Methods and compositions for isolation and growth of kidney tubule stem cells, in vitro kidney tubulogenesis and ex vivo construction of renal tubules

Abstract

Methods, including culture media conditions, which provide for isolation and purification of renal tubule stem cells and for in vitro kidney tubulogenesis are disclosed. The methods rely on culturing adult kidney cells in a culture media treated with combinations of transforming growth factor-β 1 , epidermal growth factor, and all-trans retinoic acid.

What is claimed as new and desired to be secured by Letters Patent of the United States is:  
     
         1 . An ex vivo renal tubule tissue system prepared by a process comprising culturing adult kidney cells in a culture medium containing all-trans retinoic acid and either epidermal growth factor or transforming growth factor in an amount effective for achieving tubulogenesis.  
     
     
         2 . The renal tubule tissue system of  claim 1 , wherein said cells are further treated with transforming growth factor-β 1 .  
     
     
         3 . The renal tubule tissue system of  claim 2 , wherein said culture medium contains a soluble factor selected from the group consisting of fetal calf serum, prostaglandins, hydrocortisone, triodothyronine, selenium, fibroblastic growth factor, hepatocyte growth factor and combinations thereof.  
     
     
         4 . The renal tubule tissue system of  claim 2 , wherein said culture medium contains an insoluble factor selected from the group consisting of Type I collagen, Type IV collagen, laminin, proteoglycans, fibronectin, and combinations thereof.  
     
     
         5 . The renal tubule tissue system of  claim 2 , wherein said culture medium contains a soluble factor selected from the group consisting of fetal calf serum, prostaglandins, hydrocortisone, triodothyronine, selenium, fibroblastic growth factor, hepatocyte growth factor, and combinations thereof, and an insoluble factor selected from the group consisting of Type I collagen, Type IV collagen, laminin, proteoglycans, fibronectin, and combinations thereof.  
     
     
         6 . A transformed renal tubule cell produced by the process of culturing renal tubule cells in a culture medium containing with all-trans retinoic acid and either epidermal growth factor or transforming growth factor-α in an amount effective to achieve tubulogenesis, in the presence of a cell transformation vector.  
     
     
         7 . The transformed renal tubule cell of  claim 6 , wherein said cells are further treated with transforming growth factor-β 1 .  
     
     
         8 . The transformed renal tubule cell of  claim 7 , wherein said culture medium contains a soluble factor selected from the group consisting of fetal calf serum, protaglandins, hydrocortisone, triodothyronine, selenium, fibroblastic growth factor, hepatocyte growth factor and combinations thereof.  
     
     
         9 . The transformed renal tubule cell of  claim 7 , wherein said culture medium contains an insoluble factor selected from the group consisting of Type I collagen, Type IV collagen, laminin, proteoglycans, fibronectin, and combinations thereof.  
     
     
         10 . The transformed renal tubule cell of  claim 7 , wherein said culture medium contains a soluble factor selected from the group consisting of fetal calf serum, prostaglandins, hydrocortisone, triodothyronine, selenium, fibroblastic growth factor, hepatocyte growth factor, and combinations thereof, and an insoluble factor selected from the group consisting of Type I collagen, Type IV collagen, laminin, proteoglycans, fibronectin, and combinations thereof.  
     
     
         11 . A method for growing kidney tubule stem cells ex vivo, comprising culturing adult kidney cells in the presence of all-trans retinoic acid, and either epidermal growth factor or transforming growth factor-α, in an amount effective for growing kidney tubule stem cells.  
     
     
         12 . The method of  claim 11 , wherein said cells are further treated with transforming growth factor-β 1 .  
     
     
         13 . The method of  claim 12 , wherein said culture medium contains a soluble factor selected from the group consisting of fetal calf serum, prostaglandins, hydrocortisone, triodothyronine, selenium, fibroblastic growth factor, hepatocyte growth factor and combinations thereof.  
     
     
         14 . The method of  claim 12 , wherein said culture medium contains an insoluble factor selected from the group consisting of Type I collagen, Type IV collagen, laminin, proteoglycans, fibronectin, and combinations thereof.  
     
     
         15 . The method of  claim 12 , wherein said culture medium contains a soluble factor, selected from the group consisting of fetal calf serum, prostaglandins, hydrocortisone, triodothyronine, selenium, fibroblastic growth factor, hepatocyte growth factor, and combinations thereof, and an insoluble factor selected from the group consisting of Type I collagen, Type IV collagen, laminin, proteoglycans, fibronectin, and combinations thereof.  
     
     
         16 . A method for effecting tubulogenesis in a renal cell culture, comprising culturing adult kidney cells in the presence of all-trans retinoic acid and either epidermal growth factor or transforming growth factor-α in an amount effective for achieving tubulogenesis.  
     
     
         17 . The method of  claim 16 , wherein said cells are further treated with transforming growth factor-β 1 .  
     
     
         18 . The method of  claim 17 , wherein said culture medium contains a soluble factor selected from the group consisting of fetal calf serum, prostaglandins, hydrocortisone, triodothyronine, selenium, fibroblastic growth factor, hepatocyte growth factor and combinations thereof.  
     
     
         19 . The method of  claim 17 , wherein said culture medium contains an insoluble factor selected from the group consisting of Type I collagen, Type IV collagen, laminin, proteoglycans, fibronectin, and combinations thereof.  
     
