Biofluid purification using biocompatible membranes

Abstract

To provide biological fluid purification using a biocompatible membrane.SOLUTION: An apparatus for integrated adaptive biological blood purification includes a function unit. The function unit is provided with: a membrane including a pipe surface and a filtration surface; a blood vessel channel system which is adhered to the blood vessel surface of the membrane and includes a first lumen space in fluid communication with the blood vessel surface of the membrane, a first end configured to be in fluid communication with a fluid supply part and to contact the same, and a second end configured to be in fluid communication with a filtration fluid outlet and to contact the same; and a filtration channel system which is adhered to the filtration surface of the membrane and includes a second lumen space in fluid communication with the filtration surface of the membrane and a third end configured to be in fluid communication with a filtrate outlet and to contact the same.SELECTED DRAWING: Figure 11

Description

[Background technology]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of U.S. Provisional Patent Application No. 62 / 812,239, filed February 28, 2019, the entire contents of which are incorporated herein by reference.

[0002] (Background technology) Current methods of artificial blood purification require the use of dialysate filters (in conventional hemodialysis), dialysis fluids (in both conventional dialysis and peritoneal dialysis), substitution fluids (in hemofiltration and hemodiafiltration), anticoagulants to prevent activation of the coagulation cascade by the filtration material (conventional hemodialysis), blood pumps to generate the necessary flow and hydrostatic gradients to enable filtration and dialysis (conventional hemodialysis), and, in some applications, the use of adsorbent materials to bind toxin molecules.

[0003] Currently, no technology exists for creating a blood purification system that can function continuously in a fully integrated, preferably implantable, device without the need for these components. Furthermore, bioengineering human-scale tissue and organ transplants that can address the above-mentioned problems requires the creation of matrices that provide the functional architecture necessary to enable each cell to perform its specific role and generate functional tissue constructs. Tissues and organs containing one or more epithelial structures (digestive, endocrine, nervous, lymphatic, integumentary, reproductive, respiratory, sensory, urinary, and circulatory) depend on the presence of a thin basement membrane, which allows functions such as filtration of fluids (kidneys, eyes, lymphatic, brain), diffusion of gases (lungs), secretion and absorption of electrolytes and other molecules (kidneys, intestines, liver, intestinal tissue), and diffusion of hormones (pancreas, pituitary, adrenal glands) from one lumen or compartment to another. In many cases, this basement membrane must be <1 um or <10 um thick to allow function (Rayat et al., Indian journal of pathology & microbiology 48, 453-458 (2005) and Kopf et al., Nature Immunology 16,36-44(2015)).

[0004] Currently, no technology exists to produce membranes composed of biological and / or natural matrix materials with the tunable micro- or nanoporosity and physiological thickness necessary to enable such functionality in biological tissue scaffolds, such as for artificial tissues or organs for hemodialysis. Summary of the Invention [Means for solving the problem]

[0005] The work described herein demonstrates the fabrication of biocompatible membranes with tunable thickness and pore size.

[0006] Additionally, the work herein demonstrates the design, fabrication, and use of an integrated adaptive biological blood purification (IABBP) device with a biocompatible membrane. The purpose of this device is to remove toxins and excess water from a patient's circulation, replacing the body's own purification systems, such as the kidneys and liver. In contrast to currently available systems, the function of this device does not rely on the use of external dialysate fluids (as used in hemodialysis and peritoneal dialysis), substitution fluids (as used in hemofiltration and hemodiafiltration), or adsorbent materials (as used in ambulatory dialysis and extracorporeal liver replacement). The IABBP device is connected to the patient's vascular system via direct connections to arteries and veins. The patient's cardiovascular system is used to perfuse the IABBP device with sufficient blood flow to enable its function. This can be achieved without the use of an additional mechanical pump. However, in some cases, a pump may be used to increase IABBP device function.

[0007] Connection to the patient's vascular system is established via an arteriovenous shunt or cannulation of the vena cava, or via direct anastomosis of the IABBP device vascular conduit to the patient's vasculature. The IABBP device can be used extracorporeally or implanted in the patient similar to a donor organ. The IABBP device can be used for continuous or intermittent treatment. The IABBP device produces filtrate that drains into an extracorporeal collection system or is connected to the patient's bladder via a surgical anastomosis.

[0008] IABBPs can be manufactured by creating a scaffold, which is then repopulated with cells and cultured to mature the resulting tissue for function. IABBP scaffolds are created by combining multiple functional units. Each functional unit is created by fabricating a biocompatible extracellular matrix membrane. This membrane can be porous or non-porous depending on the functional needs. Sacrificial materials are then printed onto the membrane on both sides. The entire membrane is then embedded in the matrix material. The sacrificial material is then removed, resulting in two channel systems separated by the membrane. Several functional units can be stacked to create a scaffold large enough to meet the needs of the human body. Cells can then be repopulated into both channel systems to generate functional, living tissue.

[0009] Some aspects of the present disclosure provide an apparatus for integrated adaptive biological blood purification, the apparatus comprising: a functional unit, the functional unit comprising: a membrane having a tubular surface and a filtering surface; a vascular channel system adhered to the vascular surface of the membrane and including a first luminal space in fluid communication with the vascular surface of the membrane, the vascular channel system comprising a first end configured to fluidly connect to a fluid supply and a second end configured to fluidly connect to a filtrate fluid outlet; and a filtration channel system adhered to the filtering surface of the membrane and including a second luminal space in fluid communication with the filtering surface of the membrane, the third end configured to fluidly connect to a filtrate outlet. and a system, wherein the vascular channel system and the filtration channel system are in fluid communication with each other across the membrane, and the functional unit further comprises at least three segments, the at least three segments including at least a filtration segment configured to provide ultrafiltration to produce a first ultrafiltrate, a tubular segment connected thereto configured to provide reabsorption to produce a second ultrafiltrate, and a conduit segment connected thereto configured to provide concentration to produce a tertiary ultrafiltrate, and the membrane comprises three membrane segments including at least a filtration membrane segment, a tubular membrane segment, and a conduit membrane segment.

[0010] In some embodiments, the membrane comprises a biocompatible extracellular matrix membrane separating the vascular channel system from the filtration channel system, the biocompatible extracellular matrix membrane being embedded in a matrix material. In some embodiments, the biocompatible extracellular matrix membrane comprises a collagen membrane having a thickness of 0.1 to 10 micrometers (0.1 to 10 μm) that supports cell adhesion on both the vascular and filtration surfaces of the collagen membrane. In some embodiments, the matrix material is gelatin. In some embodiments, the biocompatible extracellular matrix membrane comprises fibers, nanofibers, or other longitudinal elements. In some embodiments, the fibers, nanofibers, or other longitudinal elements increase or modify the mechanical strength of the membrane. In some embodiments, the fibers, nanofibers, or other longitudinal elements are uniformly distributed throughout the membrane, providing homogeneous mechanical reinforcement. In some embodiments, the fibers, nanofibers, or other longitudinal elements are non-uniformly distributed in the membrane, providing heterogeneous mechanical reinforcement.

[0011] In some embodiments, the filtration membrane segment allows for the production of filtrate from a first luminal space in the vascular channel system to a second luminal space of the filtration channel system, the tubular membrane segment allows for the exchange and / or diffusion of solutes and water between the vascular channel system and the filtration channel system, and the conduit membrane segment allows for the transport of water and solutes from the filtration channel system to the vascular channel system.

[0012] In some embodiments, the functional unit comprises at least one biological fluid inflow conduit fluidly connected to a first end and a first luminal space of the vascular channel system, and at least one biological fluid outflow conduit fluidly connected to a second end and a first luminal space of the vascular channel system; the functional unit comprises at least one filtrate outflow conduit fluidly connected to a third end and a second luminal space of the filtration channel system; and the functional unit further comprises one or more blood vessel segment conduits interconnecting the filtration segments, tubular segments, and conduit segments of the vascular channel system and the first luminal space, and one or more filtration segment conduits interconnecting the filtration segments, tubular segments, and conduit segments of the filtration channel system and the second luminal space.

[0013] In some embodiments, at least one biofluid inflow conduit is in fluid communication with an arterial conduit, at least one biofluid outflow conduit is in fluid communication with a vascular conduit, and at least one filtrate outflow conduit is in fluid communication with a drain conduit. In some embodiments, the device produces ultrafiltrate that is drained using the drain conduit into an extracorporeal collection system or into the patient's bladder. In some embodiments, the first luminal space and the second luminal space are embedded in the scaffold.

[0014] In some embodiments, the filtration segment of the vascular channel system comprises a vascular channel wall lined with endothelial cells selected from primary human glomerular endothelial cells, induced pluripotent stem cell (iPSC)-derived endothelial cells, and / or human umbilical endothelial cells. In some embodiments, the tubular segment of the vascular channel system comprises a vascular channel wall lined with endothelial cells selected from primary human peritubular capillary endothelial cells, iPSC-derived endothelial cells, and / or human umbilical endothelial cells. In some embodiments, the conduit segment of the vascular channel system comprises a vascular channel wall lined with endothelial cells selected from primary human renal medullary endothelial cells, iPSC-derived endothelial cells, and / or human umbilical endothelial cells. In some embodiments, the filtration segment of the filtration channel system comprises a filtration channel wall lined with epithelial cells selected from primary human podocytes and / or human iPSC-derived podocytes. In some embodiments, the tubular segments of the filtration channel system comprise filtration channel walls lined with epithelial cells selected from primary human renal tubular epithelial cells and / or iPSC-derived renal tubular epithelial cells.

[0015] In some embodiments, the tubular segments of the filtration channel system comprise filtration channel walls lined with epithelial cells selected from primary human renal tubular epithelial cells and / or iPSC-derived renal tubular epithelial cells.

[0016] In some embodiments, the device comprises a plurality of functional units, including a functional unit and an additional functional unit of the same configuration, each functional unit of the plurality of functional units having a first end and a first lumen space of a vascular channel system in fluid communication with at least one biofluid inflow conduit, a second end and a first lumen space of the vascular channel system in fluid communication with at least one biofluid outflow conduit, and a third end and a second lumen space of a filtration channel system in fluid communication with a filtrate outflow conduit, wherein each first end of the plurality of functional units individually connects in parallel to one of a plurality of manifold ports of the at least one biofluid inflow conduit, each second end of the plurality of functional units individually connects in parallel to one of a plurality of manifold ports of the at least one biofluid outflow conduit, and each third end of the plurality of functional units individually connects in parallel to one of a plurality of manifold ports of the at least one filtrate outflow conduit. In some embodiments, the device comprises a plurality of functional units, including a functional unit and an additional functional unit of the same configuration, stacked in parallel layers of functional units.

[0017] In some embodiments, the at least one biological fluid inflow conduit comprises a blood inlet conduit configured to transport blood inflow, and the at least one biological fluid outflow conduit comprises a blood outflow conduit configured to transport blood outflow, and parallel layers of functional units configured for biological blood purification.

[0018] In some embodiments, the filtration segment of the filtration channel system is configured to provide ultrafiltration to produce a primary ultrafiltrate, the capillary segment is configured to provide reabsorption by absorption of solutes and water to produce a secondary ultrafiltrate stream, and the conduit segment is configured to provide concentration by absorption of water to produce a tertiary ultrafiltrate stream.

[0019] In some embodiments, the device is configured for extracorporeal operation in a sterile, heated enclosure. In some embodiments, the device is disposed within a capsule and sized and configured for placement within the human body to replace or augment kidney or liver function. In some embodiments, the membrane comprises a porous membrane including pores arranged to interconnect the vascular surface and the filtering surface. In some embodiments, the pores have a diameter of 1 μm to 15 μm.

[0020] Some aspects of the present disclosure relate to methods of treating a patient with inadequate renal or hepatic function, including fluidly connecting a device described herein to the patient's circulatory system and passing the patient's blood through the vascular channel system of the device from a filtering member segment to a tubular member segment, from the tubular member segment to a conduit member segment, and from the conduit member segment back into the patient's circulatory system. In some embodiments, the device is implanted in the patient. In some embodiments, ultrafiltrate produced by the device is delivered extracorporeally to the patient. In some embodiments, ultrafiltrate produced by the device is delivered to the patient's bladder.

[0021] In some embodiments, the device is external to the patient.

[0022] Some aspects of the present disclosure relate to methods of manufacturing the devices described herein, including providing a plurality of membranes having sacrificial material in the form of a vascular channel network on a vascular surface and sacrificial material in the form of a filtration channel system on a filtration surface; immersing the plurality of membranes in a solution containing a scaffold material (i.e., membrane solution); gelling the scaffold material; and removing the sacrificial material, thereby forming the luminal spaces of the vascular channel system and the filtration channel system.

[0023] In some embodiments, the multiple membranes are each produced by chemical or physical thin-film deposition, spraying, atomizing, electrospinning, dip-coating, or gelling of a solution comprising liquefied or homogenized decellularized tissue, gelatin, gelatin complex, collagen, fibrin, hydrogel, hydrogel complex, chitosan, nitrocellulose, polylactic acid, or extracellular matrix in a thin-film layer, followed by curing, cross-linking, polymerization, drying, or gelling of the solution to form the membrane layer.

[0024] In some embodiments, the membrane solution further comprises a porogen homogeneously mixed therein. In some embodiments, the porogen is a self-assembling triblock copolymer. In some embodiments, the self-assembling triblock copolymer is a poloxamer formulation, preferably Pluronic F127, at a concentration of 1-40% by weight.

[0025] In some embodiments, the membrane solution further comprises one or more agents that modify the mechanical or biological properties of one or more membranes. In some embodiments, the one or more agents are selected from glycerin, sorbitol, propylene glycol, plasticizers, fibers or other longitudinal elements, and encapsulated growth factors. In some embodiments, the membrane solution further comprises fibers, nanotubes, or other longitudinally oriented materials to provide improved mechanical properties. These fibers can be mixed into the membrane solution prior to fabrication to distribute the fibers uniformly throughout the membrane. Alternatively, these fibers can be deposited or integrated onto the membrane after fabrication through techniques such as electrospinning, 3D printing, or other techniques. The fibers can be distributed homogeneously throughout the membrane or systematically to provide the membrane with heterogeneous mechanical properties. In some embodiments, the method for producing thin film layers is repeated one or more times to produce one or more membranes having two or more membrane layers. In some embodiments, the two or more layers are produced from solutions with different components, agents, and / or concentrations.

[0026] In some embodiments, at least one of the plurality of films is treated to remove the porogen, thereby forming pores in the film.

