Additive manufacturing of hydrogel tubes for biomedical applications

Abstract

[0013] Embodiments of the present disclosure include methods for simultaneously fabricating two or more hydrogel constructs (e.g., tubular hydrogel constructs). In some embodiments, the methods include one or more of the following steps: providing a vat including a bio-ink composition containing one or more monomers and / or one or more polymers; applying electromagnetic radiation from an electromagnetic radiation source to cure a layer of the hydrogel construct (e.g., a tubular hydrogel construct); and applying electromagnetic radiation from the electromagnetic radiation source one or more additional times to generate one or more additional layers of the hydrogel construct (e.g., a tubular hydrogel construct).

Description

Technical Field

[0001] Cross - Reference to Related Applications Cross - Reference to Related Applications This application claims the priority of U.S. Provisional Application No. 63 / 185,299, filed on May 6, 2021, the entire disclosure of which is incorporated herein by reference.

Background Art

[0002] Background Compositions containing hydrogels can be used to form objects for use in biocompatible structures. These objects can be formed using three - dimensional (3D) printing techniques. Cells can be attached for practical applications such as synthetic organs.

Summary of the Invention

[0003] Summary of the Invention Embodiments of the present disclosure relate to a method for simultaneously producing two or more tubular hydrogel constructs, comprising providing a vat containing a bio-ink composition comprising one or more monomers, one or more polymers, one or more UV absorbers, one or more photoinitiators, one or more natural or synthetic ECMs, and / or peptides; applying electromagnetic radiation from an electromagnetic radiation source to cure layers of the tubular hydrogel constructs; and applying electromagnetic radiation from the electromagnetic radiation source one or more times additionally to produce one or more additional layers of the tubular hydrogel constructs. In some embodiments, the electromagnetic radiation is UV radiation. In some embodiments, 10 or more tubular hydrogel constructs are produced simultaneously. In some embodiments, the vat further comprises a liquid that is miscible with the bio-ink. In some embodiments, the bio-ink comprises a poly(ethylene glycol) di-(meth)acrylate polymer. In some embodiments, the bio-ink comprises at least one photoinitiator. In some embodiments, the bio-ink comprises DI water. In some embodiments, the bio-ink further comprises UV dyes, proteins, peptides, biopharmaceuticals, pharmaceutical compounds, and / or extracellular matrix materials. In some embodiments, the tubular hydrogel constructs are substantially the same shape, size, and / or have the same relative dimensions as organs or organ fragments. In some embodiments, organs or organ fragments include blood vessels, trachea, bronchi, esophagus, ureter, renal tubule, bile duct, renal tubule, renal tubule, bile duct, hepatic duct, nerve conduit, CSF shunt, larynx, or pharynx. In some embodiments, blood vessels include pulmonary artery, renal artery, coronary artery, peripheral artery, pulmonary vein, or renal vein. In some embodiments, the tubular hydrogel construct includes a hemodialysis graft. In some embodiments, the tubular hydrogel construct allows for endothelialization of the lumen of the tubular hydrogel construct and / or cellularization of the outer surface of the tubular hydrogel construct. In some embodiments, the lumen of the tubular hydrogel construct includes a patterned surface. In some embodiments, the patterned surface includes patterning that allows for unidirectional flow through the tube.In some embodiments, the tubular hydrogel construct includes one or more branches. In some embodiments, the hydrogel construct is polymerized poly(ethylene glycol) di(meth)acrylate, polymerized poly(ethylene glycol) di(meth)acrylamide, polymerized poly(ethylene glycol) (meth)acrylate / (methacrylamide), poly(ethylene glycol)-block-poly(ε-caprolactone), polycaprolactone, polyvinyl alcohol, gelatin, methylcellulose, hydroxyethyl methylcellulose, hydroxypropyl methylcellulose, polyethylene oxide, polyacrylamide, polyacrylic acid, polymethacrylic acid, poly The following are selected from the group, including salts of acrylic acid, salts of polymethacrylic acid, poly(2-hydroxyethyl methacrylate), polylactic acid, polyglycolic acid, polyvinyl alcohol, polymethacrylic anhydride, polyacrylic anhydride, polysebacic anhydride, and other polyanhydrides, collagen, poly(hyaluronic acid), hyaluronic acid-containing polymers and copolymers, polypeptides, dextran, dextran sulfate, chitosan, chitin, agarose gel, fibrin gel, soy-derived hydrogel, alginate-based hydrogel, poly(sodium alginate), hydroxypropyl acrylate (HPA), lithium phenyl 2,4,6-trimethylbenzoyl phosphinate (LAP), sodium phenyl-2,4,6-phenyl-2,4,6-trimethylbenzoyl phosphinate (NaP), and combinations thereof.

[0004] Further embodiments include batches of tubular hydrogel constructs produced by the processes of the embodiments described above. In some embodiments, the tubular hydrogel constructs include different shapes. figure

[0005] Figure 1A shows cross-sectional views of several embodiments of hydrogel structures. Figure 1B shows the 45-degree field of view of the embodiment shown in Figure 1A. Figure 2 shows a model of a printed tube. Figure 3 is a photograph of a printed tube. Figure 4 is a photograph of a printed tube. Figures 5A-D are photographs of the printed tube being attached to the modified tube fitting. Detailed explanation

[0006] As used herein, “3D printing” refers to any technique used to create a three-dimensional object using a digital model of that object. Exemplary 3D printing techniques include [insert].

