Effects of Sox-9 gene silenced by small interfering RNA on proliferation and apoptosis of epiphysis stem cells / 中国组织工程研究

ABSTRACT

BACKGROUND:Current research has shown that many cellgrowth factors can promote the proliferation and differentiation of epiphysis stem cells, and this regulation is closely related to Sox-9. OBJECTIVE:To investigate the effects of smal interfering RNA-induced specific silence of Sox-9 gene on the proliferation and apoptosis of epiphysis stem cells. METHODS:Epiphysis stem cells were isolated from the ring of La Croix with enzyme digestion and purified with magnetic activated cellsorting, identified by immunocytochemistry assay. The cells in good conditions were selected to be transfected by Sox-9 silenced by smal interfering RNA with fluorescent marker, and then were observed under the fluorescent microscope to check the transfection efficiency. Next step, epiphysis stem cells were divided into 2 groupsgroup one was cultured normal y as control group;group two was transfected by Sox-9 silenced by smal interfering RNA through LipofectamineTM 2000 as experimental group. RESULTS AND CONCLUSION:The primary epiphysis stem cells were separated and purified, which were of stable growth and tightly attachment. The results of immunohistochemistry and immunofluorescence showed the epiphysis stem cells expressed cel-specific markers, fibroblast growth factor receptor 3. After transfection, reverse transcription-PCR results showed that Sox-9 gene expression in epiphysis stem cellwas inhibited specifical y;3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide results showed that cellproliferation in experimental group decreased significantly compared with the control group (P<0.05), and the cellapoptosis detected by flow cytometry showed that the experimental group increased as compared with the control group (P<0.05). Sox-9 gene plays an important role in regulating the proliferation and apoptosis of rat epiphysis stem cells.