computing devices, such as, for example, one or more servers, relays, routers, network access points, base stations, and / or the like.

[0108] As used herein, the term “computer-readable medium’’ refers to any medium configured to participate in providing information to a processor, including instructions for execution. Such a medium may take many forms, including, but not limited to a non-transitoiy computer-readable storage medium (for example, non-volatile media, volatile media), and transmission media. Transmission media include, for example, coaxial cables, copper wire, fiber optic cables, and carrier waves that travel through space without wires or cables, such as acoustic waves and electromagnetic waves, including radio, optical, and infrared waves. Signals include man-made transient variations in amplitude, frequency, phase, polarization, or other physical properties transmitted through the transmission media. Examples of non- transitory computer-readable media include a floppy disk, a flexible disk, hard disk, magnetic tape, any other non-transitory magnetic medium, a compact disc read only memory (CD- ROM), compact disc compact disc-rewritable (CD-RW), digital versatile disc (DVD), Blu- Ray, any other non-transitory optical medium, punch cards, paper tape, optical mark sheets, any other physical medium with patterns of holes or other optically recognizable indicia, a random access memory (RAM), a programmable read only memory (PROM), an erasable programmable read only memory (EPROM), a FLASH-EPROM, any other memory chip or cartridge, a carrier wave, or any other non-transitory medium from which a computer can read. The term computer-readable storage medium is used herein to refer to any computer-readable medium except transmission media. However, it will be appreciated that where embodiments are described to use a computer-readable storage medium, other types of computer-readable mediums may be substituted for or used in addition to the computer-readable storage medium in alternative embodiments. By way of example only, a design file for a printed article may be stored on a computer-readable medium and may be read by a computing device, such as described hereinbelow , for controlling part or all of a three-dimensional (3D) printing process and associated apparatuses and components, according to various embodiments described herein.

[0109] As used herein, the term “circuitry” refers to all of the following: (a) hardware-only circuit implementations (such as implementations in only analog and / or digital circuitry); (b) to combinations of circuits and computer program product(s) comprising software (and / or firmware instructions stored on one or more computer readable memories), such as (as applicable): (i) to a combination of processor(s) or (ii) to portions of processor(s) / software (including digital signal processor(s)), software, and memory(ies) that work together to cause36LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 an apparatus, such as a mobile phone or server, to perform various functions described herein); and (c) to circuits, such as, for example, a microprocessor(s) or a portion of a microprocessor(s), that require software or firmware for operation, even if the software or firmware is not physically present. This definition of ‘’circuitry” applies to all uses of this term in this application, including in any claims. As a further example, as used in this application, the term “circuitry” would also cover an implementation of merely a processor (or multiple processors) or portion of a processor and its (or their) accompanying software and / or firmware. The term “circuitry” would also cover, for example and if applicable to the particular claim element, a baseband integrated circuit or applications processor integrated circuit for a mobile phone or a similar integrated circuit in a server, a cellular network device, other network device, and / or other computing device.

[0110] As used herein, the term “computing device” refers to a specialized, centralized device, network, or system, comprising at least a processor and a memory device including computer program code, and configured to provide guidance or direction related to the charge transactions carried out in one or more charging networks.

[0111] Some or all of the elements, steps, or components of the approaches described herein can be carried out by a computing device or an apparatus comprising a processor and memory. Examples of such computing devices and apparatuses are described in more detail below.

[0112] Embodiments of the present invention may be implemented in various ways, including as computer program products that comprise articles of manufacture. Such computer program products may include one or more software components including, for example, software objects, methods, data structures, or the like. A software component may be coded in any of a variety of programming languages. An illustrative programming language may be a lower-level programming language such as an assembly language associated with a particular hardware architecture and / or operating system platform. A software component comprising assembly language instructions may require conversion into executable machine code by an assembler prior to execution by the hardware architecture and / or platform. Another example programming language may be a higher-level programming language that may be portable across multiple architectures. A software component comprising higher-level programming language instructions may require conversion to an intermediate representation by an interpreter or a compiler prior to execution.

[0113] Other examples of programming languages include, but are not limited to, a macro language, a shell or command language, a job control language, a script language, a database37LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 query or search language, and / or a report writing language. In one or more example embodiments, a software component comprising instructions in one of the foregoing examples of programming languages may be executed directly by an operating system or other software component without having to be first transformed into another form. A software component may be stored as a file or other data storage construct. Software components of a similar ty pe or functionally related may be stored together such as, for example, in a particular directory, folder, or library. Software components may be static (e.g., pre-established or fixed) or dynamic (e.g., created or modified at the time of execution).

[0114] A computer program product may include a non- transitory computer-readable storage medium storing applications, programs, program modules, scripts, source code, program code, object code, byte code, compiled code, interpreted code, machine code, executable instructions, and / or the like (also referred to herein as executable instructions, instructions for execution, computer program products, program code, and / or similar terms used herein interchangeably). Such non-transitoiy computer-readable storage media include all computer-readable media (including volatile and non-volatile media).

[0115] In one embodiment, a non-volatile computer-readable storage medium may include a floppy disk, flexible disk, hard disk, solid-state storage (SSS) (e.g., a solid-state drive (SSD), solid state card (SSC), solid state module (SSM), enterprise flash drive, magnetic tape, or any other non-transitory magnetic medium, and / or the like. A non-volatile computer-readable storage medium may also include a punch card, paper tape, optical mark sheet (or any other physical medium with patterns of holes or other optically recognizable indicia), compact disc read only memory' (CD-ROM), compact disc-rewritable (CD-RW), digital versatile disc (DVD), Blu-ray disc (BD), any other non-transitory optical medium, and / or the like. Such anon-volatile computer-readable storage medium may also include read-only memory (ROM), programmable read-only memory (PROM), erasable programmable read-only memory (EPROM), electrically erasable programmable read-only memory7(EEPROM), flash memory7(e.g., Serial, NAND, NOR, and / or the like), multimedia memory7cards (MMC), secure digital (SD) memory cards, SmartMedia cards, CompactFlash (CF) cards, Memory Sticks, and / or the like. Further, a non-volatile computer-readable storage medium may also include conductive-bridging random access memory (CBRAM), phase-change random access memory7(PRAM), ferroelectric random-access memory7(FeRAM), non-volatile randomaccess memory (NVRAM), magnetoresistive random-access memory (MRAM), resistive random-access memory (RRAM), Silicon-Oxide-Nitride-Oxide-Silicon memory (SONOS),38LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 floating junction gate random access memory (FJG RAM), Millipede memory, racetrack memory, and / or the like.

[0116] In one embodiment, a volatile computer-readable storage medium may include random access memory (RAM), dynamic random access memory (DRAM), static random access memory (SRAM), fast page mode dynamic random access memory (FPM DRAM), extended data-out dynamic random access memory (EDO DRAM), synchronous dynamic random access memory (SDRAM), double data rate synchronous dynamic random access memory (DDR SDRAM), double data rate type two synchronous dynamic random access memory (DDR2 SDRAM), double data rate type three synchronous dynamic random access memory (DDR3 SDRAM), Rambus dynamic random access memory (RDRAM), Twin Transistor RAM (TTRAM), Thyristor RAM (T-RAM), Zero-capacitor (Z-RAM), Rambus inline memory module (RIMM), dual in-line memory module (DIMM), single in-line memory module (SIMM), video random access memory' (VRAM), cache memory (including various levels), flash memory, register memory', and / or the like. It will be appreciated that where embodiments are described to use a computer-readable storage medium, other types of computer-readable storage media may be substituted for or used in addition to the computer- readable storage media described above.

[0117] As should be appreciated, various embodiments of the present invention may also be implemented as methods, apparatus, systems, computing devices, computing entities, and / or the like. As such, embodiments of the present invention may take the form of an apparatus, system, computing device, computing entity', and / or the like executing instructions stored on a computer-readable storage medium to perform certain steps or operations. Thus, embodiments of the present invention may also take the form of an entirely hardware embodiment, an entirely computer program product embodiment, and / or an embodiment that comprises combination of computer program products and hardware performing certain steps or operations.Example Computing Device

[0118] FIG. 4 provides a schematic of such a computing device 400 according to one embodiment of the present invention. In general, the terms computing device, computing entity, computer, entity, device, system, and / or similar words used herein interchangeably may refer to, for example, one or more computers, computing entities, desktops, mobile phones, tablets, phablets, notebooks, laptops, distributed systems, kiosks, input terminals, servers or server networks, blades, gateways, switches, processing devices, processing entities, set-top boxes, relays, routers, network access points, base stations, the like, and / or any combination of devices or entities adapted to perform the functions, operations, and / or processes described39LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 herein. Such functions, operations, and / or processes may include, for example, transmitting, receiving, operating on, processing, displaying, storing, determining, creating / generating, monitoring, evaluating, comparing, and / or similar terms used herein interchangeably. In one embodiment, these functions, operations, and / or processes can be performed on data, content, information, and / or similar terms used herein interchangeably .

[0119] As shown in FIG. 4, in one embodiment, the computing device 400 may include or be in communication with one or more processing elements 402 (also referred to as processors, processing circuitry, and / or similar terms used herein interchangeably) that communicate with other elements within the computing device 400 via a bus, for example. As will be understood, the processing element 402 may be embodied in a number of different ways. For example, the processing element 402 may be embodied as one or more complex programmable logic devices (CPLDs), microprocessors, multi-core processors, coprocessing entities, applicationspecific instructi on-set processors (ASIPs), microcontrollers, and / or controllers. Further, the processing element 402 may be embodied as one or more other processing devices or circuitry. The term circuitry may refer to an entirely hardware embodiment or a combination of hardware and computer program products. Thus, the processing element 402 may be embodied as integrated circuits, application specific integrated circuits (ASICs), field programmable gate arrays (FPGAs), programmable logic arrays (PLAs), hardware accelerators, another circuitry, and / or the like. As will therefore be understood, the processing element 402 may be configured for a particular use or configured to execute instructions stored in volatile or non-volatile media or otherwise accessible to the processing element 402. As such, whether configured by hardware or computer program products, or by a combination thereof, the processing element 402 may be capable of performing steps or operations according to embodiments of the present invention when configured accordingly.

[0120] In one embodiment, the computing device 400 may further include or be in communication with non-volatile media (also referred to as non-volatile storage, memory, memory storage, memory' circuitry' and / or similar terms used herein interchangeably). In one embodiment, the non-volatile storage or memory’ may include one or more non-volatile storage or memory media 403. including but not limited to hard disks. ROM. PROM, EPROM. EEPROM, flash memory, MMCs, SD memory cards, Memory Sticks, CBRAM, PRAM, FeRAM, NVRAM, MRAM, RRAM, SONOS, FJG RAM, Millipede memory , racetrack memory, and / or the like. As will be recognized, the non-volatile storage or memory media may store databases, database instances, database management systems, data, applications, programs, program modules, scripts, source code, object code, byte code, compiled code,40LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 interpreted code, machine code, executable instructions, and / or the like. The term database, database instance, database management system, and / or similar terms used herein interchangeably may refer to a collection of records or data that is stored in a computer-readable storage medium using one or more database models, such as a hierarchical database model, network model, relational model, entity-relationship model, object model, document model, semantic model, graph model, and / or the like.