[0007] As used herein, “printable ink” and “printable composition” refer to any composition that can be used to form an object using 3D printing technology. “Bio-ink” is a printable ink that forms a material having one or more desired biocompatible properties. For example, a bio-ink may contain one or more materials that promote adhesion and proliferation of a desired cell type. The printed object may support the adhesion, proliferation, interaction, and diffusion of primary cells and induced pluripotent stem cells. In some cases, the bio-ink may be formed on a hydrogel. Compounds in the bio-ink can be selected or modified to incorporate chemical functional groups by means of chemical synthesis, etc. Chemical functionality may enable the incorporation of modified materials as components in the bio-ink. Modification may enable the chemical bonding of desired components. The desired components may maintain their cell interaction function. Such incorporation may enable the modification of the mechanical properties of the printed object without interfering with cell adhesion.

[0008] As used herein, “extracellular matrix” and “ECM” refer to natural and synthetic ECM, as well as one or more constituent materials of ECM. For example, ECM may refer to naturally occurring ECM or ECM produced using synthetic techniques. ECM may also refer to one or more constituent materials of naturally occurring ECM, such as collagen (natural or synthetic). In some cases, “ECM material” is used to refer to a specific material. ECM can be produced using techniques including 3D printing. ECM can be produced using hydrogel materials.

[0009] As used herein, “extracellular matrix” and “ECM” refer to natural and synthetic ECM, as well as one or more constituent materials of ECM. For example, ECM may refer to naturally occurring ECM or ECM produced using synthetic techniques. ECM may also refer to one or more constituent materials of naturally occurring ECM, such as natural or synthetic collagen. In some cases, “ECM material” is used to refer to a specific material. ECM can be produced using techniques including 3D printing. ECM can be produced using hydrogel materials. ECM matrix materials such as collagen I, gelatin, elastin, and fibronectin can be functionalized with methacrylate groups to enable their incorporation into photocrosslinkable hydrogels. Incorporation of ECM materials into other materials and objects, such as 3D printed materials, can increase biocompatibility and enable cell adhesion and interaction within the materials and objects. The degree to which a material enables cell adhesion may vary based on the amount of ECM material, the availability of binding sites on or within the material, the surface charge of the material, the polarity of the material, and the mechanical properties of the material.

[0010] As used herein, the terms “object,” “construction,” and “article” may be used interchangeably and refer to items comprising the compositions of the present invention.

[0011] When used herein, the terms “comprising” or “comprises” are intended to mean that a composition and method includes the listed elements but does not exclude others. “Essentially consisting of” when used to define a composition and method means excluding other elements that are essentially important to the combination for the described purpose. Thus, a composition essentially consisting of the elements defined herein does not exclude other materials or steps that do not substantially affect the basic and novel features of the claimed invention. “Consists of” means excluding trace amounts or more of other components and substantial method steps. Embodiments defined by each of these transition terms are within the scope of the invention. Where an embodiment is defined by one of these terms (e.g., “comprising”), it should be understood that this disclosure also includes alternative embodiments. Some of these embodiments may include “essentially consisting of” and “consists of” for the aforementioned embodiment. As used herein, (meth)acrylate means methacrylate and / or acrylate. As used herein, unless otherwise specified, "molecular weight" means number-average molecular weight. Unless otherwise specified, percentages (%) refer to mass percentages. Please verify / correct based on the percentages used in the examples.

[0012] This application incorporates, by reference, the following documents in their entirety: (a) U.S. Provisional Application No. 63 / 185,293, filed on May 6, 2021, entitled “Use of Functionalized and Defunctionalized ECM, ECM Fragments, Peptides and Bioactive Components for the Preparation of Cell-Adhesive 3D Printed Objects,” (b) U.S. Provisional Application No. 63 / 185,302, filed on May 6, 2021, entitled “Modified 3D Printed Objects and Their Uses,” and the U.S. Application and / or PCT Application with the same title, filed on May 6, 2022, entitled “Photocurability Enhancement of 3D Printed Hydrogel Objects,” and the U.S. Application and / or PCT Application with the same title, filed on May 6, 2022, entitled “Hydrophilic Monomers (HDROPHILIC U.S. Provisional Application No. 63 / 185,300, filed May 6, 2021, entitled "Controlling the Size of 3D Printed Hydrogel Objects Using Monomers, Hydrophobic Monomers, and Crosslinkers," and U.S. Application and / or PCT Application (e) with the same title, filed May 6, 2022, entitled "Biomimetic 3D Printing and its Applications," and U.S. Application and / or PCT Application with the same title, filed May 6, 2022.

[0013] ECMs can be functionalized with methacrylate groups by substituting lysine residues on amine groups with methacrylate anhydride (MAA). The degree of methacrylate in an ECM can be defined by the proportion of available amine groups modified with MAA. A higher degree of methacrylate correlates with more MAA-modified amine groups and fewer free amine groups.

[0014] Embodiments of the present disclosure include a method for simultaneously producing two or more hydrogel constructs (e.g., tubular hydrogel constructs). In some embodiments, the method includes one or more of the following steps: providing a vat containing a bio-ink composition comprising one or more monomers and / or one or more polymers; curing a layer of hydrogel constructs (e.g., tubular hydrogel constructs) by applying electromagnetic radiation from an electromagnetic radiation source; and generating one or more additional layers of hydrogel constructs (e.g., tubular hydrogel constructs) by applying electromagnetic radiation from an electromagnetic radiation source one or more times.

