Directional differentiation of human adipose-derived stem cells into keratin-forming cells

Abstract

Objective To optimize the method of the directional differentiation of adipose-derived stem cells into keratinocytes. Methods Adipose-derived stem cells (ADSCs), separated by collagenase digestion method, were isolated and cultured. Then the expression of surface specific markers CD34, CD44 and CD90 were detected by flow cytometer. The effect of different induced mediums cultured for two weeks on the differentiation of ADSCs into KCs was demonstrated: Group 1, the DMEM supplied with 2% FBS and 49% supertant of KCs; group 2, KSFM medium; group 3, DMEM medium supplied with 10% FBS and 5 μM ATRA; 10% FBS DMEM as the control group. Immunofluorescene staining was applied to detect the expression of keratin CK14 and F-actin. Results A flattened fibroblast-like morphology was observed in cells, the positive expression rate of CD34 was 0.08%, while those of CD44 and CD90 were 99%. The cells that could differentiate into osteoblasts and chondrocytes, indicated that the cells were ADSCs. There was no significant change in the cell morphology in the group 1 under the induction medium; about 10% of the cells in group 2 were altered; the morphological changes were obvious in group 3, and approximately 20% of the cells showed irregular polygon. The immunofluorescene staining of the cells in group 3 indicated that the cells showed cobblestone-like phenotype and an organized cytoskeletal network with dense actin fibers at the edges; some cells were positive for CK14. Conclusions ADSCs show higher induction rate under ATRA stimulation. Key words: Adipose tissue; Stem cells; Cell differentiation; Keratinocytes