PI3K/AKT信号通路参与地塞米松促进骨髓间充质干细胞内质网应激反应的研究  

The PI3K/AKT signaling pathway involved in dexamethasone promotes the endoplasmic stress response of bone marrow mesenchymal stem cells

作　　者：程锁利[1] 巩凡[1] 牛东生[1] 孙玺淳[1] 白志刚[1] 黄朋[1] 郭崇军[1] 梁钰琪 郭舟桐[1] CHENG Suoli;GONG Fan;NIU Dongsheng;SUN Xichun;BAI Zhigang;HUANG Peng;GUO Chongjun;LIANG Yuqi;GUO Zhoutong(Department of Joint Surgery,People’s Hospital of Ningxia Hui Autonomous Region,Yinchuan 750002,China)

机构地区：[1]宁夏回族自治区人民医院关节外科,宁夏银川750002

出　　处：《宁夏医学杂志》2022年第3期245-248,共4页Ningxia Medical Journal

基　　金：宁夏自然科学基金资助项目(2018AACO3269)。

摘　　要：目的通过地塞米松(DEX)对骨髓间充质干细胞(BMSCs)细胞生物学行为的研究,探讨地塞米松在激紊性股骨头缺血性坏死(SANFH)中的作用和可能的调控通路。方法通过Counting Kit-8试验、蛋白质印迹试验(Western blot assay)和酶联免疫吸附试验评估DEX对BMSCs增殖的影响,通过流式细胞术和蛋白质印迹法检测DEX对BMSCs凋亡的影响。采用实时定量(PCR)和Western blot assay检测DEX对内质网应激(ERS)相关基因及蛋白表达的影响,并进行免疫印迹分析以检测PI3K/AKT细胞通路中的相关蛋白表达。结果CCK8实验结果提示,随着浓度的增加,DEX可以抑制BMSCs的增殖。流式细胞术分析结果表明,随着DEX浓度的增加,BMSCs的凋亡百分比逐渐增加。通过蛋白质印迹,随着DEX浓度的增加,凋亡相关蛋白TEAR19的表达增加。检测BMSCs经DEX处理后两种ERS相关因子(IRE1a和ATF6)的表达,qRT-PCR结果显示,随着DEX浓度的增加,ATF6和IRE1a的mRNA表达显著增加(P<0.05)。此外,从蛋白质印迹分析中获得了类似的结果,如ATF6、p-IRE1a/IRE1a蛋白表达有明显升高(P<0.05)。Western blot和qRT-PCR实验结果表明,BMSCs经DEX处理后PI3K、AKT蛋白的表达显著增加(P<0.05)。结论DEX能显著抑制BMSCs的增殖,促进BMSCs的凋亡,并通过或部分通过PI3K/AKT通路的激活、提高内质网应激蛋白ATF6和IRE1a的表达,导致BMSCs的凋亡。而PI3K抑制剂(BYL719)可以部分逆转DEX对BMSCs的影响。Objective To study the biological behavior of bone marrow mesenchymal stem cells(BMSCs)by dexamethasone(DEX),and to explore the effect and possible regulatory pathway of dexamethasone in steroid-induced avascular necrosis of femoral head(SANFH).Methods Counting Kit-8 test,western blot assay and enzyme-linked immunosorbent assay were used to evaluate the effect of DEX on the proliferation of BMSCs.Flow cytometry and western blotting were used to detect the effect of Dex on the apoptosis of BMSCs.Quantitative real-time(PCR)and western blot assay were used to detect the effect of DEX on the expression of endoplasmic reticulum stress(ERS)related genes.Western blot analysis was performed to detect the nuclear and cytoplasmic distribution.Results The results of the CCK8 assay indicated that DEX inhibited the proliferation of BMSCs with increasing concentration.The results of flow cytometry analysis showed that the percentage of apoptosis of BMSCs gradually increased with the increase of DEX concentration.The expression of apoptosis-related protein TEAR19 was increased with increasing DEX concentration by western blotting and the expression of two ERS-related factors(IRE1a and ATF6)in BMSCs treated with DEX was detected.qRT-PCR results showed that the mRNA expression of ATF6 and IRE1a increased significantly with the increase of DEX concentration(P<0.05).In addition,similar results were obtained from western blot analysis,such as a significant increase in ATF6,p-IRE1a/IRE1a protein expression(P<0.05).The results of Western blot and qRT-PCR experiments showed that the expressions of PI3K and AKT proteins in BMSCs were significantly increased after DEX treatment(P<0.05).Conclusion DEX can significantly inhibit the proliferation of BMSCs,promote the apoptosis of BMSCs,and lead to the apoptosis of BMSCs through or partly through the activation of the PI3K/AKT pathway and increasing the expression of endoplasmic reticulum stress proteins ATF6 and IRE1a.A PI3K inhibitor(BYL719)could partially reverse the effects of DEX on BMS

关 键 词：PI3K/AKT 地塞米松 骨髓间充质干细胞 内质网应激 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]