[0015] Among 3D printing technologies, the most efficient are digital photolithography (DLP) and stereolithography (SLA). In 3D printers using DLP or SLA, ink material is layered on a container or spread on a sheet, and a predetermined area or surface of the ink is exposed to ultraviolet-visible (UV / Vis) light controlled by a digital micromirror device or rotating mirror. In the DLP method, additional layers are laid repeatedly or continuously until the desired 3D article is formed, with each layer being cured. The SLA method differs from the DLP method in that the ink is solidified by a line of radiation beam. Other methods of 3D printing can be found in 3D Printing Techniques and Processes by Michael Degnan, December 2017, Cavendish Square Publishing LLC, the disclosure of which is incorporated herein by reference. In some embodiments, the polymerization / curing of the hydrogel construct (e.g., a tubular hydrogel construct) is carried out at a vat temperature in the range of approximately 4°C to approximately 37°C, for example, at room temperature.

[0016] In some embodiments, the electromagnetic radiation is UV radiation. For example, UV radiation may be suitable for UV-initiated polymerization, and the composition may include, for example, a UV initiator or photoinitiator compound that reacts with and absorbs light in the range of 100 to 400 nm. Examples of photoinitiators include benzophenone, phenylbis(2,4,6-trimethylbenzoyl)phosphine oxide (BAPO), 2-hydroxy-2-methyl-1-phenyl-propan-1-one, 2-hydroxy-4'-(2-hydroxyxetoxy)-2-methylpropiophenone, 2,2'-azobis[2-methyl-n-(2-hydroxyethyl)propionamide], 2,2-dimethoxy-2-phenylacetophenone, diphenyl(2,4,6-trimethylbenzoyl)phosphine oxide, lithium phenyl(2,4,6-trimethylbenzoyl)phosphine (LAP), and ethyl(2,4,6-trimethylbenzoyl)phenylphosphine, as well as sodium phenyl-2,4,6-phenyl-2,4,6-trimethylbenzoyl phosphinate (NaP).

[0017] In some embodiments, ten or more hydrogel constructs (e.g., tubular hydrogel constructs) are manufactured simultaneously. For example, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, or more hydrogel constructs (e.g., tubular hydrogel constructs) can be manufactured simultaneously. The hydrogel constructs manufactured simultaneously (e.g., tubular hydrogel constructs) may be the same or different in shape from one another.

[0018] A vat containing a bio-ink composition may also contain other components, such as a liquid that is immiscible with the bio-ink. In some embodiments, the liquid that is immiscible with the bio-ink is selected from one or more hydrophobic substances. For example, in some embodiments, the immiscible liquid is selected from mineral oil, butyl acetate, petroleum ether, and mixtures thereof. In some embodiments, the mixture contains about 25% (w / w) to about 50% (w / w) of petroleum ether (e.g., about 25%, 30%, 35%, 40%, 45%, or 50% (w / w) of petroleum ether). In some embodiments, the mixture contains about 25% (w / w) to about 50% (w / w) of butyl acetate (e.g., about 25%, 30%, 35%, 40%, 45%, or 50% (w / w) of petroleum ether). In some embodiments, the mixture comprises mineral oil, for example, about 50% (w / w) to about 90% (w / w) of mineral oil (e.g., about 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, or 90% (w / w), or in between). In some embodiments, one or more hydrophobic substances comprises an oil having a viscosity of at least 5 cP at 25°C (e.g., at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25 cP, or in between) and / or an organic solvent having a boiling point above 100°C in STP (e.g., 105, 110, 120, 130, 140, 150, 160, 200, 250, 300, 350, 400, 450, 500, 550, 600, 650°C, or in between).

[0019] During printing, the hydrogel object can be immersed in the liquid for the entire printing period. This immersion can prevent dehydration and provide buoyancy. During the method embodied herein, the vat may be reloaded from time to time with additional components of the bio-ink and / or additional liquids that are immiscible with the bio-ink.

[0020] The bio-ink of this embodiment is not particularly limited and may, for example, be suitable for forming composite structures composed of one or more different polymer monomers. Hydrogel materials that can be used in the present invention, as well as methods for producing them, may be known to those skilled in the art. For example, hydrogels such as those described by Calo et al., European Polymer Journal, Vol. 65, April 2015, pp. 252-267, can be used. In some embodiments, the hydrogel structure comprises polymerized (meth)acrylate and / or (meth)acrylamide hydrogels. In some embodiments, the structure is polymerized poly(ethylene glycol) di(meth)acrylate, polymerized poly(ethylene glycol) di(meth)acrylamide, polymerized poly(ethylene glycol) (meth)acrylate / (methacrylate), poly(ethylene glycol)-block-poly(ε-caprolactone), polycaprolactone, polyvinyl alcohol, gelatin, methylcellulose, hydroxyethyl methylcellulose, hydroxypropyl methylcellulose, polyethylene oxide, polyacrylamide, polyacrylic acid, polymethacrylic acid, polyacrylic acid It contains polymers containing salts of polymethacrylate, salts of polymethacrylate, poly(2-hydroxyethyl methacrylate), polylactic acid, polyglycolic acid, polyvinyl alcohol, poly(methacrylate) anhydride, poly(acrylic) anhydride, polysebacic acid anhydride, and other polyanhydrides, collagen, poly(hyaluronic acid), hyaluronic acid-containing polymers and copolymers, polypeptides, dextran, dextran sulfate, chitosan, chitin, agarose gel, fibrin gel, soy-derived hydrogel, alginate-based hydrogel, poly(sodium alginate), hydroxypropyl acrylate (HPA), lithium phenyl 2,4,6-trimethylbenzoyl phosphinate (LAP), and combinations thereof.In some embodiments, the Mw of the hydrogel polymer is approximately 400Da, 500Da, 700Da, 800Da, 900Da, 1000Da, 1100Da, 1200Da, 1300Da, 1400Da, 1500Da, 1600Da, 1700Da, 1800Da, 1900Da, 2000Da, 2100Da, 2200Da, 2300Da, 2400Da, 2500Da, 2600Da, 2700Da, 2800Da, 2900Da, 3000Da, 3100Da, 3200Da, 3300Da, 3 400Da, 3500Da, 3600Da, 3700Da, 3800Da, 3900Da, 4000Da, 4100Da, 4200Da, 4300Da, 4400Da, 4500Da, 4600Da, 4700Da, 4800Da, 4900Da, 50 00Da, 5100Da, 5200Da, 5300Da, 5400Da, 5500Da, 5600Da, 5700Da, 5800Da, 5900Da, 6000Da, 6100Da, 6200Da, 6300Da, 6400Da, 6500Da, 7000 These are Da, 7500Da, 8000Da, 8500Da, 9000Da, 9500Da, 10000Da, 15000Da, or 20000Da. In some embodiments, the bio-ink may contain two or more hydrogel polymers, each having a different molecular weight. In some embodiments, the concentration of the hydrogel polymer in the bio-ink may be about 5% to about 50%, or about 10% to about 40%, or about 15% to about 30%, for example, about 20%, or any value or partial range within these ranges.