[0121] In one embodiment, the computing device 400 may further include or be in communication with volatile media (also referred to as volatile storage, memory, memory storage, memory circuitry and / or similar terms used herein interchangeably). In one embodiment, the volatile storage or memory may also include one or more volatile storage or memory media 404, including but not limited to RAM, DRAM, SRAM, FPM DRAM, EDO DRAM, SDRAM, DDR SDRAM, DDR2 SDRAM, DDR3 SDRAM, RDRAM, TTRAM, T- RAM, Z-RAM, RIMM, DIMM, SIMM, VRAM, cache memory, register memoiy. and / or the like. As will be recognized, the volatile storage or memoiy media may be used to store at least portions of the databases, database instances, database management systems, data, applications, programs, program modules, scripts, source code, object code, byte code, compiled code, interpreted code, machine code, executable instructions, and / or the like being executed by, for example, the processing element 402. Thus, the databases, database instances, database management systems, data, applications, programs, program modules, scripts, source code, object code, byte code, compiled code, interpreted code, machine code, executable instructions, and / or the like may be used to control certain aspects of the operation of the computing device 400 with the assistance of the processing element 402 and operating system.

[0122] In some embodiments, the computing device 400 may also include one or more network interfaces, such as a transceiver 408 for communicating with various computing entities, such as by communicating data, content, information, and / or similar terms used herein interchangeably that can be transmitted, received, operated on, processed, displayed, stored, and / or the like. Such communication may be executed using a wired data transmission protocol, such as fiber distributed data interface (FDDI), digital subscriber line (DSL), Ethernet, asynchronous transfer mode (ATM), frame relay, data over cable service interface specification (DOCSIS), or any other wired transmission protocol. Similarly, the computing device 400 may be configured to communicate via wireless external communication networks using any of a variety of protocols, such as general packet radio service (GPRS), Universal Mobile Telecommunications System (UMTS). Code Division Multiple Access 2000 (CDMA2000), CDMA2000 IX (IxRTT), Wideband Code Division Multiple41LEGAI,02 / 46428716v1Atorney Docket No.: 049648 / 636743Access (WCDMA), Global System for Mobile Communications (GSM), Enhanced Data rates for GSM Evolution (EDGE), Time Division-Synchronous Code Division Multiple Access (TD-SCDMA), Long Term Evolution (LTE), Evolved Universal Terrestrial Radio Access Network (E-UTRAN), Evolution-Data Optimized (EVDO), High Speed Packet Access (HSPA), High-Speed Downlink Packet Access (HSDPA), IEEE 802. 11 (Wi-Fi), Wi-Fi Direct, 802.16 (WiMAX), ultra-wideband (UWB), infrared (IR) protocols, near field communication (NFC) protocols, Wibree. Bluetooth protocols, wireless universal serial bus (USB) protocols, and / or any other wireless protocol.

[0123] Although not shown, the computing device 400 may include or be in communication with one or more input elements, such as a keyboard input, a mouse input, a touch screen / display input, motion input, movement input, audio input, pointing device input, joystick input, keypad input, and / or the like. The computing device 400 may also include or be in communication with one or more output elements (not shown), such as audio output, video output, screen / display output, motion output, movement output, and / or the like.Example External Computing Device

[0124] FIG. 5 provides an illustrative schematic representative of an external computing device 500 that can be used in conjunction with embodiments of the present invention. In general, the terms device, system, computing entity, entity7, and / or similar words used herein interchangeably may refer to, for example, one or more computers, computing entities, desktops, mobile phones, tablets, phablets, notebooks, laptops, distributed systems, kiosks, input terminals, servers or server networks, blades, gateways, switches, processing devices, processing entities, set-top boxes, relays, routers, network access points, base stations, the like, and / or any combination of devices or entities adapted to perform the functions, operations, and / or processes described herein. External computing entities 500 can be operated by various parties. As shown in FIG. 5, the external computing device 500 can include an antenna 508, a transmitter 509 (e.g., radio), a receiver 510 (e.g., radio), and a processing element 502 (e.g, CPLDs, microprocessors, mult-core processors, coprocessing entities, ASIPs, microcontrollers, and / or controllers) that provides signals to and receives signals from the transmitter 509 and receiver 510, correspondingly.

[0125] The signals provided to and received from the transmiter 509 and the receiver 510, correspondingly, may include signaling information / data in accordance with air interface standards of applicable wireless systems. In this regard, the external computing device 500 may be capable of operating with one or more air interface standards, communication protocols, modulation types, and access types. More particularly, the external computing device 500 may42LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 operate in accordance with any of a number of wireless communication standards and protocols, such as those described above with regard to the computing device 400. In a particular embodiment, the external computing device 500 may operate in accordance with multiple wireless communication standards and protocols, such as UMTS, CDMA2000, IxRTT, WCDMA, GSM, EDGE, TD-SCDMA, LTE, E-UTRAN, EVDO, HSPA, HSDPA, Wi-Fi, Wi-Fi Direct, WiMAX. UWB, IR, NFC, Bluetooth, USB, and / or the like. Similarly, the external computing device 500 may operate in accordance with multiple wired communication standards and protocols, such as those described above with regard to the computing device 500 via a network interface 506.

[0126] Via these communication standards and protocols, the external computing device 500 can communicate with various other entities using concepts such as Unstructured Supplementary Service Data (USSD), Short Message Service (SMS), Multimedia Messaging Service (MMS), Dual-Tone Multi-Frequency Signaling (DTMF), and / or Subscriber Identity Module Dialer (SIM dialer). The external computing device 500 can also download changes, add-ons, and updates, for instance, to its firmware, software (e.g.. including executable instructions, applications, program modules), and operating system.

[0127] According to one embodiment, the external computing device 500 may include location determining aspects, devices, modules, functionalities, and / or similar words used herein interchangeably. For example, the external computing device 500 may include outdoor positioning aspects, such as a location module adapted to acquire, for example, latitude, longitude, altitude, geocode, course, direction, heading, speed, universal time (UTC), date, and / or various other information / data. In one embodiment, the location module can acquire data, sometimes known as ephemeris data, by identifying the number of satellites in view and the relative positions of those satellites (e.g.. using global positioning systems (GPS)). The satellites may be a variety of different satellites, including Low Earth Orbit (LEO) satellite systems, Department of Defense (DOD) satellite systems, the European Union Galileo positioning systems, the Chinese Compass navigation systems, Indian Regional Navigational satellite systems, and / or the like. This data can be collected using a variety of coordinate systems, such as the Decimal Degrees (DD); Degrees. Minutes. Seconds (DMS); Universal Transverse Mercator (UTM); Universal Polar Stereographic (UPS) coordinate systems; and / or the like. Alternatively, the location information / data can be determined by triangulating a position of the external computing entity 500 in connection with a variety of other systems, including cellular towers, Wi-Fi access points, and / or the like. Similarly, the external computing device 500 may include indoor positioning aspects, such as a location module43LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 adapted to acquire, for example, latitude, longitude, altitude, geocode, course, direction, heading, speed, time, date, and / or various other information / data. Some of the indoor systems may use various position or location technologies including RFID tags, indoor beacons or transmitters, Wi-Fi access points, cellular towers, nearby computing devices (e.g., smartphones, laptops) and / or the like. For instance, such technologies may include the iBeacons, Gimbal proximity beacons, Bluetooth Low Energy (BLE) transmitters, NFC transmitters, and / or the like. These indoor positioning aspects can be used in a variety of settings to determine the location of someone or something to within inches or centimeters.

[0128] The external computing device 500 may also comprise a user interface (that can include a display 507 coupled to the processing element 502) and / or a user input interface (coupled to the processing element 502). For example, the user interface may be a user application, browser, user interface, and / or similar words used herein interchangeably executing on and / or accessible via the external computing device 500 to interact with and / or cause display of information / data from the computing device 500, as described herein. The user input interface can comprise any of a number of devices or interfaces allowing the external computing device 500 to receive data, such as a keypad 511 (hard or soft), a touch display, voice / speech or motion interfaces, or other input device. In embodiments including a keypad 511, the keypad 511 can include (or cause display of) the conventional numeric (0-9) and related keys (#, *), and other keys used for operating the external computing device 500 and may include a full set of alphabetic keys or set of keys that may be activated to provide a full set of alphanumeric keys. In addition to providing input, the user input interface can be used, for example, to activate or deactivate certain functions, such as screen savers and / or sleep modes.

[0129] The external computing device 500 can also include volatile storage or memory 504 and / or non-volatile storage or memory 503, which can be embedded and / or may be removable. For example, the non-volatile memory may be ROM, PROM, EPROM, EEPROM, flash memory, MMCs, SD memory cards, Memory' Sticks, CBRAM, PRAM, FeRAM, NVRAM, MRAM, RRAM, SONOS. FJG RAM, Millipede memory, racetrack memory’, and / or the like. The volatile memory’ may be RAM, DRAM. SRAM, FPM DRAM, EDO DRAM, SDRAM. DDR SDRAM, DDR2 SDRAM, DDR3 SDRAM, RDRAM, TTRAM, T-RAM, Z-RAM, RIMM, DIMM, SIMM, VRAM, cache memory’, register memory’, and / or the like. The volatile and non-volatile storage or memory can store databases, database instances, database management systems, data, applications, programs, program modules, scripts, source code, object code, byte code, compiled code, interpreted code, machine code, executable instructions,44LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 and / or the like to implement the functions of the external computing device 500. As indicated, this may include a user application that is resident on the entity or accessible through a browser or other user interface for communicating with the computing entity 400 and / or various other computing entities.

[0130] In another embodiment, the external computing device 500 may include one or more components or functionality that are the same or similar to those of the computing device 400, as described in greater detail above. As will be recognized, these architectures and descriptions are provided for exemplary purposes only and are not limiting to the various embodiments.

[0131] In some embodiments, the computing device 400 can comprise the external computing device 500. the computing device 400 suitable to carry out movement of the various components of the external computing device 500, flow rates or deposition / dispersal volumes, or the like. In some embodiments, the computing device 400 or a component thereof can be configured to be in communication with the external computing device 500, which can be configured to provide instructions for printing, a design file for a printed article, printing nozzle and / or nebulizer path instructions, or the like to the computing device 400, which is configured to carry out printing.

[0132] Several methods, processes, or approaches for preparing (e.g., embedded bioprinting) of a thick structured meat construct are provided below. Some or all of the steps, sub-steps, elements, or actions of the below methods, processes, or approaches can be carried out by a computer-facilitated or computer-controlled system, device, apparatus, or component. For example, an embedded bioprinting system can comprise the computing device 400, wherein the computing device 400 is configured to initiate, provide instructions related to, control, cany’ out, or otherwise cause performance of some or all of an example methods, process, or approach such as described herein. In another example, an embedded bioprinting system can comprise the external computing device 500, wherein the external computing device 500 is configured to initiate, provide instructions related to, control, cany’ out, or otherwise cause performance of some or all of an example methods, process, or approach such as described herein. In some embodiments, an example embedded printing system or device can comprise the computing device 400, wherein the computing device 400 is configured to carry’ out some or all steps or elements of a method, process, or approach described herein in response to receiving instructions from the external computing device 500. Several example methods are described below, each of which can be at least partially initiated, controlled,45LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 performed, carried out, or caused to be performed by one or more computer-comprising devices, such as the computing device 400 or the external computing device 500.

[0133] Referring now to FIG. 6, a method 600 for preparing cellular inks (bioinks) and a cellular yield stress support matrix material is illustrated. The method 600 can comprise: mixing myoblasts and a hydrogel precursor with a porous microgel to form a myoblastmicrogel composite ink 601. The method 600 can further comprise: mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink 602. The method 600 can further comprise: mixing adipose- derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel to form a cellular yield-stress matrix material 603.