[0027] In some embodiments, the membrane solution comprises 3-35% by weight gelatin or gelatin-polymer complex. In some embodiments, the thin film layer is crosslinked using a solution containing glutaraldehyde, transglutaminase, or other crosslinking enzymes or molecules.

[0028] In some embodiments, the sacrificial material has thermoreversible gelation properties or can be dissolved in a non-polar solvent. In some embodiments, the sacrificial material is removed using a non-polar solvent or by thermally reversing the gelation. In some embodiments, the sacrificial material comprises a poloxamer formulation, preferably Pluronic F127.

[0029] In some embodiments, the scaffold material is an extracellular matrix material. In some embodiments, the extracellular matrix material is gelatin. In some embodiments, the scaffold material is gelled by cross-linking with a solution containing glutaraldehyde, transglutaminase, or other cross-linking enzymes or molecules, and / or the scaffold material is thermally cross-linked.

[0030] In some embodiments, the steps of immersing a plurality of membranes in a solution comprising a scaffold material and gelling the scaffold material include: (a) providing a bottom mold (64) having an open top reservoir, the bottom mold being configured with a vascular channel system inflow conduit structure (63) and a vascular channel system outflow conduit structure (65), each having an internal lumen filled with a sacrificial material, the reservoirs being partially filled with a gelling scaffold material partially embedding the vascular channel system inflow conduit structure and the vascular channel system outflow conduit structure; (b) providing a plurality of membranes in a frame; (c) filling the open top reservoir of the bottom mold (64) with a solution comprising a scaffold material; (d) placing the frame on top of the bottom mold such that the membranes in the frame contact the solution; (e) gelling the solution and then removing the frame from the membranes; (f) placing a spacer (62) having an interior volume around the top of the membranes; and (g) filling the interior volume of the spacer with the solution comprising the scaffold material. (h) placing a frame on top of the spacer so that the membranes in the frame contact the solution; (i) optionally repeating steps e.-h. one or more times to add additional membranes to the device; (j) placing a spacer (57) on top of the final membrane configured with a filtration channel system outflow conduit structure (56) having an internal lumen filled with a sacrificial material; (k) filling the internal volume of the spacer (57) with a solution containing a scaffold material and gelling the solution; (l) adding a shaft filled with the sacrificial material to the gelled solution fluidly connecting a first end of the plurality of membranes to a vascular channel system inflow conduit structure (63), a second end of the plurality of membranes to a vascular channel system outflow conduit structure (65), and a third end of the plurality of membranes to a filtration channel system outflow conduit structure (56); and (m) removing the sacrificial material from the construct.

[0031] In some embodiments, the method of generating the device further includes adding cells to one or more segments of the vascular channel system and / or the filtration channel system. In some embodiments, the cells are added to the segments by (a) filling the vascular channel system and the filtration channel system with fluid, (b) disposing the cells in a first volume of fluid approximately equal to the volume of fluid in the channel system of the target segment, (c) adding the first volume to the device through a first fluid supply or fluid outlet in fluid communication with the target segment, and (d) adding a second volume of fluid approximately equal to the volume of fluid contained between the target segment and the first fluid supply or fluid outlet and / or removing a third volume of fluid approximately equal to the volume of fluid contained between the target segment and a second fluid supply or fluid outlet in fluid communication with the first fluid supply or fluid outlet.

[0032] In some embodiments, cells are added to each functional unit of the device. In some embodiments, cells are added to each segment of each functional unit of the device (e.g., to both or either the vascular channel system and the filtration channel system located in each segment). In some embodiments, cells are added to both the vascular channel system and the filtration channel system. The cells are not limited and can be any cell described herein.

[0033] Some aspects of the present disclosure relate to membranes comprising a biological or synthetic matrix material and having pores with diameters of about 1 μM to 15 μM. In some embodiments, the biological or synthetic matrix material comprises decellularized tissue, gelatin, a gelatin composite, collagen, fibrin, a hydrogel, a hydrogel composite, chitosan, nitrocellulose, polylactic acid, or an extracellular matrix. In some embodiments, the membrane has a thickness of about 0.1 μM to 100 μM. The membrane can be any thickness described herein, without limitation.

[0034] Some aspects of the present disclosure relate to methods for producing the membranes described herein, comprising chemically or physically thin-film depositing, atomizing, spraying, electrospinning, dip-coating, or gelling a solution (i.e., membrane solution) containing liquefied or homogenized decellularized tissue, gelatin, gelatin complexes, collagen, fibrin, hydrogels, hydrogel complexes, chitosan, nitrocellulose, polylactic acid, or extracellular matrix in a thin-film layer, followed by curing, crosslinking, polymerizing, drying, or gelling the solution to form the membrane layer. In some embodiments, the membrane solution further comprises a porogen homogenously mixed therein. The porogen is not limited and can be any porogen described herein. In some embodiments, the porogen is a self-assembling triblock copolymer. In some embodiments, the self-assembling triblock copolymer is a poloxamer formulation, preferably Pluronic F127, at a concentration of 1-40% by weight in the membrane solution.

[0035] In some embodiments, the membrane solution further comprises one or more agents that modify the mechanical or biological properties of the membrane, hi some embodiments, the one or more agents are selected from glycerin, sorbitol, propylene glycol, plasticizers, fibers or other longitudinal elements, and growth factors (e.g., encapsulated growth factors).

[0036] In some embodiments, the method of producing a membrane further comprises adding one or more additional membrane layers to the first membrane layer by the methods disclosed herein to create a membrane of mixed composition or architecture, hi some embodiments, the two or more layers are produced from membrane solutions having different components, agents, and / or concentrations.

[0037] In some embodiments, the film is treated to remove the porogen, thereby forming pores in the film. In some embodiments, the porogen material has thermoreversible gelling properties or can be dissolved in a non-polar solvent. In some embodiments, the porogen material is Pluronic F127 and is removed by treatment with a non-polar solvent (e.g., isopropanol).

[0038] In some embodiments, the membrane solution comprises 3-35% by weight gelatin or gelatin-polymer complex.

[0039] In some embodiments, the thin film layer can be dried, gelled, crosslinked, or otherwise solidified and removed from the substrate. In some embodiments, the thin film layer is crosslinked using a solution containing glutaraldehyde, transglutaminase, or other crosslinking enzymes or molecules. In some embodiments, the concentration of the crosslinker is about 0.01 to 5 g per 10 g of scaffold material.

