Effects Of Tumor Necrosis Factor-like Weak Inducer Of Apoptosis On The Biological Behaviors Of Human Hepatic Stellate Cells And The Potential Mechanism

Background:Liver fibrosis is an intermediate process of cirrhosis in chronic liver disease.The severe complications of hepatic fibrosis bought poor living quality and prognosis.However,so far,there haven't satisfactory drugs to cure it.The activation of hepatic stellate cells(HSCs),has been known as a crucial pathogenic step in the development of liver fibrosis.The activated HSCs synthesize excessive extracellular matrix(ECM),consequently,the ECM constitutes the pseudolobules septa in liver cirrhosis.Based on the above theory,the activated HSCs turn into the critical target to reverse liver fibrosis.Tumor necrosis factor-like weak inducer of apoptosis(TWEAK)is a multifunction cytokine.It plays a role in proinflammatory,revascularization,and regulating cell growth and cell apoptosis.In the liver,the signal and function of TWEAK have mainly been explored in liver regeneration.It was reported that TWEAK promoted the proliferation of the liver progenitor cells.However,the investigation of TWEAK on liver fibrosis is limited.Objective:the objective is to explore the effects of TWEAK on proliferation,migration,cell cycle and apoptosis of human HSCs line--LX-2,and to explore the potential mechanism,which can provide a molecular basis for the therapy of liver fibrosis.Methods:LX-2 was incubated with different concentrations of TWEAK for 24 h or 48 h,and the effects of TWEAK on the proliferation of LX-2 was investigated by Cell Counting Kit-8(CCK-8).The flow cytometry was used to detect the change of cell cycles and apoptosis,and Transwell assay to detect the migration.The expression of MMP1,MMP2,MMP3,MMP7,MMP8,MMP9,MMP10,MMP11,MMP12,MMP13 gene was identified by quantitative real-time polymerase chain reaction and western blot;the activity of matrix metalloproteinases(MMPs)was tested by enzyme-linked immuno sorbent assay;small interfering RNA transfection was applied for depletion of MMP9 and p65.The phalloidin staining was used for investigating the cytoskeleton of LX-2 cells.Results:Compared with controls,20 ng/mL,40 ng/mL and 100 ng/mL TWEAK could promote LX-2 migration,and 100 ng/mL had a stronger effect;The proliferation,cell cycles and apoptosis of LX-2 was not influenced by different concentrations of TWEAK.TWEAK significantly upregulated the expression of MMP7,MMP8,MMP9,MMP13 and the activity of MMP9 in LX-2 cells.MMP9 knocking down attenuated LX-2 cells migration enhanced by TWEAK.Furthermore,we found that TWEAK activated canonical NF-?B pathway to upregulate MMP9 expression in LX-2 cells.TWEAK increased the expression of a-SMA,vimentin,desmin in LX-2 cells and changed the cell morphology.Conclusion:TWEAK promotes HSCs migration via canonical NF-?B/MMP9 pathway.TWEAK made LX-2 cells much more activated.