[0134] Some or all of the method 600 can be carried out using a system (e.g., 100), by an apparatus, or by a computing device, such as 400 or 500. For example, an apparatus can comprise at least one processor and at least one memory storing instructions thereon that, when executed by the at least one processor, cause the apparatus to perform some or all of the method 600. Additionally, a computer program product can be provided that comprises a non- transitory computer-readable storage medium storing instructions thereon that, when executed by a processor, cause a machine or apparatus to perform some or all of the method 600.Example Methods, Approaches, and Processes

[0135] Referring now to FIG. 7, a process flow diagram of a method 700 for carrying out embedded printing of thick structured meat constructs. The method 700 can comprise: mixing myoblasts and a hydrogel precursor with a porous microgel to form a myoblast-microgel composite ink 701. The method 700 can further comprise: mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink 702. The method 700 can further comprise: mixing adipose-derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel to form a cellular yield-stress matrix material 703. The method 700 can further comprise: iteratively printing, into the cellular yield-stress matrix material, the myoblast-microgel composite ink to form muscle portions, the adipocyte progenitor cell-microgel composite ink to form lipid portions, and a sacrificial ink to form vascular network portions 704. The support cells in the cellular yield-stress matrix material comprises one or more from among: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells.

[0136] Some or all of the method 700 can be carried out using a system (e.g, 100), by an apparatus, or by a computing device, such as 400 or 500. For example, an apparatus can comprise at least one processor and at least one memory storing instructions thereon that, when46LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 executed by the at least one processor, cause the apparatus to perform some or all of the method 700. Additionally, a computer program product can be provided that comprises a non- transitory computer-readable storage medium storing instructions thereon that, when executed by a processor, cause a machine or apparatus to perform some or all of the method 700.

[0137] Referring now to FIG. 8, a process flow diagram of a method 800 for carry ing out embedded printing of thick structured meat constructs. The method 800 can comprise: mixing myoblasts and a hydrogel precursor with a porous microgel to form a myoblast-microgel composite ink 801. The method 800 can further comprise: mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink 802. The method 800 can further comprise: mixing adipose-derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel to form a cellular yield-stress matrix material 803. The method 800 can further comprise: iteratively printing, into the cellular yield-stress matrix material, the myoblast-microgel composite ink to form muscle portions, the adipocyte progenitor cell-microgel composite ink to form lipid portions, and a sacrificial ink to form non-perfusable vascular network portions 804. The method 800 can further comprise: flushing the sacrificial ink from the cellular yield-stress matrix material to form a perfusable vascular network from the non-perfusable vascular network portions 805.

[0138] Some or all of the method 800 can be carried out using a system (e ., 100), by an apparatus, or by a computing device, such as 400 or 500. For example, an apparatus can comprise at least one processor and at least one memory storing instructions thereon that, when executed by the at least one processor, cause the apparatus to perform some or all of the method 800. Additionally, a computer program product can be provided that comprises a non- transitory computer-readable storage medium storing instructions thereon that, when executed by a processor, cause a machine or apparatus to perform some or all of the method 800.

[0139] Referring now to FIG. 9, a process flow diagram of a method 900 for carry ing out embedded printing of thick structured meat constructs. The method 900 can comprise: printing one or more portions of a myoblast-microgel composite ink into a cellular yield-stress matrix material according to first spatial portions of a meat construct print design, the first spatial portions of the meat construct print design being associated with one or more muscle portions of a thick structured meat construct 901. The method 900 can further comprise: printing one or more portions of the adipocyte progenitor cell microgel composite ink into the cellular yieldstress matrix material according to second spatial portions of the meat construct print design, the second spatial portions of the meat construct print design being associated with one or more47LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 lipid portions of the thick structured meat construct 902. The method 900 can further comprise: printing one or more portions of a sacrificial ink into the cellular yield-stress matrix material according to third spatial portions of the meat construct print design, the third spatial portions of the meat construct print design being associated with a perfusable vascular network within the thick structured meat construct 903. The method 900 can further comprise: and at least partially solidifying one or more of: the myoblast-microgel composite ink. the adipocyte progenitor cell-microgel composite ink. or the cellular yield-stress matrix material 904.

[0140] Some or all of the method 900 can be carried out using a system (e.g., 100), by an apparatus, or by a computing device, such as 400 or 500. For example, an apparatus can comprise at least one processor and at least one memory storing instructions thereon that, when executed by the at least one processor, cause the apparatus to perform some or all of the method 900. Additionally, a computer program product can be provided that comprises a non- transitory computer-readable storage medium storing instructions thereon that, when executed by a processor, cause a machine or apparatus to perform some or all of the method 900.

[0141] Referring now to FIG. 10, a process flow diagram of a method 1000 for carrying out embedded printing of thick structured meat constructs. The method 1000 can comprise: printing one or more portions of a myoblast-microgel composite ink into a cellular yield-stress matrix material according to first spatial portions of a meat construct print design, the first spatial portions of the meat construct print design being associated with one or more muscle portions of a thick structured meat construct 1001. The method 1000 can further comprise: printing one or more portions of the adipocyte progenitor cell microgel composite ink into the cellular yield-stress matrix material according to second spatial portions of the meat construct print design, the second spatial portions of the meat construct print design being associated with one or more lipid portions of the thick structured meat construct 1002. The method 1000 can further comprise: printing one or more portions of a sacrificial ink into the cellular yieldstress matrix material according to third spatial portions of the meat construct print design, the third spatial portions of the meat construct print design being associated with a perfusable vascular netw ork within the thick structured meat construct 1003. The method 1000 can further comprise: at least partially solidifying one or more of: the myoblast-microgel composite ink. the adipocyte progenitor cell-microgel composite ink, or the cellular yield-stress matrix material 1004. The method 1000 can further comprise: removing the one or more portions of the sacrificial ink printed into the cellular yield-stress matrix material to form the perfusable vascular network within the thick structured meat construct 1005.48LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743

[0142] Some or all of the method 1000 can be carried out using a system (e.g, 100), by an apparatus, or by a computing device, such as 400 or 500. For example, an apparatus can comprise at least one processor and at least one memory storing instructions thereon that, when executed by the at least one processor, cause the apparatus to perform some or all of the method 1000. Additionally, a computer program product can be provided that comprises a non-transitory computer-readable storage medium storing instructions thereon that, when executed by a processor, cause a machine or apparatus to perform some or all of the method 1000.

[0143] Referring now to FIG. 11, illustrates a process flow diagram of a method 1100 for carrying out embedded printing of thick structured meat constructs. The method 1100 can comprise: causing a print nozzle to move along a predefined pathway through a cellular yieldstress matrix material 1001. The method 1100 can further comprise: while causing the print nozzle to move along the predefined pathway through the yield-stress matrix material, communicating a first printing ink, a second printing ink, and a sacrificial ink into the cellular yield-stress support material at, respectively, a plurality of first points, a plurality’ of second points, and a plurality of third points along the predefined pathway through the cellular yieldstress support material to form an intermediate thick structured meat construct, wherein the printing nozzle is configured to exert shear forces on the cellular yield-stress matrix material, thereby causing at least partial liquefaction of the cellular yield-stress support matrix material during formation of the intermediate thick structured meat construct 1102, The method 1100 can further comprise: removing the sacrificial ink from the intermediate thick structured meat construct to form a thick structured meat construct, wherein the removing the sacrificial ink forms a perfusable vascular network through at least a portion of the thick structured meat construct 1103.

[0144] Some or all of the method 1100 can be carried out using a system (e.g, 100), by an apparatus, or by a computing device, such as 400 or 500. For example, an apparatus can comprise at least one processor and at least one memory storing instructions thereon that, when executed by the at least one processor, cause the apparatus to perform some or all of the method 1100. Additionally, a computer program product can be provided that comprises a non-transitory computer-readable storage medium storing instructions thereon that, when executed by a processor, cause a machine or apparatus to perform some or all of the method 1100.

[0145] Described herein are various examples of systems, methods, processes, approaches, devices, apparatuses, and computer program products, such as storage media (e.g, a non-49I.EGAI, 02 / 46428716v1Attorney Docket No.: 049648 / 636743 transitory computer-readable storage medium) for embedded printing of thick structured meat constructs and / or configured for carrying out, at least in part, embedded printing of thick structured meat constructs. The present disclosure describes a scalable three-dimensional (3D) printing method for fabricating 3D thick structured meat constructs with textures similar to those of traditional meat using embedded printing in a yield-stress support bath. The meat texture or structure is made possible by using embedded printing, and the meat volumetric size or thickness is enabled by incorporating a perfusable network in the meat construct to support the growth and maturation of the whole construct. In some embodiments, both myoblasts and adipocyte progenitor cells are individually printed into specific layouts in a yield-stress support bath to mimic the meat structure, and the cellular matrix bath is a bulky microgel-based matrix with endothelial cells and other support cells such as the adipose-derived stem cells, fibroblast cells, and / or stromal vascular fraction cells. In addition, main perfusable channels can be created in the bulky meat construct by adopting a sacrificial ink-assisted embedded printing process, which support the co-culture of endothelial cells and support cells in the matrix to selfassemble into capillary structures. Made of the main perfusable channels and capillary structures, a perfusable network is formed in the meat construct, serving as a vessel system.

[0146] As described in the example above, two different cellular microgel composite inks and one or more sacrificial ink are first printed into a designed pattern in a yield-stress support bath, which is made from endothelial cells and other support cells such as the adipose-derived stem cells, fibroblast cells, and / or stromal vascular fraction cells. The two cellular microgel composite inks are myoblast-laden microgel composite ink and adipocyte progenitor cellmicrogel composite ink. The function of the myoblast-laden microgel composite in and adipocyte progenitor cell-microgel composite ink is to construct muscle and fat structures in the final meat product, respectively.

[0147] In some embodiments, the muscle, fat, and vascular tissues are iteratively printed in a cellular yield stress matrix bath using two cellular inks and one or more sacrificial inks. The two cellular microgel composite inks are myoblast laden microgel composite ink and adipocyte progenitor cell-microgel composite ink. Generally, both the cellular inks and yield stress support bath comprise a discrete microgel phase either porous or solid microgel and a continuous hydrogel precursor phase hydrogel precursor based cellular suspension. Such microgel based materials have yield stress rheological properties providing good printability for embedded extrusion printing. The hydrogel precursor that makes up the continuous phase can be made of cross linkable hydrogels, such as gelatin. The cellular inks, including myoblasts laden and adipocyte progenitor cells laden inks are prepared by mixing living cells with porous50LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 or solid microgels and a liquid hydrogel precursor. The yield stress support bath comprises at least one of a porous or solid microgel. a liquid hydrogel precursor endothelial cell, and support cells such as adipose derived stem cells fibroblasts and / or stromal vascular fraction cells which facilitate endothelial cells to form capillary structures for microcirculation across the whole volume of the meat construct being printed during the post printing perfusion step.

[0148] In some embodiments, a cross linking agent such as transglutaminase (TG) may be added to the myoblast microgel composite ink, the adipocyte progenitor cell microgel composite ink, and the yield stress support bath for in situ cross-linking. Alternatively, the meat construct may be cross-linked using a different mechanism such as ultraviolet radiation after the printing process is completed. The sacrificial ink should be easily removable after printing and is usually made of a water-soluble polymer and rheology modifiers. In some embodiments, printed myoblast filaments are cultured to form a muscle tissue layer, where extrusion printing induced shear force and / or post printing tensioning are crucial for the development of mature myofibers. A meat construct may consist of multiple layers of myoblasts, multiple layers of adipocytes, and several sacrificial ink filaments and their spatial arrangement should be controlled per the texture and perfusable network as designed.