[0040] This patent or application file contains at least one drawing executed in color. Copies of this patent or patent application publication with color drawing(s) will be provided by the Office upon request and payment of the necessary fee. The present invention provides, for example, the following. (Item 1) 1. A device for integrated adaptive biological blood purification, comprising: a functional unit, the functional unit comprising: a membrane having a vascular surface and a filtering surface; a vascular channel system adhered to the vascular surface of the membrane and including a first luminal space in fluid communication with the vascular surface of the membrane, the vascular channel system having a first end configured to fluidly connect to a fluid supply and a second end configured to fluidly connect to a filtrate fluid outlet; a filtration channel system adhered to the filtering surface of the membrane and including a second lumen space in fluid communication with the filtering surface of the membrane, the filtration channel system having a third end configured to connect in fluid communication to a filtrate outlet; the vascular channel system and the filtration channel system are in fluid communication with each other across the membrane; the functional unit further comprises at least three segments, the at least three segments including at least a filtration segment configured to provide ultrafiltration to produce a first ultrafiltrate, a tubular segment connected thereto configured to provide reabsorption to produce a second ultrafiltrate, and a conduit segment connected thereto configured to provide concentration to produce a tertiary ultrafiltrate; The device, wherein the membrane comprises at least three membrane segments including a filtration membrane segment, a capillary membrane segment, and a conduit membrane segment. (Item 2) 2. The device of claim 1, wherein the membrane comprises a biocompatible extracellular matrix membrane separating the vascular channel system from the filtration channel system, the biocompatible extracellular matrix membrane being embedded in a matrix material. (Item 3) 3. The device of item 2, wherein the biocompatible extracellular matrix membrane comprises a collagen membrane having a thickness of 0.1 to 10 micrometers (0.1-10 μm) that supports cell adhesion on both the vascular surface and the filtration surface of the collagen membrane. (Item 4) 4. The device according to items 1 to 3, wherein the membrane is configured to have a minimum shear stress of 5 dynes / cm^2 and / or a minimum transmembrane pressure difference of 10 mmHg. (Item 5) the filtration membrane segment permits the production of filtrate from the first luminal space in the vascular channel system to the second luminal space of the filtration channel system; the tubular membrane segments permit exchange and / or diffusion of solutes and water between the vascular channel system and the filtration channel system; 5. The device according to items 1 to 4, wherein the tubular membrane segment allows transport of water and solutes from the filtration channel system to the vascular channel system. (Item 6) The device according to items 1 to 5, wherein the functional unit comprises at least one biological fluid inflow conduit fluidly connected to the first end and the first lumen space of the vascular channel system, and at least one biological fluid outflow conduit fluidly connected to the second end and the first lumen space of the vascular channel system, the functional unit comprises at least one filtrate outflow conduit fluidly connected to the third end and the second lumen space of the filtration channel system, and the functional unit further comprises one or more blood vessel segment conduits interconnecting the filtration segment, the tubular segment, and the conduit segment of the vascular channel system and the first lumen space, and one or more filtration segment conduits interconnecting the filtration segment, the tubular segment, and the conduit segment of the filtration channel system and the second lumen space. (Item 7) 7. The device of claim 6, wherein the at least one biofluid inflow conduit is in fluid communication with an arterial conduit, the at least one biofluid outflow conduit is in fluid communication with a vascular conduit, and the at least one filtrate outflow conduit is in fluid communication with a drain conduit. (Item 8) 8. The device of claim 7, wherein the device produces ultrafiltrate that is drained into an extracorporeal collection system using a drain conduit or into the patient's bladder using a drain conduit. (Item 9) 9. The device according to items 1 to 8, wherein the first luminal space and the second luminal space are embedded in a scaffold. (Item 10) 10. The device of items 1 to 9, wherein the filtration segment of the vascular channel system comprises a vascular channel wall lined with endothelial cells selected from primary human glomerular endothelial cells, induced pluripotent stem cell (iPSC)-derived endothelial cells, and / or human umbilical cord endothelial cells. (Item 11) 11. The device of items 1 to 10, wherein the tubular segments of the vascular channel system comprise vascular channel walls lined with endothelial cells selected from primary human peritubular capillary endothelial cells, iPSC-derived endothelial cells, and / or human umbilical endothelial cells. (Item 12) 12. The device of items 1 to 11, wherein the conduit segments of the vascular channel system have vascular channel walls lined with endothelial cells selected from primary human renal medullary endothelial cells, iPSC-derived endothelial cells, and / or human umbilical endothelial cells. (Item 13) 13. The device of items 1 to 12, wherein the filtration segment of the filtration channel system comprises a filtration channel wall lined with epithelial cells selected from primary human podocytes and / or human iPSC-derived podocytes. (Item 14) 14. The device of items 1 to 13, wherein the tubular segments of the filtration channel system have filtration channel walls lined with epithelial cells selected from primary human renal tubular epithelial cells and / or iPSC-derived renal tubular epithelial cells. (Item 15) 15. The device of items 1 to 14, wherein the tubular segments of the filtration channel system have filtration channel walls lined with epithelial cells selected from primary human renal tubular epithelial cells and / or iPSC-derived renal tubular epithelial cells. (Item 16) 16. The device according to items 1 to 15, wherein the endothelial cells and / or the epithelial cells are allogeneic or autologous to the patient using the device. (Item 17) The apparatus comprises a plurality of functional units including the functional unit and an additional functional unit of the same configuration, each functional unit of the plurality of functional units comprising: a first end of a vascular channel system in fluid communication with the at least one biological fluid inflow conduit and a first lumen space; a second end of the vascular channel system in fluid communication with the at least one biological fluid outflow conduit and a first lumen space; a third end of the filtration channel system in fluid communication with the filtrate outlet conduit and a second lumen space; a first end of each of the plurality of functional units individually connected in parallel to one of a plurality of manifold ports of the at least one biological fluid inlet conduit; a second end of each of the plurality of functional units individually connected in parallel to one of a plurality of manifold ports of the at least one biological fluid outlet conduit; 17. The apparatus according to items 1 to 16, wherein the third ends of the plurality of functional units are individually connected in parallel to one of the plurality of manifold ports of the at least one filtrate outlet conduit. (Item 18) Item 18. The apparatus of item 17, wherein the apparatus comprises the plurality of functional units including the functional unit and additional functional units of the same configuration stacked in parallel layers of functional units. (Item 19) 19. The device according to items 1 to 18, wherein the at least one biological fluid inflow conduit comprises a blood inlet conduit configured to transport blood inflow, and the at least one biological fluid outflow conduit comprises a blood outflow conduit configured to transport blood outflow, and parallel layers of functional units configured for biological blood purification. (Item 20) 20. The apparatus of claim 19, wherein the filtration segment of the filtration channel system is configured to provide ultrafiltration to produce a primary ultrafiltrate, the capillary segment is configured to provide reabsorption to produce a secondary ultrafiltrate stream by absorption of solutes and water, and the conduit segment is configured to provide concentration to produce a tertiary ultrafiltrate stream by absorption of water. (Item 21) 21. The apparatus according to items 1 to 20, wherein the apparatus is configured for extracorporeal manipulation in a sterile, heated enclosure. (Item 22) 22. The device of claim 21, wherein blood or dialysis fluid from the peritoneal cavity is delivered to the device using a mechanical pump. (Item 23) 21. The device of items 1 to 20, wherein the device is disposed within a capsule and is sized and configured for placement within the human body to replace or augment kidney or liver function. (Item 24) 24. The device of items 1 to 23, wherein the membrane comprises a porous membrane comprising pores arranged to interconnect the vascular surface and the filtering surface. (Item 25) Item 25. The device according to item 24, wherein the pores have a diameter of 1 μm to 15 μm. (Item 26) 26. A method of treating a patient having insufficient renal or hepatic function, comprising: fluidly connecting the device of items 1 to 25 to the patient's circulatory system; and passing the patient's blood through the vascular channel system of the device from the filtering member segment to the tubular member segment, from the tubular member segment to the conduit member segment, and from the conduit member segment back into the patient's circulatory system. (Item 27) 27. The method of claim 26, wherein the device is implanted in the patient. (Item 28) 28. The method of claim 27, wherein the ultrafiltrate produced by the device is delivered extracorporeally to the patient. (Item 29) 28. The method of claim 27, wherein the ultrafiltrate produced by the device is delivered to the patient's bladder. (Item 30) 27. The method of claim 26, wherein the device is extracorporeal to the patient. (Item 31) A method for producing the device according to items 1 to 25, providing a plurality of membranes having sacrificial material on said vascular surface in the form of a vascular channel network and having sacrificial material on said filtering surface in the form of said filtration channel system; immersing the plurality of membranes in a solution containing a scaffold material; gelling the scaffold material; and removing the sacrificial material, thereby forming the luminal spaces of the vascular channel system and the filtration channel system. (Item 32) 32. The method of claim 31, wherein the plurality of membranes are each produced by chemical or physical thin-film deposition, spraying, atomizing, electrospinning, dip-coating, or gelling of a solution comprising liquefied or homogenized decellularized tissue, gelatin, gelatin complex, collagen, fibrin, hydrogel, hydrogel complex, chitosan, nitrocellulose, polylactic acid, or extracellular matrix in a thin-film layer, followed by curing, cross-linking, polymerization, drying, or gelling of the solution to form a membrane layer. (Item 33) 33. The method of claim 32, wherein the solution further comprises a porogen homogeneously mixed therein. (Item 34) 34. The method of claim 33, wherein the porogen is a self-assembling triblock copolymer. (Item 35) 35. The method according to item 34, wherein the self-assembling triblock copolymer is a poloxamer formulation, preferably Pluronic F127 at a concentration of 1 to 40% by weight. (Item 36) 36. The method according to items 32 to 35, wherein the solution further comprises one or more agents that modify the mechanical or biological properties of the one or more membranes. (Item 37) 37. The method of claim 36, wherein the one or more agents are selected from glycerin, sorbitol, propylene glycol, plasticizers, fibers or other longitudinal elements, and encapsulated growth factors. (Item 38) 38. The method according to any one of items 32 to 37, further comprising repeating the method according to item 32 one or more times to produce one or more membranes having two or more membrane layers. (Item 39) 39. The method of claim 38, wherein the two or more layers are formed from solutions having different components, agents, and / or concentrations. (Item 40) 40. The method of claim 32, wherein at least one of the plurality of films is treated to remove the porogen, thereby forming pores in the film. (Item 41) 41. The method according to items 32 to 40, wherein the solution contains 3 to 35% by weight of gelatin or a gelatin-polymer complex. (Item 42) 42. The method according to any one of claims 32 to 41, wherein the thin film layer is cross-linked using a solution containing glutaraldehyde, transglutaminase, or other cross-linking enzymes or molecules. (Item 43) 43. The method according to any one of items 31 to 42, wherein the sacrificial material has thermoreversible gelling properties or can be dissolved in a non-polar solvent. (Item 44) 44. The method according to any one of items 31 to 43, wherein the scaffold material is an extracellular matrix material. (Item 45) Item 45. The method of item 44, wherein the extracellular matrix material is gelatin. (Item 46) 46. ​​The method according to items 31 to 45, wherein the scaffold material is gelled by cross-linking using a solution containing glutaraldehyde, transglutaminase, or other cross-linking enzymes or molecules, and / or the scaffold material is thermally cross-linked. (Item 47) 47. The method of claim 31, wherein the sacrificial material is removed using a non-polar solvent or by thermally reversing gelation. (Item 48) 48. The method according to items 31 to 47, wherein the sacrificial material comprises a poloxamer formulation, preferably Pluronic F127. (Item 49) the step of immersing the plurality of membranes in a solution containing a scaffold material and the step of gelling the scaffold material a. providing a bottom mold (64) having an open top reservoir, the bottom mold being configured with a vascular channel system inlet conduit structure (63) and a vascular channel system outlet conduit structure (65), each having an internal lumen filled with a sacrificial material, the reservoir being partially filled with a gelling scaffold material partially embedding the vascular channel system inlet conduit structure and the vascular channel system outlet conduit structure; b. providing a plurality of membranes in a frame; c. filling the open top reservoir of the bottom mold (64) with a solution containing the scaffold material; d. placing a frame on top of the bottom mold so that the membrane in the frame contacts the solution; e. allowing the solution to gel and then removing the frame from the membrane; f. disposing a spacer (62) having an interior volume around the top of said membrane; g. filling the interior volume of the spacer with a solution comprising the scaffold material; h. placing a frame on top of the spacer so that the membrane in the frame contacts the solution; i. optionally repeating steps e.-h. one or more times to add additional membranes to the device; j. placing a spacer (57) on top of a final membrane configured with a filtration channel system outflow conduit structure (56) having an internal lumen filled with a sacrificial material; k. filling the interior volume of the spacer (57) with a solution containing the scaffold material and gelling the solution, thereby embedding the final membrane; l. adding a shaft filled with a sacrificial material to the gelled solution fluidly connecting the first ends of the plurality of membranes to the vascular channel system inlet conduit structure (63), adding a shaft filled with a sacrificial material to the gelled solution fluidly connecting the second ends of the plurality of membranes to the vascular channel system outlet conduit structure (65), and adding a shaft filled with a sacrificial material to the gelled solution fluidly connecting the third ends of the plurality of membranes to the filtration channel system outlet conduit structure (56); m. removing the sacrificial material from the construct. (Item 50) 50. The method according to items 31 to 49, further comprising adding cells to one or more segments of the vascular channel system and / or filtration channel system of the membrane. (Item 51) The cells a. providing a fluid-filled vascular channel system and a filtration channel system; b. placing the cells in a first volume of fluid approximately equal to the volume of fluid in the channel system of a target segment; c. adding the first volume to the device through a first fluid supply or fluid outlet in fluid communication with the target segment; d. The method of claim 50, wherein fluid is added to the segment by adding a second volume of fluid approximately equal to the volume of fluid contained between the target segment and the first fluid supply or fluid outlet, and / or removing a third volume of fluid approximately equal to the volume of fluid contained between the target segment and a second fluid supply or fluid outlet that is in fluid communication with the first fluid supply or fluid outlet. (Item 52) 52. The method according to items 50 to 51, wherein the cells are added to a plurality of segments of a vascular channel system and / or a filtration channel system. (Item 53) A membrane comprising a biological or synthetic matrix material and having pores with diameters of approximately 1 μM to 15 μM. (Item 54) 54. The membrane of item 53, wherein the biological or synthetic matrix material comprises decellularized tissue, gelatin, a gelatin complex, collagen, fibrin, a hydrogel, a hydrogel complex, chitosan, nitrocellulose, polylactic acid, or an extracellular matrix. (Item 55) 55. The membrane according to items 53 to 54, having a thickness of about 0.1 μM to 100 μM. (Item 56) 56. A method for producing the membrane of items 53 to 55, comprising chemically or physically thin-film depositing, spraying, atomizing, electrospinning, dip-coating, or gelling a solution comprising liquefied or homogenized decellularized tissue, gelatin, gelatin complex, collagen, fibrin, hydrogel, hydrogel complex, chitosan, nitrocellulose, polylactic acid, or extracellular matrix in a thin-film layer, followed by curing, cross-linking, polymerizing, drying, or gelling the solution to form a membrane layer. (Item 57) 57. The method of claim 56, wherein the solution further comprises a porogen homogeneously mixed therein. (Item 58) 58. The method of claim 57, wherein the porogen is a self-assembling triblock copolymer. (Item 59) 59. The method of claim 58, wherein the self-assembling triblock copolymer is a poloxamer formulation, preferably Pluronic F127 at a concentration of 1 to 40% by weight. (Item 60) 60. The method according to items 56 to 59, wherein the solution further comprises one or more agents that modify the mechanical or biological properties of the membrane. (Item 61) 61. The method of claim 60, wherein the one or more agents are selected from glycerin, sorbitol, propylene glycol, plasticizers, fibers or other longitudinal elements, and encapsulated growth factors. (Item 62) 62. The method according to any one of items 56 to 61, further comprising repeating the method according to item 56 one or more times to produce a membrane having two or more membrane layers. (Item 63) Item 63. The method of item 62, wherein the two or more layers are formed from solutions having different components, agents, and / or concentrations. (Item 64) 64. The method of claim 56, wherein the film is treated to remove the porogen, thereby forming pores in the film. (Item 65) 65. The method according to items 56 to 64, wherein the solution contains 3 to 35% by weight of gelatin or a gelatin-polymer complex. (Item 66) 66. The method according to items 56 to 65, wherein the thin film layer is cross-linked using a solution containing glutaraldehyde, transglutaminase, or other cross-linking enzymes or molecules. [Brief explanation of the drawings]

[0041] [Figure 1] 1 shows microscopic imaging of thin films fabricated with varying amounts of sacrificial porogen material to facilitate control of porosity and pore size. [Figure 2] Microscopic imaging of a thin film fabricated with the sacrificial porogen then removed to create a porous thin film is shown. [Figure 3] Shown is a porous thin film seeded with layers of fluorescently labeled endothelial and epithelial cells on opposite sides of the membrane. [Figure 4]A schematic diagram of a single functional unit of an IABBP device is shown. Blood flows from a recipient's artery into the filtration segment, where primary filtrate is produced. The blood and filtrate then flow into the tubular segments in their respective channels. Absorption and secretion occur in the tubular segments. The blood and secondary filtrate then flow into the conduit segments in their respective channels. The secondary filtrate is concentrated and drained through the conduit, while the blood is returned to the recipient's circulation. (1) Blood inflow, (2) Blood inflow conduit, (3) filtration segment blood channel system, (4) Blood flow into the tubular segment, (5) tubular segment blood channel system, (6) Blood flow into the conduit segment, (7) conduit segment channel system, (8) Blood outflow conduit, (9) Blood outflow, (10) filtration, (11) filtration segment filtrate channel, (12) filtrate flow into the tubular segment, (13) Secretion of solutes and water, (14) ) tubular segment filtrate channel system, (15) solute and water absorption, (16) filtrate flow into the tubular segment, (17) concentration (water absorption), (18) tubular segment filtrate channel system, (19) filtrate outflow conduit, (20) filtrate outflow, (21) embedding matrix, (22) capsule, (23) filtration segment, (24) tubular segment, (25) tubular segment, (26) filtration membrane, (27) tubular membrane, (28) tubular membrane. [Figure 5] A schematic diagram of the channel architecture of a single functional IABBP unit is shown. Corresponding segments of the nephron are shown to illustrate the sequential functions of each segment: (2) inflow conduit, (8) outflow conduit, (23) filtration segment, (24) tubule segment, (25) duct segment, and (30) corresponding segments of a human nephron. [Figure 6]A schematic diagram of each segment of a functional IABBP unit with relocated cells is shown. Each channel system within each segment is separated by specialized membranes and lined with specialized cells to enable higher-level function. (26) filtration segment membrane, (31) filtration segment blood channel, (32) filtration segment endothelial cell, (27) tubule segment membrane, (33) tubule segment blood channel, (34) tubule segment endothelial cell, (28) conduit segment membrane, (35) conduit segment blood channel, (36) conduit segment endothelial cell, (37) filtration segment epithelial cell, (38) filtration segment filtrate channel, (39) tubule segment epithelial cell, (40) tubule segment filtrate channel, (41) conduit segment epithelial cell, (42) conduit segment filtrate channel. [Figure 7] A three-dimensional rendering of a cross section of a functional IABBP unit is shown. The vascular space is separated from the filtrate space by specialized membranes: (41) vascular channel, (42) matrix membrane, (43) filtrate channel, and (44) main vascular channel. [Figure 8] Schematic diagram of stacking of functional units in an IABBP device with parallel perfusion and drainage: (45) graft blood inlet, (46) graft blood inlet conduit, (47) stacked functional unit, (48) stacked functional unit, (49) stacked functional unit, (50) stacked functional unit, (51) graft blood outflow conduit, (52) graft blood outflow, (53) graft filtrate outflow conduit, (54) graft filtrate outflow. [Figure 9] A 3D rendering of a multilayer IABBP device is shown. Five layers of functional units are stacked and connected in parallel to be perfused with blood and produce filtrate, which is discharged through a filtrate discharge conduit: (46) graft blood inlet conduit, (51) graft blood outlet conduit, (53) graft filtrate outlet conduit, (47) stacked functional units, and (55) connecting channel. [Figure 10]3D rendering of the fabrication mold for creating a multilayer IABBP device. The base and top molds house the blood vessels and filtrate conduits. Each cassette allows for the addition of embedding material and supports the patterned membrane. (56) Notch for top conduit, (57) Top mold, (58) Spacer 1, (59) Spacer 2, (60) Spacer 3, (61) Spacer 4, (62) Spacer 5, (63) Groove for bottom conduit 1, (64) Bottom mold, (65) Groove for bottom conduit 2. [Figure 11] Figure 1 shows a membrane testing apparatus and its schematic diagram. A shows a photograph of the membrane testing apparatus. B shows a schematic diagram of the membrane testing apparatus and its components. (66) Plasma or blood inlet, (67) Plasma or blood inlet port, (68) Top chamber vascular space, (69) Solute and fluid exchange, (70) Plasma or blood outlet, (71) Plasma or blood outlet port, (72) Membrane, (73) Bottom chamber, (74) Bottom mold, (75) Filtrate outlet port, (76) Filtrate outlet, (77) O-ring. [Figure 12] 1 shows the pore size and area distribution of membranes made from a mixture of gelatin and 0.25% Pluronic F127. [Figure 13] (Top panel) shows Pluronic F127 concentration versus flow rate. (Bottom panel) shows fluid pressure versus flow rate. DETAILED DESCRIPTION OF THE INVENTION

[0042] Biofluid Purification Some aspects of the present disclosure relate to an apparatus for adaptive biological blood purification (AIBBP), the apparatus comprising: (1) a membrane having a vascular surface and a filtering surface; (2) a vascular channel system adhered to the vascular surface of the membrane and including a first luminal space in fluid communication with the vascular surface of the membrane, the vascular channel system having a first end configured to fluidly connect to a fluid supply and a second end configured to fluidly connect to a filtrate outlet; and (3) a filtration channel system adhered to the filtering surface of the membrane and including a second luminal space in fluid communication with the filtering surface of the membrane, the filtration channel system having a third end configured to fluidly connect to a filtrate outlet. The vascular channel system and the filtration channel system are in fluid communication with each other across the membrane. Furthermore, the functional unit further comprises at least three segments, the at least three segments including at least a filtration segment configured to provide ultrafiltration to produce a first ultrafiltrate, a tubular segment connected thereto configured to provide reabsorption to produce a second ultrafiltrate, and a conduit segment connected thereto configured to provide concentration to produce a tertiary ultrafiltrate, and the membrane comprises three membrane segments including at least a filtration membrane segment, a tubular membrane segment, and a conduit membrane segment.

