Use of adipose tissue-derived stromal cells for chondrocyte differentiation and cartilage repair
AssigneeArtecel Sciences, Inc.
InventorsYuan-Di C. Halvorsen, William O. Wilkison, Jeffrey Martin Gimble
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Abstract
Methods and compositions for directing adipose-derived stromal cells cultivated in vitro to differentiate into cells of the chondrocyte lineage are disclosed. The invention further provides a variety of chondroinductive agents which can be used singly or in combination with other nutrient components to induce chondrogenesis in adipose-derived stromal cells either in cultivating monolayers or in a biocompatible lattice or matrix in a three-dimensional configuration. Use of the differentiated chondrocytes for the therapeutic treatment of a number of human conditions and diseases including repair of cartilage in vivo is disclosed.
Claims18
A medium for differentiating adipose tissue derived stromal cells into chondrocyte cells, 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, wherein the nutrient supplement comprises 1-20% fetal bovine serum or 1-20% horse serum;
- (iv) ascorbate or vitamin C analogue;
- and
- (v) a glucocorticoid or other chemical agent capable of activating the cellular glucocorticoid receptor.
A method for differentiating adipose tissue derived stromal cells into chondrocytic cells, comprising:
- a) pelleting said stromal cells by centrifuging between 50,000 to 5 million cells at 500×g for 2 to 20 minutes in sterile tubes containing a medium selected form the group consisting of Dulbecco's Modified Eagle's Medium (DMEM), alpha modified Minimal Essential Medium (αMEM), and Roswell Park Memorial Institute media 1640 (RPMI Media 1640);
- b) plating said isolated stromal cells 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 supplemented with 1 to 20% fetal bovine serum or 1 to 20% horse serum;
- (iv) ascorbate or vitamin C analog;
- and
- (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 in with 5% CO2 and between 1% and 20% oxygen.
A method for differentiating adipose tissue derived stromal cells into chondrocytic cells, comprising:
- a) suspending stromal cells 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 said suspended 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 supplemented with 1 to 20% fetal bovine serum or 1 to 20% horse serum;
- (iv) ascorbate or vitamin C analog;
- and
- (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.
Dependencies are inferred from the claim text.