Compositions and methods of functionally enhanced in vitro cell culture system

Abstract

Compositions and methods described herein provide a cell culture system in which cells are in high metabolic states from the onset of the culture. Combinations of various cell culture components disclosed and employed herein allow cells to be in high metabolic states useful for drug testing immediately after the start of cell culture.

What is claimed is: 
     
         1 . A cell culture system comprising at least one compartment for culturing cells, a culture of a first population of metabolically active cells and at least two cell culture components selected from:
 (i) a cell culture environment with an oxygen concentration higher than the atmospheric concentration;   (ii) a second cell population for co-culture with said first population of cells and/or a structure configured for three-dimensional culture of said first population of cells with or without a second cell population for co-culture with said first population of cells;   (iii) a serum-free culture medium or culture medium with a low concentration of serum; and   (iv) a cell culture medium that is configured to contact or come into proximity with said first population of cells under at least one condition of actuated perfusion or flow.   
     
     
         2 . The cell culture system of  claim 1 , wherein said first population of cells comprise hepatocytes. 
     
     
         3 . The cell culture system of  claim 1 , wherein said structure for three-dimensional culturing comprises a gel sandwich culture, a gel overlay culture, a micropatterned overlay culture, a scaffold, a tissue slice, a tissue segment culture, or an artificial tissue construct. 
     
     
         4 . The cell culture system of  claim 1 , wherein said second cell population comprises fibroblasts, glial cells, endothelial cells, non-parenchymal cells, or stromal cells. 
     
     
         5 . The cell culture system of  claim 1 , wherein said cell culture environment comprises about 95% oxygen and about 5% CO 2 . 
     
     
         6 . The cell culture system of  claim 1  comprising a cell culture environment comprising about 95% oxygen and about 5% CO 2 , a three-dimensional culture comprising hepatocytes and fibroblasts and a serum-free culture media. 
     
     
         7 . The cell culture system of  claim 1  further comprising a cell binding material on said compartment for attachment of said cells. 
     
     
         8 . The cell culture system of  claim 1  further comprising at least one subcellular component contained in the at least one compartment or another separate compartment. 
     
     
         9 . The cell culture system of  claim 1  wherein said culture medium is circulated to said at least one compartment under actuated flow or perfusion through at least one microfluidic channel. 
     
     
         10 . A method of culturing a first population of metabolically active cells comprising culturing said cells in the presence of at least two cell culture components selected from:
 (i) a cell culture environment with an oxygen concentration higher than the atmospheric concentration;   (ii) a second cell population for co-culture with said first population of cells and/or a structure configured for three-dimensional culture of said first population of cells with or without a second cell population for co-culture with said first population of cells;   (iii) a serum-free culture medium or culture medium with a low concentration of serum; and   (iv) a cell culture medium that is configured to contact or come into proximity with said first population of cells under at least one condition of actuated perfusion or flow.   
     
     
         11 . The method of  claim 10 , wherein said first population of cells comprises hepatocytes. 
     
     
         12 . The method of  claim 10  wherein said first population comprises kidney cells or keratinocytes. 
     
     
         13 . The method of  claim 10 , wherein said oxygen concentration is higher than atmospheric for the seeding of said first population of cells. 
     
     
         14 . The method of  claim 10 , wherein said structure for three-dimensional culturing comprises a gel sandwich culture, a gel overlay culture, a micropatterned overlay culture, a scaffold, a tissue slice, a tissue segment culture, or an artificial tissue construct. 
     
     
         15 . The method of  claim 10 , wherein said second cell population comprises stromal cells, non-parenchymal cells or immune cells. 
     
     
         16 . The method of  claim 10 , wherein said cell culture environment comprises about 95% oxygen and about 5% CO 2 . 
     
     
         17 . The method of  claim 10 , wherein primary cell hepatocytes are seeded onto a three-dimensional cell culture structure together with secondary cells comprising fibroblasts in an environment comprising about 95% oxygen and about 5% CO 2  and cultured in a serum-free medium. 
     
     
         18 . The method of  claim 10 , wherein said compartment further comprises a coating of a binding material for attachment of said cells. 
     
     
         19 . A method of screening a material for its pharmacologic, metabolic, pharmacokinetic, or toxicological properties comprising culturing a first population of metabolically active cells in the presence of at least two cell culture components selected from:
 (i) a cell culture environment with an oxygen concentration higher than the atmospheric concentration;   (ii) a second cell population for co-culture with said first population of cells and/or a structure configured for three-dimensional culture of said first population of cells with or without a second cell population for co-culture with said first population of cells;   (iii) a serum-free culture medium or culture medium with a low concentration of serum; and   (iv) a cell culture medium that is configured to contact or come into proximity with said first population of cells under at least one condition of actuated perfusion or flow.   
     
     
         20 . The method of  claim 19 , further comprising measuring an activity of said first population of cells. 
     
     
         21 . The method of  claim 19 , wherein said primary cells comprise hepatocytes. 
     
     
         22 . The method of  claim 19 , wherein said primary cells comprise kidney cells or keratinocytes. 
     
     
         23 . The method of  claim 20 , wherein said activity of said first cells comprises a metabolite, a biomarker, gene expression, or protein activity. 
     
     
         24 . The method of  claim 19 , wherein said oxygen concentration is higher than atmospheric for the seeding of said primary cells. 
     
     
         25 . The method of  claim 19 , wherein said structure for three-dimensional culturing comprises a gel sandwich culture, a gel overlay culture, a micropatterned overlay culture, a scaffold, a tissue slice, a tissue segment culture, or an artificial tissue construct. 
     
     
         26 . The method of  claim 19  further comprising contacting said first cells or the cell culture media from said first cells with a subcellular component. 
     
     
         27 . The method of  claim 26 , further comprising measuring an activity of said subcellular component. 
     
     
         28 . A kit for culturing cells comprising:
 (i) a device for culturing cells containing at least one compartment, each compartment filled with serum-free cell culture medium;   (ii) a vial of a first population of cryopreserved metabolically active cells;   (iii) a vial of a second population of cryopreserved cells; and   (iv) a canister of gases comprising 95% oxygen and 5% CO 2 .   
     
     
         29 . The kit of  claim 28 , wherein said first population of cells comprise hepatocytes. 
     
     
         30 . The kit of  claim 28 , further comprising a scaffold for three-dimensional cell growth at least one compartment. 
     
     
         31 . The kit of  claim 28 , wherein said device comprises a microtiter plate. 
     
     
         32 . The kit of  claim 28 , wherein said device comprises a chip with at least one microfluidic channel adapted to flow culture media through said at least one compartment.