[0043] In some embodiments, the filtration membrane segment allows for the generation of filtrate from a first luminal space in the vascular channel system to a second luminal space of the filtration channel system. In some embodiments, the tubular membrane segment allows for the exchange and / or diffusion of solutes and water between the vascular channel system and the filtration channel system. In some embodiments, the tubular membrane segment allows for the transfer of water and solutes from the filtration channel system to the vascular channel system.

[0044] In some embodiments, the functional unit comprises at least one biological fluid inflow conduit in fluid communication with a first end and a first luminal space of the vascular channel system and at least one biological fluid outflow conduit in fluid communication with a second end and a first luminal space of the vascular channel system. In some embodiments, the functional unit comprises at least one filtrate outflow conduit in fluid communication with a third end and a second luminal space of the filtration channel system. In some embodiments, the functional unit further comprises one or more blood vessel segment conduits interconnecting the filtration segments, tubular segments, and conduit segments of the vascular channel system and the first luminal space, and one or more filtration segment conduits interconnecting the filtration segments, tubular segments, and conduit segments of the filtration channel system and the second luminal space.

[0045] In some embodiments, at least one biological fluid inflow conduit is in fluid communication with an arterial conduit, at least one biological fluid outflow conduit is in fluid communication with a vascular conduit, and at least one filtrate outflow conduit is in fluid communication with a drain conduit.

[0046] In some embodiments, the membrane comprises a porous membrane comprising pores arranged to interconnect the vascular surface and the filtering surface. In some embodiments, the pores have a diameter of less than about 15 μm, 14 μm, 13 μm, 12 μm, 11 μm, 10 μm, 9 μm, 8 μm, 7 μm, 6 μm, 5 μm, 4 μm, 3 μm, 2 μm, or 1 μm. In some embodiments, the pores have an average or mean diameter of about 15 μm, 14 μm, 13 μm, 12 μm, 11 μm, 10 μm, 9 μm, 8 μm, 7 μm, 6 μm, 5 μm, 4 μm, 3 μm, 2 μm, or 1 μm. In some embodiments, the pore-containing membrane has a flow rate of about 0.2-2.0 mL / min, about 0.5-1.5 mL / min, or about 0.8-1.2 mL / min when subjected to a fluid pressure of 40 mmHg. In some embodiments, the membrane is manufactured by a process comprising mixing an extracellular matrix material (e.g., gelatin) with a porogen (pore-forming agent). In some embodiments, the pore-forming agent is Pluronic F127. In some embodiments, the membrane is manufactured by the process disclosed herein.

[0047] In some embodiments, the membrane is as described in PCT Application No. PCT / US2017 / 67141 (filed December 18, 2017, incorporated herein by reference in its entirety). In some embodiments, the membrane can be constructed from any biological, synthetic, or composite material suitable for thin film deposition and capable of maintaining mechanical viability and barrier integrity between compartments. The membrane can include pores, slits, surface roughness, or other functional properties imparted during fabrication using techniques known in the art designed to improve the membrane's function, biocompatibility, or other qualities. The membrane can be fabricated from biological, synthetic, or composite materials such as collagen, gelatin, other hydrogels, cellulose, or other materials that can be thinly deposited and then crosslinked, dried, gelled, hardened, or otherwise stabilized to form a coherent, mechanically stable membrane. The membrane can undergo further processing or manipulation to provide enhanced function or mechanical structure. The membrane can be of uniform or variable thickness ranging from 0.01 μm to 100 μm or greater. In some embodiments, the membrane has a thickness of about 0.1 μM to about 100 μM, about 0.1 μM to about 100 μM, about 0.5 μM to about 50 μM, about 1.0 μM to about 40 μM, about 5.0 μM to about 30 μM, or about 10 μM to about 20 μM, or any range therebetween. In some embodiments, the membrane has a thickness of about 10 μM or less. In some embodiments, the membrane has a thickness of about 1 to 8 μM. In some embodiments, the membrane has a thickness of about 5 μM or less.

[0048] In some embodiments, the biocompatible extracellular matrix membrane comprises fibers, nanofibers, or other longitudinal elements. In some embodiments, the fibers, nanofibers, or other longitudinal elements are attached to the exterior surface of the membrane. In some embodiments, the fibers, nanofibers, or other longitudinal elements increase or modify the mechanical strength of the membrane. In some embodiments, the fibers, nanofibers, or other longitudinal elements form a mesh (e.g., an ordered mesh, a disordered mesh), or are oriented in a substantially single direction or substantially in two directions (e.g., a mesh).

[0049] In some embodiments, the fibers, nanofibers, or other longitudinal elements increase the mechanical strength of the membrane or portion thereof by at least about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 99% or more compared to an identical membrane without the fibers, nanofibers, or other longitudinal elements. In some embodiments, the fibers, nanofibers, or other longitudinal elements increase the mechanical strength of the membrane or portion thereof by at least about 1.1-fold, 1.5-fold, 2-fold, 3-fold, 5-fold, 10-fold, or more compared to an identical membrane without the fibers, nanofibers, or other longitudinal elements. In some embodiments, the fibers, nanofibers, or other longitudinal elements are uniformly distributed throughout the membrane, providing homogeneous mechanical reinforcement. In some embodiments, the fibers, nanofibers, or other longitudinal elements are non-uniformly distributed in the membrane, providing heterogeneous mechanical reinforcement. In some embodiments, the fibers, nanofibers, or other longitudinal elements provide resistance to cell infiltration and maintain separation of different cell populations on each side of the membrane. In some embodiments, the fibers, nanofibers, or other longitudinal elements form a mesh, providing resistance to cell infiltration and maintaining separation of different cell populations on each side of the membrane.

[0050] The fibers, nanofibers, and other longitudinal elements disclosed herein can be made from a variety of materials, including (but not limited to) Dyneema®, an extremely strong polyethylene manufactured by DSM High Performance Fibers, a subsidiary of DSM NV. Fibers can also be combined with fibers or wires of other materials, such as Nitinol (a version of a shape-memory nickel-titanium alloy), to help control the expanded shape of the filter. Other viable materials for use as fibers, nanofibers, and other longitudinal elements include those known in the fiber art, such as carbon, glass, ceramic, metals and metal alloys (including the aforementioned Nitinol), natural and synthetic polymers (including ultra-high molecular weight, highly oriented polymers and silk), or combinations thereof. In some embodiments, the fibers, nanofibers, or other longitudinal elements comprise silk or polymers (e.g., polycarbonate). In some embodiments, the fibers, nanofibers, or other longitudinal elements form a mesh (e.g., polycarbonate mesh). Furthermore, fibers, nanofibers, and other longitudinal elements can be made of monofilaments or multifilaments and can be configured with all kinds of cross sections and orientations. Fibers can be made of round, flat, or differently shaped monofilaments or multifilaments. In some embodiments, the fibers are non-immunogenic.

[0051] In some embodiments, the membrane comprises, consists essentially of, or consists of a biocompatible extracellular matrix membrane that separates the vascular channel system from the filtration channel system. In some embodiments, the biocompatible extracellular matrix membrane comprises a collagen membrane having a thickness of about 0.1 to 10 micrometers (e.g., 0.3 to 10 μm) that supports cell adhesion on both the vascular and filtration surfaces of the collagen membrane.

[0052] In some embodiments, the biocompatible extracellular matrix membrane is embedded in a matrix material (i.e., a scaffold material). In some embodiments, the scaffold comprises a hydrogel, such as gelatin, PLA, chitosan, a composite of hydrogel or other hydrogel materials, and composites of various concentrations and compositions. In some embodiments, varying the concentration and composition of the hydrogel materials and composites allows for tuning of the mechanical and biological properties of the scaffold, which can enhance and further specialize the tissue construct for a desired biological application. In some embodiments, the scaffold may include the addition of glycerin, sorbitol, propylene glycol, or other plasticizers to gelatin or gelatin composite hydrogels. The composition of the scaffold is not limited and can be any suitable scaffold material known in the art.

[0053] In some embodiments, the device comprises at least one biological fluid inflow conduit in fluid communication with a first end and a first luminal space of the vascular channel system, and at least one biological fluid outflow conduit in fluid communication with a second end and a first luminal space of the vascular channel system, and the functional unit comprises at least one filtrate outflow conduit in fluid communication with a third end and a second luminal space of the filtration channel system.

[0054] In some embodiments, the vascular and filtration channel systems are fabricated by the method disclosed in PCT Application No. PCT / US2017 / 67141 (filed December 18, 2017, which is incorporated herein by reference in its entirety). Briefly, a sacrificial material is layered onto a membrane, followed by a scaffold material, and then the sacrificial material is removed, leaving a luminal space bounded by the scaffold material and the membrane. The membrane is then inverted, and a sacrificial material is layered onto the membrane, followed by the scaffold material, and then the sacrificial material is removed, leaving a second luminal space bounded by the scaffold material and the membrane. In some embodiments, the vascular and filtration channel systems are in partial or complete fluid communication across the membrane (e.g., are opposite mirror images of each other across the membrane). In some embodiments, the area of ​​the membrane where the first and second luminal spaces are in fluid communication across the membrane is at least 30 cm. 2 , at least 60cm 2 , at least 90cm 2 , at least 100cm 2 , at least 150cm 2 , at least 200cm 2 , at least 250cm 2 , at least 300cm 2 , at least 450cm 2 , at least 600 cm 2 , at least 800 cm 2 , at least 1000 cm 2 , or at least 1200 cm 2 In some embodiments, at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 99%, or more of the first and / or second luminal spaces are in fluid communication across the membrane with the other luminal spaces.

[0055] In some embodiments, at least one biofluid inflow conduit is in fluid communication with an arterial conduit, in some embodiments, at least one biofluid outflow conduit is in fluid communication with a vascular conduit, and in some embodiments, at least one filtrate outflow conduit is in fluid communication with a drain conduit.

[0056] In some embodiments, the device produces filtrate and / or ultrafiltrate that is drained into an extracorporeal collection system (e.g., a waste container) using a drain conduit. In some embodiments, the filtrate and / or ultrafiltrate is drained into a plumbing system or a sewage treatment system. In some embodiments, the filtrate and / or ultrafiltrate is drained into a patient's bladder or digestive system using a drain conduit.

[0057] In some embodiments, the vascular channel system comprises vascular channel walls lined with endothelial and / or epithelial cells, and / or the filtration channel comprises filtration channel walls lined with endothelial and / or epithelial cells. In some embodiments, the cells are at confluence on the vascular channel walls and / or filtration channel walls. In some embodiments, the vascular channel walls and / or filtration channel walls are at a density of at least 2×10 6 In some embodiments, the vascular channel wall and / or the filtration channel wall comprises at least 1 x 10 cells. 7 In some embodiments, the vascular channel walls and / or filtration channel walls contain at least 2×10 cells. 7 In some embodiments, the cells comprise at least 30 cm 2 , at least 60cm 2 , at least 90cm 2 , at least 100cm 2 , at least 150cm 2 , at least 200cm 2 , at least 250cm 2 , at least 300cm 2 , at least 450cm 2 , at least 600 cm 2 , at least 800 cm 2 , at least 1000 cm 2 , or at least 1200 cm 2 or more on the vascular channel wall and / or filtration channel wall.

[0058] In some embodiments, the epithelial cell type is selected from prostate cells, breast cells, hepatocytes, pancreatic islet cells including beta cells, lung epithelial cells, kidney cells, bladder cells, gastric epithelial cells, large and small intestinal epithelial cells, urethral epithelial cells, testicular epithelial cells, ovarian epithelial cells, cervical epithelial cells, thyroid cells, parathyroid cells, adrenal cells, thymus cells, gallbladder cells, and pituitary cells. In some embodiments, the endothelial cells are brain endothelial cells, vascular endothelial cells, primary human peritubular capillary endothelial cells, iPSC-derived endothelial cells, human umbilical cord endothelial cells, primary human renal medullary endothelial cells, human podocytes, or human iPSC-derived podocytes. In some embodiments, the cells are allogeneic to the patient using the device. In some embodiments, the cells are autologous to the patient using the device. In some embodiments, the cells are derived from a cell line. In some embodiments, the cells are derived from autologous stem cells. In some embodiments, the autologous stem cells are derived from induced pluripotent stem cells.

[0059] In some embodiments, the filtration segment of the vascular channel system comprises vascular channel walls lined with endothelial cells selected from primary human glomerular endothelial cells, induced pluripotent stem cell (iPSC)-derived endothelial cells, and / or human umbilical cord endothelial cells.

[0060] In some embodiments, cells are obtained from kidneys, as described below in the Examples section entitled "Primary Cell Isolation from Discarded Kidneys."

[0061] In some embodiments, the tubular segments of the vascular channel system comprise vascular channel walls (e.g., walls comprising a scaffold material and a membrane) lined with endothelial cells selected from primary human peritubular capillary endothelial cells, iPSC-derived endothelial cells, and / or human umbilical endothelial cells.

[0062] In some embodiments, the tubular segments of the vascular channel system comprise vascular channel walls lined with endothelial cells selected from primary human renal medullary endothelial cells, iPSC-derived endothelial cells, and / or human umbilical cord endothelial cells.

[0063] In some embodiments, the filtration segment of the filtration channel system comprises filtration channel walls lined with epithelial cells selected from primary human podocytes and / or human iPSC-derived podocytes.

[0064] In some embodiments, the tubular segments of the filtration channel system comprise filtration channel walls (e.g., walls comprising a scaffold material and a membrane) lined with epithelial cells selected from primary human renal tubular epithelial cells and / or iPSC-derived renal tubular epithelial cells.

[0065] In some embodiments, the tubular segments of the filtration channel system comprise filtration channel walls lined with epithelial cells selected from primary human renal tubular epithelial cells and / or iPSC-derived renal tubular epithelial cells.