[0149] In some embodiments, the sacrificial patterns removed after the printing process is completed and the meat construct is sufficiently cross-linked. Depending on the type of sacrificial materials used, some sacrificial patterns may be simply removed by immersing the printed meat construct in phosphate buffered saline until the sacrificial material becomes diluted and much less viscous and then followed by flushing gently using a syringe. Once the sacrificial material is completely removed, main perfusable channels are formed for post printing perfusion.

[0150] In some embodiments, the printed meat construct in connected to a perfusion system and post printing tensioning setup for cell growth and maturation as well as capillary structure formation by perfusing cell media through the created main perfusable channels. The main perfusable channels and capillary structures form a perfusable network in the meat construct enabling it to be thick.

[0151] Various different bioinks, either alone or in combination, can be used in some or all of the systems, apparatuses, devices, methods, processes, approaches, and / or computer program products described herein for performing embedded printing of thick structured meat constructs. Bioinks can be formed from mixing myoblasts and a hydrogel precursor with a porous microgel to form a myoblast-microgel composite ink and from mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor51LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 cell-microgel composite ink. A cellular yield-stress support matrix material can be formed by mixing adipose-derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel. A thick, structured, meat construct can be formed by iteratively printing, into the cellular yield-stress matrix material, the myoblast-microgel composite ink to form muscle portions, the adipocyte progenitor cell-microgel composite ink to form lipid portions, and a sacrificial ink to form vascular network portions. Sacrificial ink is then removed to define a perfusable vascular network through the thick structured meat construct.

[0152] According to an embodiment, a method of preparing a thick structured meat construct can be carried out, the method comprising: mixing myoblasts and a hydrogel precursor with a porous microgel to form a myoblast-microgel composite ink. In some embodiments, the method can further comprise: mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink. In some embodiments, the method can further comprise: mixing adipose- derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel to form a cellular yield-stress matrix material. In some embodiments, the method can further comprise: iteratively printing one or more portions of the myoblast-microgel composite ink into the cellular yield-stress matrix material according to first spatial portions of a meat construct print design, the first spatial portions of the meat construct print design being associated with one or more muscle portions of the thick structured meat construct, one or more portions of the adipocyte progenitor cell-microgel composite ink into the cellular yield-stress matrix material according to second spatial portions of the meat construct print design, the second spatial portions of the meat construct print design being associated with one or more lipid portions of the thick structured meat construct, and one or more portions of a sacrificial ink into the cellular yield-stress matrix material according to third spatial portions of the meat construct print design, the third spatial portions of the meat construct print design being associated with a perfusable vascular network within the thick structured meat construct. In some embodiments, the method can further comprise: at least partially solidifying one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, or the cellular yield-stress matrix material. In some embodiments, the method can further comprise: removing the one or more portions of the sacrificial ink printed into the cellular yield-stress matrix material to form the perfusable vascular network within the thick structured meat construct.

[0153] In some embodiments, the method can further comprise: disposing a cross-linking agent into one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cell-52LEGAI.02 / 46428716v1Attorney Docket No.: 049648 / 636743 microgel composite ink, or the cellular yield-stress matrix material. In some embodiments, the cross-linking agent comprises transglutaminase. In some embodiments, the hydrogel precursor comprises a hydrogel precursor-based cellular suspension. In some embodiments, the hydrogel precursor-based cellular suspension comprises gelatin. In some embodiments, the support cells in the cellular yield-stress matrix material comprises one or more from among: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, the support cells facilitate the endothelial cells in the cellular yield-stress matrix material to form one or more capillary structures for microcirculation across the muscle portions of the thick structured meat construct.

[0154] In some embodiments, the porous microgel in the myoblast-microgel composite ink comprises gelatin. In some embodiments, the method further comprises: loading one or more of differentiation media or culturing media into the porous microgel. In some embodiments, the porous microgel in the adipocyte progenitor microgel composite ink comprises gelatin. In some embodiments, the method further comprises: loading one or more of differentiation media or culturing media into the porous microgel. In some embodiments, the cellular yield-stress matrix material is retained within a yield-stress support bath. In some embodiments, the printing of the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, and the sacrificial ink into the cellular yield-stress matrix material comprises: adjacently printing the myoblast-microgel composite ink, the adipocyte progenitor cellmicrogel composite ink. and the sacrificial ink into the cellular yield-stress matrix material from a first side of the yield-stress support bath to a second side of the yield-stress support bath based on the meat construct print design associated with the thick structured meat construct.

[0155] In some embodiments, the sacrificial ink comprises one or more water-soluble polymers. In some embodiments, the thick structured meat construct comprises multiple layers of myoblasts, multiple layers of adipocytes, and multiple sacrificial ink filaments, and wherein a spatial arrangement of the multiple layers of myoblasts, adipocytes, and sacrificial ink filaments is controlled during the printing based upon the meat construct print design associated with the thick structured meat construct. In some embodiments, the printing of the myoblast- microgel composite ink into the cellular yield-stress support material to form the one or more muscle portions of the thick structured meat construct is carried out using one or more extrusion printing nozzles. In some embodiments, the one or more extrusion printing nozzles are dimensioned and configured to exert extrusion printing-induced shear forces on the myoblast- microgel composite ink, thereby facilitating mature myofiber formation therefrom.53LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743

[0156] In some embodiments, the method further comprises: tensioning the myoblastmicrogel composite ink to further facilitate myofiber formation therefrom. In some embodiments, the tensioning comprises: performing mechanical tensioning of the myoblasts in the myoblast-microgel composite ink to facilitate one or more of: cell alignment, cell fusion, myotubule formation, an increased number of cell nuclei, increased myogenic protein expression, or increased cell differentiation. In some embodiments, the mechanical tensioning of the myoblasts comprises exerting on the muscle tissues one or more of: shear strain, volumetric strain, longitudinal strain, tensile strain, compressive strain, bending strain, stretching strain, multiaxial strain, uniaxial strain, randomized iterative strain, periodic strain, cyclic strain, or continuous strain.

[0157] According to another embodiment, an apparatus can be provided that comprises at least one processor and at least one memory storing instructions thereon that, when executed by the at least one processor, cause the apparatus to perform a method of preparing a thick structured meat construct. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, cause the apparatus to perform: mixing myoblasts and a hydrogel precursor with a porous microgel to form a myoblast-microgel composite ink. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, can further cause the apparatus to perform: mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, can further cause the apparatus to perform: mixing adipose-derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel to form a cellular yield-stress matrix material. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, can further cause the apparatus to perform: iteratively printing one or more portions of the myoblast-microgel composite ink into the cellular yield-stress matrix material according to first spatial portions of a meat construct print design, the first spatial portions of the meat construct print design being associated with one or more muscle portions of the thick structured meat construct, one or more portions of the adipocyte progenitor cellmicrogel composite ink into the cellular yield-stress matrix material according to second spatial portions of the meat construct print design, the second spatial portions of the meat construct print design being associated with one or more lipid portions of the thick structured meat construct, and one or more portions of a sacrificial ink into the cellular yield-stress matrix material according to third spatial portions of the meat construct print design, the third spatial54LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 portions of the meat construct print design being associated with a perfusable vascular network within the thick structured meat construct. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, can further cause the apparatus to perform: at least partially solidifying one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cell-mi crogel composite ink, or the cellular yield-stress matrix material. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, can further cause the apparatus to perform: removing the one or more portions of the sacrificial ink printed into the cellular yield-stress matrix material to form the perfusable vascular network within the thick structured meat construct.

[0158] In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, further cause the apparatus to perform: disposing a cross-linking agent into one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, or the cellular yield-stress matrix material. In some embodiments, the cross-linking agent comprises transglutaminase. In some embodiments, the hydrogel precursor comprises a hydrogel precursor-based cellular suspension. In some embodiments, the hydrogel precursor-based cellular suspension comprises gelatin. In some embodiments, the support cells in the cellular yield-stress matrix material comprises one or more from among: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, the support cells facilitate the endothelial cells in the cellular yieldstress matrix material to form one or more capillary structures for microcirculation across the muscle portions of the thick structured meat construct.

[0159] In some embodiments, the porous microgel in the myoblast-microgel composite ink comprises gelatin. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, further cause the apparatus to perform: loading one or more of differentiation media or culturing media into the porous microgel. In some embodiments, the porous microgel in the adipocyte progenitor microgel composite ink comprises gelatin. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, further cause the apparatus to perform: loading one or more of differentiation media or culturing media into the porous microgel. In some embodiments, the cellular yield-stress matrix material is retained within a yield-stress support bath. In some embodiments, the printing of the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink. and the sacrificial ink into the cellular yieldstress matrix material comprises: adjacently printing the myoblast-microgel composite ink, the55LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 adipocyte progenitor cell-microgel composite ink, and the sacrificial ink into the cellular yieldstress matrix material from a first side of the yield-stress support bath to a second side of the yield-stress support bath based on the meat construct print design associated with the thick structured meat construct.

[0160] In some embodiments, the sacrificial ink comprises one or more water-soluble polymers. In some embodiments, the thick structured meat construct comprises multiple layers of myoblasts, multiple layers of adipocytes, and multiple sacrificial ink filaments, and wherein a spatial arrangement of the multiple layers of myoblasts, adipocytes, and sacrificial ink filaments is controlled during the printing based upon the meat construct print design associated with the thick structured meat construct. In some embodiments, the printing of the myoblastmicrogel composite ink into the cellular yield-stress support material to form the one or more muscle portions of the thick structured meat construct is carried out using one or more extrusion printing nozzles. In some embodiments, the one or more extrusion printing nozzles are dimensioned and configured to exert extrusion printing-induced shear forces on the myoblastmicrogel composite ink, thereby facilitating mature myofiber formation therefrom.

[0161] In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, further cause the apparatus to perform: tensioning the myoblast-microgel composite ink to further facilitate myofiber formation therefrom. In some embodiments, the tensioning comprises: performing mechanical tensioning of the myoblasts in the myoblast-microgel composite ink to facilitate one or more of: cell alignment, cell fusion, myotubule formation, an increased number of cell nuclei, increased myogenic protein expression, or increased cell differentiation. In some embodiments, the mechanical tensioning of the myoblasts comprises exerting on the muscle tissues one or more of: shear strain, volumetric strain, longitudinal strain, tensile strain, compressive strain, bending strain, stretching strain, multiaxial strain, uniaxial strain, randomized iterative strain, periodic strain, cyclic strain, or continuous strain.

[0162] According to another embodiment, a computer program product can be provided that comprises anon-transitory computer-readable storage medium storing instructions thereon that, when executed by at least one processor of an apparatus, cause the apparatus to perform at least a portion of a method of preparing a thick structured meat construct. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, cause the apparatus to perform: mixing my oblasts and a hydrogel precursor with a porous microgel to form a myoblast-microgel composite ink. In some embodiments, the instructions stored on the non-transitory computer-readable storage56LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 medium, when executed by the at least one processor, can further cause the apparatus to perform: mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: mixing adipose- derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel to form a cellular yield-stress matrix material. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: iteratively printing one or more portions of the myoblast-microgel composite ink into the cellular yield-stress matrix material according to first spatial portions of a meat construct print design, the first spatial portions of the meat construct print design being associated with one or more muscle portions of the thick structured meat construct, one or more portions of the adipocyte progenitor cell-microgel composite ink into the cellular yield-stress matrix material according to second spatial portions of the meat construct print design, the second spatial portions of the meat construct print design being associated with one or more lipid portions of the thick structured meat construct, and one or more portions of a sacrificial ink into the cellular yield-stress matrix material according to third spatial portions of the meat construct print design, the third spatial portions of the meat construct print design being associated with a perfusable vascular network within the thick structured meat construct; at least partially solidifying one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, or the cellular yield-stress matrix material. In some embodiments, the instructions stored on the non-transitory computer- readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: removing the one or more portions of the sacrificial ink printed into the cellular yield-stress matrix material to form the perfusable vascular network within the thick structured meat construct.

