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目的探讨补体C5a/C5aR通路在结肠癌发病中对纤维介素蛋白-2凝血酶原酶(fibrinogen-like protein 2,FGL2)的调节机制,明确补体系统在结肠癌发病中的功能和作用。方法收集2013年12月至2015年7月第三军医大学西南医院普外科、新桥医院消化科15例经病理活检确诊的住院临床结肠癌患者癌及癌旁组织。采用免疫组织化学方法检测结肠癌患者癌及癌旁组织中补体活化片段C5b-9、活化C3及C5aR的表达,佐证补体系统在结肠癌中的活化状态;进一步通过体内外实验(Western blot)验证结肠癌患者癌及癌旁组织MAPK信号通路中P38的磷酸化水平及FGL2的沉积,明确结肠癌发病中补体C5a/C5aR通路与二者的调节关系。结果免疫组织化学及Western blot实验证实结肠癌患者癌组织C5b-9、活化C3及C5aR的表达明显高于癌旁组织,FGL2的沉积亦显著高于癌旁组织;体外结果提示C5a能促进巨噬细胞系Raw264.7MAPK通路中P38的磷酸化水平(P=0.0013)及FGL2的产生能力(P=0.0071),加入C5aR阻断剂之后,二者表达随之减弱。结论 C5a/C5aR通路能通过MAPK信号途径中的P38信号完成对FGL2的调节作用进而参与结肠癌的发病进程。
Objective To investigate the regulatory mechanism of complement C5a / C5aR pathway in fibroinogen-like protein 2 (FGL2) in the pathogenesis of colon cancer and to clarify the function and role of complement system in the pathogenesis of colon cancer. Methods From December 2013 to July 2015, 15 cases of pathological biopsy in hospitalized patients with clinical colon cancer and adjacent tissues were collected from the Department of General Surgery, Xinqiao Hospital, General Surgery, Southwest Hospital, Third Military Medical University. The expressions of C5b-9, C3 and C5aR were detected by immunohistochemical method in colorectal cancer tissues and adjacent normal tissues to confirm the activation status of complement system in colon cancer. The results of in vitro and in vivo experiments The phosphorylation level of P38 and the deposition of FGL2 in MAPK signal pathways of colon cancer and paracancerous tissues of patients with colon cancer confirm the regulation of complement C5a / C5aR pathway and the regulation of both. Results Immunohistochemistry and Western blot confirmed that the expression of C5b-9, C3 and C5aR in colorectal cancer tissues was significantly higher than that in paracancerous tissues, and the deposition of FGL2 was also significantly higher than that in paracancerous tissues. In vitro results suggest that C5a can promote macrophages Phosphorylation level of P38 (P = 0.0013) and FGL2 production ability (P = 0.0071) in Raw264.7MAPK cell line were weaker than those in control group. The C5aR blocker decreased the expression of P38. Conclusion C5a / C5aR pathway can regulate FGL2 through P38 signal in MAPK signaling pathway and participate in the pathogenesis of colon cancer.