[0066] In some embodiments, the vascular channel system comprises a vascular channel diameter between 1 mm and 10 μm. In some embodiments, the filtration channel system comprises a filtration channel diameter between 1 mm and 10 μm. In some embodiments, the channel system (e.g., the vascular channel system and / or the filtration channel system) comprises more than one channel (e.g., at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 40, 50, 100, 200, 500, 750, 1000, 2000, 10,000 channels). In some embodiments, the channel system comprises a branched channel network having one or more branches of gradually decreasing diameter (e.g., at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 40, 50, 100, 200, 500, 750, 1000, 2000, 10,000 branches). In some embodiments, the channel diameters can include, but are not limited to, about 10 cm, 5 cm, 2 cm, 1 cm, 500 mm, 250 mm, 100 mm, 50 mm, 10 mm, 5 mm, 1 mm, 500 μm, 50 μm, 10 μm, 5 μm, 3 μm, 1 μm, 0.5 μm, 0.1 μm, 0.05 μm, 0.02 μm, or 0.01 μm. In some embodiments, the vascular channel system and the filtration channel system are embedded in a scaffold (eg, a biocompatible matrix material).

[0067] In some embodiments, the device includes a plurality of functional units including a functional unit and an additional functional unit of the same configuration (see, for example, FIG. 8 ), each functional unit of the plurality of functional units having a first end and a first lumen space of a vascular channel system in fluid communication with at least one biofluid inflow conduit, a second end and a first lumen space of the vascular channel system in fluid communication with at least one biofluid outflow conduit, and a third end and a second lumen space of a filtration channel system in fluid communication with a filtrate outflow conduit, wherein each first end of the plurality of functional units is individually connected in parallel to one of a plurality of manifold ports of the at least one biofluid inflow conduit, each second end of the plurality of functional units is individually connected in parallel to one of a plurality of manifold ports of the at least one biofluid outflow conduit, and each third end of the plurality of functional units is individually connected in parallel to one of a plurality of manifold ports of the at least one filtrate outflow conduit. In some embodiments, the plurality of functional units including a functional unit and an additional functional unit of the same configuration are stacked in parallel layers of functional units.

[0068] In some embodiments, the at least one biological fluid inflow conduit comprises a blood inlet conduit configured to transport blood inflow, and the at least one biological fluid outflow conduit comprises a blood outflow conduit configured to transport blood outflow, and parallel layers of functional units configured for biological blood purification.

[0069] In some embodiments, the filtration segment of the filtration channel system is configured to provide ultrafiltration to produce a primary ultrafiltrate, the capillary segment is configured to provide reabsorption by absorption of solutes and water to produce a secondary ultrafiltrate stream, and the conduit segment is configured to provide concentration by absorption of water to produce a tertiary ultrafiltrate stream.

[0070] In some embodiments, the device is configured for extracorporeal operation in a sterile, heated enclosure. In some embodiments, blood is delivered to the device using a mechanical pump. In some embodiments, the device is disposed within a capsule and sized and configured for placement within the human body to replace or augment tissue or organ function (e.g., liver and / or kidney function).

[0071] Some aspects of the present disclosure relate to methods of treating a patient with inadequate renal or hepatic function, the method including fluidly connecting a device described herein to the patient's circulatory system and passing the patient's blood through a vascular channel system of the device from a filtering member segment to a tubular member segment, from the tubular member segment to a conduit member segment, and from the conduit member segment back into the patient's circulatory system. In some embodiments, the device comprises multiple functional units described herein, and the patient's blood is passed through multiple functional units. In some embodiments, the patient's blood is passed through the functional units in parallel. In some embodiments, the patient's blood is passed through the functional units in series. In some embodiments, the device is implanted in the patient. In some embodiments, ultrafiltrate produced by the device is delivered extracorporeally to the patient. In some embodiments, ultrafiltrate produced by the device is delivered to the patient's bladder.

[0072] In some embodiments, the device is external to the patient, for example, the device may be stationary or the device may be configured to be carried by the patient, such as in a backpack or waist pack, allowing the patient to be mobile while using the device.

[0073] In some embodiments, the device is extracorporeal to the patient and configured for use in peritoneal dialysis (e.g., as a peritoneal dialysis adjunct device). Peritoneal dialysis (PD) is a type of dialysis that uses the peritoneal membrane in the patient's abdomen as a membrane through which fluid and dissolved substances are exchanged with the blood. PD is used in individuals with renal failure to remove excess fluid, correct electrolyte problems, and remove toxins. Continuous ambulatory peritoneal dialysis (CAPD) requires the patient to add and remove dialysate multiple times per day through a catheter in the abdomen. During the period when dialysate resides in the peritoneal cavity, the patient may be mobile (i.e., ambulatory). However, the longer the dialysate resides in the peritoneal cavity, the less effective the dialysate becomes at removing waste products as the dialysate approaches equilibrium. In continuous-flow peritoneal dialysis (CFPD), dialysate is continuously added and removed from the patient's peritoneal cavity, typically during sleep. CFPD requires a large dialysate reservoir and a total dialysate volume of 6 to 12 liters per sleep period.

[0074] In some embodiments, the device is configured to be connected to the peritoneal cavity so that fluid from the peritoneal cavity circulates through the device (e.g., via a pump) and at least a portion of it is returned to the peritoneal cavity. In some embodiments, the device is configured to remove a portion of the fluid from the peritoneal cavity as waste. In some embodiments, the device includes a waste outlet or waste storage receptacle that can be optionally drained or replaced (e.g., hot-swapped during operation) for a new waste storage receptacle if needed. In some embodiments, the device is part of a peritoneal dialysis system (CAPD and / or CFPD) that includes a pump that circulates dialysate solution from the patient's peritoneal cavity through the device and back to the peritoneal cavity, thereby removing toxins and / or excess fluid from the dialysis solution. In some embodiments, use of the device reduces the volume of dialysis fluid required for effective dialysis (e.g., by at least 10%, 25%, 50%, or more). In some embodiments, use of the device increases the interval between dialysis fluid exchanges in CAFD (e.g., by at least 10%, 25%, 50%, or more) without loss of effectiveness compared to CAFD without the device. In some embodiments, the device comprises kidney cells (eg, cells from discarded kidneys as detailed in the Examples section below).

[0075] Manufacturing method Some aspects of the present disclosure relate to methods of manufacturing the devices disclosed herein, including providing a plurality of membranes having sacrificial material in the form of a vascular channel network on a vascular surface and sacrificial material in the form of a filtration channel system on a filtration surface; immersing the plurality of membranes in a solution including a scaffold material (e.g., a scaffold material in a sol state); gelling the scaffold material; and removing the sacrificial material, thereby forming the luminal spaces of the vascular channel system and filtration channel system described herein.

[0076] In some embodiments, the multiple membranes are each produced by chemical or physical thin-film deposition, spraying, atomizing, electrospinning, dip-coating, or gelling of a solution (membrane solution) comprising liquefied or homogenized decellularized tissue, gelatin, gelatin complex, collagen, fibrin, hydrogel, hydrogel complex, chitosan, nitrocellulose, polylactic acid, or extracellular matrix in a thin-film layer on a substrate, followed by curing, cross-linking, polymerization, drying, or gelling of the solution to form the membrane layer.

[0077] In some embodiments, the membrane solution further includes fibers, nanotubes, or other longitudinally oriented materials to provide improved mechanical properties. The fibers, nanotubes, or other longitudinally oriented materials are not limited and can be any of the fibers, nanotubes, or other longitudinally oriented materials disclosed herein. These fibers, nanotubes, or other longitudinally oriented materials can be mixed into the membrane solution prior to fabrication to distribute the fibers uniformly throughout the membrane. Alternatively, these fibers, nanotubes, or other longitudinally oriented materials can be deposited or integrated onto the membrane after fabrication through techniques such as electrospinning, 3D printing, or other techniques. In some embodiments, the membrane can be bonded to the fibers, nanotubes, or other longitudinally oriented materials. The fibers, nanotubes, or other longitudinally oriented materials can be distributed homogeneously throughout the membrane or systematically to provide the membrane with heterogeneous mechanical properties.

[0078] In some embodiments, the membrane solution further comprises a porogen homogeneously mixed therein. In some embodiments, the porogen is in the form of micelles in the solution (e.g., the porogen is at a concentration sufficient to form micelles in the solution). In some embodiments, the porogen is incorporated into the solution via mixing or sonication. In some embodiments, the porogen is a self-assembling triblock copolymer. In some embodiments, the self-assembling triblock copolymer is a poloxamer formulation. In some embodiments, the porogen is Pluronic F127. In some embodiments, the porogen is at a concentration of 1-40% by weight. In some embodiments, the porogen is at a concentration of about 1 wt%, about 2 wt%, about 3 wt%, about 4 wt%, about 5 wt%, about 6 wt%, about 7 wt%, about 8 wt%, about 9 wt%, about 10 wt%, about 11 wt%, about 12 wt%, about 13 wt%, about 14 wt%, about 15 wt%, about 16 wt%, about 17 wt%, about 18 wt%, about 19 wt%, about 20 wt%, about 21 wt%, about 22 wt%, about 23 wt%, about 24 wt%, about 25 wt%, about 26 wt%, about 27 wt%, about 28 wt%, about 29 wt%, about 30 wt%, about 31 wt%, about 32 wt%, about 33 wt%, about 34 wt%, about 35 wt%, about 36 wt%, about 37 wt%, about 38 wt%, about 39 wt%, or about 40 wt% in solution. Pore ​​size in the resulting membranes can be controlled by using different polymers, but also by varying concentrations, solution properties, and processing techniques to control micelle size and aggregation. Varying concentrations and compositions of sacrificial porogen materials can allow substantial opportunities for tuning of mechanical and biological properties, including, but not limited to, porosity, pore size, permeability, sieving, filtration, and other functions that can enhance and further specialize tissue constructs for desired biological applications.

[0079] In some embodiments, the membrane solution further comprises one or more agents that modify the mechanical or biological properties of one or more membranes. Examples of agents include, but are not limited to, glycerin, sorbitol, propylene glycol, or other plasticizers for gelatin or gelatin composite hydrogels (see FMVanina et al., Food Hydrocolloids 19, 899-907 (2005)). In some embodiments, the agent comprises a growth factor (e.g., an encapsulated growth factor). In some embodiments, the one or more agents are selected from glycerin, sorbitol, propylene glycol, plasticizers, fibers or other longitudinal elements, and encapsulated growth factors.

[0080] In some embodiments, the method of producing a membrane further comprises adding one or more additional membrane layers to the first membrane layer by the methods disclosed herein to create a membrane of mixed composition or architecture, hi some embodiments, the two or more layers are produced from membrane solutions having different components, agents, and / or concentrations.

[0081] In some embodiments, the film is treated to remove the porogen, thereby forming pores in the film. The sacrificial porogen material is removed passively or forcibly in conjunction with dissolution, phase transition, reversal of thermal gelation, or other techniques known in the art. In some embodiments, the sacrificial material has thermally reversible gelation properties or can be dissolved in a non-polar solvent.

[0082] In some embodiments, the porogen material is Pluronic F127 and is removed by treatment with a non-polar solvent (eg, isopropanol).

[0083] In some embodiments, the membrane solution comprises 3-35% by weight of gelatin or gelatin-polymer complex, hi some embodiments, the solution comprises about 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, or 35% by weight of gelatin or gelatin-polymer complex.

[0084] In some embodiments, the thin film layer can be dried, gelled, crosslinked, or otherwise solidified and removed from the substrate. In some embodiments, the thin film layer is crosslinked using a solution containing glutaraldehyde, transglutaminase, or other crosslinking enzymes or molecules. In some embodiments, a crosslinking agent is added to the film solution, for example, just before applying the solution to the thin film on the substrate. In some embodiments, the crosslinking agent is contacted with the thin film layer after contact with the substrate. In some embodiments, the concentration of crosslinking agent is about 0.01 to 5 g per 10 g of gelatin.

[0085] In some embodiments, the solution comprising the scaffold material comprises an extracellular matrix material, hi some embodiments, the extracellular matrix material is gelatin.

[0086] In some embodiments, the scaffolding material is gelled by cross-linking with a solution containing glutaraldehyde, transglutaminase, or other cross-linking enzymes or molecules, and / or the scaffolding material is thermally cross-linked.

[0087] In some embodiments, the steps of immersing a plurality of membranes in a solution comprising a scaffold material and gelling the scaffold material include: (a) providing a bottom mold (64) having an open top reservoir, the bottom mold being configured with a vascular channel system inflow conduit structure (63) and a vascular channel system outflow conduit structure (65), each having an internal lumen filled with a sacrificial material, the reservoirs being partially filled with a gelling scaffold material partially embedding the vascular channel system inflow conduit structure and the vascular channel system outflow conduit structure; (b) providing a plurality of membranes in a frame; (c) filling the open top reservoir of the bottom mold (64) with a solution comprising a scaffold material; (d) placing the frame on top of the bottom mold such that the membranes in the frame contact the solution; (e) gelling the solution and then removing the frame from the membranes; (f) placing a spacer (62) having an interior volume around the top of the membranes; and (g) filling the interior volume of the spacer with the solution comprising the scaffold material. (h) placing a frame on top of the spacer so that the membranes in the frame contact the solution; (i) optionally repeating steps e.-h. one or more times to add additional membranes to the device; (j) placing a spacer (57) on top of the final membrane configured with a filtration channel system outflow conduit structure (56) having an internal lumen filled with a sacrificial material; (k) filling the internal volume of the spacer (57) with a solution containing a scaffold material and gelling the solution; (l) adding a shaft filled with the sacrificial material to the gelled solution fluidly connecting a first end of the plurality of membranes to a vascular channel system inflow conduit structure (63), a second end of the plurality of membranes to a vascular channel system outflow conduit structure (65), and a third end of the plurality of membranes to a filtration channel system outflow conduit structure (56); and (m) removing the sacrificial material from the construct. In some embodiments, methods for fabricating the devices described herein include the methods described in the Examples below under "Fabrication of Multilayer Devices."

[0088] In some embodiments, the method of manufacturing the device further includes adding cells to one or more segments of the vascular channel system and / or the filtration channel system. In some embodiments, the cells are added to each functional unit of the device. In some embodiments, the cells are added to each segment of each functional unit of the device (e.g., to both or either the vascular channel system and the filtration channel system located in each segment). In some embodiments, the cells are added to both the vascular channel system and the filtration channel system. The cells are not limited and can be any cell described herein.

[0089] In some embodiments, cells are added to the segment by (a) filling the vascular channel system and the filtration channel system with fluid, (b) placing the cells in a first volume of fluid approximately equal to the volume of fluid in the channel system of the target segment, (c) adding the first volume to the device through a first fluid supply or fluid outlet in fluid communication with the target segment, and (d) adding a second volume of fluid approximately equal to the volume of fluid contained between the target segment and the first fluid supply or fluid outlet, and / or removing a third volume of fluid approximately equal to the volume of fluid contained between the target segment and a second fluid supply or fluid outlet in fluid communication with the first fluid supply or fluid outlet. In some embodiments, cells are added (e.g., seeded) by methods described in the Examples contained herein.

