摘要
目的观察Rhopaladins类似物(RPDPB)对宫颈Hela细胞增殖、凋亡的影响,并探讨其作用机制。方法取对数生长期Hela细胞,将细胞分为实验组、对照组、空白组,实验组又分为不同剂量组,不同剂量组分别以3.125、6.25、12.5、25、50、100μmol/L RPDPB干预,对照组只加DMSO培养液,空白组加入不含细胞的MEM培养液,分别于24、48、72 h采用CCK-8法检测OD值,计算细胞增殖抑制率。取对数生长期Hela细胞,将细胞分为实验组、对照组,实验组又分为不同剂量组,不同剂量组分别以12.5、25、50μmol/L RPDPB干预,对照组只加DMSO不加药,48 h后采用AnnexinV-FITC/PI双染法检测Annexin V-FITC和PI,计算细胞凋亡率。取对数生长期Hela细胞,将细胞分为实验组、对照组,实验组又分为不同剂量组,不同剂量组分别以12.5、25、50μmol/L RPDPB干预,对照组只加DMSO不加药,48 h后采用RT-PCR法检测细胞人乳头瘤病毒18 E6(HPV18 E6)、人乳头瘤病毒18 E7(HPV18 E7)、WNT2B、β-catenin mRNA和miR-145 RNA。取对数生长期Hela细胞,将细胞分为实验组、对照组,实验组又分为不同剂量组,不同剂量组分别以12.5、25、50μmol/L RPDPB干预,对照组只加DMSO不加药,48 h后采用Western blot法检测细胞WNT2B、β-catenin蛋白。结果与对照组比较,实验组细胞增殖抑制率增加,且呈时间和剂量依赖性(P均<0.05)。与对照组比较,实验组细胞早期凋亡率和晚期凋亡率增加,且呈剂量依赖性(P均<0.05)。与对照组比较,实验组HPV18 E6 mRNA、HPV18 E7 mRNA、WNT2B mRNA、β-catenin mRNA相对表达量降低,miR-145 RNA相对表达量升高,且呈剂量依赖性(P均<0.05)。与对照组比较,实验组WNT2B、β-catenin蛋白相对表达量降低,且呈剂量依赖性(P均<0.05)。结论RPDPB可抑制宫颈癌Hela细胞增殖,并诱导细胞凋亡,呈时间和剂量依懒性,作用机制可能是其通过下调HPVE6/E7 mRNA表达,导致miR-145 RNA表达上调,进而抑制下游WNT2B/β-catenin通路中WNT2B、β-catenin mRNA和蛋白表达。
Objective To observe the effects of Rhopaladins analogues(RPDPB)on the proliferation and apoptosis of cervical cancer Hela cells,and to explore the mechanism.Methods The Hela cells in the logarithmic growth phase were randomly divided into three groups:the experimental group,control group and blank group.The experimental group was then randomly divided into different dose groups,which were intervened with 3.125,6.25,12.5,25,50 and100μmol/L RPDPB,respectively.DMSO nutrient solution was added to the cells of the control group,and MEM nutrient solution without cells was added in the blank group,the OD values were detected by CCK-8 at 24,48 and 72 h,and we calculated the cell proliferation inhibition rates.Hela cells in the logarithmic growth phase were divided into the experimental group and the control group.The experimental group was divided into different dose groups,which were intervened with12.5,25 and 50μmol/L RPDPB,respectively.The cells in the control group were only added with DMSO medium.After48 h,the AnnexinV-FITC/PI double staining was used to determine AnnexinV-FITC and PI,and we calculated the apoptosis rate.The Hela cells in the logarithmic growth phase were divided into the experimental group and control group.The experimental group was then divided into different dose groups,which were intervened with 12.5,25 and 50μmol/L RPDPB,respectively.The cells in the control group were only added with DMSO medium.After 48 h,the human papilloma virus 18 E6(HPV18 E6),HPV18 E7,WNT2 B,β-catenin mRNA,and microRNA145(miR-145)RNA were detected by RT-PCR.Hela cells in the logarithmic growth phase were divided into the experimental group and control group.The experimental group was divided into different dose groups,which were intervened with 12.5,25 and 50μmol/L RPDPB,respectively.The cells in the control group were only added with DMSO medium.After 48 h,Western blotting was applied to detect WNT2 B andβ-catenin proteins.Results Compared with the control group,the inhibition rate of cell proliferation in the experimental group increased in a time-and dose-dependent manner(P<0.05).Compared with the control group,the early and late apoptosis rates in the experimental group increased in a dose-dependent manner(both P<0.05).The relative expression of HPV18 E6 mRNA,HP18 E7 mRNA,WNT2 B mRNA andβ-catenin mRNA in the experimental group decreased,and the relative expression of miR-145 mRNA increased in the experimental group in a dose-dependent manner(all P<0.05).Compared with the control group,the relative expression levels of WNT2 B andβ-catenin proteins in the experimental group were reduced in a dose-dependent manner(both P<0.05).Conclusions RPDPB can inhibit the proliferation and induce the apoptosis of Hela cells of cervical cancer in a time-and dose-dependent manner.The mechanism may be that it could up-regulate the expression of miR-145 RNA,and furthermore inhibit the expression of WNT2 B andβ-catenin mRNA and the expression of WNT2 B andβ-catenin protein expression in the WNT2 B/β-catenin pathway via down-regulating the expression of HPV18 E6/E7 mRNA.
作者
王彦娇
曾小华
柯丽娜
朱秀莲
陈琴华
李斌
WANG Yanjiao;ZENG Xiaohua;KE Lina;ZHU Xiulian;CHEN Qinhua;LI Bin(Graduate Training Base,Chinese Medicine Dongfeng General Hospital,Jinzhou Medical University,Shiyan 442008,China;不详)
出处
《山东医药》
CAS
2022年第12期30-34,共5页
Shandong Medical Journal
基金
国家自然科学基金资助项目(81872509)
湖北省卫生健康委员会中医药专硕面上项目(2Y202111051)
湖北省教育厅科学技术研究项目(B2020106)
作者简介
第一作者:王彦娇(1994-),女,在读硕士,主要研究方向为妇科肿瘤。E-mail:2604540563@qq.com;通信作者:李斌(1962-),男,主任医师,教授,主要研究方向为妇科肿瘤。E-mail:1562096135@qq.com