[0163] In some embodiments, the instructions stored on the non-transitory computer- readable storage medium, when executed by the at least one processor, further cause the apparatus to perform: disposing a cross-linking agent into one or more of: the myoblast- microgel composite ink, the adipocyte progenitor cell-microgel composite ink, or the cellular yield-stress matrix material. In some embodiments, the cross-linking agent comprises transglutaminase. In some embodiments, the hydrogel precursor comprises a hydrogel precursor-based cellular suspension. In some embodiments, the hydrogel precursor-based cellular suspension comprises gelatin. In some embodiments, the support cells in the cellular57LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 yield-stress matrix material comprises one or more from among: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, the support cells facilitate the endothelial cells in the cellular yield-stress matrix material to form one or more capillary structures for microcirculation across the muscle portions of the thick structured meat construct.

[0164] In some embodiments, the porous microgel in the myoblast-microgel composite ink comprises gelatin. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, further cause the apparatus to perform: loading one or more of differentiation media or culturing media into the porous microgel. In some embodiments, the porous microgel in the adipocyte progenitor microgel composite ink comprises gelatin. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, further cause the apparatus to perform: loading one or more of differentiation media or culturing media into the porous microgel. In some embodiments, the cellular yield-stress matrix material is retained within a yield-stress support bath. In some embodiments, the printing of the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, and the sacrificial ink into the cellular yield-stress matrix material comprises: adjacently printing the myoblast-microgel composite ink, the adipocyte progenitor cellmicrogel composite ink. and the sacrificial ink into the cellular yield-stress matrix material from a first side of the yield-stress support bath to a second side of the yield-stress support bath based on the meat construct print design associated with the thick structured meat construct.

[0165] In some embodiments, the sacrificial ink comprises one or more water-soluble polymers. In some embodiments, the thick structured meat construct comprises multiple layers of myoblasts, multiple layers of adipocytes, and multiple sacrificial ink filaments, and wherein a spatial arrangement of the multiple layers of myoblasts, adipocytes, and sacrificial ink filaments is controlled during the printing based upon the meat construct print design associated with the thick structured meat construct. In some embodiments, the printing of the myoblast- microgel composite ink into the cellular yield-stress support material to form the one or more muscle portions of the thick structured meat construct is carried out using one or more extrusion printing nozzles. In some embodiments, the one or more extrusion printing nozzles are dimensioned and configured to exert extrusion printing-induced shear forces on the myoblast- microgel composite ink, thereby facilitating mature myofiber formation therefrom.

[0166] In some embodiments, the instructions stored on the non-transitory computer- readable storage medium, when executed by the at least one processor, further cause the58LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 apparatus to perform: tensioning the myoblast-microgel composite ink to further facilitate myofiber formation therefrom. In some embodiments, the tensioning comprises: performing mechanical tensioning of the myoblasts in the myoblast-microgel composite ink to facilitate one or more of: cell alignment, cell fusion, myotubule formation, an increased number of cell nuclei, increased myogenic protein expression, or increased cell differentiation. In some embodiments, the mechanical tensioning of the myoblasts comprises exerting on the muscle tissues one or more of: shear strain, volumetric strain, longitudinal strain, tensile strain, compressive strain, bending strain, stretching strain, multiaxial strain, uniaxial strain, randomized iterative strain, periodic strain, cyclic strain, or continuous strain.

[0167] According to another embodiment, a system or device can be provided that is configured to perform at least a portion of a method of preparing a thick structured meat construct. In some embodiments, the system or device comprises: means for mixing myoblasts and a hydrogel precursor with a porous microgel to form a myoblast-microgel composite ink. In some embodiments, the system or device further comprises: means for mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink. In some embodiments, the system or device further comprises: means for mixing adipose-derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel to form a cellular yield-stress matrix material. In some embodiments, the system or device further comprises: means for iteratively printing one or more portions of the myoblast-microgel composite ink into the cellular yield-stress matrix material according to first spatial portions of a meat construct print design, the first spatial portions of the meat construct print design being associated with one or more muscle portions of the thick structured meat construct, one or more portions of the adipocyte progenitor cellmicrogel composite ink into the cellular yield-stress matrix material according to second spatial portions of the meat construct print design, the second spatial portions of the meat construct print design being associated with one or more lipid portions of the thick structured meat construct, and one or more portions of a sacrificial ink into the cellular yield-stress matrix material according to third spatial portions of the meat construct print design, the third spatial portions of the meat construct print design being associated with a perfusable vascular network within the thick structured meat construct. In some embodiments, the system or device further comprises: means for at least partially solidifying one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, or the cellular yield-stress matrix material. In some embodiments, the system or device further comprises: means for removing the one or more portions of the sacrificial ink printed into the cellular yield-stress59LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 matrix material to form the perfusable vascular network within the thick structured meat construct.

[0168] In some embodiments, the system or device can further comprise: means for disposing a cross-linking agent into one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, or the cellular yield-stress matrix material. In some embodiments, the cross-linking agent comprises transglutaminase. In some embodiments, the hydrogel precursor comprises a hydrogel precursor-based cellular suspension. In some embodiments, the hydrogel precursor-based cellular suspension comprises gelatin. In some embodiments, the support cells in the cellular yield-stress matrix material comprises one or more from among: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, the support cells facilitate the endothelial cells in the cellular yield-stress matrix material to form one or more capillary structures for microcirculation across the muscle portions of the thick structured meat construct.

[0169] In some embodiments, the porous microgel in the my oblast-microgel composite ink comprises gelatin. In some embodiments, the system or device can further comprise: means for loading one or more of differentiation media or culturing media into the porous microgel. In some embodiments, the porous microgel in the adipocyte progenitor microgel composite ink comprises gelatin. In some embodiments, the system or device can further comprise: means for loading one or more of differentiation media or culturing media into the porous microgel. In some embodiments, the cellular yield-stress matrix material is retained within a yield-stress support bath. In some embodiments, the printing of the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, and the sacrificial ink into the cellular yieldstress matrix material comprises: adjacently printing the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, and the sacrificial ink into the cellular yieldstress matrix material from a first side of the yield-stress support bath to a second side of the yield-stress support bath based on the meat construct print design associated with the thick structured meat construct.

[0170] In some embodiments, the sacrificial ink comprises one or more water-soluble polymers. In some embodiments, the thick structured meat construct comprises multiple layers of myoblasts, multiple layers of adipocytes, and multiple sacrificial ink filaments, and wherein a spatial arrangement of the multiple layers of myoblasts, adipocytes, and sacrificial ink filaments is controlled during the printing based upon the meat construct print design associated with the thick structured meat construct. In some embodiments, the printing of the myoblast- microgel composite ink into the cellular yield-stress support material to form the one or more60LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 muscle portions of the thick structured meat construct is carried out using one or more extrusion printing nozzles. In some embodiments, the one or more extrusion printing nozzles are dimensioned and configured to exert extrusion printing-induced shear forces on the myoblastmicrogel composite ink, thereby facilitating mature myofiber formation therefrom.

[0171] In some embodiments, the system or device can further comprise: means for tensioning the myoblast-microgel composite ink to further facilitate myofiber formation therefrom. In some embodiments, the means for tensioning comprise means for performing mechanical tensioning of the myoblasts in the myoblast-microgel composite ink to facilitate one or more of: cell alignment, cell fusion, myotubule formation, an increased number of cell nuclei, increased myogenic protein expression, or increased cell differentiation. In some embodiments, the means for mechanical tensioning of the myoblasts comprises means for exerting, on the muscle tissues, one or more of: shear strain, volumetric strain, longitudinal strain, tensile strain, compressive strain, bending strain, stretching strain, multiaxial strain, uniaxial strain, randomized iterative strain, periodic strain, cyclic strain, or continuous strain.

[0172] According to another embodiment, a method of preparing a thick structured meat construct can be carried out, the method comprising: mixing a plurality of first living cells with one or more porous microgels and a hydrogel precursor to form a first ink. In some embodiments, the method can further comprise: mixing a plurality of second living cells with one or more porous microgels and a hydrogel precursor to form a second ink. In some embodiments, the method can further comprise: mixing one or more porous microgels, a plurality of endothelial cells, and a plurality of support cells with a hydrogel precursor to form a cellular yield-stress matrix material. In some embodiments, the method can further comprise: adding a crosslinking agent to at least one of the first ink, the second ink, or the cellular yieldstress matrix material; forming a third ink comprising a sacrificial material. In some embodiments, the method can further comprise: selectively and iteratively dispensing one or more volumes of the first ink, one or more volumes of the second ink, and one or more volumes of the third ink into the cellular yield-stress matrix material to form an uncross-linked printed tissue construct. In some embodiments, the method can further comprise: exposing the uncrosslinked printed tissue construct to ultraviolet radiation or a to cross-link the cellular yield-stress matrix material and the cellular microgel composite ink, thereby forming a cross-linked printed tissue construct. In some embodiments, the method can further comprise: immersing the crosslinked printed tissue construct in phosphate-buffered saline to at least partially dilute or dissolve the third ink. In some embodiments, the method can further comprise: flushing the third ink and phosphate-buffered saline from the cross-linked printed tissue construct to form61LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 one or more channels of a perfusable network within the cross-linked printed tissue construct. In some embodiments, the method can further comprise: after flushing the sacrificial ink and phosphate-buffered saline from the cross-linked printed tissue construct, connecting the perfusable network of the cross-linked printed tissue construct to a peristaltic pump-based perfusion system together with a post-printing tensile-force-based tensioning system applied to the tissue construct. In some embodiments, the method can further comprise: perfusing a cellular media through the one or more channels of the perfusable network within the crosslinked printed tissue construct.

[0173] In some embodiments, the cellular yield-stress matrix comprises one or more solid porous microgels, a liquid hydrogel precursor, a plurality of endothelial cells, and a plurality of support cells. In some embodiments, the plurality of support cells comprises one or more of: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, the crosslinking agent comprises transglutaminase. In some embodiments, the first ink is a first cellular microgel composite ink. In some embodiments, the method further comprises: mixing a plurality of living cells with one or more solid porous microgels and a liquid hydrogel precursor to form the second ink. wherein the second ink is a second cellular microgel composite ink.

[0174] In some embodiments, the first cellular microgel composite ink is a myoblast-laden microgel composite ink comprised of at least a plurality of living cells, solid porous microgels, and a liquid hydrogel precursor. In some embodiments, the plurality of living cells comprises animal stem cells. In some embodiments, the solid porous microgels comprise one or more of: polyethylene glycol (PEG), hyaluronic acid, chitosan, or gelatin. In some embodiments, the liquid hydrogel precursor is comprised of at least collagen, gelatin, gellan, alginate, fibrin, silk, chitosan, or hyaluronic acid.

[0175] In some embodiments, the second cellular microgel composite in is an adipocyte progenitor cell-laden microgel composite ink comprised of at least a plurality of living cells, solid porous microgels, and a liquid hydrogel precursor. In some embodiments, the third ink comprises one or more of: a hydrophilic non-ionic surfactant, a poloxamer, a material comprising a central hydrophobic block of polypropylene glycol flanked by two hydrophilic blocks of polyethylene glycol, C572H1146O259, methylcellulose, polyvinyl alcohol, xanthan gum, or carbohydrates.