[0090] Pore-containing membrane and method for producing same Some aspects of the present disclosure relate to membranes comprising a biological or synthetic matrix material and having pores with diameters of approximately 1 μM to 15 μM. In some embodiments, the biological or synthetic matrix material comprises decellularized tissue, gelatin, a gelatin composite, collagen, fibrin, a hydrogel, a hydrogel composite, chitosan, nitrocellulose, polylactic acid, or an extracellular matrix. In some embodiments, the membrane may comprise any extracellular matrix material or scaffold material described herein. In some embodiments, the membrane has a thickness of approximately 0.1 μM to 100 μM. The membrane may be any thickness described herein, without limitation.

[0091] Some aspects of the present disclosure relate to methods for producing the membranes described herein, including chemically or physically thin-film depositing, spraying, atomizing, electrospinning, dip-coating, or gelling a solution (i.e., membrane solution) containing liquefied or homogenized decellularized tissue, gelatin, gelatin complex, collagen, fibrin, hydrogel, hydrogel complex, chitosan, nitrocellulose, polylactic acid, or extracellular matrix in a thin-film layer, followed by curing, crosslinking, polymerizing, drying, or gelling the solution to form the membrane layer. The method for producing (i.e., manufacturing) the membrane is not limited and can be any method described herein or known in the art. In some embodiments, the membrane solution further comprises a porogen homogenously mixed therein. The porogen is not limited and can be any porogen described herein. In some embodiments, the porogen is a self-assembling triblock copolymer. In some embodiments, the self-assembling triblock copolymer is a poloxamer formulation, preferably Pluronic F127, at a concentration of 1-40% by weight in the membrane solution. The concentration of the porogen in the membrane solution is not limited and can be any concentration disclosed herein.

[0092] In some embodiments, the membrane solution further comprises one or more agents that modify the mechanical or biological properties of the membrane. The one or more agents are not limited and can be any agent that modifies the mechanical or biological properties of the membrane described herein. In some embodiments, the one or more agents are selected from glycerin, sorbitol, propylene glycol, plasticizers, fibers or other longitudinal elements, and growth factors (e.g., encapsulated growth factors).

[0093] In some embodiments, the method of producing a membrane further comprises adding one or more additional membrane layers to the first membrane layer by the methods disclosed herein to create a membrane of mixed composition or architecture, hi some embodiments, the two or more layers are produced from membrane solutions having different components, agents, and / or concentrations.

[0094] In some embodiments, the film is treated to remove the porogen, thereby forming pores in the film. The sacrificial porogen material is removed passively or forcibly in conjunction with dissolution, phase transition, reversal of thermal gelation, or other techniques known in the art. In some embodiments, the sacrificial material has thermally reversible gelation properties or can be dissolved in a non-polar solvent.

[0095] In some embodiments, the porogen material is Pluronic F127 and is removed by treatment with a non-polar solvent (eg, isopropanol).

[0096] In some embodiments, the membrane solution comprises 3-35% by weight of gelatin or gelatin-polymer complex, hi some embodiments, the solution comprises about 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, or 35% by weight of gelatin or gelatin-polymer complex.

[0097] In some embodiments, the thin film layer can be dried, gelled, crosslinked, or otherwise solidified and removed from the substrate. In some embodiments, the thin film layer is crosslinked using a solution containing glutaraldehyde, transglutaminase, or other crosslinking enzymes or molecules. In some embodiments, a crosslinking agent is added to the film solution, for example, just before applying the solution to the thin film on the substrate. In some embodiments, the crosslinking agent is contacted with the thin film layer after contact with the substrate. In some embodiments, the concentration of crosslinking agent is about 0.01 to 5 g per 10 g of gelatin.

[0098] In some aspects of the present disclosure, membranes are used in tissue or biological constructs incorporating membranes (e.g., basement membranes) produced by the methods described herein. In some embodiments, tissue or biological constructs containing membranes produced as described herein include hydrogels such as gelatin, collagen, PLA, chitosan, or hydrogel composites, or other hydrogel materials and compounds. Without limitation, 3-35 wt% gelatin and gelatin-polymer composites can be used using various thin film deposition techniques. Sacrificial porogen materials of varying concentrations and compositions can allow for substantial opportunities for tuning of mechanical and biological properties, including, but not limited to, porosity, pore size, permeability, sieving, filtration, and other functions that can enhance and further specialize tissue constructs for desired biological applications.

[0099] In some embodiments, the membranes described herein are modified via techniques such as divalent metal ion removal or other techniques known in the art to obtain tunable mechanical and biological properties (see Qi et al., Scientific Reports 4:4706 (2013)).

[0100] In some embodiments of the methods for producing membranes described herein, a second polymer or hydrogel material is produced that provides a support matrix for the membrane material. This second hydrogel or polymer may be constructed of the same material as the membrane or may be constructed of a complementary hydrogel or polymer.

[0101] In some embodiments, the membranes are constructed partially or completely of gelatin or other hydrogel materials that have been modified to be photocurable using various wavelengths of ultraviolet light, such as gelatin methacrylate. Various concentrations of such materials can be made using published protocols or techniques known in the art.

[0102] Those skilled in the art will readily appreciate that the present invention is well adapted to carry out the objects and obtain the ends and advantages mentioned, as well as those inherent therein. The details of the description and examples herein are representative of particular embodiments and are exemplary and are not intended as limitations on the scope of the invention. Modifications therein and other uses will occur to those skilled in the art. These modifications are encompassed within the spirit of the invention. It will be readily apparent to those skilled in the art that various substitutions and modifications can be made to the invention disclosed herein without departing from the scope and spirit of the invention.

[0103] As used herein in the specification and claims, the articles "a" and "an" should be understood to include plural referents unless clearly dictated to the contrary. A claim or description including "or" between one or more members of a group is deemed to be satisfied when one, more than one, or all of the group members are present in, employed in, or otherwise relevant to a given product or process, unless dictated to the contrary or otherwise evident from context. The invention includes embodiments in which exactly one member of a group is present in, employed in, or otherwise relevant to a given product or process. The invention also includes embodiments in which more than one, or all of the group members are present in, employed in, or otherwise relevant to a given product or process. Furthermore, it should be understood that the present invention provides for all variations, combinations, and permutations in which one or more limitations, elements, clauses, descriptive terms, etc. from one or more of the enumerated claims are introduced into another claim (or any other claim, if relevant) dependent on the same base claim, unless otherwise indicated or unless it is apparent to one of ordinary skill in the art that a contradiction or inconsistency would result. All embodiments described herein are contemplated as being applicable to all different aspects of the invention, where appropriate. It is also contemplated that any embodiment or aspect may be freely combined with one or more other such embodiments or aspects, as appropriate. When elements are presented as lists, e.g., in a Markush group or similar format, it is understood that each subgroup of elements is also disclosed, and that any element(s) may be removed from the group. In general, when the invention, or aspects of the invention, are referred to as including particular elements, features, etc., it should be understood that the particular embodiment or aspect of the invention consists of or consists essentially of such elements, features, etc. For purposes of brevity, those embodiments will not in all instances be specifically described in as much language herein.It should also be understood that any embodiment or aspect of the invention may be explicitly excluded from the claims, regardless of whether a specific exclusion is set forth herein. For example, any one or more active agents, additives, ingredients, optional drugs, organism types, disorders, subjects, or combinations thereof may be excluded.

[0104] Where a claim or description is directed to a composition of matter, it is understood that methods of making or using the composition of matter by any method disclosed herein, and methods of using the composition of matter for any purpose disclosed herein, are aspects of the invention, unless otherwise indicated or it is apparent to one of ordinary skill in the art that a contradiction or inconsistency would arise. Where a claim or description is directed to a method, it is understood that, for example, methods of making compositions useful in practicing the method, and products produced according to the method, are aspects of the invention, unless otherwise indicated or it is apparent to one of ordinary skill in the art that a contradiction or inconsistency would arise.

[0105] When ranges are given herein, the invention includes embodiments in which the endpoints are included, embodiments in which both endpoints are excluded, and embodiments in which one endpoint is included and the other is excluded. Unless otherwise indicated, both endpoints should be presumed to be included. Furthermore, unless otherwise indicated or apparent from the context and the understanding of one of ordinary skill in the art, it should be understood that values ​​expressed as ranges can take any specific value or subrange within the stated range in different embodiments of the invention, down to the tenth of the unit of the lower limit of the range, unless the context clearly dictates otherwise. It should also be understood that when a series of numerical values ​​is described herein, the invention similarly includes embodiments relating to any intermediate value or range defined by any two of the values ​​in the series, with the smallest value being the minimum and the largest value being the maximum. Numerical values ​​used herein include values ​​expressed as percentages. For any embodiment of the invention in which a numerical value is prefaced by "about" or "approximately," the invention includes embodiments in which the exact value is recited. For any embodiment of the invention where a numerical value is not prefaced by "about" or "approximately," the invention includes embodiments where the value is prefaced by "about" or "approximately."

[0106] As used herein, "A and / or B," when A and B are different claim terms, generally means at least one of A, B, or both A and B. For example, a sequence that is complementary to and / or hybridizes to another sequence includes (i) a sequence that is complementary to the other sequence, even though the sequence may not necessarily hybridize to the other sequence under all conditions, (ii) a sequence that hybridizes to the other sequence, even though the sequence is not fully complementary to the other sequence, and (iii) a sequence that is both complementary to and hybridizes to the other sequence.

[0107] "Approximately" or "about" generally includes numbers in either direction (greater or lesser than the number) within 1%, or in some embodiments, within 5% of the number, or in some embodiments, within 10% of the number, unless otherwise stated or clear from the context (unless such number would unacceptably exceed 100% of possible values). Unless expressly indicated to the contrary, in any method claimed herein that includes more than one act, the order of the method acts is not necessarily limited to the order in which the method acts are recited, although it should be understood that the invention includes embodiments in which the order is so limited. It should also be understood that any product or composition described herein can be considered "isolated" unless otherwise stated or clear from the context.

[0108] As used herein, the terms "comprising" or "comprises" are used in reference to compositions, methods, and their respective component(s) that are essential to the present invention, and still allow for the inclusion of unspecified elements, whether essential or not.

[0109] As used herein, the term "consisting essentially of" refers to elements required for a given embodiment. The term permits the presence of additional elements that do not materially affect the basic and novel or functional characteristic(s) of that embodiment of the invention.

[0110] The term "consisting of" refers to compositions, methods, and their respective components described herein, excluding any element not recited in that description of an embodiment. [Example]

[0111] Example 1 porous membrane manufacturing In step 1, a solution of gelatin, collagen, fibrin, or other biological or synthetic matrix material is made.

[0112] In step 2, the solution containing the sacrificial porogen material is combined with the matrix solution made in step 1 and mixed thoroughly.

[0113] In step 3, the matrix-porogen solution is deposited onto a substrate via spin coating, dip coating, or other such thin film deposition techniques and allowed to dry, gel, or otherwise solidify. This deposition technique allows for precise control over the thickness of the thin film.

[0114] In step 4, additional layers of similar or different composition are optionally deposited onto the first layer to create a composite or layered thin film, or the additional layers are deposited in a manner that allows for patterning or other spatial organization within the film.

[0115] In step 5, the sacrificial porogen material is removed through dissolution, decomposition, or other destructive techniques, leaving behind empty spaces in the thin film that act as pores.

[0116] In one example of the present invention, the sacrificial porogen material is composed of a self-assembling triblock copolymer (Pluronic), such as F127 or other poloxamer formulations that form micelles above a certain concentration. These micelles are incorporated into the matrix solution by mixing, sonication, or other methods to produce a sacrificial porogen homogeneously dispersed within the bulk matrix solution. Pore size can be controlled by using different polymers, but also by varying concentration, solution properties, and processing techniques to control micelle size and aggregation, as shown in Figure 1.

[0117] This matrix-porogen solution is then deposited onto a substrate via spin coating or other thin film deposition techniques, and depending on the technique and parameters, the thickness of the thin film can range anywhere from 0.1 to 100 μm. The thin film can be dried, gelled, crosslinked, or otherwise solidified and removed from the substrate. Once removed from the substrate, the sacrificial porogen material is passively or forcibly removed via dissolution, phase transition, thermal gelation reversal, or other techniques known in the art, creating an open pore structure in the thin film. Figure 2 shows one such example of this type of thin film.

[0118] Specific examples of applications of the present invention include tissues or biological constructs incorporating basement membranes produced by the described methods. Further examples include tissues or biological constructs containing membranes produced as described using hydrogels such as gelatin, collagen, PLA, chitosan, or hydrogel composites, or other hydrogel materials and compounds. Without limitation, gelatin and gelatin-polymer composites may be used in concentrations ranging from 3 to 35% by weight using various thin film deposition techniques. Varying concentrations and compositions of sacrificial porogen materials may allow for substantial opportunities for tuning mechanical and biological properties, including, but not limited to, porosity, pore size, permeability, sieving, filtration, and other features that can enhance and further specialize tissue constructs for desired biological applications.

[0119] Further examples of the present invention include membranes described in the previous examples that include composites of hydrogels, polymers, and materials that have been modified via techniques such as divalent metal ion removal or other techniques known in the art to provide tunable mechanical and biological properties (see Qi et al., Scientific Reports 4:4706 (2013)).

[0120] Further examples of the present invention include membranes described in the previous examples that include hydrogels, polymers, and composites of materials that have been modified through the addition of enhancers or compounds to provide tunable mechanical and biological properties. Exemplary techniques include, but are not limited to, the addition of glycerin, sorbitol, propylene glycol, or other plasticizers to gelatin or gelatin composite hydrogels (see F.M. Vanina et al., Food Hydrocolloids 19, 899-907 (2005)).

[0121] Further examples of the present invention include the membranes described in the previous examples with the addition of a second polymer or hydrogel material that provides a support matrix for the basement membrane material. This second hydrogel or polymer may be constructed of the same material as the basement membrane, or may be constructed of a complementary hydrogel or polymer.

[0122] Further examples of the present invention include films described in the previous examples where the thin films are composed partially or completely of gelatin or other hydrogel materials, such as gelatin methacrylate, that have been modified to be photocurable using various wavelengths of ultraviolet light. Various concentrations of such materials can be made using published protocols or techniques known in the art.

