Hepatobiliary Surgery Hospital and Institute, Southwest Hospital, Third Military Medical University, 400038, Chongqing, China.
Dig Dis Sci. 2010 Oct;55(10):2844-52. doi: 10.1007/s10620-009-1121-9. Epub 2010 Jan 26.
Cancer cells are thought to possess immune evasion properties due to FasL overexpression in many types of human tumors. In the present study, we set out to investigate the role of MAPK-ERK pathway in 67-kDa laminin receptor induced FasL expression and FasL-mediated apoptosis in human cholangiocarcinoma cells.
The expression of FasL and its promoter activity in cultured cholangiocarcinoma cells were examined after treatment with laminin or transfection with plasmids containing siRNA targeted to 67-kDa laminin receptor. The effects of MAPK-ERK cascade inhibitor and c-Myc inhibition by siRNA on 67-kDa laminin receptor-induced FasL expression were determined. Apoptosis assay was performed to analyze the apoptosis of lymphocytes cocultured with cholangiocarcinoma cells treated with or without MAPK-ERK cascade inhibitor.
Our results revealed that the specific MAPK-ERK cascade inhibitor, PD98059, significantly attenuated phosphorylation of c-Myc on Ser-62 and FasL upregulation in QBC-939 cells and these cells showed decreased cytotoxicity against Fas-sensitive Jurkat T cells. A luciferase reporter assay revealed that FasL promoter activity was significantly reduced in cells treated with PD98059 or transfected with c-Myc siRNA.
Based on these results, we conclude that 67LR induces FasL expression and cytotoxicity against Fas-sensitive Jurkat T cells in human cholangiocarcinoma cells through the phosphorylation of c-Myc on Ser-62 and the subsequent activation of the FasL promoter through the ERK pathway.
由于许多类型的人类肿瘤中 FasL 的过表达，癌细胞被认为具有免疫逃逸特性。在本研究中，我们着手研究 MAPK-ERK 通路在 67-kDa 层粘连蛋白受体诱导的 FasL 表达和 FasL 介导的人胆管癌细胞凋亡中的作用。
用层粘连蛋白处理或用含有靶向 67-kDa 层粘连蛋白受体的 siRNA 的质粒转染培养的胆管癌细胞后，检测 FasL 的表达及其启动子活性。测定 MAPK-ERK 级联抑制剂和 siRNA 抑制 c-Myc 对 67-kDa 层粘连蛋白受体诱导的 FasL 表达的影响。用凋亡测定法分析用或不用 MAPK-ERK 级联抑制剂处理的胆管癌细胞与淋巴细胞共培养后的淋巴细胞凋亡。
我们的结果表明，特异性 MAPK-ERK 级联抑制剂 PD98059 显著减弱了 QBC-939 细胞中 c-Myc 的丝氨酸 62 磷酸化和 FasL 的上调，这些细胞对 Fas 敏感的 Jurkat T 细胞的细胞毒性降低。荧光素酶报告基因检测显示，用 PD98059 处理或转染 c-Myc siRNA 的细胞中 FasL 启动子活性显著降低。
基于这些结果，我们得出结论，67LR 通过 Ser-62 上 c-Myc 的磷酸化和随后通过 ERK 通路激活 FasL 启动子，诱导人胆管癌细胞中 FasL 的表达和对 Fas 敏感的 Jurkat T 细胞的细胞毒性。
