沉默小鼠成纤维细胞活化蛋白表达对原代胰腺癌细胞增殖及凋亡的影响  

Effect of fibroblast activation protein expression silencing of mouse fibroblast cells on the proliferation of mouse pancreatic cancer cells

作　　者：邵叶波[1] 靳大勇[1] 戎叶飞[1] 许雪峰[1] 

机构地区：[1]复旦大学普外科研究所复旦大学附属中山医院普外科,上海200032

出　　处：《中华胰腺病杂志》2012年第4期242-245,共4页Chinese Journal of Pancreatology

基　　金：复旦大学附属中山医院青年基金资助项目（科补-261）

摘　　要：目的应用RNA干扰技术沉默小鼠胰腺癌相关成纤维细胞的成纤维活化蛋白（FAP）表达，观察其对小鼠原代胰腺癌细胞增殖及凋亡的影响。方法构建靶向小鼠FAP基因的重组表达质粒siFAP及对照质粒siMOCK，分别转染小鼠原代胰腺癌相关成纤维细胞mPCa-FCs．1212，采用定量RT-PCR法及蛋白质印迹法检测转染细胞的FAPmRNA及蛋白表达。将该细胞与小鼠原代胰腺癌细胞mPCa-1212按1：1的比例共培养，应用MTr比色法检测mPCa-1212细胞的增殖抑制率，应用AnnexinV．FITC／PI染色及流式细胞仪检测细胞凋亡。结果转染重组质粒siFAP的mPCa-FCs-1212细胞的FAPmRNA及蛋白表达较转染对照质粒siMOCK的细胞的表达明显下调[0．584±0．029比1．052±0．281，P：0．0213；（27．18±3．23）％比（61．58±4．72）％，P=0．0317]。转染siFAP和转染siMOCK的mPCa-FCs-1212分别与mPCa．1212细胞共培养3d后，mPCa-1212的增殖抑制率分别为（23．02±3．32）％和（1．11±0．23）％；细胞凋亡率分别为（42．31±5．34）％和（7．38±2．09）％，两组细胞的差异具有统计学意义（P=0．000）。结论沉默FAP基因的mPCa-FCs-1212在体外可有效抑制mPCa-1212细胞的增殖，诱导细胞凋亡，可能是-种潜在的基因治疗新方法。Objective Small interfering RNA （siRNA） was used to silence the fibroblast activation protein （FAP） expression of mouse pancreatic cancer related fibroblast cells （mPCa-FCs-1212）, and to observe the effects of mPCa-FCs-1212 silencing FAP gene on mouse pancreatic cancer cells （mPCa-1212） proliferation and apoptosis. Methods The small interfering RNA targeting FAP gene was designed; the recombinant siRNA plasmid siFAP and control plasmid siMOCK was constructed, which were transfected into mPCa-FCs-1212, respectively. The FAP mRNA and protein expression in transfected cells were examined by real-time PCR and Western blotting. The mPCa-1212 and transfected mPCa-FCs-1212 were co-cultured with a 1：1 ratio in vitro. The growth inhibitory rates and apoptosis rates of mPCa-1212 were detected by MTT assay and Annexin V-FITC/PI staining and FCbl assay. Results The mRNA and protein expressions of FAP in siFAP transfected mPCa-FCs- 1212 were significantly down-regulated when compared with that in siMOCK transfected mPCa-FCs-1212 [ 0.584 ± 0.029 vs. 1.052±0.281, P=0.0213; （27.18 ±3.23）% vs. （61.58 ±4.72）%,P=0.0317]. The mPCa- 1212 was co-cultured with the mPCa-FCs-1212 transfected with siFAP or siMOCK for 3 d, and the inhibitory rates of mPCa-1212 were （23.02 ±3.32）% and （1.11 ±0.23）%, and the apoptosis rates were （42.31 ±5.34）% and （7.38 ±2.09 ）%, the difference between the two groups was statistically significant （P = 0. 000 ）. Conclusions mPCa-FCs-1212 silencing FAP gene can inhibit the proliferation of mPCa-1212 in vitro and induce cell apoptosis, and may be a potential new approach to gene therapy.

关 键 词：RNA 小分子干扰 基因沉默 成纤维细胞活化蛋白 胰腺肿瘤 细胞增殖 细胞凋亡 

分 类 号：R735.9[医药卫生—肿瘤]