     
         20 . The method of  claim 17 , wherein said culture medium contains a soluble factor selected from the group consisting of fetal calf serum, prostaglandins, hydrocortisone, triodothyronine, selenium, fibroblastic growth factor, hepatocyte growth factor, and combinations thereof, and an insoluble factor, selected from the group consisting of Type I collagen, Type IV collagen, laminin, proteoglycans, fibronectin, and combinations thereof.  
     
     
         21 . A method for constructing and maintaining an ex vivo renal tubule tissue system, comprising culturing adult kidney cells in the presence of all-trans retinoic acid and either epidermal growth factor or transforming growth factor-α in an amount effective for tubulogenesis.  
     
     
         22 . The method of  claim 21 , wherein said cells are further treated with transforming growth factor-β 1 .  
     
     
         23 . The method of  claim 22 , wherein said culture medium contains a soluble factor selected from the group consisting of fetal calf serum, prostaglandins, hydrocortisone, triodothyronine, selenium, fibroblastic growth factor, hepatocyte growth factor and combinations thereof.  
     
     
         24 . The method of  claim 22 , wherein said culture medium contains an insoluble factor selected from the group consisting of Type I collagen, Type IV collagen, laminin, proteoglycans, fibronectin, and combinations thereof.  
     
     
         25 . The method of  claim 22 , wherein said culture medium contains a soluble factor selected from the group consisting of fetal calf serum, prostaglandins, hydrocortisone, triodothyronine, selenium, fibroblastic growth factor, hepatocyte growth factor, and combinations thereof, and an insoluble factor selected from the group consisting of Type I collagen, Type IV collagen, laminin, proteoglycans, fibronectin, and combinations thereof.  
     
     
         26 . A method for effecting kidney repair, comprising administering to a patient in need thereof all-trans retinoic acid and either epidermal growth factor or transforming growth factor-α in an amount effective for achieving kidney repair.  
     
     
         27 . The method of  claim 26 , wherein said patient is further treated with transforming growth factor-β 1 .  
     
     
         28 . The method of  claim 27 , wherein said patient is treated with a soluble factor selected from the group consisting of fetal calf serum, prostaglandins, hydrocortisone, triodothyronine, selenium, fibroblastic growth factor, hepatocyte growth factor and combinations thereof.  
     
     
         29 . The method of  claim 27 , wherein said patient is further treated with an insoluble factor, selected from the group consisting of Type I collagen, Type IV collagen, laminin, proteoglycans, fibronectin, and combinations thereof.  
     
     
         30 . The method of  claim 27 , wherein said patient is further treated with a soluble factor, selected from the group consisting of fetal calf serum, prostaglandins, hydrocortisone, triodothyronine, selenium, fibroblastic growth factor, hepatocyte growth factor and combinations thereof, and an insoluble factor selected from the group consisting of Type I collagen, Type IV collagen, laminin, proteoglycans, fibronectin, and combinations thereof.  
     
     
         31 . A method for effecting gene therapy upon renal tubule tissue, comprising culturing adult kidney cells in a culture medium containing all-trans retinoic acid and either epidermal growth factor or transforming growth factor-α in an amount effective to achieve tubulogenesis in the presence of a transforming vector.  
     
     
         32 . The method of  claim 31 , wherein said cells are further treated with transforming growth factor-β 1 .  
     
     
         33 . The method of  claim 32 , wherein said culture medium contains a soluble factor selected from the group consisting of fetal calf serum, prostaglandins, hydrocortisone, triodothyronine, selenium, fibroblastic growth factor, hepatocyte growth factor and combinations thereof.  
     
     
         34 . The method of  claim 32 , wherein said culture medium contains an insoluble factor selected from the group consisting of Type I collagen, Type IV collagen, laminin, proteoglycans, fibronectin, and combinations thereof.  
     
     
         35 . The method of  claim 32 , wherein said culture medium contains a soluble factor selected from the group consisting of fetal calf serum, prostaglandins, hydrocortisone, triodothyronine, selenium, fibroblastic growth factor, hepatocyte growth factor, and combinations thereof, and an insoluble factor selected from the group consisting of Type I collagen, Type IV collagen, laminin, proteoglycans, fibronectin, and combinations thereof.  
     
     
         36 . A method for effecting kidney repair, comprising removing renal cells from a patient in need of treatment, culturing said renal cells in a culture medium containing all-trans retinoic acid and either epidermal growth factor or transforming growth factor-α in an amount effective for achieving a tubule tissue system and returning said tubule tissue to said patient.  
     
     
         37 . The method of  claim 36 , wherein said cells are further treated with transforming growth factor-β 1 .  
     
     
         38 . The method of  claim 37 , wherein said culture medium contains a soluble factor selected from the group consisting of fetal calf serum, prostaglandins, hydrocortisone, triodothyronine, selenium, fibroblastic growth factor, hepatocyte growth factor and combinations thereof.  
     
     
         39 . The method of  claim 37 , wherein said culture medium contains insoluble factor selected from the group consisting of Type I collagen, Type IV collagen, laminin, proteogiycans, fibronectin, and combinations thereof.  
     
     
         40 . The method of  claim 37 , wherein said culture medium contains a soluble factor selected from the group consisting of fetal calf serum, prostaglandins, hydrocortisone, triodothyronine, selenium, fibroblastic growth factor, hepatocyte growth factor and combinations thereof, and an insoluble factor selected from the group consisting of Type I collagen, Type IV collagen, laminin, proteoglycans, fibronectin, and combinations thereof.