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目的探讨多房棘球绦虫原头蚴对体外培养宿主肝细胞丝裂素活化蛋白激酶(mitogen activated protein kinase,MAPK)信号通路的影响。方法采用宿主大鼠肝细胞BRL3A和人源肝癌细胞系HepG2,分别与多房棘球绦虫原头蚴体外无血清共培养。实验组(与多房棘球绦虫原头蚴共培养)每瓶细胞接种约3×103个原头蚴;处理组(U0126)每瓶细胞先用20μmol/L U0126处理3 h,再接种约3×103个原头蚴,继续共培养,分别在12、24、48、72、96和120 h收集细胞,同时收集对照组(无血清培养基)细胞。利用免疫印迹技术(Western blot)检测p-ERK1/2、p-p38和p-JNK的变化。结果在48 h,肝细胞BRL3A共培养组p-ERK1/2表达活性与对照组相比差异有统计学意义(P<0.05),在12、24和48 h,p-JNK和p-p38表达活性分别有微弱升高;在72和96 h肝细胞HepG2共培养组p-ERK1/2表达活性与对照组相比差异有统计学意义(P<0.05),p-JNK和p-p38表达活性无明显变化;U0126处理组p-ERK1/2活性受抑制。结论多房棘球绦虫原头蚴可能自身分泌细胞因子EmIns、EmBMP1/2、EmAct或egfd,通过与宿主表面受体结合激活宿主肝细胞MAPK信号通路。
Objective To investigate the effect of Echinococcus multilocularis protoscolex on mitogen activated protein kinase (MAPK) signaling pathway in cultured hepatocytes in vitro. Methods Host rat hepatocytes BRL3A and human hepatocellular carcinoma cell line HepG2 were co-cultured with serum-free cultures of Echinococcus multilocularis in vitro. In the experimental group (co-cultured with Echinococcus multilocularis), each bottle of cells was inoculated with about 3 × 103 progenitor cells. In the untreated group (U0126), each bottle of cells was treated with 20 μmol / L U0126 for 3 h and then inoculated with about 3 × 103 prototheca and continued to co-culture. The cells were harvested at 12, 24, 48, 72, 96 and 120 h respectively and the control (serum-free medium) cells were collected. The changes of p-ERK1 / 2, p-p38 and p-JNK were detected by Western blot. Results Compared with the control group, the expression of p-ERK1 / 2 in BRL3A co-cultured hepatocytes at 48 h was significantly different (P <0.05). At 12, 24 and 48 h, the expression of p-JNK and p-p38 The activity of p-ERK1 / 2 in HepG2 cells co-cultured with HepG2 at 72 and 96 h was significantly lower than that of the control group (P <0.05). The expression of p-JNK and p-p38 No significant change; U0126 treatment group p-ERK1 / 2 activity was inhibited. Conclusion Echinococcus multilocularis may secrete the cytokines EmIns, EmBMP1 / 2, EmAct or egfd by itself and bind to the host surface receptors to activate the MAPK signaling pathway of host hepatocytes.