[0123] Further examples of the present invention include the membranes described in the previous examples, where the membranes are manufactured in a multi-step process to create membranes of mixed composition or architecture.

[0124] Further examples of the present invention include the membranes described in the previous examples, in which a hardening solution or compound is applied before, during, or after membrane formation, which acts to polymerize, gel, cure, or otherwise solidify the polymer or hydrogel material. For example, a membrane is produced and then subjected to a cross-linking solution, which may contain, but is not limited to, glutaraldehyde, transglutaminase, or other cross-linking enzymes or molecules at a concentration of 0.01 to 5 g per 10 g of gelatin. Alternatively, the cross-linking agent may be incorporated into the solution prior to membrane formation.

[0125] Further examples of the present invention include membranes as described in the previous examples, including hydrogels and polymers that are encapsulated or loaded with biological factors to promote cell and tissue growth.

[0126] Example 2 Implantable IABBP device An implantable IABBP device for renal replacement contains one or more functional units. Each of these functional units contains two separate channel systems (Figures 4 and 6) separated by extracellular matrix material and lined with cells. One channel system (referred to herein as the vascular channel or vascular channel system) is lined with endothelial cells and perfused with blood, which is gradually purified as it passes through the device. The second channel system (referred to herein as the filtrate channel or filtrate channel system) is lined with epithelial cells and perfused with filtrate, which is gradually processed. Blood flows from the artery into the vascular channel system and returns to the vein via a vascular conduit. Filtrate is produced by the device and flows through the filtrate channel. Once fully processed, the filtrate is drained into an extracorporeal collection system via a conduit and a surgically created fistula or into the patient's bladder via a conduit and a surgical anastomosis. Within the IABBP device, the channel network is composed of three segments that serve different functions. The segments are arranged in series based on the blood passing through the device so that the blood is processed sequentially by each segment before returning to the cardiovascular system.

[0127] In the first segment (referred to herein as the filtration segment), the IABBP device generates a primary ultrafiltrate through cell-enhanced ultrafiltration. A fraction of the blood (the filtrate fraction) is filtered from the vascular channel to the filtrate channel. Blood cells and larger molecules are retained in the vascular channel via cell- and matrix-mediated sieving, while water, glucose, urea toxins such as blood urea nitrogen (BUN), and other solutes are freely filtered.

[0128] The blood flow through the filtration segment is Q bf The arterial inflow Q is defined as A and filtered fraction FF f depends on (Q bf =Q A -(Q A *FF f The filtrate flow from the filtration segment is the primary filtrate flow (Q fI ), which is defined as the blood flow Q bf and filtered fraction FF f depends on (Q ff =Q bf *FF f ). The blood pressure during the filtration segment is P bf is defined as the blood flow Q bf , channel architecture, inflow blood pressure P bA , and back pressure P from the downstream vascular network bt It is a dynamic parameter determined by (Figure 5).

[0129] Similar to blood pressure, filtrate pressure during the filtration segment is a dynamic parameter determined by the channel architecture, filtrate flow, and backpressure from the downstream filtrate channel system (Figure 5).

[0130] In the second segment (referred to herein as the tubular segment), this primary ultrafiltrate then undergoes further modification via cell-enhanced solute secretion (active and passive transport into the filtrate via cells and extracellular matrix materials) and absorption (active and passive removal from the filtrate via cells and extracellular matrix materials) to produce the secondary filtrate. In this segment, cells also contribute to active metabolic regulation via bicarbonate synthesis.

[0131] The blood flow through the tubular segment is Q bt is defined as the inflow blood flow Q from the filtration segment. bf minus the relative filtration fraction of the tubular segment (Q bt =Q bf -(FF t *Q bf )). The filtrate flow from the tubular segment is the secondary filtrate flow Q fII is defined as the primary filtrate flow, the tubular filtration fraction, FF t , and canalicular blood flow Q bt depends on (Q fII =Q fI +(Q bt *FF t )). The blood pressure in the tubular segment is proportional to the inflow blood flow Q bf , channel architecture, inflow blood pressure P from the filtration segment bf , and back pressure P from the downstream vascular network bd It is a dynamic parameter determined by (Figure 5).

[0132] Similar to blood pressure, filtrate pressure in the tubular segments is P ft is defined as the influent filtrate flow rate, Q fI , channel architecture, and backpressure P from the downstream filtrate channel system fd It is a dynamic parameter determined by (Figure 5).

[0133] In the third segment (referred to herein as a conduit segment or conduit segment system), this secondary filtrate is then concentrated (removal of water from the filtrate via the cells and extracellular matrix) to produce a tertiary filtrate, which is then discharged from the device as described above.

[0134] The blood flow through the ductal segment is Q bd is defined as the inflow blood flow Q through the tubule segment. bt The absorption fraction of the ductal segment AF d is equal to the sum of (Q bd =Q bt +(AF d *Q fII )). Since the ductal segment is the terminal segment, the blood flow through the ductal segment, Q bd is the venous blood outflow Q bv is equal to (Q bd =Q bv ) The filtrate flow in the tubular segment is the tertiary filtrate flow Q fIII which is defined as the secondary filtrate flow Q fII and ductal absorption fraction AF d depends on (Q fII =Q fI +(Q bt *FF t )). The blood pressure in the ductal segment is proportional to the inflow blood flow Q bt , channel architecture, inflow blood pressure P from the tubular segment bt , and venous back pressure P bV It is a dynamic parameter determined by (Figure 5).

[0135] Similar to blood pressure, filtrate pressure in a ductal segment is P fd The channel architecture, defined as filtrate inflow Q from the tubular segment, ft , and back pressure P from the filtrate discharge system fb It is a dynamic parameter determined by (Figure 5).

[0136] In each segment, cells line various channels to support their respective functions (Figure 6).

[0137] In the filtration segment, endothelial cells (e.g., primary human glomerular endothelial cells, induced pluripotent stem cell (iPSC)-derived endothelial cells, and / or human umbilical cord endothelial cells) line the vascular channel system. These endothelial cells may form a fenestrated lining to enable filtration and sieving functions. The filtration channel system in the filtration segment is lined with epithelial cells (e.g., primary human podocytes, human iPSC-derived podocytes), which may further enhance filtration and sieving functions.

[0138] In the tubular segments, endothelial cells (e.g., primary human peritubular capillary endothelial cells, iPSC-derived endothelial cells, human umbilical endothelial cells) line the vascular channel system. The filtration channel system in the tubular segments is lined by epithelial cells (e.g., primary human renal tubular epithelial cells and / or iPSC-derived renal tubular epithelial cells) that allow the absorption and secretion of solutes and water.

[0139] In the ductal segments, endothelial cells (e.g., primary human renal medullary endothelial cells, iPSC-derived endothelial cells, and / or human umbilical endothelial cells) line the vascular channel system. The filtration channel system in the ductal segments is lined by epithelial cells (e.g., primary human renal tubular epithelial cells and / or iPSC-derived renal tubular epithelial cells) that allow for water reabsorption and concentration of the secondary filtrate to form the tertiary filtrate.

[0140] In each segment, the vascular channel and the filtrate channel are separated by a membrane that supports the function of each segment. (Figures 4 and 7) In the filtration segment, this membrane allows the formation of filtrate from the vascular space to the filtration space. In the canalicular segment, this membrane allows the exchange of solutes and water between the vascular channel system and the filtration channel system. In the conduit segment, this membrane allows the transfer of water and solutes from the filtration channel system to the vascular channel system. The membrane separating each channel system can be in the form of a collagen membrane 0.1 to 10 (e.g., 0.3 to 10) micrometers thick that supports cell adhesion on both sides and resists membrane fouling. The membrane can be porous to facilitate higher filtration or resorption rates or increased solute exchange. The membrane can be crosslinked to various degrees to alter its physical and biological properties.

[0141] The respective channel system architecture in each segment is tailored to provide a specific resistance and thus control the hydrostatic pressure in the vascular and filtrate channels to enable their respective functions. The channel system architecture is designed to minimize turbulence to reduce the risk of clot formation, cell activation, or blockage. The channel system may include one or more branching networks and sub-networks to increase its surface area. The channels may have varying diameters to provide uniform pressure across each segment and its membrane.

[0142] To increase functional capacity and meet the needs of various patients, functional units of the purification device can be stacked and function in parallel (Figure 8).

[0143] Table 1 lists examples of target functional specifications for clinically available IABBP devices. [Table 1]

[0144] Extracorporeal IABBP device In one example, the IABBP is not implanted but is maintained in a sterile, heated enclosure (bioreactor) and connected to the patient's blood circulation via an arteriovenous fistula or central venous cannulation. Blood can be delivered to the device with or without the assistance of a mechanical pump.

[0145] Example 3 - Membrane Fabrication In one example, to produce a membrane for hemofiltration, a gelatin matrix solution is prepared at a concentration of 5-30% by weight. This solution can be prepared by dissolving gelatin in water with PBS, cell culture medium, growth factors, or other enhancers. The solution is heated to 45°C to maintain the gelatin in a sol state.

[0146] To generate porous membranes, a specific volume of porogen solution containing 1-40 wt% of a self-assembling triblock copolymer (Pluronic F127) is added to a gelatin matrix solution. This combined matrix-porogen solution is then mixed via sonication to disperse F127 into the gelatin solution to a final concentration above the critical micelle concentration (CMC) and critical micelle temperature (CMT), allowing F127 to form micelles and micellar aggregates that can act as a sacrificial porogen. This combined matrix-porogen solution is then subsequently deposited onto a substrate via spin coating or other thin film deposition techniques; depending on the technique and parameters, film thicknesses can range anywhere from 0.1 to 10 μm. The precise concentration of F127 in the final membrane is used to control the membrane's pore size and bulk porosity, which directly correlates with the diffusion and filtration capabilities of the acellular membrane. This, in turn, determines the membrane's functional capabilities when cellularized. For example, to create a membrane that provides a functional filtration rate (0.2-2 ml per minute) in the context of the first segment (filtration segment) of the IABBP, a membrane was fabricated using 25 ml of 30 wt. % gelatin mixed with 3.15 ml of 35 wt. % F127 stock solution, providing a concentration of approximately 4.00 wt. % F127 in the working solution. This solution was then thoroughly mixed, degassed, and deposited onto a substrate via spin coating, allowing it to dry before removal and subsequent crosslinking. This membrane provides a range of filtration rates suitable for filtration function within the context of the IABBP (Figure 12), and the precise rate and overall porosity can be increased or decreased to achieve the desired function. Pore size and distribution can be analyzed using experimental methods such as fluorescent bead analysis using the membrane testing system described below, or in a more quantitative manner using pore image analysis, which can provide distribution and other data not available through experimental methods (Figure 13).

[0147] The thin film is then dried and removed from the substrate. At this point, the thin film is dry and uncrosslinked and can be incorporated into a scaffold or other such biological structure and crosslinked. This can be done by mounting the thin film in a frame, which allows for 3D printing or other deposition of sacrificial materials on either side of the membrane to create opposing channel networks. The thin film and channel construct can then be embedded in a scaffold and crosslinked. Once crosslinked, the sacrificial material can be removed and the channels perfused. Isopropanol or other nonpolar solvents that disrupt the micelle cores can be used to dissolve and remove the Pluronic F127 micelles in the membrane, which can be perfused through the scaffold to open pores in the membrane. Alternatively, the dried thin film can be crosslinked by immersion in glutaraldehyde solution or other crosslinking agents (such as transglutaminase). The sacrificial porogen material can then be removed by further immersion of the thin film in a solvent. Once the porogen material is removed, the thin film can be rinsed with PBS or other solution to remove any residual crosslinker or solvent, after which the cells can be repositioned as desired.

[0148] Cellularized or acellular membranes can be tested in an in vitro isolated membrane device (Figures 12A-12B). Briefly, an isolated membrane is placed between two support strips or mesh screens of high porosity and low resistance, designed to expose substantially the entire membrane surface area and allow unimpeded filtration or flow. These support strips or mesh screens are clamped in place between two halves of a chamber containing ports that allow perfusion with fluids, gases, or a combination of both to simulate in vivo membrane function. This type of chamber and test system allows for high-throughput, short- or long-term testing of cellularized or acellular membranes. Data generated by this system can be correlated with known membrane surface areas to provide functional data normalized to surface area, which can inform device design.

[0149] Example 4 - Fabrication of various channel patterns on thin films Channel architecture can be precisely controlled using extrusion-based 3D printing techniques, which print channels from a sacrificial material on either side of a membrane. The entire device is then embedded in an extracellular matrix material (e.g., a scaffold), and the sacrificial material is removed, resulting in two channel networks, one on either side of the membrane. The architecture and dimensions of the channels that make up these networks are designed using computer-aided design (CAD) software and then converted into G-code to control the 3D printing process. Channel diameter can be adjusted by varying the 3D printer motor speed, the amount of pressure-driven extrusion, and the temperature of the extruder head, all of which are controlled through the G-code and associated 3D printer electronics. For example, to create tapered channels of increasing diameter, the motor speed driving the extruder print head can be controlled to move incrementally slower along the length of the channel while holding all other parameters constant. Alternatively, the rate of printed material extrusion can be increased by increasing pressure-driven extrusion along the length of the channel while holding all other parameters constant. The interplay of these variable parameters can be calibrated and used to print desired channel configurations. Precise control of the channel geometry and architecture allows for control of hydrostatic pressure and flow patterns throughout the channel network and across the membrane.

