青藤碱联合二甲双胍对肝癌SMMC-7721细胞系裸鼠模型的促存活、抗凋亡和对TGF-p/P38/HsP27通路的抑制

Synergistic Effect of Sinomeninein Combination with Metformin on Hepatocellular Carcinoma Cells and Its Mechanism in Rat Models

目的探究青藤碱(Sinomenine)联合二甲双胍(Metformin)在裸鼠肝癌(hepatocellular carcinoma,HCC)模型中对肿瘤的效用及作用机制。方法建立裸鼠HCC细胞系SMMC-7721皮下模型,采用100mg/kg Sinomenine与100 mg/kg Metformin单独或联合处理并测量肿瘤体积,裸鼠体重以及最终裸鼠的生存曲线。通过TUNEL染色检测肿瘤细胞凋亡,免疫组化检测肿瘤组织中Ki67、Caspase-3和VEGF表达情况。利用蛋白质免疫印迹技术检测肿瘤增殖、凋亡及其下游靶基因等相关分子表达。结果与SMMC-7721空白对照组相比,Sinomenine或Metformin组裸鼠生存率增加,肿瘤体积及重量降低(P<0. 05 )。与Metformin单独用药组相比,联合用药组裸鼠生存率也获得提高,肿瘤体积及重量亦有所降低(P<0.05)。肿瘤细胞凋亡增加,增殖减少。凋亡基因B血表达上升,凋亡抑制因子Bcl-2,增殖因子Ki67、PCNA表达降低。血管生成生长因子(VEGF)、基质金属蛋白酶(MMP-14)降低。以及TGF-β-p-P38-p-HSP27信号通路活性降低。结论Sinomenine可增强Metformin对肝癌移植瘤模型肝癌细胞增殖抑制和凋亡促进能力,其机制可能与抑制TGF-β/P-P38通路活化有关。

Objective This study investigated the synergistic effect of Sinomenine combined with Metformin on Hepatocellular carcinoma cells and its possible mechanism in rat models.Methods The levels of proliferation-and apoptosis-related proteins were determined by Western blotting.Immunohistochemisty and TUNEL staining were performed to investigate the possible mechanisms.The in vivo activity of all bio-molecules were determined by using a mouse xenograft model.Results Compared with blank control,the survival rates in Sinomenine-or Metformin-treated rats was increased and the tumor sizes were reduced(P<0.05).Compared with rats treated with Sinomenine or Metformin alone,the rats treated by Sinomenine combined with Metformin had higher survival rate and the tumor size and weight were decreased(P<0.05).Moreover,in the Sinomenine+Metformin group,Bax gene expression was up-regulated;Bcl-2,Ki67 and PCNA expressions were down-regulated.VEGF,MMP-14 levels were lowered.The activity of the TGF-β/p-P38/p-HSP27 signaling pathways were decreased.Conclusion Sinomenine could enhance the hepato-cyte-proliferation-pn)moting effect and hepatocyte apoptosis-inhibiting effect of Metformin.The possi ble mechanism might be associated with the activation of TGF-β/P38 signaling pathway.