溴苯基姜黄素对胆管癌细胞凋亡、迁移和侵袭的影响及机制研究  

Study on the Effect and Its Mechanism of Bromophenylcurcumin on Apoptosis,Migration and Invasion of Cholangiocarcinoma Cells

作　　者：赵冀安[1] 马艳荣 聂文佳[3] 董梁[3] 刘文聪[4] 谷敬锋[1] ZHAO Ji’an;MA Yanrong;NIE Wenjia;DONG Liang;LIU Wencong;GU Jingfeng(Dept.of General Surgery,the First Hospital of Hebei Medical University,Shijiazhuang 050013,China;Dept.of Emergency,the First Hospital of Hebei Medical University,Shijiazhuang 050013,China;Dept.of Medical Service,the First Hospital of Hebei Medical University,Shijiazhuang 050013,China;Dept.of Ultrasonography,the First Hospital of Hebei Medical University,Shijiazhuang 050013,China)

机构地区：[1]河北医科大学第一医院普通外科,石家庄050013 [2]河北医科大学第一医院急诊科,石家庄050013 [3]河北医科大学第一医院医务处,石家庄050013 [4]河北医科大学第一医院超声科,石家庄050013

出　　处：《中国药房》2021年第4期467-474,共8页China Pharmacy

基　　金：河北省医学科学研究课题计划项目(No.20190485)。

摘　　要：目的:研究溴苯基姜黄素(GL63)对人胆管癌RBE细胞凋亡、迁移和侵袭的影响,并基于Janus激酶(JAK)/信号转导及转录激活因子(STAT)信号通路探讨其作用机制。方法:采用MTT法检测不同浓度GL63[0(空白对照,下同)、1.25、2.5、5、10、20、40μmol/L]作用48 h对RBE细胞增殖的影响,并计算其半数抑制浓度(IC_(50));采用结晶紫染色法检测不同浓度GL63(0、5、10、20μmol/L)作用48 h对细胞集落形成的影响;分别采用流式细胞术、Hoechst 33342染色法、细胞划痕实验、Transwell小室侵袭实验检测不同浓度GL63(0、5、10、20μmol/L)作用24 h对细胞周期分布、凋亡情况以及细胞迁移和侵袭能力的影响;采用Western blotting法检测不同浓度GL63(0、5、10、20μmol/L)作用24 h对细胞中JAK2/STAT3信号通路肿瘤相关蛋白表达的影响。结果:不同浓度GL63组(1.25~40μmol/L)RBE细胞的增殖抑制率均较空白对照组显著升高(P<0.01),并呈剂量依赖趋势;其IC_(50)为(8.46±1.30)μmol/L。与空白对照组比较,不同浓度GL63组(5、10、20μmol/L)RBE细胞的集落形成抑制率均显著降低(P<0.01);G0/G1期细胞占比均显著升高,S期细胞占比均显著降低(P<0.01);细胞凋亡率均显著升高(P<0.01),且细胞核呈浓染致密的固缩形态并可见凋亡小体;细胞迁移愈合率均显著降低(P<0.01),穿过基底膜的侵袭细胞数均显著减少(P<0.01);细胞中磷酸化JAK2、磷酸化STAT3、B淋巴细胞瘤-2(Bcl-2)、基质金属蛋白酶2(MMP-2)、MMP-9、前体胱天蛋白酶9(Pro-caspase-9)、Pro-caspase-3蛋白表达水平均显著降低,Bcl-2相关x蛋白(Bax)、细胞色素C、裂解Caspase-9(Cleaved-caspase-9)、Cleaved-caspase-3蛋白表达水平均显著升高(P<0.01)。结论:GL63能通过抑制JAK2/STAT3信号通路来实现对人胆管癌RBE细胞增殖、迁移和侵袭的抑制并诱导其发生凋亡。OBJECTIVE:To study the effects of bromophenylcurcumin(GL63)on the apoptosis,migration and invasion of human cholangiocarcinoma RBE cells,and to investigate its mechanism based on JAK/STAT signaling pathway.METHODS:MTT assay was used to detect the effects of different concentrations of GL63[0(blank control,similarly hereinafter),1.25,2.5,5,10,20,40μmol/L]on the proliferation of RBE cells after 48 h treatment;the IC_(50) was calculated.The effects of different concentrations of GL63(0,5,10,20μmol/L)on colony formation were detected by crystal violet staining after 48 h treatment.Flow cytometry,Hoechst 33342 staining,cell scratch test and Transwell chamber invasion test were used to detect the effects of different concentrations of GL63(0,5,10,20μmol/L)on cell cycle distribution,apoptosis,migration and invasion ability after 24 h treatment.Western blotting assay was adopted to detect the effects of different concentration of GL63(0,5,10,20μmol/L)on the expression of JAK2/STAT3 signal pathway associated proteins.RESULTS:The proliferation inhibition rates of RBE cells in different concentrations of GL63 groups(1.25-40μmol/L)were significantly increased,compared with blank control group(P<0.01),and showed a dose-dependent trend,with IC_(50) of(8.46±1.30)μmol/L.Compared with blank control group,inhibition rates of RBE cell colony formation were significantly decreased in different concentrations(5,10,20μmol/L)of GL63 groups(P<0.01).The percentage of RBE cells at G0/G1 phase increased significantly,while that at S phase decreased significantly(P<0.01).The apoptotic rate increased significantly(P<0.01),and the nucleus showed dense pyknosis and apoptotic bodies.The rate of cell migration and healing was significantly decreased(P<0.01),and the number of invasive cells through basement membrane was significantly decreased(P<0.01).The protein expression of p-JAK2,p-STAT3,Bcl-2,MMP-2,MMP-9,Pro-caspase-9 and Pro-caspase-3 were down-regulated significantly while the expression of Bax,Cyt-c,Cleaved-caspase-9 and Cleaved-c

关 键 词：溴苯基姜黄素 胆管癌 RBE细胞 Janus激酶2/信号转导及转录激活因子3信号通路 迁移 侵袭 凋亡 

分 类 号：R735.8[医药卫生—肿瘤] R965[医药卫生—临床医学]