[0021] The dimensions of the hydrogel construct (e.g., tubular hydrogel construct) are not particularly limited and can be changed depending on the application. In some embodiments, the hydrogel construct (e.g., tubular hydrogel construct) comprises multiple layers having a thickness of 200 μm to 500 μm. In some embodiments, the tubular hydrogel construct has a wall thickness of up to about 1 mm, about 2 mm, about 3 mm, about 4 mm, or about 5 mm. For example, the wall thickness may be about 0.05 mm, about 0.1 mm, about 0.2 mm, about 0.3 mm, about 0.4 mm, about 0.5 mm, about 0.6 mm, about 0.7 mm, about 0.8 mm, about 0.9 mm, about 1 mm, about 1.1 mm, about 1.2 mm, about 1.3 mm, about 1.4 mm, or about 1.5 mm (or in the range in between). In some embodiments, the tubular hydrogel construct has a maximum length of approximately 250 mm (for example, approximately 10 mm, 20 mm, 30 mm, 40 mm, 50 mm, 60 mm, 70 mm, 80 mm, 90 mm, 100 mm, 110 mm, 120 mm, 130 mm, 140 mm, 150 mm, 160 mm, 170 mm, 180 mm, 190 mm, 200 mm, 210 mm, 220 mm, 230 mm, 240 mm, or 250 mm (or a range in between)).

[0022] In some embodiments, the bio-ink contains a poly(ethylene glycol) di-(meth)acrylate polymer. In some embodiments, the poly(ethylene glycol) di-(meth)acrylate polymer is present in concentrations of about 400 to about 20,000 (e.g., about 400, 500, 100, 1500, 2000, 2500, 3000, 3500, 4000, 4500, 5000, 5500, 6000, 6500, 7000, 7500, 8000, 8500, 9000, 9500). The bio-ink has a weight-average molecular weight (Mw) of 10000, 10500, 11000, 11500, 12000, 12500, 13000, 13500, 14000, 14500, 15000, 15500, 16000, 16500, 17000, 17500, 18000, 18500, 19000, 19500, or 2000, or a range in between. In some embodiments, the bio-ink may contain two or more poly(ethylene glycol) di-(meth)acrylate polymers, each having a different molecular weight. In some embodiments, the concentration of poly(ethylene glycol) di-(meth)acrylate polymer in the bio-ink may be about 5% to about 50%, or about 10% to about 40%, or about 15% to about 30%, for example, about 20%, or any value or partial range within these ranges.

[0023] In some embodiments, the bio-ink comprises one or more hydroxy C1-2 alkyl (meth)acrylates, poly(alkylene oxide) alkyl ether (meth)acrylates, N-hydroxy C1-2 alkyl (meth)acrylamides, a polyethylene glycol methyl ether acrylate (PEGMEA), poly(ethylene glycol) methyl ether methacrylate, poly(propylene glycol) methyl ether methacrylate, poly(propylene glycol) methyl ether methacrylate, hydroxyethyl acrylate (HEA), N-hydroxyethyl acrylamide (HEA), N-hydroxyethyl acrylamide (HEAA), hydroxyethyl methacrylate, hydroxypropyl acrylate (HPA 3-hydroxypropyl acrylate and / or 2-hydroxypropyl acrylate), hydroxypropyl methacrylate, hydroxybutyl acrylate (HBA), hydroxybutyl methacrylate, poly(alkylene oxide) di(meth)acrylates, diethylene glycol di(meth)acrylates, tetraethylene glycol di(meth)acrylates, N,N´-methylenebis(acrylamide), (poly)lactic acid di(meth)acrylates, (poly)glycolic acid di(meth)acrylates, (poly)lactic acid-co-glycolide di(meth)acrylates, (poly)caprolactone di(meth)acrylates, (poly)dioxanone di(meth)acrylates, (poly)fumarate di(meth)acrylates, (carboxy)(methyl)cellulose di(meth)acrylates, hyaluronic acid di(meth)acrylates, heparan sulfate di(meth)acrylates, dextran di(meth)acrylates, alginate di(meth)acrylates, pectin di(meth)acrylates, or collagen di(meth)acrylates or mixtures thereof. In some embodiments, the bio-ink can comprise one or more poly(ethylene glycol) di-(meth)acrylate polymers, and one or more additional polymers such as an alginate-based hydrogel. In some embodiments, the concentration of one or more additional polymers in the bio-ink can be from about 0.5% to about 10%, or from about 1% to about 8%, or from about 1.5% to about 5%, such as about 2.5%, or any value or sub-range within these ranges.

