Method of printing a tissue construct with embedded vasculature

Abstract

A printed tissue construct comprises one or more tissue patterns, where each tissue pattern comprises a plurality of viable cells of one or more predetermined cell types. A network of vascular channels interpenetrates the one or more tissue patterns. An extracellular matrix composition at least partially surrounds the one or more tissue patterns and the network of vascular channels. A method of printing a tissue construct with embedded vasculature comprises depositing one or more cell-laden filaments, each comprising a plurality of viable cells, on a substrate to form one or more tissue patterns. Each of the one or more tissue patterns comprises one or more predetermined cell types. One or more sacrificial filaments, each comprising a fugitive ink, are deposited on the substrate to form a vascular pattern interpenetrating the one or more tissue patterns. The vascular pattern and the one or more tissue patterns are at least partially surrounded with an extracellular matrix composition. The fugitive ink is then removed to create vascular channels in the extracellular matrix composition, thereby forming an interpenetrating vascular network in a tissue construct.

1 .- 24 . (canceled) 
     
     
         25 . A three-dimensionally (3D) printed tissue construct with embedded vasculature, the printed tissue construct comprising:
 one or more tissue patterns, each tissue pattern comprising a plurality of viable cells of one or more predetermined cell types produced by depositing on a substrate one or more cell-laden filaments each comprising a plurality of viable cells:   a network of interconnected vascular channels of various sizes interpenetrating the one or more tissue patterns, wherein the lamest channels of the network provide a single inlet and a single outlet for perfusion, while the smallest channels of the network reduce the characteristic diffusion distance between adjacent conduits; and   an extracellular matrix composition at least partially surrounding the one or more tissue patterns and the interconnected network of vascular channels.   
     
     
         26 . The 3D printed tissue construct of  claim 25 , comprising up to n predetermined cell types, where n is a positive integer satisfying 2≤n≤300. 
     
     
         27 . The 3D printed tissue construct of  claim 25 , wherein the viable cells are mammalian cells selected from the group consisting of: germ cells, somatic cells and stem cells. 
     
     
         28 . The 3D printed tissue construct of  claim 25 , wherein the extracellular matrix composition comprises a synthetic or naturally derived biomaterial. 
     
     
         29 . The 3D printed tissue construct of  claim 28 , wherein the extracellular matrix composition comprises a hydrogel. 
     
     
         30 . The 3D printed tissue construct of  claim 28 , wherein the extracellular matrix composition comprises at least one of gelatin, fibrin, or gelatin methacrylate. 
     
     
         31 . The 3D printed tissue construct of  claim 30 , wherein the extracellular matrix composition further comprises a microgel. 
     
     
         32 . The 3D printed tissue construct of  claim 25 , wherein the extracellular matrix composition fully surrounds the network of vascular channels. 
     
     
         33 . The 3D printed tissue construct of  claim 25 , wherein the viable cells are distributed uniformly throughout each of the one or more tissue patterns. 
     
     
         34 . The 3D printed tissue construct of  claim 25 , wherein at least some of the viable cells of each of the one or more tissue patterns are disposed at a distance of no greater than about 1 mm from one or more of the vascular channels. 
     
     
         35 . The 3D printed tissue construct of  claim 34 , wherein the distance is no greater than about 300 microns. 
     
     
         36 . The 3D printed tissue construct of  claim 25 , wherein each of the one or more tissue patterns is defined by an arrangement of one or more cell-laden filaments comprising the viable cells. 
     
     
         37 . The 3D printed tissue construct of  claim 36 , wherein the one or more cell-laden filaments further comprise an extracellular matrix material. 
     
     
         38 . The 3D printed tissue construct of  claim 37 , wherein the extracellular matrix material comprises one or more of gelatin, fibrin, and gelatin methacrylate. 
     
     
         39 . The 3D printed tissue construct of  claim 25 , wherein at least one of the one or more cell-laden filaments further comprise one or more functional chemical substances selected from the group consisting of: drugs, small molecules, toxins, proteins, and hormones. 
     
     
         40 . The 3D printed tissue construct of  claim 39 , wherein the extracellular matrix composition comprises the one or more additives, the one or more additives having diffused from the one or more cell-laden filaments. 
     
     
         41 . The 3D printed tissue construct of  claim 25 , wherein the interconnected network of vascular channels has a branching structure, the interconnected network comprising one or more bifurcations. 
     
     
         42 . The 3D printed tissue construct of  claim 25 , wherein one or more of the vascular channels is curvilinear. 
     
     
         43 . The 3D printed tissue construct of  claim 25 , wherein one or more of the vascular channels has a nonuniform diameter along the length thereof. 
     
     
         44 . The 3D printed tissue construct of  claim 25 , wherein one or more of the vascular channels comprise an endothelial layer thereon, the endothelial layer comprising at least about 70% confluency. 
     
     
         45 . The 3D printed tissue construct of  claim 44 , wherein the endothelial layer comprises 100% confluency. 
     
     
         46 . The 3D printed tissue construct of  claim 45 , wherein one or more of the vascular channels further comprise a stromal layer or a smooth muscle cell layer on the endothelial layer. 
     
     
         47 . The 3D printed tissue construct of  claim 25 , further comprising one or more functional channels in the extracellular matrix composition. 
     
     
         48 . The 3D printed tissue construct of  claim 47 , wherein the one or more functional channels comprise an epithelial layer thereon. 
     
     
         49 . The 3D printed tissue construct of  claim 48 , wherein the one or more functional channels further comprise a stromal layer on the epithelial layer. 
     
     
         50 . The 3D printed tissue construct of  claim 25 , further comprising an interface structure at least partially surrounding the extracellular matrix composition and comprising flow channels in fluid communication with the interconnected network of vascular channels. 
     
     
         51 - 56 . (canceled) 
     
     
         57 . The 3D printed tissue construct of  claim 25 , further comprising one or more functional channels comprising an epithelial layer thereon, wherein the extracellular matrix composition at least partially surrounds the one or more functional channels. 
     
     
         58 . The 3D printed tissue construct of  claim 57 , wherein the one or more functional channels further comprises a stromal layer on the epithelial layer. 
     
     
         59 . (canceled) 
     
     
         60 . The 3D printed tissue construct of  claim 27 , wherein the viable cells and the one or more predetermined cell types comprise epithelial cells. 
     
     
         61 . (canceled) 
     
     
         62 . The 3D printed tissue construct of  claim 57 , wherein the tissue construct is an epithelial tissue construct selected from the group consisting of nephron, intestine, milk duct, urethra, pancreatic duct, and lymph.