[0176] In some embodiments, the first ink, second ink, or cellular matrix are not exposed to a chemical or enzymatic crosslinking agent. In some embodiments, the method further comprises: exposing the thick structured meat construct to ultraviolet radiation to cause at least62LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 partial crosslinking of one or more of the first ink, the second, the third ink, or the cellular yield-stress matrix material. In some embodiments, the phosphate buffered saline solution has a pH level of between about 5.8 and about 7.4. In some embodiments, the phosphate buffered saline comprises one or more of: sodium chloride, potassium chloride, potassium dihydrogen phosphate, or disodium hydrogen phosphate.

[0177] In some embodiments, the method further comprises: culturing the thick structured meat construct under physiologically relevant medium perfusion conditions and necessary tensioning conditions to mature the myoblasts and adipocytes while endothelial cells selfassemble into capillary’ structures around the main perfusable channel. In some embodiments, the method further comprises: when selectively and iteratively dispensing the one or more volumes of the first ink and the one or more volumes of the second ink through the printing nozzle into the cellular yield-stress matrix material to form the uncross-linked printed tissue construct, selectively and iteratively dispensing one or more volumes of the second cellular microgel composite ink through the printing nozzle into the cellular yield-stress matrix material. In some embodiments, the method further comprises: printing myoblast filaments into the cellular yield-stress matrix material; and enculturing the myoblast filaments to form a muscle tissue layer.

[0178] According to another embodiment, an apparatus can be provided that comprises at least one processor and at least one memory storing instructions thereon that, when executed by the at least one processor, cause the apparatus to perform a method of preparing a thick structured meat construct. In some embodiments, the instructions stored on the at least one memoty. when executed by the at least one processor, cause the apparatus to perform: mixing a plurality of first living cells with one or more porous microgels and a hydrogel precursor to form a first ink. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, can further cause the apparatus to perform: mixing a plurality of second living cells with one or more porous microgels and a hydrogel precursor to form a second ink. In some embodiments, the instructions stored on the at least one memory', when executed by the at least one processor, can further cause the apparatus to perform: mixing one or more porous microgels, a plurality of endothelial cells, and a plurality of support cells with a hydrogel precursor to form a cellular yield-stress matrix material. In some embodiments, the instructions stored on the at least one memory', when executed by the at least one processor, can further cause the apparatus to perform: adding a crosslinking agent to at least one of the first ink. the second ink, or the cellular yield-stress matrix material. In some embodiments, the instructions stored on the at least one memory’, when executed by the at least63LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 one processor, can further cause the apparatus to perform: forming a third ink comprising a sacrificial material. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, can further cause the apparatus to perform: selectively and iteratively dispensing one or more volumes of the first ink, one or more volumes of the second ink, and one or more volumes of the third ink into the cellular yield-stress matrix material to form an uncross-linked printed tissue construct. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, can further cause the apparatus to perform: exposing the uncross-linked printed tissue construct to ultraviolet radiation or a to cross-link the cellular yield-stress matrix material and the cellular microgel composite ink, thereby forming a cross-linked printed tissue construct. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, can further cause the apparatus to perform: immersing the cross-linked printed tissue construct in phosphate-buffered saline to at least partially dilute or dissolve the third ink; flushing the third ink and phosphate-buffered saline from the cross-linked printed tissue construct to form one or more channels of a perfusable network within the cross-linked printed tissue construct. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, can further cause the apparatus to perform: after flushing the sacrificial ink and phosphate-buffered saline from the cross-linked printed tissue construct, connecting the perfusable network of the cross-linked printed tissue construct to a peristaltic pump-based perfusion system together with a post-printing tensile-force-based tensioning system applied to the tissue construct. In some embodiments, the instructions stored on the at least one memory’, when executed by the at least one processor, can further cause the apparatus to perform: perfusing a cellular media through the one or more channels of the perfusable network within the cross-linked printed tissue construct.

[0179] In some embodiments, the cellular yield-stress matrix comprises one or more solid porous microgels, a liquid hydrogel precursor, a plurality of endothelial cells, and a plurality of support cells. In some embodiments, the plurality of support cells comprises one or more of: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, the crosslinking agent comprises transglutaminase. In some embodiments, the first ink is a first cellular microgel composite ink. In some embodiments, the instructions stored on the at least one memory’, when executed by the at least one processor, further cause the apparatus to perform: mixing a plurality of living cells with one or more solid porous microgels and a liquid hydrogel precursor to form the second ink, wherein the second ink is a second cellular microgel composite ink.64LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743

[0180] In some embodiments, the first cellular microgel composite ink is a myoblast-laden microgel composite ink comprised of at least a plurality of living cells, solid porous microgels, and a liquid hydrogel precursor. In some embodiments, the plurality of living cells comprises animal stem cells. In some embodiments, the solid porous microgels comprise one or more of: polyethylene glycol (PEG), hyaluronic acid, chitosan, or gelatin. In some embodiments, the liquid hydrogel precursor is comprised of at least collagen, gelatin, gellan, alginate, fibrin, silk, chitosan, or hyaluronic acid.

[0181] In some embodiments, the second cellular microgel composite in is an adipocyte progenitor cell-laden microgel composite ink comprised of at least a plurality of living cells, solid porous microgels, and a liquid hydrogel precursor. In some embodiments, the third ink comprises one or more of: a hydrophilic non-ionic surfactant, a poloxamer, a material comprising a central hydrophobic block of polypropylene glycol flanked by two hydrophilic blocks of polyethylene glycol, C572H1146O259, methylcellulose, polyvinyl alcohol, xanthan gum, or carbohydrates.

[0182] In some embodiments, the first ink, second ink, or cellular matrix are not exposed to a chemical or enzymatic crosslinking agent. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, further cause the apparatus to perform: exposing the thick structured meat construct to ultraviolet radiation to cause at least partial crosslinking of one or more of the first ink, the second, the third ink, or the cellular yield-stress matrix material. In some embodiments, the phosphate buffered saline solution has a pH level of between about 5.8 and about 7.4. In some embodiments, the phosphate buffered saline comprises one or more of: sodium chloride, potassium chloride, potassium dihydrogen phosphate, or disodium hydrogen phosphate.

[0183] In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, further cause the apparatus to perform: culturing the thick structured meat construct under physiologically relevant medium perfusion conditions and necessary' tensioning conditions to mature the myoblasts and adipocytes while endothelial cells self-assemble into capillary structures around the main perfusable channel. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, further cause the apparatus to perform: when selectively and iteratively dispensing the one or more volumes of the first ink and the one or more volumes of the second ink through the printing nozzle into the cellular yield-stress matrix material to form the uncrosslinked printed tissue construct, selectively and iteratively dispensing one or more volumes of the second cellular microgel composite ink through the printing nozzle into the cellular yield-65I.EGAI, 02 / 46428716v1Attorney Docket No.: 049648 / 636743 stress matrix material. In some embodiments, the instructions stored on the at least one memory, when executed by the at least one processor, further cause the apparatus to perform: printing myoblast filaments into the cellular yield-stress matrix material; and enculturing the myoblast filaments to form a muscle tissue layer.

[0184] According to another embodiment, a computer program product can be provided that comprises anon-transitory computer-readable storage medium storing instructions thereon that, when executed by at least one processor of an apparatus, cause the apparatus to perform a a method of preparing a thick structured meat construct. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, cause the apparatus to perform: mixing a plurality of first living cells with one or more porous microgels and a hydrogel precursor to form a first ink. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: mixing a plurality7of second living cells with one or more porous microgels and a hydrogel precursor to form a second ink. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: mixing one or more porous microgels, a plurality of endothelial cells, and a plurality7of support cells with a hydrogel precursor to form a cellular yield-stress matrix material. In some embodiments, the instructions stored on the non-transitory computer- readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: adding a crosslinking agent to at least one of the first ink, the second ink, or the cellular yield-stress matrix material. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: forming a third ink comprising a sacrificial material; selectively and iteratively dispensing one or more volumes of the first ink, one or more volumes of the second ink, and one or more volumes of the third ink into the cellular yield-stress matrix material to form an uncross-linked printed tissue construct. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: exposing the uncross-linked printed tissue construct to ultraviolet radiation or a to cross-link the cellular yield-stress matrix material and the cellular microgel composite ink, thereby forming a cross-linked printed tissue construct. In some embodiments, the instructions stored on the non-transitory7computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: immersing the cross-linked printed tissue66LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 construct in phosphate-buffered saline to at least partially dilute or dissolve the third ink; flushing the third ink and phosphate-buffered saline from the cross-linked printed tissue construct to form one or more channels of a perfusable network within the cross-linked printed tissue construct. In some embodiments, the instructions stored on the non-transitory computer- readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: after flushing the sacrificial ink and phosphate-buffered saline from the cross-linked printed tissue construct, connecting the perfusable network of the cross-linked printed tissue construct to a peristaltic pump-based perfusion system together with a postprinting tensile-force-based tensioning system applied to the tissue construct. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor, can further cause the apparatus to perform: perfusing a cellular media through the one or more channels of the perfusable network within the cross-linked printed tissue construct.

[0185] In some embodiments, the cellular yield-stress matrix comprises one or more solid porous microgels, a liquid hydrogel precursor, a plurality of endothelial cells, and a plurality of support cells. In some embodiments, the plurality of support cells comprises one or more of: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, the crosslinking agent comprises transglutaminase. In some embodiments, the first ink is a first cellular microgel composite ink. In some embodiments, the instructions stored on the non-transitory computer-readable storage medium, when executed by the at least one processor of the apparatus, further cause the apparatus to perform: mixing a plurality of living cells with one or more solid porous microgels and a liquid hydrogel precursor to form the second ink. wherein the second ink is a second cellular microgel composite ink.

[0186] In some embodiments, the first cellular microgel composite ink is a myoblast-laden microgel composite ink comprised of at least a plurality of living cells, solid porous microgels, and a liquid hydrogel precursor. In some embodiments, the plurality of living cells comprises animal stem cells. In some embodiments, the solid porous microgels comprise one or more of: polyethylene glycol (PEG), hyaluronic acid, chitosan, or gelatin. In some embodiments, the liquid hydrogel precursor is comprised of at least collagen, gelatin, gellan. alginate, fibrin, silk, chitosan, or hyaluronic acid.

[0187] In some embodiments, the second cellular microgel composite in is an adipocyte progenitor cell-laden microgel composite ink comprised of at least a plurality of living cells, solid porous microgels, and a liquid hydrogel precursor. In some embodiments, the third ink comprises one or more of: a hydrophilic non-ionic surfactant, a poloxamer, a material67LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 comprising a central hydrophobic block of polypropylene glycol flanked by two hydrophilic blocks of polyethylene glycol, C572H1146O259, methylcellulose, polyvinyl alcohol, xanthan gum, or carbohydrates.

[0188] In some embodiments, the first ink, second ink, or cellular matrix are not exposed to a chemical or enzy matic crosslinking agent. In some embodiments, the instructions stored on the non-transi lory computer-readable storage medium, when executed by the at least one processor of the apparatus, further cause the apparatus to perform: exposing the thick structured meat construct to ultraviolet radiation to cause at least partial crosslinking of one or more of the first ink, the second, the third ink, or the cellular yield-stress matrix material. In some embodiments, the phosphate buffered saline solution has a pH level of between about 5.8 and about 7.4. In some embodiments, the phosphate buffered saline comprises one or more of: sodium chloride, potassium chloride, potassium dihydrogen phosphate, or disodium hydrogen phosphate.