[0024] In some embodiments, the bio-ink further comprises a photoinitiator. The photoinitiator is not particularly mimicked, and examples of suitable photoinitiators include lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP), trimethylbenzoyl-based photoinitiators, diphenyl(2,4,6-trimethylbenzoyl)phosphine oxide (TPO nanoparticles), photoinitiators of the Irgacure class, ruthenium, riboflavin, phenyl-2,4,6 phenyl-2,4,6-trimethylbenzoylphosphinate sodium (NaP), or mixtures thereof. In some embodiments, the concentration of the photoinitiator in the bio-ink can be from about 0.1% to about 5%, or from about 0.2% to about 3%, or from about 0.5% to about 2%, such as about 1%, or any value or sub-range within these ranges.

[0025] In some embodiments, the bio-ink further comprises a solvent such as water. In some embodiments, the water is deionized. In certain embodiments, the bio-ink comprises from about 50 to about 90% DI water (e.g., about 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90% DI water, or a range therebetween).

[0026] In some embodiments, the bio-ink further comprises UV dyes, proteins, peptides, biopharmaceuticals, pharmaceutical compounds, and / or extracellular matrix materials. In some embodiments, the peptide is selected from RGD, KQAGDV, YIGSR, REDV, IKVAV, RNIAEIIKDI, KHIFSDDSSE, VPGIG, FHRRIKA, KRSR, APGL, VRN, AAAAAAAAA, GGLGPAGGK, GVPGI, LPETG(G)n, and IEGR. Other examples of suitable additional components include ECM or ECM-like substances such as protease-sensitive amino acid sequences. The proteases can be selected from Arg-C proteinase, Asp-N endopeptidase, BNPS-skatole, caspase 1-10, chymotrypsin high specificity (from C-term to [FYW], not before P), chymotrypsin low specificity (up to C-term [FYWML], not before P), clostripine (clostridiopeptidase B), CNBr, enterokinase, factor Xa, formic acid, glutamyl endopeptidase, granzyme B, hydroxylamine, iodosobenzoic acid, LysC, neutrophil elastase, NTCB (2-nitro-5-thiocyanobenzoic acid), pepsin, proline-endopeptidase, proteinase K, staphylococcal peptidase I, thermolysin, thrombin, and trypsin. In some embodiments, the concentration of UV dye in the bio-ink may be about 0.02% to about 2%, or about 0.03% to about 1.5%, or about 0.05% to about 1%, for example, about 0.2%, or any value or sub-range within these ranges.

[0027] In some embodiments, the hydrogel scaffold remains immersed or immersed (or partially immersed) in a liquid that is immiscible with the bio-ink during the process. In some embodiments, the hydrogel scaffold is immersed in a container. In some embodiments, the hydrogel scaffold is immersed in a container. In some embodiments, the method further includes adding a liquid that is miscible with bio-ink to replace at least a portion of the bio-ink that is consumed or otherwise lost during printing. In some embodiments, a liquid that is miscible with the bio-ink is placed inside the container to prevent the bio-ink from evaporating.

[0028] In some embodiments, the hydrogel constructs (e.g., tubular hydrogel constructs) are substantially the same shape, size, and / or have the same relative dimensions as the organ or organ fragment. For example, the organ or organ fragment may include blood vessels, trachea, bronchi, esophagus, ureter, renal tubule, bile duct, renal tubule, renal tubule, bile duct, hepatic duct, nerve conduit, CSF shunt, larynx, or pharynx. For example, in some embodiments, the hydrogel constructs (e.g., tubular hydrogel constructs) described herein are formed into structures that mimic or replicate a portion of the lung structure, for example, by using 3D printing technology. Using the hydrogel constructs (e.g., tubular hydrogel constructs), a scaffold for cell adhesion and proliferation can be formed, resulting in structures having one or more desired properties of an organ, such as a structure that can perform the gas exchange function of the lung. These objects may contain hydrogels. The organ or portion of an organ may, in a preferred embodiment, be a human lung.

[0029] In some embodiments, the tubular hydrogel construct comprises a hemodialysis graft. In some embodiments, the tubular hydrogel construct allows for endothelialization of the lumen of the tubular hydrogel construct and / or cytosis of the outer surface of the tubular hydrogel construct. In some embodiments, the lumen of the tubular hydrogel construct comprises a patterned surface. In some embodiments, the patterned surface comprises patterning that allows for unidirectional flow through the tube. In some embodiments, the tubular hydrogel construct comprises one or more branches. In some embodiments, the bio-ink may be exposed to electromagnetic radiation, such as UV radiation, to generate a tubular structure. In some embodiments, the intensity of the electromagnetic radiation, such as UV radiation, may be 1 mW / cm² to 100 mW / cm², or 2 mW / cm² to 80 mW / cm², or 5 mW / cm to 50 mW / cm, or any value or sub-range within these ranges. In some embodiments, the exposure time to electromagnetic radiation, such as UV radiation, may be 0.1 seconds to 100 seconds, or 0.1 seconds to 50 seconds, or 0.2 seconds to 30 seconds, or any value or sub-range within these ranges. In some embodiments, a tubular hydrogel structure can be attached to a pump, such as a peristaltic pump. For such attachment, an adhesive, such as a cyanoacrylate adhesive, may be used. The examples described herein are illustrative and not intended to limit the present invention. Various embodiments of the present invention have been described in accordance with the present invention. Many modifications and changes can be made to the techniques described and illustrated herein without departing from the spirit and scope of the present invention. Therefore, it should be understood that the examples are illustrative and not intended to limit the scope of the present invention. [Examples]

