从HNRNPC基因角度探讨胰岛素抵抗发生机制

Mechanism of IR from the perspective of HNRNPC gene

目的以HNRNPC基因为研究对象,在细胞水平上,探讨HNRNPC基因、CaSR、PI3K/Akt信号通路与胰岛素抵抗(IR)之间的关系。方法建立人肝癌HepG2细胞IR模型,采用基因沉默/过表达技术干扰IR细胞及正常HepG2细胞中HNRNPC的表达。利用qRTPCR、Western blot技术检测HNRNPC表达水平,以检测沉默及过表达效率;通过Western blot技术检测各组细胞内CaSR蛋白表达量,同时检测IR相关信号通路PI3K/Akt的磷酸化水平;通过GOD-POD实验检测各组细胞葡萄糖消耗量。结果IR-HepG2组HNRNPC基因及蛋白表达升高,CaSR蛋白表达降低,PI3K/Akt信号通路被抑制,细胞葡萄糖消耗量降低;成功沉默及过表达HNRNPC后,IR-HNRNPC-siRNA组CaSR蛋白表达升高,PI3K/Akt信号通路抑制状态有所改善,细胞葡萄糖消耗量提高;HepG2-HNRNPC-OE组CaSR蛋白表达降低,PI3K/Akt信号通路被抑制,细胞葡萄糖消耗量降低。结论HNRNPC基因的高表达可下调CaSR表达,从而降低PI3K/Akt信号通路活性,这可能是IR的发生机制之一。

Objective To explore the relationship between HNRNPC,CaSR,PI3K/Akt signaling pathways and insulin resistance(IR)from the perspective of HNRNPC at the cellular level.Methods The IR model was established based on human liver cancer HepG2 cells,and gene silencing/overexpression techniques were used to interfere with the expression of HNRNPC in IR cells and normal HepG2 cells.qRT-PCR and Western blot were used to detect the expression level of HNRNPC to determine the silencing and overexpression efficiency.Western blot was used to detect the expression level of CaSR protein in cells,as well as the phosphorylation level of PI3K/Akt,an IR related signaling pathway.GOD-POD experiment was used to detect the glucose consumption of cells in each group.Results In the IR-HepG2 group,HNRNPC gene and protein expressions were increased,CaSR protein expression was reduced,PI3K/Akt signaling pathway was inhibited,and cell glucose consumption was reduced.After the successful silencing and overexpression of HNRNPC,the expression of CaSR protein in the IR-HNRNPC-siRNA group was increased,the inhibition of PI3K/Akt signaling pathway was improved,and the glucose consumption of cells was increased.In the HepG2-HNRNPC-OE group,the expression of CaSR protein was reduced,the PI3K/Akt signaling pathway was inhibited,and the cell glucose consumption was reduced.Conclusion The expression of CaSR can be downregulated by high expression of the HNRNPC gene,thereby reducing the activity of the PI3K/Akt signaling pathway,a possible mechanism of IR.