Engineered Tissues for in vitro Research Uses, Arrays Thereof, and Methods of Making the Same

Abstract

Disclosed are living, three-dimensional tissue constructs for in vitro scientific and medical research, arrays thereof, and methods of making said tissues and arrays.

1 - 52 . (canceled) 
     
     
         53 . An in vitro living, three-dimensional tissue construct comprising:
 (a) at least one adherent cell type;   (b) a vascular or pseudo vascular network and cells that are not part of the vascular or pseudo vascular network; and   (c) a multi-layered architecture, wherein each layer of the multi-layered architecture is in direct contact with at least one other layer, and wherein the layers are cohered to form the construct;   wherein the tissue construct is free of pre-formed scaffold at the time of bioprinting and at the time of use in an in vitro assay.   
     
     
         54 . The tissue construct of  claim 53 , wherein the tissue construct comprises at least one layer comprising a plurality of cell types arranged relative to each other to create a planar geometry. 
     
     
         55 . The tissue construct of  claim 53 , wherein the tissue construct comprises at least one layer that is compositionally or architecturally distinct from at least one other layer to create a laminar geometry. 
     
     
         56 . The tissue construct of  claim 53 , further comprising a non-adherent cell type. 
     
     
         57 . The tissue construct of  claim 53 , wherein the tissue construct is at least about 25 μm in its smallest dimension at the time of bioprinting, wherein the tissue construct is no greater than about 3 cm in its largest dimension at the time of bioprinting, or both. 
     
     
         58 . The tissue construct of  claim 53 , wherein the at least one adherent cell type is a non-differentiated cell. 
     
     
         59 . The tissue construct of  claim 53 , wherein the at least one adherent cell type originated from a tissue selected from the group consisting of: liver, gastrointestinal, pancreatic, kidney, lung, tracheal, vascular, skeletal muscle, cardiac, skin, smooth muscle, connective tissue, corneal, genitourinary, breast, reproductive, endothelial, epithelial, fibroblast, neural, Schwann, adipose, bone, bone marrow, cartilage, pericytes, mesothelial, endocrine, stromal, lymph, blood, endoderm, ectoderm, and mesoderm. 
     
     
         60 . The tissue construct of  claim 53 , wherein the tissue construct is a vascular wall segment. 
     
     
         61 . An array of living, three-dimensional tissue constructs, wherein each tissue construct in the array is a tissue construct of  claim 53 . 
     
     
         62 . A method of producing a living, three-dimensional tissue construct for in vitro use comprising:
 a. bioprinting multiple layers by extrusion of at least one semi-solid or solid bio-ink comprising at least one adherent cell type, wherein at least one semi-solid or solid bio-ink comprises endothelial cells, and wherein each layer is in direct contact with at least one other layer; and   b. cohering the layers to form a living, three-dimensional tissue construct comprising a vascular or pseudo vascular network and cells that are not part of the vascular or pseudo vascular network;   wherein the tissue construct is free of pre-formed scaffold at the time of bioprinting and at the time of use in an in vitro assay.   
     
     
         63 . The method of  claim 62 , wherein the bio-ink comprises multicellular aggregates comprising mammalian cells. 
     
     
         64 . The method of  claim 62 , wherein the bioprinting further comprises arranging a plurality of cell types relative to each other to create a planar geometry in at least one layer. 
     
     
         65 . The method of  claim 62 , comprising bioprinting at least one layer that is compositionally or architecturally distinct from at least one other layer to create a laminar geometry. 
     
     
         66 . The method of  claim 62 , wherein the tissue construct is at least about 25 μm in its smallest dimension at the time of bioprinting, wherein the tissue construct is no greater than about 3 cm in its largest dimension at the time of bioprinting, or both. 
     
     
         67 . The method of  claim 62 , wherein the at least one adherent cell type, the endothelial cells, or both are differentiated cells. 
     
     
         68 . The method of  claim 62 , wherein the at least one adherent cell type, the endothelial cells, or both are non-differentiated cells. 
     
     
         69 . The method of  claim 62 , comprising isolating the at least one adherent cell type from a tissue selected from the group consisting of: liver, gastrointestinal, pancreatic, kidney, lung, tracheal, vascular, skeletal muscle, cardiac, skin, smooth muscle, connective tissue, corneal, genitourinary, breast, reproductive, endothelial, epithelial, fibroblast, neural, Schwann, adipose, bone, bone marrow, cartilage, pericytes, mesothelial, endocrine, stromal, lymph, blood, endoderm, ectoderm, mesoderm. 
     
     
         70 . The method of  claim 62 , comprising isolating the endothelial cells from a tissue selected from the group consisting of: blood, blood vessel, lymphatic vessel, tissue of the digestive tract, tissue of the genitourinary tract, adipose tissue, tissue of the respiratory tract, tissue of the reproductive system, mesoderm-derived tissue, bone marrow, and umbilical tissue. 
     
     
         71 . The method of  claim 62 , wherein the tissue construct is a vascular wall segment. 
     
     
         72 . The method of  claim 62 , comprising producing multiple tissue constructs and constructing an array from the multiple tissue constructs.