DcR 3基因对胰腺癌裸鼠移植瘤化疗敏感性的影响

Effect of DcR3 gene on chemo-sensitivity of pancreatic cancer xenografts in nude mice

目的探讨RNA干扰沉默诱骗受体-3(DcR3)对人胰腺癌细胞裸鼠移植瘤化疗敏感性的影响及其可能机制。方法取对数生长期的AsPC-1细胞1×106个/ml,接种于5周龄裸鼠右后肢腹股沟,当皮下肿瘤直径为8mm,随机分为4组(n=10):对照组、化疗组、阴性质粒+化疗组、DcR3-siRNA+化疗组。观察DcR3-siRNA联合化疗药物对人胰腺癌AsPC-1细胞裸鼠移植瘤的治疗效果。应用ELISA和Westernblot检测DcR3蛋白的变化;TUNEL分析肿瘤细胞凋亡;Westernblot和RT-PCR检测FasL、Caspase-8和Caspase-3等凋亡因子的表达。结果化疗组、阴性质粒+化疗及DcR3-siRNA+化疗组3组均可抑制移植瘤的生长,与对照组比较,3组抑瘤率平均值分别为(35.87±4.58)%、(40.68±4.16)%和(90.25±2.53)%,4组比较差异有统计学意义(F=47.736,P=0.000),DcR3siRNA+化疗组抑瘤率较化疗组升高;对照组、化疗组、阴性质粒+化疗以及DcR3-siRNA+化疗组移植瘤重量平均值分别为(0.95±0.03)、(0.63±0.04)、(0.67±0.02)和(0.17±0.06)g,4组比较差异有统计学意义(F=85.531,P=0.000),DcR3-siRNA+化疗组瘤重较化疗组减轻;对照组、化疗组、阴性质粒+化疗组、DcR3-siRNA+化疗组凋亡率平均值分别为(6.3±2.21)%、(14.8±2.65)%、(14.5±3.06)%和(54.6±3.23)%,4组比较差异有统计学意义(F=104.225,P=0.000),DcR3-siRNA+化疗组凋亡率较化疗组提高;DcR3-siRNA+化疗组的FasL、Caspase-8和Caspase-3蛋白表达较其他各组上升(P=0.000)。结论沉默DcR3可激活FasL/Caspase凋亡途径,促进细胞凋亡,增加胰腺癌细胞移植瘤对化疗的敏感性。

Objective To investigate the effect of Decoy Receptor 3 gene(DcR3)on chemo-sensitivity of human pancreatic cancer cells.Methods Totally 1×106 pancreatic cancer AsPC-1 cells in log-growth phase were harvested and subcutaneously injected in nude mouse.Seven days post injection,tumor-bearing mice were then randomly divided into 4 groups(n=10):control group,chemotherapy group,negativeplasmid+chemotherapy group and DcR3-siRNA+chemotherapy group.Expression of DcR3 was detected by ELISA and RT-PCR.Tumor apoptosis rate was identified by TUNEL.Expressions of FasL,Caspase-8,and Caspase-3 protein were measured by Western blot.Results Tumor growth was inhibited in chemotherapy group,negative plasmid+chemotherapy group and DcR3-siRNA+chemotherapy group compared with control group[(35.87±4.58)%vs(0±0)%,(40.68±4.16)%vs(0±0)%,(90.25±2.53)%vs(0±0)%,P=0.000,respectively].Inhibitive effect inDcR3-siRNA+chemotherapy group was more obvious compared with chemotherapy only group.Tumor weight was decreased significantly in chemotherapy group,negative plasmid+chemotherapy and DcR3-siRNA+chemotherapy group when compared with control group[(0.63±0.04)g vs(0.95±0.03)g,(0.67±0.02)g vs(0.95±0.03)g,(0.17±0.06)g vs(0.95±0.03)g,F=85.531,P=0.000,respectively].Treatment of DcR3-siRNA+chemotherapy dramatically reduced tumor weight compared with the chemotherapy only group(P<0.05).Apoptosis rate in chemotherapy group,negative plasmid+chemotherapy group,and DcR3 siRNA+chemotherapy group were upregulated when compared with control group[(14.8±2.65)%vs(6.3±2.21)%,(14.5±3.06)%vs(6.3±2.21)%,(54.6±3.23)%vs(6.3±2.21)%,F=104.225,P=0.000,respectively].Apoptosis rate in DcR3-siRNA+chemotherapy group was significantly higher than that of the chemotherapy group.The expressions of FasL,Caspase-8 and Caspase-3 protein in DcR3-siRNA+chemotherapy group was enhanced compared with remaining 3 groups(P=0.000).Conclusion DcR3-siRNA gene increases the chemo-sensitivity of human pancreatic cancer by promoting cells apoptosis.