Mixed population of cells deriving from adipose tissue and methods of isolating and using the same

Abstract

Adipose tissue-derived stromal cells and methods of isolating and using the same. In at least one embodiment of isolated adipose tissue-derived stromal cells of the present disclosure, the cells are isolated by performing adipose tissue resection or suction on a mammalian patient, dissecting tissue obtained from said tissue resection or suction and dissociating said tissue into a cell suspension, removing adipocytes from the cell suspension, culturing the adipocyte-depleted cell suspension in EGM-2-MV media, and isolating adipose tissue-derived stromal cells secreting vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF), and granulocyte-colony stimulating factor (G-CSF).

1 . A mixed population of cells, obtained by:
 a. isolating subcutaneous adipose tissue from a mammal;   b. dissociating the isolated subcutaneous adipose tissue into a cell suspension;   c. removing adipocytes from the cell suspension, resulting in isolated cells; and   d. culturing the isolated cells in EGM-2-MV media such that a mixed population of cells comprising a first population of CD34+/VE-cadherin− cells and a second /population of CD34+/VE-cadherin+ endothelial cells are obtained.   
     
     
         2 . The cells of  claim 1 , wherein the culturing is in EGM-2-MV media on matrigel. 
     
     
         3 . The cells of  claim 1 , wherein the mixed population of cells are further cultured in the presence of 5-azacytidine. 
     
     
         4 . The cells of  claim 1 , wherein the mixed population of cells are further cultured in in EBM-2 media. 
     
     
         5 . The cells of  claim 4 , which secrete a factor selected from the group consisting of pro-angiogenic factors, anti-apoptotic factors, vasculoprotective factors, and cardioprotective factors. 
     
     
         6 . The cells of  claim 1 , wherein the first population is a larger quantity than the second population. 
     
     
         7 . The cells of  claim 1 , wherein the dissociating is performed using agitation, sonic energy, thermal energy, or a combination thereof. 
     
     
         8 . The cells of  claim 1 , wherein the second population comprises a population of cells having an endothelial phenotype. 
     
     
         9 . The cells of  claim 1 , wherein the dissociating is performed using collagenase, dispase, trypsin, or a combination thereof. 
     
     
         10 . The cells of  claim 1 , further cultured in DMEM and 10% fetal bovine serum. 
     
     
         11 . A mixed population of cells, obtained by:
 isolating subcutaneous adipose tissue using liposuction or surgery;   dissecting the subcutaneous adipose tissue and dissociating said tissue into a cell suspension;   removing adipocytes from the cell suspension;   isolating cells from the cell suspension after removal of the adipocytes; and   culturing the isolated cells in EGM-2-MV media such that a mixed population of cells comprising a first population of CD34+/VE-cadherin− cells and a second population of CD34+/VE-cadherin+ endothelial cells are obtained.   
     
     
         12 . The cells of  claim 11 , wherein the dissociating is performed using agitation, sonic energy, thermal energy, or a combination thereof. 
     
     
         13 . The cells of  claim 11 , wherein the dissociating is performed using collagenase, dispase, trypsin, or a combination thereof. 
     
     
         14 . The cells of  claim 11 , wherein the cells comprise an exogenous nucleic acid selected from the group of nucleic acids encoding VEGF, bFGF, IGF1, IGF2, HGF, cardiotrophin, myotrophin, nitric oxide synthase 1, nitric oxide synthase 2, nitric oxide synthase 3, fibroblast growth factor, pleotrophin, endothelin, and angiopoietin. 
     
     
         15 . The cells of  claim 11 , wherein the second population comprises a population of cells having an endothelial phenotype. 
     
     
         16 . The cells of  claim 11 , further comprising the step of culturing the mixed population of cells in EBM-2 media. 
     
     
         17 . The cells of  claim 16 , wherein the mixed population of cells secrete at least one factor selected from the group consisting of a vascular endothelial growth factor (VEGF), a hepatocyte growth factor (HGF), granulocyte-macrophage colony stimulating factor (GM-CSF), and a granulocyte-colony stimulating factor (G-CSF). 
     
     
         18 . A mixed population of cells, obtained by:
 a) isolating subcutaneous adipose tissue from a mammal;   b) dissecting the subcutaneous adipose tissue;   c) dissociating the dissected subcutaneous tissue into a cell suspension;   d) removing adipocytes from the cell suspension;   e) isolating cells from the cell suspension after removal of the adipocytes;   f) culturing the cells from step e) in EGM-2-MV media; and   g) culturing the cells from step f) such that a mixed population of cells comprising a first population of CD34+/VE-cadherin− cells and a second population of CD34+/VE-cadherin+ endothelial cells are obtained.   
     
     
         19 . The cells of  claim 18 , wherein the mixed population of cells contain pluripotent cells capable of differentiating into another cell type. 
     
     
         20 . The cells of  claim 18 , wherein the mixed population of cells includes a population of cells having an endothelial phenotype.