18α-GA通过上调蛋白酶体活性促进晚期骨髓间充质干细胞增殖  

Upregulation of proteasome activity by 18α-GA promotes proliferation of late-passage BMSCs in vitro

作　　者：杨佳超[1] 赵云鹤[1] 蒋蓉[1] 陆利[1] 

机构地区：[1]山西医科大学人体解剖学教研室,山西太原030001

出　　处：《中国病理生理杂志》2015年第12期2183-2187,共5页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.81200254);山西省回国留学人员科研基金资助项目(No.2014-033);山西省研究生创新基金资助项目(No.20133075)

摘　　要：目的:明确18α-甘草次酸(18α-glycyrrhetinic acid,18α-GA)对晚期骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMSCs)衰老标志物和增殖能力的影响。方法:晚期BMSCs(≥14代)应用2.0 mg/L的18α-GA持续作用30 d,比较18α-GA组与DMSO组蛋白酶体活性;衰老相关β-半乳糖苷酶(senescence-associatedβ-galactosidase,SA-β-Gal)染色和Western blot检测衰老相关蛋白p53、p21和p16的表达变化;CCK-8、Brd U掺入试验、流式细胞术和Western blot分别检测细胞增殖能力、细胞周期分布及细胞周期相关分子表达变化。结果:应用18α-GA后BMSCs蛋白酶体活性较DMSO组增高约0.2倍(P<0.01)。18α-GA组SA-β-Gal阳性衰老细胞较DMSO组减少,且细胞着色浅淡;衰老相关蛋白p53和p21表达水平分别较对照组降低(P<0.05)。CCK-8实验及流式细胞术结果显示18α-GA组细胞增殖能力较DMSO组增高,S期细胞显著增多(P<0.05),18α-GA组Brd U染色阳性细胞率较DMSO对照组增高(P<0.05)。18α-GA组细胞周期相关蛋白cyclin D1及CDK4表达水平分别较DMSO组增高(P<0.05)。结论:18α-GA可激活晚期BMSCs蛋白酶体活性延缓细胞衰老,并且可能通过上调细胞周期相关蛋白表达水平促进晚期BMSCs增殖。AIM： To investigate the effect of 18 alpha-glycyrrhetinic acid（ 18α-GA） on delaying the senescent progress and promoting the proliferation in late-passage bone marrow mesenchymal stem cells（ BMSCs）. METHODS：Late-passage BMSCs were incubated with 2. 0 mg / L 18α-GA or the same volume of DMSO for 30 d,and the cells were harvested to determine the proteasome activity. The expression of senescence-related proteins p53,p21 and p16 was detected by senescence-associated β-galactosidase（ SA-β-Gal） staining and Western blot. The cell proliferation,the expression level of cell cycle-related proteins and cell cycle distribution of the cells were measured by CCK-8 assay,Brd U incorporation,Western blot and flow cytometry. RESULTS： Compared with DMSO group,the proteasome activity in 18α-GA group increased significantly by about 0. 2 times（ P〈0. 01）. SA-β-Gal-positive cells in 18α-GA group decreased,and cell staining was lighter. The contents of p53 and p21 in 18α-GA group were decreased（ P〈0. 05）. The results of CCK-8 assay showed that the A value in 18α-GA group was 0. 3 times higher than that in DMSO group（ P〈0. 01）. Brd U incorporation showed the increased proliferation in 18α-GA group compared with DMSO group（ P〈0. 05）. The cells in G1 phase in18α-GA group decreased significantly compared with DMSO group,while the cells in S phase increased significantly（ P〈0. 05）. The expression level of cyclin D1 in 18α-GA group was 2. 8 times higher than that in DMSO group（ P〈0. 01）,and the CDK4 level was 1. 4 times higher than that in DMSO group（ P〈0. 05）. CONCLUSION： Activation of the proteasome activity by 18α-GA delays the aging process in the BMSCs and promotes the cell proliferation via up-regulation of the cell cycle-related proteins.

关 键 词：18α-甘草次酸 蛋白酶体 骨髓间充质干细胞 细胞增殖 

分 类 号：R363[医药卫生—病理学]