黄连素对人卵巢癌SKOV3细胞内质网应激-自噬途径的调节作用  

Effect of berberine on endoplasmic reticulum stress-autophagy pathway in human ovarian cancer SKOV3 cells

作　　者：李松岩 于洋[2] 刘师兵[2] 崔万丽 徐路 孙奇 徐冶 LI Song-yan;YU Yang;LIU Shi-bing;CUI Wan-li;XU Lu;SUN Qi;XU Ye(School of Basic Medical Science,Jilin Medical University,Jilin 132013,China;Medical Research Laboratory,Jilin Medical University,Jilin 132013,China)

机构地区：[1]吉林医药学院基础医学院,吉林吉林132013 [2]吉林医药学院科研实验室,吉林吉林132013

出　　处：《中国病理生理杂志》2019年第10期1869-1873,共5页Chinese Journal of Pathophysiology

基　　金：吉林省教育厅十三五科学技术研究项目(No.2016238);吉林省教育厅十三五科学技术研究项目(No.JJKH20191064KJ)

摘　　要：目的:观察黄连素对人卵巢癌SKOV3细胞内质网应激-自噬途径的调节作用。方法:体外培养人卵巢癌SKOV3细胞,分别加入12.5、25、50、100、200和400μmol/L黄连素,作用12 h、24 h和48 h后,MTT法检测黄连素对SKOV3细胞活力的影响;将细胞分为对照组、黄连素(50μmol/L)组、黄连素(100μmol/L)组和黄连素(200μmol/L)组,处理SKOV3细胞24 h后,倒置相差显微镜观察黄连素对SKOV3细胞形态的影响;间接免疫荧光法检测自噬相关蛋白微管相关蛋白1轻链3(LC3)和泛素结合蛋白p62的表达;采用Western blot法检测beclin-1、LC3、p62、CCAAT/增强子结合蛋白同源蛋白(CHOP)和葡萄糖调节蛋白78(GRP78)蛋白的表达。结果:MTT结果显示,黄连素(12.5、25、50、100、200和400μmol/L)作用SKOV3细胞12 h、24 h和48 h,SKOV3细胞活力明显降低,12 h、24 h和48 h的IC 50分别为(764.7±0.3)μmol/L、(231.6±0.1)μmol/L和(96.2±0.1)μmol/L。激光共聚焦显微镜检测发现,LC3和p62蛋白散在分布,且荧光强度增加的同时呈现点状聚集,黄连素(200μmol/L)组可见明显增强,且有明显的共定位现象。Western blot检测结果表明,与空白对照组相比,黄连素(200μmol/L)组内质网应激相关蛋白GRP78和CHOP及自噬相关蛋白beclin-1、LC3和p62表达显著上升(P<0.05)。结论:黄连素可能通过调节细胞自噬促进SKOV3细胞内质网应激。AIM:To investigate the regulatory effect of berberine on the endoplasmic reticulum stress-auto-phagy pathway in human ovarian cancer SKOV3 cells.METHODS:Human ovarian cancer SKOV3 cells were cultured in vitro,and berberine at doses of 12.5,25,50,100,200 and 400μmol/L were added.After exposure for 12 h,24 h and 48 h,the viability of the SKOV3 cells was measured by MTT assay.The cells were divided into control group,berberine(50μmol/L)group,berberine(100μmol/L)group,and berberine(200μmol/L)group.After treatment with berberine for 24 h,the effects of berberine on the morphological changes of SKOV3 cells were observed under inverted phase-contrast microscope.The protein expression of microtubule-associated protein 1 light chain 3(LC3)and ubiquitin-binding protein p62 was observed by indirect immunofluorescence method under laser confocal microscope.The protein expression of beclin-1,LC3,p62,CCAAT/lenhancer binding protein homologous protein(CHOP)and glucose-regulated protein 78(GRP78)was determined by Western blot.RESULTS:Berberine at 12.5,25,50,100,200 and 400μmol/L significantly decreased the viability of SKOV3 cells at 12 h,24 h and 48 h,and the IC 50 values of 12 h,24 h and 48 h were(764.7±0.3)μmol/L,(231.6±0.1)μmol/L and(96.2±0.1)μmol/L,respectively.Laser confocal microscopy showed that the LC3 and p62 proteins were scattered and the fluorescence intensity was increased,while the point-like aggregation was also observed.Berberine at 200μmol/L obviously enhanced the co-localization of LC3 and p62 proteins.Compared with control group,the expression of endoplasmic reticulum stress-related proteins GRP78 and CHOP,and autophagy-related proteins beclin-1,LC3 and p62 in berberine(200μmol/L)group was increased significantly(P<0.05).CONCLUSION:Berberine may promote endoplasmic reticulum stress in SKOV3 cells by regulating autophagy.

关 键 词：黄连素 卵巢癌 内质网应激 自噬 

分 类 号：R737.31[医药卫生—肿瘤] R285.5[医药卫生—临床医学]