HIGH CHOLESTEROL DIETS EXACERBATE MOUSE LIVER FIBROSIS THROUGH BUILDUP OF FREE CHOLESTEROL IN HEPATIC STELLATE CELLS

HSC activation.Manganese superoxide dismutase (MnSOD) is an antioxidant enzyme in mitochondria, the major source of superoxide anions.CuZnSOD is mainly responsible for dismutation of superoxide anions in the cytosol.The function of SODs in the activation of HSCs has not been investigated yet.Aim: To determine the expression and role of superoxide dismutases in HSC activation.Methods: Experiments were performed in the human HSC cell line (LX-2) and in rat hepatic stellate cells.MnSOD and CuZnSOD were downregulated in LX-2 cells using siRNA.Expression of mRNA levels was determined using qPCR.Protein was quantified using Western Blot and localized by immunofluorescence.Results: MnSOD expression in cultured rat HSCs declined rapidly during activation, while CuZnSOD expression remained constant.MnSOD and CuZnSOD mRNA levels were effectively downregulated 24 up to 96 hrs after transfection in LX-2 cells.Reduced expression of MnSOD was also confirmed at protein level.MnSOD downregulation resulted in a morphological change of LX-2 cells to a stretched morphology.mRNA levels of activation markers (a-SMA, TGF-b and collagen type 1) were induced after downregulation of only MnSOD, not CuZnSOD.Conclusion: Our results suggest that downregulation of MnSOD is linked to the activation of HSCs.Apparently, mitochondria-derived superoxide anions are the most important for HSC activation, since downregulating CuZnSOD shows no effect on LX-2 activation.As the reduced expression of MnSOD may be the driving force of HSC activation, interventions to increase MnSOD levels in HSCs may be considered as a therapeutic target for liver fibrosis.