miR-17对人包皮成纤维细胞衰老的影响

Effects of miR-17 on the senescence of human foreskin fibroblasts

目的探讨miR-17对人包皮成纤维细胞(HFF)衰老的影响。方法 miR-17重组慢病毒感染细胞,qRT-PCR确定感染效率,CCK-8法观察miR-17对细胞增殖的影响,衰老相关β-半乳糖苷酶染色观察miR-17对细胞衰老的影响,流式细胞法检测miR-17对细胞周期的影响,Western blotting检测cyclin D1和p21蛋白的表达。结果成功分离得到HFF,并建立了稳定表达miR-17的HFF-miR-17细胞系。且HFF-miR-17细胞增殖能力明显升高,β-半乳糖苷酶染色阳性率降低;S期细胞群比例明显增高,细胞周期蛋白cyclin D1表达显著上调,p21蛋白明显下调。结论 miR-17可通过上调cyclin D1蛋白、下调p21蛋白促进原代细胞的增殖,抑制原代细胞的衰老。

Objective To study the effect of miR-17 on the senescence of human foreskin fibroblasts （HFF）. Methods Recombinant lentivirus-expressed miR-17 infected HFF and rniR-17 expression was examined with qRT-PCR. The proliferation and senescence of I-IFF were detected with CCK-8 and SA-13-Gal kit, respectively. Cell cycle profile was observed with flow cytometry. The protein levels of cyclin D1 and p21 were detected with Western blotting. Results HFF were successfully isolated. Compared with HFF-NC, the proliferation ability of HFF-miR-17 cells was obviously increased while the expression of SA-β-Gal in senescent cells decreased. S phase cells of HFF-miR-17 increased signifi- cantly and more HFF-NC ceils arrested in the G1 phase. Expression of cyclin D1 was significantly up-regulated in HFF- miR-17, while p21 expression was attenuated. Conclusion miR-17 promotes the proliferation and inhibits senescence of primary cells by upregulating cyclin D1 and downregulating p21 proteins.