白细胞介素-6对胰腺癌小鼠癌组织的细胞增殖及上皮间质转化的影响

Influence of interleukin-6 in cell proliferation and epithelial mesenchymal transition in carcinoma tissues in mice with pancreatic cancer

目的 探讨白细胞介素-6(IL-6)在胰腺癌裸鼠模型癌组织增殖与上皮间质化中的作用,并分析其分子机制。方法 将18只成功构建胰腺癌细胞荷瘤的BALB/c裸鼠模型随机分为模型对照组、IL-6过表达组和免疫球蛋白G(IgG)抗体组,每组6只。模型对照组于肿瘤部位注射生理盐水进行干预,IL-6过表达组于肿瘤部位注射IL-6过表达质粒进行干预,IgG抗体组予肿瘤部位注射IgG抗体进行干预。3组均于末次给药24 h后处死小鼠,并收集胰腺肿瘤组织。对各组肿瘤组织进行离体称重并计算抑瘤率。通过苏木精-伊红(HE)染色观察胰腺组织的病理学改变。采用酶联免疫吸附试验(ELISA)检测各组裸鼠肿瘤组织中IL-6、p-STAT3、NF-κB p65和VEGF蛋白的表达水平。采用蛋白质印迹法(Western blot)检测各组肿瘤组织中E-cadherin、Vimentin、MMP-9蛋白的表达水平。采用实时荧光定量聚合酶链反应(qRT-PCR)检测各组IL-6、p-STAT3、SOCS3基因的表达水平。结果 3组裸鼠胰腺肿瘤组织的平均瘤重及抑瘤率比较,差异均无统计学意义(P>0.05)。HE染色结果显示,模型对照组肿瘤组织细胞体积增大,癌细胞排列呈巢状;IL-6过表达组可见肿瘤组织轻度纤维化,少量炎症浸润;IgG抗体组可见核分裂象,大量肿瘤细胞坏死,局灶组织纤维化。IL-6过表达组E-cadherin蛋白、SOCS3基因表达水平低于模型对照组和IgG抗体组,而Vimentin、MMP-9、IL-6、p-STAT3、NF-κB p65、VEGF蛋白的表达水平及IL-6、NF-κB p65基因的表达水平高于模型对照组和IgG抗体组,差异均有统计学意义(P<0.05);IgG抗体组Vimentin、MMP-9、IL-6、p-STAT3、NF-κB p65、VEGF蛋白的表达水平及IL-6、NF-κB p65基因的表达水平低于模型对照组,而E-cadherin和SOCS3基因水平高于模型对照组,差异均有统计学意义(P<0.05)。结论 IL-6可以通过p-STAT3和(或)NF-κB p65信号通路调节胰腺癌裸鼠模型肿瘤组织中的相关细胞因子,影响肿瘤组织的增殖及上皮间质化。

Objective To explore the role of interleukin-6(IL-6)in the proliferation and epithelial mesenchymalization of cancerous tissues in a nude mouse model of pancreatic cancer,and to analyze its molecular mechanism.Methods Eighteen BALB/c nude mice,who were successfully constructed with pancreatic cancer cell-loaded tumors,were randomly divided into model control group,IL-6 overexpression group and immunoglobulin G(IgG)antibody group,six mice in each group.The control group was injected with saline at the tumor site for intervention,the IL-6 overexpression group was injected with IL-6 overexpression plasmid at the tumor site for intervention and the IgG antibody group was injected with IgG antibody at the tumor site for intervention.The mice in the three groups were executed 24 h after the last intervention,and the pancreatic tumor tissues were collected.The tumor tissues of each group were weighed ex vivo and the tumor inhibition rate was calculated.The pathological changes of pancreatic tissues were observed by hematoxylin-eosin(HE)staining.The expression levels of IL-6,p-STAT3,NF-κB p65 and VEGF proteins in the tumor tissues of nude mice in each group were detected by enzyme-linked immunoso rbent assay(ELISA).The expression levels of E-cadherin,Vimentin,and MMP-9 proteins in tumor tissues of each group were detected by Western blot.The expression levels of IL-6,p-STAT3 and SOCS3 genes in each group were detected by qRT-PCR.Results Comparison of the average tumor weight and tumor inhibition rate of pancreatic tumor tissues in the three groups of nude mice showed no statistically significant difference(P>0.05).HE staining results showed that the tumor tissues were with an increase in the cell volume and the cancer cells were arranged in a nested shape in the model control group;tumor tissues could be seen to be mildly fibrotic,with a small amount of inflammatory cell infiltration in the IL-6 overexpression group;nuclear schizophrenia was seen,with a large number of tumor cells necrotic and focal tissue fibrosis in the IgG antibody group.In the IL-6 overexpression group,the expression levels of E-cadherin protein and SOCS3 gene were lower than those in the model control group and the IgG antibody group,while the expression levels of Vimentin,MMP-9,IL-6,p-STAT3,NF-κB p65 and VEGF proteins,as well as IL-6 and NF-κB p65 genes,were higher than those in the model control group and the IgG antibody group,and the differences were statistically significant(P<0.05);in the IgG antibody group,the expression levels of Vimentin,MMP-9,IL-6,p-STAT3,NF-κB p65,VEGF proteins and the expression levels of IL-6 and NF-κB p65 genes were lower than those in the model control group,while the levels of E-cadherin and SOCS3 genes were higher than those in the model control group,and the differences were all statistically different(P<0.05).Conclusion IL-6 can regulate related cytokines in tumor tissues of nude mouse model of pancreatic cancer through p-STAT3 and/or NF-κB p65 signaling pathway,affecting proliferation and epithelial mesenchymal transition of tumor tissues.