高尔基磷蛋白3对脂肪干细胞生物学特性的影响

Effect of Golgi phosphoprotein 3 on biological characteristics of adipose stem cells

目的探讨高尔基磷蛋白3(GOLPH3)对人脂肪来源间充质干细胞(ADSC)增殖、迁移及成脂分化的影响。方法分离培养人原代ADSC细胞。采用慢病毒载体在ADSC中过表达GOLPH3(GOLPH3过表达组,GPLPH3-over),以空载体慢病毒作为对照(NC-over)。细胞计数试剂盒-8(CCK-8)、Transwell细胞迁移实验分别检测两组细胞的增殖和迁移能力,并经14 d成脂分化诱导后行油红O染色,观察脂滴数目。蛋白质印迹法(Western blot)检测两组细胞中特定成脂标志物过氧化物酶增殖物激活受体γ(PPAR-γ)和增强子结合蛋白α(C/EBPα)蛋白水平的表达。组间比较采用t检验。结果成功获得稳定过表达GOPLH3的ADSC。CCK-8结果显示,GOLPH3-over组增殖能力强于NC-over组(0.579±0.142比0.466±0.081,t=3.127,P<0.05)。Transwell细胞迁移实验显示,GOLPH3-over组迁移能力强于NC-over组(39.000±2.646比25.000±3.000,t=5.563,P<0.05)。油红O染色结果表明,GOLPH3-over组脂滴多于NC-over组(136919±12748比67837±2665,t=9.187,P<0.05)。Western blot结果表明,GOLPH3-over组PPAR-γ(1.061±0.175比0.521±0.082,t=4.904,P<0.05)和C/EBPα(3.120±0.171比2.196±0.174,t=6.548,P<0.05)蛋白表达水平高于NC-over组高,差异均有统计学意义。结论过表达GOLPH3可以促进体外ADSC的增殖、迁移及其向脂肪细胞的分化。

Objective To investigate the effect of Golgi phosphoprotein 3(GOLPH3)on the proliferation,migration and adipogenic differentiation of human adipose derived mesenchymal stem cells(ADSCs).Methods Human primary ADSCs were isolated and cultured.The lentivirus vector was used to overexpress COLPH3(GOLPH3 overexpression group,GPLPH3-over)in ADSCs,and the empty vector lentivirus was used as the control(NC-over).Cell counting kit-8(CCK-8)and Transwell cell migration tests were used to detect the proliferation and migration ability of the two groups,and Oil red O staining was performed after 14 days of adipogenic differentiation induction to observe the number of lipid droplets.Western blotting was used to detect the expression levels of peroxidase proliferator-activated receptor(PPAR-)and enhancer binding proteinα(C/EBPα).The measurement data were tested by test be-tween groups.Results ADSCs stably overexpressing GOPLH3 were successfully obtained.CCK-8 results showed that the proliferation ability of GOLPH3-over group was enhanced(0.579±0.142 vs.0.466±0.081,t=3.127,P<0.05).Transwell cell migration experiment showed that the migration ability of GOLPH3-over group was stronger than that of NC-over group(39.000±2.646 vs.25.000±3.000,t=5.563,P<0.05).The results of Oil red O staining showed that the lipid drops in GOLPH3-over group were more than those in NC-over group(136919±12748 vs.67837±2665,t=9.187,P<0.05).The results of Western blotting showed that PPAR-(1.061±0.175 vs.0.521±0.082,t=4.904,P<0.05)and C/EBPα(3.120±0.171 vs.2.196±0.174,t=6.548,P<0.05)in GOLPH3-over group was higher expressed than that in NC-over group with the difference being statistically significant.Conclusion Overexpression of GOLPH3 can promote the proliferation,migration and differentiation of ADSCs into adipocytes in vitro.