Use of adipose tissue-derived stromal cells for chondrocyte differentiation and cartilage repair

A method for differentiating adipose tissue derived stromal cells into chondrocytic cells, comprising:

• a) pelleting said stromal cells and preadipocytes by centrifuging between 50,000 to 5 million cells at 500×g for 2 to 20 minutes in sterile tubes containing a medium;
• b) plating isolated stromal cells and preadipocytes at a density of 500 to 20,000 cells/cm2 in a differentiating medium;
• c) supplementing said medium with:
• i) a chondroinductive agent capable of activating any cellular signal transduction pathway leading to the mature chondrocyte phenotype
• ii) an antibiotic
• iii) a nutrient supplement with 1 to 20% fetal bovine serum or 1 to 20% horse serum or any other biological or synthetic equivalent combination of proteins
• iv) ascorbate or related vitamin C analog
• v) a glucocorticoid or other chemical agent capable of activating the cellular glucocorticoid receptor;
• and
• d) incubating said cells at about 31° C. to 37° C. for about 3-4 weeks with 5% CO2 and between 1% and 20% oxygen;
• and,
• e) determining the extent of differentiation of the isolated cells.

A method for differentiating adipose tissue derived stromal cells into chondrocytic cells, comprising:

• a) suspending stromal cells and preadipocytes at a concentration of 0.5 to 10 million cells per ml in calcium alginate or any other biocompatible lattice or matrix capable of supporting chondrogenesis in a three-dimensional configuration;
• b) transferring cells to 35 mm culture dishes and plating cells at a density of 500 to 20,000 cells/cm2 in a differentiating medium comprising a chemically defined culture medium having or supplemented with:
• i) a chondroinductive agent capable of activating any cellular transduction pathway leading to the mature chondrocyte phenotype;
• ii) an antibiotic;
• iii) a nutrient supplement with 1 to 20% fetal bovine serum or 1 to 20% horse serum or any other biological or synthetic equivalent combination of proteins;
• iv) ascorbate or related vitamin C analog;
• v) a glucocorticoid or other chemical agent capable of activating the cellular glucocorticoid receptor;
• and
• c) incubating said cells at about 31 to 37° C. for about 3-4 weeks in an incubator with 5% CO2 and between 1% and 20% oxygen, and
• d) determining the extent of differentiation of the isolated cells.