The Role Of Nucleophosmin In The Activation Of Hepatic Stellate Cells And Its Mechanism

Research objects:Nucleophosmin is a multifunctional and widely expressed phosphoprotein that mainly located in the nucleolus,and is involved in a variety of cell activities,especially for cell survival,and the phenomenon of NPM overexpression has been confirmed in many types of cancer.Liver fibrosis is a long-term interaction with a variety of stimulating factors in the liver disease,and its morbidity and mortality have an increasing trend in the world.However,there is no clear conclusion about the molecular mechanism of hepatic fibrosis.This paper is based on TGF-beta stimulates hepatic stellate cells to activate,therewith,the expression of NPM and related liver fibrosis markers proteins such as ACTA2 were increased.Then,we studied the role of NPM in the development of hepatic fibrosis and its preliminary molecular mechanism to provide theoretical basis for exploring effective treatment of liver fibrosis.Methods:Firstly,we studied the role of NPM in the development of liver fibrosis by stimulating LX-2 cells with TGF-beta.Secondly,knockout or silence endogenous NPM by CRISPAR-Cas9/SiRNA,and verified by Western blot and RT-qPCR.Finally,we used NSC348884(NPM oligonucleotids inhibitor)and MK2206(AKT phosphorylation inhibitor)to stimulate LX-2 cells,and verify whether NPM is involved in the development of liver fibrosis by Akt pathway.Simultaneously,we detected the effect of inhibitor on reactive oxygen levels,cell cycle and apoptosis by flow cytometry.Results:TGF-beta stimulated LX-2 cells to activate,then NPM and ACTA2 gene expression levels were increased,indicating that NPM involved in the occurrence of liver fibrosis.During TGF-beta treat LX-2,p-AKT protein expression level also increased.In knockout NPM expression of hepatic stellate cells,the expression of liver fibrosis marker proteins such as ACTA2,Collagen I and MMP9 are decrease significantly,and the expression levels of p-AKT/p-mTOR is inhibited.NSC348884(NPM oligonucleotids inhibitor)and MK2206(AKT phosphorylation inhibitor)treated LX-2 cells,the expression levels of liver fibrosis marker proteins-ACTA2,Collagen I and MMP9 decreased with the reduction of NPM.In addition,the level of the reactive oxygen species treated with TGF-beta in 5h have significantly improved.Conclusion:NPM is involved in the activation of hepatic stellate cells,which is the central event of hepatic fibrosis,and plays a role in the development of hepatic fibrosis.Knockout or silence NPM expression will inhibit the expression of hepatic fibrosis marker(ACTA2,Collagenl and MMP9).NPM is likely involved in the regulation of liver fibrosis through the Akt pathway.