通过PI3K/SGK1信号通路周期性牵张应力促进尿路上皮细胞增殖  

Promoting the proliferation of urothelial cells by periodic stretch stress of PI3K/SGK1 signaling pathway

作　　者：韦堂墙 伍季[1] 王安果[1] 张宗平[1] 李云祥[1] WEI Tang-qiang;WU Ji;WANG An-guo;ZHANG Zong-ping;LI Yun-xiang(Department of Urology,Second Clinical Medical School,North Sichuan Medical College,Nanchong 637000,Sichuan,China)

机构地区：[1]川北医学院第二临床医学院泌尿外科,四川南充637000

出　　处：《生物医学工程与临床》2021年第4期475-481,共7页Biomedical Engineering and Clinical Medicine

基　　金：国家自然科学基金资助项目(81900617);南充市科技基金资助项目(18SXHZ0494)。

摘　　要：目的探索周期性牵张应力促进尿路上皮细胞(HUC)增殖的响应机制。方法选择HUC与硅胶膜复合培养24 h。将其置于BOSE BioDynamic生物反应器中,加载产生特定大小的周期性牵张应力,流式细胞术检测细胞周期分布并计算细胞增殖指数。实时荧光定量聚合酶链式反应(PCR)及Western blot检测血清-糖皮质激素诱导型蛋白激酶1基因(SGK1)及蛋白激酶B基因(Akt)等基因的表达变化。进行siRNA转染实验(无牵张应力-空白siRNA组,无牵张应力-SGK1 siRNA组,牵张应力组-空白siRNA组,牵张应力-SGK1 siRNA组)和SGK1药物抑制剂实验(牵张应力组及牵张应力-SGK1抑制剂组)。结果实时荧光定量PCR检测结果发现,周期性牵张应力促进SGK1表达升高(4.04±0.51)倍(P<0.01),而Akt的转录水平未发生明显变化;进一步行蛋白免疫印迹检测,SGK1的蛋白表达升高(1.59±0.12)倍(P<0.01),SGK1的磷酸化水平升高(2.53±0.15)倍(P<0.01);而P-Akt(Thr308/Ser473)/Akt表达未明显受到牵张应力影响(P>0.05)。运用siRNA成功下调SGK1的表达后,周期性牵张应力所诱导的HUC增殖效应得到显著抑制,细胞增殖指数从(56.56±4.25)%降至(43.31±4.69)%(P<0.05);使用SGK1抑制剂后,牵张应力所诱导的HUC增殖效应同样得到显著抑制,细胞增殖指数从(56.92±3.22)%降至(42.24±1.85)%(P<0.05)。结论通过PI3K/SGK1信号通路而非PI3K/Akt信号通路周期性牵张应力促进HUC增殖。Objective To explore the response mechanisms of human urothelial cells(HUC)proliferation induced by cyclic stretch.Methods The HUC was co-cultured with silica gel membrane for 24 hours and placed in Bose BioDynamic bioreactor,and special cyclic stretch was loaded.Flow cytometry was used to detect cell cycle distribution and calculate cell proliferation index(CPI).The real-time quantitative polymerase chain reaction(PCR)and Western blot were used to detect expression levels of serum and glucocorticoid-induced protein kinase 1 gene(SGK1)and protein kinase B gene(Akt).The siRNA transfection experiment(non-stretch stress-blank siRNA group,non-stretch stress-SGK1 siRNA group,stretch stress group-blank siRNA group,stretch stress-SGK1 siRNA group)and SGK1 drug inhibitor experiment(stretch stress group and stretch stressSGK1 inhibitor group)were carried out.Results The fluorescence quantitative PCR detection results showed that periodic stretch stress promoted increase of SGK1 expression by(4.04±0.51)times(P<0.01),while transcription level of Akt remained unchanged significantly.Western blot detection results showed that protein expression of SGK1 increased(1.59±0.12)times(P<0.01),and phosphorylation level of SGK1 increased(2.53±0.15)times(P<0.01),while P-Akt(Thr308/Ser473)/Akt expression was not significantly changed(P>0.05).After siRNA successfully down-regulated SGK1 expression,the proliferation effect of HUC induced by periodic stretch stress was significantly inhibited,and cell proliferation index decreased from(56.56±4.25)%to(43.31±4.69)%(P<0.05).After SGK1 was inhibited,HUC proliferation effect induced by stretch stress was also significantly inhibited,and cell proliferation index decreased from(56.92±3.22)%to(42.24±1.85)%(P<0.05).Conclusion It is demonstrated that periodic stretch stress can promote HUC proliferation by PI3K/SGK1 signaling pathway but not PI3K/Akt signaling pathway.

关 键 词：尿路上皮细胞 牵张应力 细胞增殖 机械信号转导 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学] Q813.5[医药卫生—基础医学]