[0030] Example 1 3D printing using polyethylene glycol diacrylate (PEG-DA) A PEG-DA 6k solution was prepared using PEG-DA 6k; LAP; UV386A (a UV dye from QCR Solutions Corp.) and DI water. The tube was 3D printed using PED-GA 6k solution. The printed tube model had an outer diameter (OD) of 12.5 mm, an inner diameter (ID) of 7.5 mm, and a height of 10 mm (see Figure 2). Figure 3 is a photograph of a printed tube. Example 2 A pair of 5mm long inner diameter tubes with wall thicknesses of 1.5mm and 2mm were printed. Based on this, numerous 5mm ID tubes with the desired two wall thicknesses were printed. Figure 4 is a photograph of a printed tube. Example 3 Short (3-10 cm) 3D printed tubes were secured at both ends to the tube and attached to the peristaltic pump. The attachment was achieved using medical-grade mesh and cyanoacrylate adhesive. After curing, the fluid was allowed to pass through the tubes for as long as possible until leakage was observed. Seven out of eight tubes printed successfully. The unsuccessful tubes printed as several spirals. After printing was complete, the tubes were rinsed with tap water for 5 minutes and then immersed in phosphate-buffered saline (PBS) for 45 minutes. I poured the excess ink material back into the amber jug. Next, two of the tubes were tested for installation. The first tube was successfully connected to a tube with an outer diameter of 14 mm using a DERMABOND® PRINEO® skin closure system. Instructions were followed in writing. Both sides. A curing time of 60 seconds was used. Subsequently, the sample was accidentally torqued and ruptured in the middle. The ends were left attached. The sample was filled with tap water and stained red with food coloring. The second tube was attached using the DERMABOND® PRINEO® skin closure system, following the modified instructions. This time, two wraps of polyethylene (PE) mesh were placed on each side. The wraps were not completely overlapping but were staggered to provide ample coverage over the joint between the sample end and the tube. This configuration was run with a CP peristaltic pump at maximum flow rate over the weekend. The samples were immersed in DI water to prevent drying over the weekend, and leakage could be evaluated by observing changes in the color of the bath water. The connections remained intact, and the flow rate was reduced from 170 ml / min to 70 ml / min for about 3 hours. The pump motor light went out, indicating the power was cut off. Next, the flow rate was increased back to 120 ml / min to give the connections a greater challenge. The sample remained running continuously throughout the day without any leakage. The water appeared to be stained pink, as if stained water had diffused through the sample. The sample continued running overnight. The sample connection remained intact until the following morning. The water bath still appeared pinkish, but there was no apparent leakage. Example 4 We attempted to connect previously printed tubing to a modified connector. The previous connector model did not have room for screw coupling around the swollen printed tubing. The design has been modified to accommodate the increased wall thickness due to swelling. Furthermore, once the tubing connector piece was in place, the tubing connector piece was further sealed. material: A previously printed 2.5 cm tube section. The sample, stored in PBS in a dark drawer. Loctite Pro Line Marine Fast Cure Adhesive Sealant. E-ZFuse Tape - Self-Fusing, Waterproof, Airtight Seal - Black Silicone Tape. FiberFix - Flex Patch. Fast Fuse DAP. Cyanoacrylate Adhesive - Rapid Fuse. Rubber Cement. Methods and results: In each case, excess buffer was gently removed from the tube surface. The product was applied as indicated in the provided instructions. It was pressed into the newly designed tube fitting. The sample did not stay in place; it continued to slide down. After holding it in place for a short time, the tube was split. Applications of silicone tape - EZ fuse - Figure 5A The initial application to the printed structure was unsuccessful. The tape adhered very well to itself and, in fact, appeared to form a seal in later attempts. It is difficult to perfectly conform to the shape of the container without applying significant stretching and pressure that would cause our printed structure to crack. It is also impossible to see what is happening underneath through the tape. However, it is one of the better candidates and should not be ignored. Fiber Flex Patch - Figure 5B This is a very difficult job. It sticks to gloves and other parts of the glove. Like the black tape, it didn't adhere to the sample. The sample quickly slipped out from under the tape. Transparent silicone sealant It was applied generously to the surface and filled in the cracks in the tube. Curing time: ~24 hours. It was not set to be useful. It was pulled cleanly from the material surface. Rapid-fuse foam It appears to stick fragments together, but it's used very haphazardly, making it difficult to control the application area. It never solidified to a usable state. Cyanoacrylate adhesive Figure 5C-D Two pieces of scrap were glued together. After about 30 seconds, they clearly bonded. The tube was then glued to a 15 mL Falcon tube. Initially, it was thought that the connection had been successfully sealed, but later it was realized that the seal might have been due only to the press-fitting. See Figure 5C. The adhesive color did not adhere to the printed object. No leakage was observed. To address this, a surgical mesh was applied and adhesive was layered on top of the mesh (basically, replacing the counter-adhesive with Dermabond as described in the surgical instructions) (see Figure 5D). This structure was attached to the end of a tubing section installed inside a peristaltic pump. Water could be flowed through at a considerable flow rate. The pump was attached to both ends, and an attempt was made to circulate the fluid continuously within the component. The initial attachment appeared fine, but when the printed portion bent slightly, it completely split in two. It was extremely brittle and could not withstand the displacement. This may be due to aging of the printed piece or the inherent brittleness of the material. I tried to reattach a new piece. There may have been a small gap in the mesh material. I tried to use the smallest mesh possible to save the remaining small amount. I was able to start the pumping process, but the leak started rapidly at the interface between the tube and the printed piece. After circulating a small amount of water, I attempted to remove the printed tube from the plastic tube. The adhesive and mesh held the printed piece so tightly that it broke rather than simply slipped out of the tube. conclusion 1. Surgical mesh combined with cyanoacrylate adhesive appears to be a viable option for attaching printed materials to the pump. 2. Silicone tape can be a backup approach.