[0189] In some embodiments, the instructions stored on the non-transitory computer- readable storage medium, when executed by the at least one processor of the apparatus, further cause the apparatus to perform: culturing the thick structured meat construct under physiologically relevant medium perfusion conditions and necessary tensioning conditions to mature the myoblasts and adipocytes while endothelial cells self-assemble into capillary7structures around the main perfusable channel. In some embodiments, the instructions stored on the non-transitory7computer-readable storage medium, when executed by the at least one processor of the apparatus, further cause the apparatus to perform: when selectively and iteratively dispensing the one or more volumes of the first ink and the one or more volumes of the second ink through the printing nozzle into the cellular yield-stress matrix material to form the uncross-linked printed tissue construct, selectively and iteratively dispensing one or more volumes of the second cellular microgel composite ink through the printing nozzle into the cellular yield-stress matrix material. In some embodiments, the instructions stored on the non- transitory computer-readable storage medium, when executed by the at least one processor of the apparatus, further cause the apparatus to perform: printing myoblast filaments into the cellular yield-stress matrix material; and enculturing the myoblast filaments to form a muscle tissue layer.

[0190] According to another embodiment, a system or device can be provided that is configured for carrying out at least a portion of a method of preparing a thick structured meat construct. In some embodiments, the system or device can comprise: means for mixing a plurality of first living cells with one or more porous microgels and a hydrogel precursor to68LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 form a first ink; mixing a plurality of second living cells with one or more porous microgels and a hydrogel precursor to form a second ink. In some embodiments, the system or device can further comprise: means for mixing one or more porous microgels, a pl urality of endothelial cells, and a plurality of support cells with a hydrogel precursor to form a cellular yield-stress matrix material. In some embodiments, the system or device can further comprise: means for adding a crosslinking agent to at least one of the first ink, the second ink, or the cellular yieldstress matrix material. In some embodiments, the system or device can further comprise: means for forming a third ink comprising a sacrificial material. In some embodiments, the system or device can further comprise: means for selectively and iteratively dispensing one or more volumes of the first ink, one or more volumes of the second ink, and one or more volumes of the third ink into the cellular yield-stress matrix material to form an uncross-linked printed tissue construct. In some embodiments, the system or device can further comprise: means for exposing the uncross-linked printed tissue construct to ultraviolet radiation or a to cross-link the cellular yield-stress matrix material and the cellular microgel composite ink, thereby forming a cross-linked printed tissue construct. In some embodiments, the system or device can further comprise: means for immersing the cross-linked printed tissue construct in phosphate-buffered saline to at least partially dilute or dissolve the third ink. In some embodiments, the system or device can further comprise: means for flushing the third ink and phosphate-buffered saline from the cross-linked printed tissue construct to form one or more channels of a perfusable network within the cross-linked printed tissue construct. In some embodiments, the system or device can further comprise: means for after flushing the sacrificial ink and phosphate-buffered saline from the cross-linked printed tissue construct, connecting the perfusable network of the cross-linked printed tissue construct to a peristaltic pump-based perfusion system together with a post-printing tensile-force-based tensioning system applied to the tissue construct. In some embodiments, the system or device can further comprise: means for perfusing a cellular media through the one or more channels of the perfusable network within the cross-linked printed tissue construct.

[0191] In some embodiments, the cellular yield-stress matrix comprises one or more solid porous microgels, a liquid hydrogel precursor, a plurality of endothelial cells, and a plurality of support cells. In some embodiments, the plurality of support cells comprises one or more of: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, the crosslinking agent comprises transglutaminase. In some embodiments, the first ink is a first cellular microgel composite ink. In some embodiments, the system or device can further comprise: means for mixing a plurality of living cells with one or more solid porous69LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 microgels and a liquid hydrogel precursor to form the second ink, wherein the second ink is a second cellular microgel composite ink.

[0192] In some embodiments, the first cellular microgel composite ink is a myoblast-laden microgel composite ink comprised of at least a plurality of living cells, solid porous microgels, and a liquid hydrogel precursor. In some embodiments, the plurality of living cells comprises animal stem cells. In some embodiments, the solid porous microgels comprise one or more of: polyethylene glycol (PEG), hyaluronic acid, chitosan, or gelatin. In some embodiments, the liquid hydrogel precursor is comprised of at least collagen, gelatin, gellan, alginate, fibrin, silk, chitosan, or hyaluronic acid.

[0193] In some embodiments, the second cellular microgel composite in is an adipocyte progenitor cell-laden microgel composite ink comprised of at least a plurality of living cells, solid porous microgels, and a liquid hydrogel precursor. In some embodiments, the third ink comprises one or more of: a hydrophilic non-ionic surfactant, a poloxamer, a material comprising a central hydrophobic block of polypropylene glycol flanked by two hydrophilic blocks of polyethylene glycol, C572H1146O259, methylcellulose, polyvinyl alcohol, xanthangum, or carbohydrates.

[0194] In some embodiments, the first ink, second ink, or cellular matrix are not exposed to a chemical or enzymatic crosslinking agent. In some embodiments, the system or device can further comprise: means for exposing the thick structured meat construct to ultraviolet radiation to cause at least partial crosslinking of one or more of the first ink, the second, the third ink, or the cellular yield-stress matrix material. In some embodiments, the phosphate buffered saline solution has a pH level of between about 5.8 and about 7.4. In some embodiments, the phosphate buffered saline comprises one or more of: sodium chloride, potassium chloride, potassium dihydrogen phosphate, or disodium hydrogen phosphate.

[0195] In some embodiments, the system or device can further comprise: means for culturing the thick structured meat construct under physiologically relevant medium perfusion conditions and necessary tensioning conditions to mature the myoblasts and adipocytes while endothelial cells self-assemble into capillary structures around the main perfusable channel. In some embodiments, the system or device can further comprise: means for when selectively and iteratively dispensing the one or more volumes of the first ink and the one or more volumes of the second ink through the printing nozzle into the cellular yield-stress matrix material to form the uncross-linked printed tissue construct, selectively and iteratively dispensing one or more volumes of the second cellular microgel composite ink through the printing nozzle into the cellular yield-stress matrix material. In some embodiments, the system or device can further70LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 comprise: means for printing myoblast filaments into the cellular yield-stress matrix material; and enculturing the myoblast filaments to form a muscle tissue layer.

[0196] According to another embodiment, a thick structured meat construct can be printed, formed, or provided. In some embodiments, the thick structured meat construct comprises: a first ink comprising a plurality of living cells, one or more solid porous microgels, and a liquid hydrogel precursor; and a cellular yield-stress matrix material comprising one or more solid porous microgels, a liquid hydrogel precursor, a plurality of endothelial cells, and a plurality of support cells. In some embodiments, the plurality of support cells comprise one or more of: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells. In some embodiments, at least one of the first ink or the cellular yield-stress matrix material further comprises a crosslinking agent. In some embodiments, the first ink and the second ink are disposed within the cellular yield-stress matrix material according to a 3D printing design / printing pathway and cured using ultraviolet radiation to form the thick structured meat / tissue construct.

[0197] In some embodiments, the crosslinking agent comprising transglutaminase. In some embodiments, the first ink comprises a cellular microgel composite. In some embodiments, the second ink comprises a sacrificial material. In some embodiments, the third ink is configured to be diluted or dissolved via phosphate-buffered saline to form a perfusable network within the thick structured meat / tissue construct. In some embodiments, the perfusable network is configured to be placed in fluidic communication with a perfusion system or a post-printing tensioning system to facilitate perfusion of the thick structured meat / tissue construct. In some embodiments, the perfusable network is configured to fluidically receive a cellular media from the perfusion system or post-printing tensioning system to enhance cellular survival within the thick structured meat / tissue construct.

[0198] According to another embodiment, an apparatus can be provided that comprises: at least one processor; and at least one memory storing instructions thereon that, when executed by the at least one processor, cause the apparatus to perform at least: mixing myoblasts and a hydrogel precursor with a porous microgel to form a my oblast-mi crogel composite ink; mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink; mixing adipose-derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel to form a cellular yield-stress matrix material; iteratively printing one or more portions of the myoblast-microgel composite ink into the cellular yield-stress matrix material according to first spatial portions of a meat construct print design, the first spatial portions of the meat construct print design being71LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 associated with one or more muscle portions of the thick structured meat construct, one or more portions of the adipocyte progenitor cell-microgel composite ink into the cellular yield-stress matrix material according to second spatial portions of the meat construct print design, the second spatial portions of the meat construct print design being associated with one or more lipid portions of the thick structured meat construct, and one or more portions of a sacrificial ink into the cellular yield-stress matrix material according to third spatial portions of the meat construct print design, the third spatial portions of the meat construct print design being associated with a perfusable vascular network within the thick structured meat construct; at least partially solidifying one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, or the cellular yield-stress matrix material; and removing the one or more portions of the sacrificial ink printed into the cellular yield-stress matrix material to form the perfusable vascular network within the thick structured meat construct.

[0199] According to another embodiment, a computer program product can be provided that comprises a non-transitory computer-readable storage medium storing computerexecutable instructions thereon that, when executed by at least one processor, cause an apparatus to perform at least: mixing myoblasts and a hydrogel precursor with a porous microgel to form a myoblast-microgel composite ink; mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink; mixing adipose-derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel to form a cellular yield-stress matrix material; iteratively printing one or more portions of the myoblast-microgel composite ink into the cellular yield-stress matrix material according to first spatial portions of a meat construct print design, the first spatial portions of the meat construct print design being associated with one or more muscle portions of the thick structured meat construct, one or more portions of the adipocyte progenitor cell-microgel composite ink into the cellular yield-stress matrix material according to second spatial portions of the meat construct print design, the second spatial portions of the meat construct print design being associated with one or more lipid portions of the thick structured meat construct, and one or more portions of a sacrificial ink into the cellular yield-stress matrix material according to third spatial portions of the meat construct print design, the third spatial portions of the meat construct print design being associated with a perfusable vascular network within the thick structured meat construct; at least partially solidifying one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cell- microgel composite ink, or the cellular yield-stress matrix material; and removing the one or72LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 more portions of the sacrificial ink printed into the cellular yield-stress matrix material to form the perfusable vascular network within the thick structured meat construct.Examples Provided Herein are Inexhaustive

[0200] It should be appreciated that the systems, apparatuses, methods, devices, components, processes, approaches, materials, compositions of matter, computer program products, non-transitory computer-readable storage media, and thick structured meat constructs described herein are provided as select, inexhaustive examples only and do not constitute a full or complete set of examples contemplated by, and know to, the inventors. Combinations, variations, hybrids, or alternatives to the example systems, apparatuses, methods, devices, components, processes, approaches, materials, compositions of matter, computer program products, non-transitory computer-readable storage media, and thick structured meat constructs described herein are contemplated by, and known to, the inventors, such that the present disclosure includes within its scope of disclosure, both in the written description and figures illustrated in the drawing sheets, any and all such combinations, variations, hybrids, or alternatives. A person having ordinary skill in the relevant art, in view of the present disclosure, will further understand and appreciate that the disclosure includes these combinations, variations, hybrids, or alternatives to the examples and embodiments provided herein and that the inventors have suitably described the examples and embodiments herein such that said person having ordinary skill in the relevant art can make and use the disclosed invention as embodied in these combinations, variations, hybrids, or alternatives to the examples and embodiments expressly described herein.

[0201] It should also be appreciated that all combinations of the foregoing concepts and additional concepts discussed in greater detail below (provided such concepts are not mutually inconsistent) are contemplated as being part of the inventive subject matter disclosed herein. In particular, the combinations of claimed subject matter appearing at the end of this disclosure are contemplated as being part of the inventive subject matter disclosed herein. It should be appreciated that terminology explicitly employed herein that also may appear in any disclosure incorporated by reference should be accorded a meaning consistent with the particular concepts disclosed herein.

