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目的探讨重组人结缔组织生长因子(hCTGF)对大鼠脂肪源性干细胞(ADSCs)分化及Ⅰ型胶原蛋白表达的影响。方法 2010年9月至2011年9月在广州医学院第一附属医院采用贴壁培养法分离纯化大鼠ADSCs,流式细胞仪分析第3代大鼠ADSCs表面抗原标记物;含100μg/L重组hCTGF的培养基作为刺激组,无重组hCTGF为对照组,分别培养7、14、28d后,RT-PCR和western-blot检测大鼠ADSCs中Ⅰ型胶原蛋白表达;观察重组hCTGF对大鼠ADSCs成骨、成脂方向分化的影响。结果大鼠ADSCs表面抗原CD44、CD90、CD105、CD49d阳性,CD34、CD45、CD106阴性;Ⅰ型胶原蛋白mRNA表达量刺激组与对照组7、14、28d时分别为1.069±0.324vs.0.298±0.029、2.162±0.038vs.0.652±0.024、3.360±0.058vs.0.815±0.041,差异有统计学意义(P<0.05),Ⅰ型胶原蛋白的表达量7、14、28d时分别为0.761±0.041vs.0.679±0.029、0.834±0.115vs.0.708±0.061、0.774±0.180vs.0.720±0.026,差异有统计学意义(P<0.05)。结论重组hCTGF可上调大鼠ADSCsⅠ型胶原蛋白的表达,抑制大鼠ADSCs向成脂、成骨分化。推测重组hCTGF可能使大鼠ADSCs发生了向成纤维细胞定向分化。
Objective To investigate the effect of recombinant human connective tissue growth factor (hCTGF) on the differentiation of adipose derived stem cells (ADSCs) and the expression of type Ⅰ collagen in rats. Methods ADSCs were isolated and purified from adherent culture in the First Affiliated Hospital of Guangzhou Medical College from September 2010 to September 2011. The surface antigens of ADSCs in the third generation rats were analyzed by flow cytometry. hCTGF as stimulation group and no recombinant hCTGF as control group. After cultured for 7, 14 and 28 days respectively, the expression of collagen Ⅰ in ADSCs was detected by RT-PCR and western-blot. The effect of recombinant hCTGF on ADSCs Bone, adipogenic direction of differentiation. Results The expression of surface antigen CD44, CD90, CD105 and CD49d in ADSCs was negative and CD34, CD45 and CD106 were negative. The expression of type Ⅰ collagen mRNA was 1.069 ± 0.324vs.0.298 ± 0.029 , 2.162 ± 0.038vs.0.652 ± 0.024,3.360 ± 0.058vs.0.815 ± 0.041, the difference was statistically significant (P <0.05). The expression of type Ⅰ collagen was 7. 7 and 28 d, respectively, 0.761 ± 0.041 vs. 0.679 ± 0.029,0.834 ± 0.115vs.0.708 ± 0.061,0.774 ± 0.180vs.0.720 ± 0.026, the difference was statistically significant (P <0.05). Conclusion Recombinant hCTGF can upregulate the expression of type Ⅰ collagen in ADSCs and inhibit the differentiation of ADSCs into adipogenic and osteogenic potential in rats. It is speculated that recombinant hCTGF may induce the ADSCs to differentiate into fibroblasts.