[0031] As used herein, the singular terms “a,” “an,” and “the” can refer to multiple objects unless the context clearly indicates otherwise. Thus, for example, a reference to an object can refer to multiple objects unless the context clearly indicates otherwise.

[0032] When used herein, the terms “substantially” and “about” are used to describe and explain minor variations. When used in conjunction with an event or situation, the terms may refer to instances in which the event or situation occurs exactly, as well as instances in which the event or situation occurs in a close approximation. When used with a numerical value, the terms may refer to a range of variation of that value of ±10% or less, for example, ±5%, ±4%, ±3%, ±2%, ±1%, ±0.5%, ±0.1%, or ±0.05%. When referring to a first numerical value as “substantially” or “about” the same as a second numerical value, the terms may refer to the first numerical value being within a range of variation of the second numerical value of ±10% or less, for example, ±5%, ±4%, ±3%, ±2%, ±1%, ±0.5%, ±0.1%, or ±0.05%.

[0033] While this disclosure has been described with reference to its specific embodiments, it will be understood by those skilled in the art that various modifications may be made and equivalents may be substituted without departing from the true spirit and scope of this disclosure as defined by the appended claims. In addition, many modifications may be made to adapt specific situations, materials, composition of substances, methods, operations, or behaviors to the purpose, spirit, and scope of this disclosure. All such modifications are intended to be within the scope of the claims appended herein. In particular, while specific methods have been described with reference to specific actions performed in a specific order, it will be understood that these actions may be combined, subdivided, or rearranged without departing from the teachings of this disclosure to form equivalent methods. Therefore, unless specifically indicated herein, the order and grouping of operations are not limitations of this disclosure. All publications, patent applications and patents referenced herein are incorporated herein in their entirety by standard. This disclosure further includes the following aspects: 《Aspect 1》 A method for simultaneously producing two or more tubular hydrogel constructs: To provide a vat containing a bio-ink composition containing one or more monomers and / or one or more polymers, Applying electromagnetic radiation from an electromagnetic radiation source to harden the layers of a tubular hydrogel structure, and Applying electromagnetic radiation from an electromagnetic radiation source one or more times to generate one or more additional layers of a tubular hydrogel structure, A method that includes this. 《Aspect 2》 The bio-ink composition comprising a monomer, according to the method according to embodiment 1. 《Aspect 3》 The bio-ink composition comprises one or more polymers, according to the method according to embodiment 1. Appearance 4 The method according to any one of embodiments 1 to 3, wherein the electromagnetic radiation is UV radiation. 《Aspect 5》 The method according to any one of embodiments 1 to 4, wherein 10 or more tubular hydrogel constructs are manufactured simultaneously. 《Aspect 6》 The method according to any one of embodiments 1 to 5, wherein the vat further comprises a liquid that is immiscible with the bio-ink. Appearance 7 The bio-ink composition comprises a poly(ethylene glycol) di-(meth)acrylate polymer, according to any one of embodiments 1 and 3 to 6. 《Aspect 8》 The bio-ink composition according to any one of embodiments 1 to 7, comprising at least one photoinitiator. 《Aspect 9》 The bio-ink composition comprises DI water, according to any one of embodiments 1 to 8. 《Aspect 10》 The method according to any one of embodiments 1 to 9, further comprising a UV dye, a protein, a peptide, a biopharmaceutical, a pharmaceutical compound, and / or an extracellular matrix material. 《Aspect 11》 The method according to any one of embodiments 1 to 10, wherein the tubular hydrogel constructs are substantially the same in shape and size and / or have the same relative dimensions as the organ or organ fragment. 《Aspect 12》 The method according to embodiment 11, wherein the organ or fragment of the organ includes a blood vessel, trachea, bronchi, esophagus, ureter, renal tubule, bile duct, renal tubule, renal tubule, bile duct, hepatic duct, nerve conduit, CSF shunt, larynx, or pharynx. 《Aspect 13》 The method according to embodiment 12, wherein the blood vessel includes a pulmonary artery, renal artery, coronary artery, peripheral artery, pulmonary vein, or renal vein. Appearance 14 The method according to any one of embodiments 1 to 13, wherein the tubular hydrogel structure includes a hemodialysis graft. 《Aspect 15》 The method according to any one of embodiments 1 to 14, wherein the tubular hydrogel structure enables endothelialization of the lumen of the tubular hydrogel structure and / or cellularization of the outer surface of the tubular hydrogel structure. 《Aspect 16》 The method according to any one of embodiments 1 to 15, wherein the lumen of the tubular hydrogel structure includes a patterned surface. 《Aspect 17》 The method according to embodiment 16, wherein the patterned surface includes patterning that allows for unidirectional flow through the tube. 《Aspect 18》 The method according to any one of embodiments 1 to 15, wherein the tubular hydrogel structure includes one or more branches. 《Aspect 19》 The hydrogel constructs include polymerized poly(ethylene glycol) di(meth)acrylate, polymerized poly(ethylene glycol) di(meth)acrylamide, polymerized poly(ethylene glycol) (meth)acrylate / (methacrylamide), poly(ethylene glycol)-block-poly(ε-caprolactone), polycaprolactone, polyvinyl alcohol, gelatin, methylcellulose, hydroxyethyl methylcellulose, hydroxypropyl methylcellulose, polyethylene oxide, polyacrylamide, polyacrylic acid, polymethacrylic acid, and polyacrylic acid. The method according to any one of embodiments 1 to 18, selected from the group comprising salts of polymethacrylate, salts of polymethacrylate, poly(2-hydroxyethyl methacrylate), polylactic acid, polyglycolic acid, polyvinyl alcohol, polymethacrylate anhydride, polyacrylic anhydride, polysebacate anhydride, and other polyanhydrides, collagen, poly(hyaluronic acid), hyaluronic acid-containing polymers and copolymers, polypeptides, dextran, dextran sulfate, chitosan, chitin, agarose gel, fibrin gel, soy-derived hydrogel, alginate-based hydrogel, poly(sodium alginate), hydroxypropyl acrylate (HPA), lithium phenyl 2,4,6-trimethylbenzoyl phosphinate (LAP), and combinations thereof. 《Appearance 20》 A batch of tubular hydrogel constructs produced by the process described in Embodiment 1. 《Aspect 21》 The batch according to embodiment 20 includes different shapes of the tubular hydrogel constructs.

