γ-氨基丁酸抑制胆管癌细胞株QBC939生长的分子机制实验研究  

Molecular mechanism of y-aminobutyric acid inhibitory on the growth of cholangiocarcinoma QBC939 cell line

作　　者：朱成林[1] 黄强[1] 刘臣海[1] 谢放[1] 朱凯[1] 

机构地区：[1]安徽医科大学附属省立医院普外科,合肥230001

出　　处：《中华外科杂志》2013年第3期261-265,共5页Chinese Journal of Surgery

基　　金：安徽省教育厅基金资助项目（KJ2012A151）;安徽医科大学校科研基金资助项目（2011xkj067）

摘　　要：目的探讨1-氨基丁酸（GABA）抑制胆管癌细胞株QBC939增殖及凋亡的分子机制。方法分别以GABA、GABA＋荷包牡丹碱（A受体拮抗剂）、GABAphaelon（B受体拮抗剂）孵育胆管癌细胞株QBC93948h，采用四甲基偶氮唑蓝（MTT）法检测细胞的增殖活性，AnnexinV—FITC／PI双染流式细胞术检测细胞的凋亡，Westernblot法检测信号转导和转录活化因子3（STAT3）、磷酸化STAT3（p-STAT3）蛋白的表达。构建人胆管癌裸鼠皮下种植瘤模型，随机分为对照组和GABA治疗组，用药5周后观察裸鼠体重和肿瘤体积，采用免疫组化及Westernblot法检测种植瘤p-STAT3蛋白的表达。结果M，rr法及流式细胞术检测结果显示phaelon能有效拮抗GABA所导致的增殖抑制（15．30％±0．80％比2．66％±0．74％，t=23．15，P=0．00）和凋亡效应（23．15％±0．21％比4．30％±0．69％，t=52．40，P=0．00），而荷包牡丹碱无此效应。Westernblot法检测结果显示，STAT3、p-STAT3蛋白在QBC939中均表达，GABA可明显下调p-STAT3蛋白的表达（0．77±0．00比0．45±0．ol，t=63．14，P=0．00）且此效应可被phaclofn部分拈抗（0．45±0．01比0．76±0．01，t=56．25，P=0．00）；与对照组比较，治疗组的种植瘤体积明显缩小[（0．62±0．03）cm3比（0．34±0．03）cm3，t=13．45，P=0．00]，p-STAT3蛋白的表达亦下降。结论GABA抑制胆管癌细胞株QBC939的生长是通过GABA。受体介导的，下调p-STAT3蛋白的表达可能是其抗肿瘤的作用机制之一。Objective To explore the molecular mechanism of -aminobutyric acid （GABA） inhibitory on the growth of cholangiocarcinoma cell line （QBC939）. Methods QBC939 cells were cultured in different groups and treated with GABA,Y GABA ＋ bicuculine （A receptor antagonist）, GABA ＋ phaclofen （ B receptor antagonist） for 48 hours. MTT assay was used to determine the proliferation of QBC939 cells. Annexin V-FITC/PI binding assay was used to detect apoptosis in the QBC939 cells. Western blot was applied to check the expression of signal transducer and activator of transcription 3 （STAT3） and phosphorylation-STAT3 （p-STAT3） proteins in different groups of QBC939 cells. Animal models of cholangiocarcinoma bearing nude mice were established by subcutaneous injection of QBC939 cells and randomized into 2 groups： control and GABA-treated groups. The effect of GABA was evaluated after 5 weeks, including the body weight and tumor volume. The expression of p-STAT3 was detected by immunohistochemistry and Western blot in xenograft tumors. Results MTT and FCM assays both showed that the effect of GABA inhibitory on the proliferation （ 15.30% ±0. 80% vs. 2. 66% ±0. 74%, t =23. 15, P=0.00） and induced apoptosis （23.15% ±0.21% vs. 4.30%± 0.69%, t =52.40, P=0.00） of QBC939 cells could be antagonized by phaclofen, but not bicueuline. The expression of STAT3 and p-STAT3 proteins were all observed in the QBC939 cells and GABA significantly down-regulated p-STAT3 protein expression （0. 77 ± 0. 00 vs. 0. 45 ±O. 01, t = 63. 14, P = 0. 00）, this action was also antagonized by phaclofen （0. 45 ±0. 01 vs.0. 76 ± 0.01, t = 56. 25, P = 0. 00）. Xenograft tumor volume （ （0. 62 ± 0.03） cm3 vs. （0.34 ±0.03） cm3, t = 13.45, P =0.00） and the expression of p-STAT3 protein weresignificantly decreased in GABA-treated group as compared with control group. Conclusions GABA may inhibit the growth of cholangiocarcinoma ceils QBC939 through GABAs receptor, and down-regulation of the p-STAT3 expressio

关 键 词：胆管肿瘤 y氨基丁酸 STAT3转录因子 

分 类 号：R735.8[医药卫生—肿瘤]