[0202] In some embodiments, one or more of the operations, steps, elements, or processes described herein may be modified or further amplified as described below. Moreover, in some embodiments, additional optional operations may also be included. It should be appreciated that each of the modifications, optional additions, and / or amplifications described herein may73LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 be included with the operations previously described herein, either alone or in combination, with any others from among the features described herein.

[0203] The provided method description, illustrations, and process flow diagrams are provided merely as illustrative examples and are not intended to require or imply that the steps of the various embodiments must each or all be performed and / or should be performed in the order presented or described. As will be appreciated by one of skill in the art, the order of steps in some or all of the embodiments described may be performed in any order. Words such as “thereafter,” “then,” “next,” etc. are not intended to limit the order of the steps; these words are simply used to guide the reader through the description of the methods. Further, any reference to claim elements in the singular, for example, using the articles “a,” “an,” or “the” is not to be construed as limiting the element to the singular. Further, any reference to dispensing, disposing, depositing, dispersing, conveying, injecting, inserting, communicating, and other such terms of art are not to be constmed as limiting the element to any particular means or method or apparatus or system, and is taken to mean conveying the material within the receiving vessel, solution, conduit, or the like by way of any suitable method.

[0204] Many modifications and other embodiments of the inventions set forth herein will come to mind to one skilled in the art to which these inventions pertain having the benefit of teachings presented in the foregoing descriptions and the associated drawings. Although the figures only show certain components of the apparatus and systems described herein, it is understood that various other components may be used in conjunction with the system. Therefore, it is to be understood that the inventions are not to be limited to the specific embodiments disclosed and that modifications and other embodiments are intended to be included within the scope of the appended claims. Moreover, the steps in the method described above may not necessarily occur in the order depicted in the accompanying diagrams, and in some cases one or more of the steps depicted may occur substantially simultaneously, or additional steps may be involved. Although specific terms are employed herein, they are used in a generic and descriptive sense only and not for purposes of limitation. Specific equipment and materials described in the examples are for illustration only and not for purposes of limitation. For instance, any and all articles, portions of articles, structures, bulk materials, and / or the like, having any form factor, scale, dimensions, aesthetic attributes, material properties, internal structures, and / or mechanical properties, which are formed according to any of the disclosed methods, approaches, processes, or variations thereof, using any devices, equipment, apparatuses, systems, or variations thereof, using any of the build material, printing mixture, ink, yield-stress support material, or other material compositions described herein or74LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 variations thereof, are all contemplated and covered by the present disclosure. None of the examples provided are intended to, nor should they, limit in any way the scope of the present disclosure.

[0205] Every document cited or referenced herein, including any cross referenced or related patent or application is hereby incorporated herein by reference in its entirety unless expressly excluded or otherwise limited. The citation of any document and / or the mention of methods or apparatuses as being conventional, typical, usual, or the like is not. and should not be taken as an acknowledgement or any form of suggestion that the reference or mentioned method / apparatus is prior art with respect to any invention disclosed or claimed herein or that it alone, or in any combination with any other reference or references, teaches, suggests or discloses any such invention or forms part of the common general knowledge in any country in the world. Further, to the extent that any meaning or definition of a term in this document conflicts with any meaning or definition of the same term in a document incorporated by reference, the meaning or definition assigned to that term in this document shall govern.

[0206] The various portions of the present disclosure, such as the Background. Summary, Brief Description of the Drawings, and Abstract sections, are provided to comply with requirements of the MP EP and are not to be considered an admission of prior art or a suggestion that any portion or part of the disclosure constitutes common general knowledge in any country in the world. The present disclosure is provided as a discussion of the inventor’s own work and improvements based on the inventor’s own work. See. e.g. , Riverwood Int 7 Corp. v. R.A. Jones Co., 324 F.3d 1346, 1354 (Fed. Cir. 2003).75LEGAI,02 / 46428716v1

Claims

Attorney Docket No.: 049648 / 636743ClaimsWhat is claimed is:

1. A method of preparing a thick structured meat construct, the method comprising: printing a volume of a myoblast-microgel composite ink into a cellular yield-stress matrix material to form muscle tissue for a thick structured meat construct, wherein the myoblast-microgel composite ink comprise myoblasts, a hydrogel precursor, and a first porous microgel, wherein the cellular yield-stress matrix material comprises adipose-derived stem cells, endothelial cells, support cells, a hydrogel precursor, and a second porous microgel; printing a volume of a adipocyte progenitor cell-microgel composite ink into the cellular yield-stress matrix material to form lipid tissue for the thick structured meat construct, wherein the adipocyte progenitor cell-microgel composite ink comprises adipocyte progenitor cells, a hydrogel precursor, and a solid microgel: printing a volume of a sacrificial ink into the cellular yield-stress matrix material to form a vascular network within the thick structured meat construct; at least partially solidifying the myoblast-microgel composite ink and the adipocyte progenitor cell-microgel composite ink; and removing the sacrificial ink from the cellular yield-stress matrix material to make the vascular network within the thick structured meat construct a perfusable vascular network.

2. The method of claim 1 , further comprising: mixing the myoblasts and the hydrogel precursor with the first porous microgel to form the myoblast-microgel composite ink; mixing the adipocyte progenitor cells and the hydrogel precursor with the solid microgel to form the adipocyte progenitor cell-microgel composite ink; and mixing the adipose-derived stem cells, the endothelial cells, the support cells, and the hydrogel precursor with the second porous microgel to form the cellular yield-stress matrix material.76LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 6367433. The method of claim 1, further comprising: disposing a cross-linking agent into one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink, or the cellular yield-stress matrix material.

4. The method of claim 3, wherein the cross-linking agent compnses transglutaminase.

5. The method of claim 1, wherein the hydrogel precursor comprises a hydrogel precursor-based cellular suspension.

6. The method of claim 5, wherein the hydrogel precursor-based cellular suspension comprises gelatin.

7. The method of claim 1, wherein the support cells in the cellular yield-stress matrix material comprises one or more from among: adipose-derived stem cells, fibroblasts, or stromal vascular fraction cells.

8. The method of claim 1, wherein the support cells facilitate the endothelial cells in the cellular yield-stress matrix material to form one or more capillary structures for microcirculation across the muscle portions of the thick structured meat construct.

9. The method of claim 1, wherein the porous microgel in the myoblast-microgel composite ink comprises gelatin, and wherein the method further comprises: loading one or more of differentiation media or culturing media into the porous microgel.

10. The method of claim 1, wherein the porous microgel in the adipocyte progenitor microgel composite ink comprises gelatin, and wherein the method further comprises: loading one or more of differentiation media or culturing media into the porous microgel.

11. The method of claim 1, wherein the cellular yield-stress matrix material is retained within a yield-stress support bath.77LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 63674312. The method of claim 11, wherein the printing of the my oblast-mi crogel composite ink, the adipocyte progenitor cell-microgel composite ink, and the sacrificial ink into the cellular yield-stress matrix material comprises: adjacently printing the my oblast-mi crogel composite ink, the adipocyte progenitor cell-microgel composite ink. and the sacrificial ink into the cellular yield-stress matrix material from a first side of the yield-stress support bath to a second side of the yield-stress support bath based on the meat construct print design associated with the thick structured meat construct.

13. The method of claim 1, wherein the thick structured meat construct comprises multiple layers of myoblasts, multiple layers of adipocytes, and multiple sacrificial ink filaments, and wherein a spatial arrangement of the multiple layers of myoblasts, adipocytes, and sacrificial ink filaments is controlled during the printing based upon the meat construct print design associated with the thick structured meat construct.

14. The method of claim 1, wherein the printing of the myoblast-microgel composite ink into the cellular yield-stress support material to form the one or more muscle portions of the thick structured meat construct is carried out using one or more extrusion printing nozzles.

15. The method of claim 14, wherein the one or more extrusion printing nozzles are dimensioned and configured to exert extrusion printing-induced shear forces on the myoblast- microgel composite ink, thereby facilitating mature myofiber formation therefrom.

16. The method of claim 1, further comprising: tensioning the myoblast-microgel composite ink to further facilitate myofiber formation therefrom.

17. The method of claim 1 , wherein the tensioning comprises: performing mechanical tensioning of the myoblasts in the myoblast-microgel composite ink to facilitate one or more of: cell alignment, cell fusion, myotubule formation, an increased number of cell nuclei, increased myogenic protein expression, or increased cell differentiation.78LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 63674318. The method of claim 17, wherein the mechanical tensioning of the myoblasts comprises exerting on the muscle tissues one or more of shear strain, volumetric strain, longitudinal strain, tensile strain, compressive strain, bending strain, stretching strain, multiaxial strain, uniaxial strain, randomized iterative strain, periodic strain, cyclic strain, or continuous strain.

19. A method comprising: providing a cellular yield-stress matrix material comprising adipose-derived stem cells, endothelial cells, support cells, a hydrogel precursor, and a first porous microgel: disposing a plurality of volumes of a myoblast-microgel composite ink into the cellular yield-stress matrix material at a first plurality of locations within the cellular yieldstress matrix material to form muscle tissue for a thick structured meat construct, wherein the myoblast-microgel composite ink comprises myoblasts, a hydrogel precursor, and a first porous microgel. wherein the cellular yield-stress matrix material comprises adipose-derived stem cells, endothelial cells, support cells, a hydrogel precursor, and a second porous microgel; disposing a plurality of volumes of a adipocyte progenitor cell-microgel composite ink into the cellular yield-stress matrix material at a second plurality of locations to form lipid tissue for the thick structured meat construct, wherein the adipocyte progenitor cell-microgel composite ink comprises adipocyte progenitor cells, a hydrogel precursor, and a solid microgel; disposing a plurality of volumes of a sacrificial ink into the cellular yield-stress matrix material at a third plurality of locations to form a vascular network within the thick structured meat construct; at least partially solidifying the cellular yield-stress matrix material, the myoblast- microgel composite ink, and the adipocyte progenitor cell-microgel composite ink; and at least partially removing the sacrificial ink to make the vascular network within the thick structured meat construct a perfusable vascular network.

20. A method of preparing a thick structured meat construct, the method comprising: mixing myoblasts and a hydrogel precursor with a porous microgel to form a myoblast-microgel composite ink;79LEGAI,02 / 46428716v1Attorney Docket No.: 049648 / 636743 mixing adipocyte progenitor cells and a hydrogel precursor with a solid microgel to form an adipocyte progenitor cell-microgel composite ink; mixing adipose-derived stem cells, endothelial cells, support cells, and a hydrogel precursor with a porous microgel to form a cellular yield-stress matrix material; iteratively printing one or more portions of the myoblast-microgel composite ink into the cellular yield-stress matrix material according to first spatial portions of a meat construct print design, the first spatial portions of the meat construct print design being associated with one or more muscle portions of the thick structured meat construct, one or more portions of the adipocyte progenitor cell-microgel composite ink into the cellular yield-stress matrix material according to second spatial portions of the meat construct print design, the second spatial portions of the meat construct print design being associated with one or more lipid portions of the thick structured meat construct, and one or more portions of a sacrificial ink into the cellular yield-stress matrix material according to third spatial portions of the meat construct print design, the third spatial portions of the meat construct print design being associated with a perfusable vascular network within the thick structured meat construct; at least partially solidifying one or more of: the myoblast-microgel composite ink, the adipocyte progenitor cell-microgel composite ink. or the cellular yield-stress matrix material; and removing the one or more portions of the sacrificial ink printed into the cellular yieldstress matrix material to form the perfusable vascular network within the thick structured meat construct.80LEGAI,02 / 46428716v1

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