[0034] While this disclosure has been described with reference to its specific embodiments, it will be understood by those skilled in the art that various modifications may be made and equivalents may be substituted without departing from the true spirit and scope of this disclosure as defined by the appended claims. In addition, many modifications may be made to adapt specific situations, materials, composition of substances, methods, operations, or behaviors to the purpose, spirit, and scope of this disclosure. All such modifications are intended to be within the scope of the claims appended herein. In particular, while specific methods have been described with reference to specific actions performed in a specific order, it will be understood that these actions may be combined, subdivided, or rearranged without departing from the teachings of this disclosure to form equivalent methods. Therefore, unless specifically indicated herein, the order and grouping of operations are not limitations of this disclosure. All publications, patent applications and patents referenced herein are incorporated herein in their entirety by standard.

Claims

1. A method for simultaneously producing two or more tubular hydrogel constructs: To provide a vat containing a bio-ink composition containing one or more monomers and / or one or more polymers, Applying electromagnetic radiation from an electromagnetic radiation source to harden the layers of a tubular hydrogel structure, and Applying electromagnetic radiation from an electromagnetic radiation source one or more times to generate one or more additional layers of a tubular hydrogel structure, It includes, The tubular hydrogel structures are independent of each other and / or not connected to each other. The aforementioned tubular hydrogel structure has a wall thickness of 0.1 mm to 5 mm. The tubular hydrogel structure has a length of 5 mm to 250 mm, and The bio-ink composition comprises a poly(ethylene glycol) di(meth)acrylate polymer, and each layer of each of the tubular hydrogel structures comprises a cured poly(ethylene glycol) di(meth)acrylate polymer. method.

2. The method according to claim 1, wherein the electromagnetic radiation is UV radiation.

3. The method according to claim 1, wherein 10 or more tubular hydrogel constructs are produced simultaneously.

4. The method according to claim 1, wherein the vat further comprises a liquid that is immiscible with the bio-ink.

5. The method according to claim 1, wherein the bio-ink composition comprises at least one photoinitiator.

6. The bio-ink composition according to claim 1, wherein the bio-ink composition comprises DI water.

7. The method according to claim 1, wherein the bio-ink composition further comprises a UV dye, a protein, a peptide, a biopharmaceutical, a pharmaceutical compound, and / or an extracellular matrix material.

8. The method according to claim 1, wherein the tubular hydrogel constructs are substantially the same shape and size and / or have the same relative dimensions as the organ or organ fragment.

9. The method according to claim 8, wherein the organ or fragment of the organ includes a blood vessel, trachea, bronchi, esophagus, ureter, renal tubule, bile duct, renal tubule, renal tubule, bile duct, hepatic duct, nerve conduit, CSF shunt, larynx, or pharynx.

10. The method according to claim 9, wherein the blood vessel includes a pulmonary artery, renal artery, coronary artery, peripheral artery, pulmonary vein, or renal vein.

11. The method according to claim 1, wherein the tubular hydrogel structure includes a hemodialysis graft.

12. The method according to claim 1, wherein the tubular hydrogel structure enables endothelialization of the lumen of the tubular hydrogel structure and / or cellularization of the outer surface of the tubular hydrogel structure.

13. The method according to claim 1, wherein the lumen of the tubular hydrogel structure includes a patterned surface.

14. The method according to claim 13, wherein the patterned surface includes patterning that allows for unidirectional flow through the tube.

15. The method according to claim 1, wherein the tubular hydrogel structure includes one or more branches.

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