苍术素通过调节RhoA/ROCK1信号通路对膀胱癌细胞增殖、凋亡和血管生成拟态的影响  

Impacts of Atractylonin on Proliferation,Apoptosis and Vasculogenic Mimicry of Bladder Cancer Cells by Regulating RhoA/ROCK1 Signal Pathway

作　　者：赖海燕[1] 李茜[1] 陆一菱 刘英[1] 秦小莉 魏仁波[2] LAI Haiyan;LI Qian;LU Yiling;LIU Ying;QIN Xiaoli;WEI Renbo(Department of Pharmaceutical,Chengdu Third People's Hospital,Chengdu 610014,China;Department of Urology,Chengdu Third People's Hospital,Chengdu 610014,China)

机构地区：[1]成都市第三人民医院药学部,成都610014 [2]成都市第三人民医院泌尿外科,成都610014

出　　处：《中国细胞生物学学报》2024年第8期1497-1505,共9页Chinese Journal of Cell Biology

摘　　要：该文探讨苍术素(ATR)调节Ras同源基因家族成员A(RhoA)/Rho激酶1(ROCK1)信号通路对膀胱癌(BC)细胞增殖、凋亡和血管生成拟态(VM)的影响。利用CCK-8法检测不同浓度(0、10、20、40、80、160mg/L)的ATR对T24细胞增殖率的影响,筛选ATR干预浓度。在此基础上将T24细胞分为NC组(正常培养)、L-ATR组(20 mg/L ATR)、M-ATR组(40 mg/L ATR)、H-ATR组(80 mg/L ATR)、H-ATR+血管紧张素II(Ang II)组(80 mg/L ATR+100 nmol/L的RhoA/ROCK1通路激活剂Ang II)。MTT检测、Edu实验检测T24细胞增殖情况;流式细胞术、TUNEL法测定T24细胞凋亡情况;Matrigel基质胶法测定T24细胞血管生成情况;鬼笔环肽染色观察T24细胞骨架;Western blot测定RhoA/ROCK1通路蛋白表达情况。随着ATR浓度的增加,T24细胞增殖率呈ATR剂量依赖性降低(P<0.05),选择ATR浓度为20 mg/L、40 mg/L、80 mg/L进行后续研究。与NC组对比,L-ATR组、M-ATR组、H-ATR组T24细胞D450值、Edu阳性细胞率及管腔数量均显著降低,凋亡率及F-肌动蛋白荧光强度显著升高,且呈ATR剂量依赖性变化(P<0.05);与H-ATR组对比,HATR+Ang II组T24细胞D450值、Edu阳性细胞率及管腔数量均显著升高,凋亡率及F-肌动蛋白荧光强度显著降低(P<0.05)。与NC组对比,L-ATR组、M-ATR组、H-ATR组T24细胞RhoA、ROCK1蛋白表达水平均显著降低,且呈ATR剂量依赖性变化(P<0.05);与H-ATR组对比,H-ATR+Ang II组T24细胞RhoA、ROCK1蛋白表达水平均显著升高(P<0.05)。ATR可能通过抑制RhoA/ROCK1信号通路激活进而抑制T24细胞增殖,影响VM的形成,诱导细胞凋亡。The aim of this study was to investigate the impacts of ATR(atractylonin)on the proliferation,apoptosis and VM(vasculogenic mimicry)of BC(bladder cancer)cells by regulating the RhoA(Ras homologous gene family member A)/ROCK1(Rho kinase 1)signaling pathway.CCK-8 method was used to detect the effect of different concentrations(0,10,20,40,80,160 mg/L)of ATR on the proliferation rate of T24 cells to screen for ATR intervention concentrations.On this basis,T24 cells were separated into NC group(normal culture),L-ATR group(20 mg/L ATR),M-ATR group(40 mg/L ATR),H-ATR group(80 mg/L ATR),and H-ATR+Ang II group(80 mg/L ATR+100 nmol/L RhoA/ROCK1 pathway activator angiotensin Ang II).MTT detection and Edu assay were applied to measure T24 cell proliferation.Flow cytometry and TUNEL assay were applied to determine T24cell apoptosis.Matrigel matrix gel method was applied to measure angiogenesis in T24 cells.Phalloidin staining method was used to observe the cytoskeleton of T24.Western blot was applied to determine the expression of RhoA/ROCK1 pathway proteins.With the increase of ATR concentration,the proliferation rate of T24 cells decreased in a dose-dependent manner(P<0.05),and the ATR concentration was selected as 20 mg/L,40 mg/L,80 mg/L for follow-up studies.Compared with the NC group,the D450 value,Edu positive cell rate,and lumen number of T24 cells in the L-ATR group,M-ATR group,and H-ATR group were greatly reduced,while the apoptosis rate and fluorescence intensity of F-actin were greatly increased,showing a dose-dependent change in ATR(P<0.05).Compared with the H-ATR group,the D450 value,Edu positive cell rate,and lumen number of T24 cells in the HATR+Ang II group were greatly increased,while the apoptosis rate and fluorescence intensity of F-actin were greatly reduced(P<0.05).Compared with the NC group,the expression of RhoA and ROCK1 proteins in T24 cells in the L-ATR group,M-ATR group,and H-ATR group were greatly reduced,and showed a dose-dependent change in ATR(P<0.05).Compared with the H-ATR group,the expression

关 键 词：苍术素 膀胱癌 Ras同源基因家族成员A/Rho激酶1 增殖 凋亡 血管生成拟态 

分 类 号：R285[医药卫生—中药学]