CAJ | 학술논문

目的观察洛铂对胆管癌RBE细胞抗肿瘤作用及B细胞淋巴瘤/白血病-2相关X蛋白(bax)、B细胞淋巴瘤/白血病-2(bcl-2)、actived-半胱氨酰天冬氨酸特异性蛋白酶(Caspase)-3、Caspase-8和细胞色素C(Cyt-C)转录及表达的影响.方法不同浓度洛铂干预胆管癌RBE细胞一定时间,噻唑蓝(MTT)比色法检测RBE细胞的生长抑制,流式细胞法检测细胞周期,膜联蛋白V(Annexin V)/碘化丙锭(PI)双染检测凋亡率,Western blot及实时定量聚合酶链反应(Real-time PCR)检测bax、bcl-2、actived-Caspase-3、Caspase-8、Cyt-C蛋白及mRNA表达的变化.结果 MTT结果显示随浓度升高及时间延长,药物对细胞增殖抑制作用增强(P＜0.05),24 h的半数抑制剂量(IC50)值为11.82 mg/L.随药物浓度增加G0/G1期与G2/M期细胞增加,S期细胞比例减少(P＜0.05).Annexin V-异硫氰酸荧光素(FITC)/PI检测显示洛铂能诱导RBE细胞凋亡,与对照组比较有更高的凋亡率(Pp＜0.05).Western blot及Real-time PCR检测结果显示bax、actived-Caspase-3、Caspase-8和胞质内Cyt-C转录和表达升高,bcl-2转录水平降低.结论洛铂对胆管癌RBE细胞的增殖具有明显的抑制作用,其可能通过增加bax、抑制bcl-2的转录和表达,释放Cyt-C激活Caspase-8、Caspase-3诱导RBE细胞凋亡.
목적 관찰락박대담관암RBE세포항종류작용급B세포림파류/백혈병-2상관X단백(bax)、B세포림파류/백혈병-2(bcl-2)、actived-반광안선천동안산특이성단백매(Caspase)-3、Caspase-8화세포색소C(Cyt-C)전록급표체적영향.방법 불동농도락박간예담관암RBE세포일정시간,새서람(MTT)비색법검측RBE세포적생장억제,류식세포법검측세포주기,막련단백V(Annexin V)/전화병정(PI)쌍염검측조망솔,Western blot급실시정량취합매련반응(Real-time PCR)검측bax、bcl-2、actived-Caspase-3、Caspase-8、Cyt-C단백급mRNA표체적변화.결과 MTT결과현시수농도승고급시간연장,약물대세포증식억제작용증강(P＜0.05),24 h적반수억제제량(IC50)치위11.82 mg/L.수약물농도증가G0/G1기여G2/M기세포증가,S기세포비례감소(P＜0.05).Annexin V-이류청산형광소(FITC)/PI검측현시락박능유도RBE세포조망,여대조조비교유경고적조망솔(Pp＜0.05).Western blot급Real-time PCR검측결과현시bax、actived-Caspase-3、Caspase-8화포질내Cyt-C전록화표체승고,bcl-2전록수평강저.결론 락박대담관암RBE세포적증식구유명현적억제작용,기가능통과증가bax、억제bcl-2적전록화표체,석방Cyt-C격활Caspase-8、Caspase-3유도RBE세포조망.
Objective The object of this study was to determine the anti-tumor effect of lobaplatin in cholangiocarcinoma (CCA) cell line RBE and to investigate the mechanisms by determines the transcription and expression levels of B cell lymphoma/leukemia-2 associated X protein (bax),B cell lymphoma/leukemia-2 (bcl-2),Actived-Caspase-3,Caspase-8 and Cytochrome C (Cyt-C).Methods The RBE cells were cultured in vitro,followed by various concentration of lobaplatin for several times.The inhibition rates of cells were determined by methyl thiazol tetrazolium (MTT) assay and cell cycle changes were detected by flow cytometric analysis.Then,Annexin V/propidium iodide (PI) double-staining assay was employed to measure the apoptosis of cells.The change in mRNA and protein levels of bax,bcl-2,activedCaspase-3,Caspase-8 and Cyt-C were determined by Western blotting and real-time quantitative polymerase chain reaction (Real-time PCR).Results Lobaplatin was shown to be more Inhibition of cell proliferation in RBE cells,with the drug concentration and the role of time increasing.The proportion of G0/G1 and G2/M increased (P ＜0.05).In Annexin V/PI assay,the percentages of apoptosis cells were higher in lobaplatin groups with statistically significant (P ＜ 0.05).Both the Western blotting and Real-time PCR assays shown bax,Actived-Caspase-3,Caspase-8 and Intracytoplasmic Cyt-C were up-regulated at both the mRNA and protein levels,while bcl-2 was decreased at mRNA level.Conclusion Lobaplatin inhibits proliferation of RBE cell and induces apoptosis by up-regulating the bax,decreasing the bcl-2,in transcription and expression levels,releasing Cyt-C,and activating Caspase-3,Caspase-8.