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

摘要:

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8 mm，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Western blot 检测DcR3 蛋白的变化；TUNEL 分析肿瘤细胞凋亡；Western blot 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），DcR3 siRNA+ 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。

Abstract:

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.