[0150] Example 5 - Fabrication of a multilayer device Multiple layers of channel networks can be stacked to increase the functional surface area of ​​the IABBP device (Figure 9). The device's support scaffold is first assembled by pouring 20 mL of 20% gelatin, 10% transglutaminase (20 U / mL) into the bottom mold (i.e., the extracellular matrix material). The prepared inflow and outflow conduits are then filled with Pluronic F127 and placed into the recessed grooves of the bottom mold (Figure 10). The conduits are half-immersed in the gelatin. The gelatin in the mold is thermally and enzymatically crosslinked. While gelatin can be crosslinked at lower temperatures, this crosslinking can be reversed by warming the gelatin. Crosslinking using enzymes such as transglutaminase or glutaraldehyde is permanent. The thin film is sandwiched between two frames. The framed membrane is printed on one side with Pluronic F127 in the desired pattern using air from a pressure-controlled extrusion assembly. Once dry, the framed membrane is lifted, inverted, and printed on the other side with Pluronic F127 in a mirror image pattern, creating opposing channel networks across the membrane. The printed membrane is then air-dried. Once the printed membrane is dry, an additional 10 mL of 20% gelatin, 10% transglutaminase is poured into the mold. This gelatin layer should completely cover the conduits. The membrane is placed on the gelatin without entrapped air, for example by mechanical means, aligning the channel network with the conduits to create a continuous network. Once lowered into place, the membrane bonds to the gelatin. After the gelatin solidifies, excess membrane is cut from the frame, and the frame is removed. Another membrane is framed, printed, and dried, and a spacer is placed on top of the first layer. At this point, shafts or pillars of F127 can be added to the channel pattern on the lower membrane, aligning with and connecting to the upper membrane channel network, thereby creating a continuous multilayer network. If necessary, holes can be punched, dissolved, or otherwise removed from the membrane to allow for interlayer connection. Fill this next spacer with 10 mL of 20% gelatin, 10% transglutaminase and lower the next membrane into place. Lower the membrane onto the next layer, ensuring that the networks on each layer are aligned.This process can be repeated for as many layers as necessary. During assembly as described above, or after placing the final membrane, each layer may be connected via a small shaft, which may be filled with Pluronic F127 or another fluid or gel that can be removed. To connect the channel networks of the various layers, these connections are made along the height of the implant. Care is taken to connect allogeneic to allogeneic (e.g., blood vessels to blood vessels and filtrate to filtrate). A prepared conduit filled with Pluronic F127 is placed on the top membrane to contact the vascular or filtration channel system and provide a location for anastomosis or cannulation of the implant into the vasculature. A taller spacer with a notch for the conduit is then placed on top of the top membrane, and the mold is filled with 20 mL of 20% gelatin and 10% transglutaminase to seal in the channel network and complete the multilayer implant. Alternatively, internal connections between membrane layers can be made once the scaffold is fully assembled by using a punch or other instrument to remove the gelatin and create hollow shafts or other connections between layers. This space may be filled with Pluronic F127 and then plugged, filled, or otherwise sealed with gelatin or other material that can bond, crosslink, glue, or otherwise adhere in place to close any remaining holes, maintaining the integrity of the channel network.

[0151] Example 6 - Primary cell isolation from discarded kidneys Primary cells are isolated from human kidneys deemed unsuitable for transplantation. The renal cortex is manually separated from the medulla and minced into small pieces less than 2 mm in diameter. The minced tissue is digested in a type IV collagenase solution at a concentration of 200 U / mL for 1 hour at 37°C under constant agitation (150 RPM). After digestion, the collagenase in the tissue slurry is neutralized by adding fetal bovine serum to a final concentration of 10%, and the digest is filtered sequentially through sieves with pore sizes of 250 μm and 125 μm. Glomeruli are collected from the top of the 125 μm sieve, and tubules are collected from the flow-through. Both portions are deposited onto gelatin-coated plastic. The media formulation is designed to promote the maintenance of epithelial and endothelial cells and is identified in Table # (co-culture medium). After one week in culture, cells are harvested by EDTA treatment to obtain a single-cell suspension and further separated into specific cell types. Glomerular endothelial cells and podocytes are obtained from the glomerular portion by immunoisolation using CD31 and nephrin antibodies, respectively. Contaminating fibroblasts are depleted by Thy1 immunoisolation. From the flow-through, peritubular endothelial cells are separated from cortical epithelial cells (proximal and distal tubules) by CD31 and CK18 immunoisolation, respectively. Contaminating fibroblasts are also depleted from these portions by Thy1 immunoisolation. The medullary tissue is processed in a similar manner to collect ductal cells, which are isolated by L1CAM immunoisolation and depleted of fibroblasts using Thy1 immunoisolation. Media formulations for specific cell types are listed in Tables 2–4. [Table 2] [Table 3]

[0152] Example 7 - Cell seeding onto IABBP scaffolds The vascular and filtration channels are lined with a confluent monolayer of endothelial or epithelial cells, respectively, to provide their respective functions. To seed the cells into the channels, the inlet and outlet conduits to both networks are cannulated, and a suspension of the appropriate cell type is injected into the inlet conduit for either the vascular or filtration channel. Fluid is simultaneously drawn from each outlet conduit at the same rate as the cell suspension is injected until the entire network is filled with the cell suspension. The cells are allowed to attach to the channel walls in static culture in a 37°C incubator for at least 1 hour, after which the entire device is rotated 180° and both the vascular and filtration networks are reseeded with the same respective cell suspension to ensure complete channel lumen seeding. After this time, the networks are perfused with culture medium and left for at least 24 hours to achieve full confluence. Direct flow of cell culture medium, blood, or serum may be introduced into each channel network to provide nutrients and oxygen to the cells and enhance cellular function for both endothelial and epithelial cells. Flow can be controlled using either a pump or gravity-driven flow.

[0153] To seed multiple cell types into a single channel network, each type must be seeded sequentially based on its location in the scaffold. It is important to note that the scaffold channel network can be perfused in either direction, allowing for this sequential seeding. To seed podocytes and other glomerular epithelial cell types in segment 1, followed by tubular cells in segment 2 and distal and collecting duct cells in segment 3, each cell type or mixture of cell types must be suspended in a volume proportional to the volume of the segment they occupy.

[0154] Once the cells are in dense suspension, the cells intended for segment 1 can be injected through the cannula adjacent to segment 1, resulting in the entirety of segment 1 being filled with the cell suspension and segment 2 being filled with the cell-free fluid previously located in segment 1. The cells are allowed to adhere for 30 minutes, up to 3 hours, or longer, and a volume of fluid equal to the suspension volume is injected through the cannula adjacent to segment 3 to flush out any remaining cell suspension media and any cells that have not attached in segment 1. The scaffold is then inverted and the procedure repeated to provide complete coverage of segment 1 with the desired cell type.

[0155] A second cell suspension containing cells for Segment 2, a volume equal to the volume of the Segment 2 channel, is then injected into the scaffold through the cannula adjacent to Segment 1. A volume of solution equal to the volume of the Segment 1 channel without cells is then injected into the cannula adjacent to Segment 1, such that the cell suspension for Segment 2 is forced through the channel, filling Segment 2 but not entering Segment 3. The cells are allowed to adhere for 30 minutes, up to 3 hours. A volume of cell-free solution equal to or greater than the combined volume of Segments 1 and 2 is then injected through the cannula adjacent to Segment 3 to flush out any remaining cell suspension media and unattached cells. The scaffold is then inverted, and the procedure is repeated to provide complete coverage of Segment 2 with the desired cell type.

[0156] A third cell suspension containing cells for Segment 3, a volume equal to the volume of the Segment 3 channel, is then injected into the scaffold through the cannula adjacent to Segment 1. A volume of cell-free solution equal to the combined volume of the Segment 1 and Segment 2 channels is then injected into the cannula adjacent to Segment 1 so that the cell suspension for Segment 3 is forced through the channels to fill Segment 3. The cells are allowed to adhere for 30 minutes, up to 3 hours. A volume of cell-free solution equal to or greater than the combined volume of Segments 1, 2, and 3 is then injected through the cannula adjacent to Segment 3 to flush out any remaining cell suspension media and unattached cells. The scaffold is then inverted, and the procedure is repeated to provide complete coverage of Segment 3 with the desired cell type.

[0157] At this point, the scaffold channel network is fully lined with the desired cell types arranged in the desired segments to provide coordinated function throughout the device. This process is repeatable and can be expanded to include additional segments. The order in which the segments are filled with cells can also be rearranged as needed. Furthermore, the order of inversion of the scaffold can also be changed as needed. For example, all the cells in each segment can be incrementally attached to a membrane or matrix (e.g., a gelatin matrix), followed by a single inversion of the membrane to attach cells to each of the other segments. This type of sequential cell seeding can be achieved in other ways using known channel volumes and not necessarily in the order or manner described herein; cells can be added in sequential order via unidirectional perfusion without reverse perfusion, allowing the cell suspension and unattached cells to be backflushed.

[0158] Example 8 - In silico modeling of the IABBP device and its function, and an in vitro model of the IABBP membrane A virtual model was generated using SolidWorks (SW). The 3D point cloud was transferred from the 3D printer to Geomagic and then to SW. The model in SolidWorks allowed us to simulate the device under various clinical conditions to better understand its performance. For example, applying 65 mmHg to the inflow conduit allowed us to calculate the ultrafiltration rate of the device on the outflow side. Membranes were inserted into the device and given performance characteristics dictated by an in vitro membrane testing device, which allows benchtop testing of various acellular and cellular membranes (Figures 12A-12B). These key characteristics drove and validated the output numbers obtained from the model. This allowed us to adjust the surface area and the number of repeated layers, two key parameters of the device for parallel blood purification.

[0159] Example 9 - Fiber membrane production In step 1, a solution of gelatin, collagen, fibrin, or other biological or synthetic matrix material is prepared. In step 2, dried silk nano- and microscale fibers are added to the matrix solution and mixed by stirring. The size of the fibers depends on the preparation, and the size composition of the fiber components can be adjusted to produce desired mechanical properties. The fiber components may be crosslinked to themselves or may be attached to the matrix components in a subsequent step to create an interpenetrating network of matrix and fibers. In step 3, a solution containing a sacrificial porogen material is combined with the matrix and fiber solution prepared in step 1 and thoroughly mixed. In step 4, the matrix-porogen solution is deposited onto a substrate via spin coating, dip coating, or other such thin film deposition techniques and allowed to dry, gel, or otherwise solidify. This deposition technique allows for precise control over the thickness of the thin film. In step 5, additional layers of similar or different composition are optionally deposited on the first layer to create a composite or layered thin film. Alternatively, additional layers are deposited in a manner that allows for patterning or other spatial organization within the film. In step 6, the sacrificial porogen material is removed through dissolution, decomposition, or other destructive techniques, leaving behind empty spaces in the thin film that act as pores.

Claims

1. 1. A device for restoring or supplementing pancreatic function, comprising: The apparatus comprises a functional unit: The functional unit comprises: a membrane having a first surface and a second surface; a first channel system comprising a first luminal space, the first luminal space being adhered to and in fluid communication with the first surface of the membrane, the first channel system comprising a first end configured to fluidly connect to a fluid supply and a second end configured to fluidly connect to a first fluid outlet; a second channel system comprising a second luminal space, the second luminal space being adhered to and in fluid communication with the second surface of the membrane, the second channel system comprising a third end configured to fluidly connect to a second fluid outlet; Equipped with the first channel system and the second channel system are in fluid communication with each other across the membrane; each of the first channel system and the second channel system includes a first channel system wall and a second channel system wall; The second channel system is populated with pancreatic islet cells, including beta cells, to generate functional viable pancreatic tissue adhered to the second channel system walls.

2. 10. The device of claim 1, wherein the membrane comprises a biocompatible extracellular matrix membrane separating the first channel system from the second channel system, the biocompatible extracellular matrix membrane being embedded in a matrix material.

3. The device of claim 2 , wherein the biocompatible extracellular matrix membrane comprises a collagen membrane, the collagen membrane supporting cell adhesion on both a first surface and a second surface of the collagen membrane.

4. 4. The device of claim 3, wherein the membrane has a thickness of between 0.1 micrometers and 20 micrometers (0.1 μm and 20 μm).

5. 10. The device of claim 1, wherein the membrane is configured to have a minimum shear stress of 0.5 Pascals and / or a minimum transmembrane pressure difference of 1333.22 Pascals.

6. 2. The device of claim 1, wherein the functional unit comprises at least one biological fluid inflow conduit in fluid communication with the first end of the first channel system and the first lumen space, and at least one biological fluid outflow conduit in fluid communication with the second end of the first channel system and the first lumen space, wherein the at least one biological fluid inflow conduit is in fluid communication with an arterial conduit and the at least one biological fluid outflow conduit is in fluid communication with a vascular conduit.

7. The device of claim 1 , wherein the first luminal space and the second luminal space are embedded in a scaffold.

8. 10. The device of claim 1, wherein the first channel system walls are lined with endothelial cells selected from induced pluripotent stem cell (iPSC)-derived endothelial cells and / or human umbilical cord endothelial cells.

9. The device of claim 8 , wherein the endothelial cells are allogeneic or autologous to the patient using the device.

10. The device of claim 1 , wherein the pancreatic islet cells are allogeneic or autologous to the patient using the device.

11. the apparatus comprises a plurality of functional units, the plurality of functional units including the functional unit and an additional functional unit of the same configuration; Each functional unit of the plurality of functional units comprises: a first end of a first channel system and a first lumen space in fluid communication with the at least one biological fluid inflow conduit; a second end of the first channel system in fluid communication with the at least one biological fluid outflow conduit and a first lumen space; a third end of the second channel system in fluid communication with the second outflow conduit and the second lumen space; and a first end of each of the plurality of functional units individually connected in parallel to one of a plurality of manifold ports of the at least one biological fluid inlet conduit; a second end of each of the plurality of functional units individually connected in parallel to one of a plurality of manifold ports of the at least one biological fluid outlet conduit; 7. The apparatus of claim 6, wherein the third ends of each of the plurality of functional units are individually connected in parallel to one of a plurality of manifold ports of the second outlet conduit.

12. 12. The apparatus of claim 11, wherein the apparatus comprises the plurality of functional units, the plurality of functional units including the functional unit and the additional functional unit of the same configuration stacked in parallel layers of functional units.

13. 10. The device of claim 1, wherein the device is configured for extracorporeal manipulation in a sterile, heated enclosure.

14. The device of claim 1 , wherein blood is delivered to the device using a mechanical pump.

15. 10. The device of claim 1, wherein the device is disposed within a capsule, the device being sized and configured for placement within a human body to replace or augment pancreatic function.

16. The device of claim 1 , wherein the membrane comprises a porous membrane including pores arranged to interconnect the first surface and the second surface.

17. 17. The device of claim 16, wherein the pores have a diameter of between 1 μm and 15 μm.

18. The device of claim 8 , wherein the device is fluidly connected to a patient's circulatory system.

19. 10. A method for manufacturing the device of claim 1, comprising: providing a membrane having a sacrificial material in the form of a first channel system on a first surface and a sacrificial material in the form of a second channel system on a second surface; soaking the membrane in a solution containing a scaffold material; gelling the scaffold material; removing the sacrificial material to form luminal spaces and walls of the first channel system and the second channel system; and populating said second channel system with pancreatic islet cells, including beta cells. A method comprising:

20. 20. The method of claim 19, wherein the method further comprises populating the first channel system of the membrane with endothelial cells.

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