TRPM7通过STAT3途径参与IL-6诱导的肝细胞增殖

TRPM7 is involved in IL-6-induced hepatocyte proliferation via the STAT3 pathway

目的探讨M型瞬时受体电位通道7(TRPM7)在小鼠肝脏部分切除手术(PHx)术后的肝再生过程中表达变化和对肝细胞(L-02)增殖的影响。方法采用小鼠70%PHx肝再生模型,应用苏木精-伊红(HE)染色观察肝脏病理及ELASA法检测术后不同时间(0、12、24、36、48、72 h,5、7 d)时谷氨酸氨基转移酶(ALT)及天门冬氨酸氨基转移酶(AST)蛋白水平,应用Western blot法检测术后不同时间肝组织TRPM7和增殖细胞核抗原(PCNA)表达。用重组人白细胞介素-6(IL-6)诱导L-02细胞增殖,Western blot法检测TRPM7的表达变化。用2-氨基乙酯二苯基硼酸(2-APB)阻断TRPM7通道,MTT法检测L-02细胞活力,Western blot法检测PCNA及磷酸化-信号转导因子和转录激活因子3(p-STAT3)的蛋白表达变化。结果 PHx术后可见组织增生,细胞体积变大,增生的肝细胞索排列有不同程度的紊乱,并伴有炎性细胞的浸润。ALT及AST酶蛋白水平均在PHx术后上升,并在12 h达到高峰,而后逐渐降低至正常水平。小鼠肝组织中TRPM7的蛋白水平在PHx术后上升并且持续至72 h (P<0.05),且与细胞增殖蛋白PCNA表达水平变化具有相关性。IL-6诱导L-02细胞活力增加,且TRPM7蛋白水平也上升。而阻断TRPM7通道之后,IL-6诱导的肝细胞活力上升被2-APB抑制,并且呈浓度依赖性,同时PCNA及p-STAT3水平也降低。结论 TRPM7在肝再生和肝细胞增殖期间表达增加,并且使用2-APB阻断TRPM7通道通过减少STAT3磷酸化来抑制肝细胞增殖,表明TRPM7在肝细胞增殖中起重要作用。

Objective To investigate the expression changes of transient receptor potential melastatin 7(TRPM7) during liver regeneration after partial hepatectomy(PHx) in mice and the effect on proliferation of hepatocytes(L-02). Methods Using 70% PHx liver regeneration model in mice, hematoxylin-eosin(HE) staining was used to observe liver pathology,ELASA was used to detect alanine aminotransferase( ALT) and aspartate aminotransferase( AST) protein levels at different times( 0 h,12 h,24 h,36 h,48 h,72 h,5 d,7 d) after surgery. Western blot analysis was used to detect the expression of TRPM7 and PCNA in liver tissues at different times after operation. L-02 cells proliferation was induced by recombinant human interleukin 6( IL-6),and the expression of TRPM7 was detected by Western blot method. TRPM7 channel was blocked by 2-Aminoethoxydiphenyl borate( 2-APB),L-02 cells viability was detected by MTT assay,and protein expressions of proliferating cell nuclear antigen( PCNA),phosphorylated signal transducers and activators of transcription 3( p-STAT3) were detected by Western blot method. Results After PHx,it was observed that tissue hyperplasia was obvious,the cell volume became larger,the proliferating hepatocyte cord was arranged with different degrees of disorder while inflammatory cell infiltration was accompanied. ALT and AST protein levels increased significantly after PHx and peaked at 12 h,and then gradually decreased to normal levels. The protein level of TRPM7 in mouse liver tissue increased significantly after PHx and lasted for 72 h( P < 0. 05),which was correlated with changes in cell proliferation protein PCNA expression levels.IL-6 induced a significant increase in L-02 cell viability and TRPM7 protein levels. IL-6-induced increase of hepatocyte viability was inhibited by 2-APB in a concentration-dependent manner,and PCNA and P-STAT3 levels were also decreased after blocking TRPM7 channel. Conclusion TRPM7 increased expression during liver regeneration and hepatocyte proliferation,and inhibition of TRPM7 channel by 2-APB inhibited hepatocyte proliferation through reducing STAT3 phosphorylation,indicating that TRPM7 plays an important role in hepatocyte proliferation.