Curcumin Inhibits Hepatic Fibrosis By Inhibiting Monocyte Infiltration And Reducing Monocyte-derived GRN Secretion

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Monocyte-secret GRN promots hepatic fibrosisby activating HSCsObjectives:Hepatic fibrosis is a common pathological process of the progress of many chronic liver diseases and a serious public health problem in China.Macrophages,especially monocyte-derived macrophages,can promote liver fibrosis,but the mechanism is still unclear.Recent studies have shown that GRN secreted macrophages is able to promote fibrosis.The purpose of this study is to reveal the mechanism of GRN secreted macrophages promoting liver fibrosis,and to seek new targets for the treatment of liver fibrosis.Methods:First of all,we detected GRN expression in the liver of 121 CLD patients,and analyzed the relationship between GRN and liver inflammation,fibrosis,clinical liver disease indicators,as well as the expression differences in patients with different causes of liver disease.Then,we established mouse liver fibrosis model by repeated intraperitoneally CCl4-injection and bile duct ligation,and detected GRN expression in different stages of fibrosis model.Establishment of liver fibrosis model with GRN-/-mice was used to explore the effect of GRN on liver fibrosis;Co-culturing of GRN with hepatic stellate cells is to detect ACTa2 and Collal expression by q-PCR.and ERK1/2,P-38,JNK,AKT and NF-kB protein expression and phosphorylation levels by WB.The expression of GRN in liver during hepatic fibrosis was observed by co-staining of GRN with macrophage surface molecules CD68,F4/80 and CD11b;the expression of GRN in liver of mice after removing macrophages was observed by tail vein injection of liposome;and the co-culture of bone marrow cells and primary hepatic HSCs of wild mice or GRN-/-mice was used to observe whether monocytes-derived macrophages could activate hepatic stellate cells.Results:The expression of GRN in the liver of CLD patients was significantly higher than that of normal people,and was positively correlated with liver inflammation.There was no significant difference between different causes.The correlation analysis of liver clinical indicators in patients with CLD showed that the number of GRN + cells in liver tissue was positively correlated with ALT,AST,GGT.Tbill and Child-Pugh scores,but not with albumin level.In fibrosis models of intraperitoneally repeated CCl4-injection and bile duct ligation,GRN expression increased significantly with the increase of CCl4-injection times and the prolongation of bile duct ligation time.The expression of alpha-SMA and Col-1 in the liver of GRN-/-mice was significantly reduced.After co-culture of GRN and LX-2,the expression of ACTa2 and Collal in LX-2 increased significantly,and the phosphoric acid levels of ERK1/2 and STAT3 increased significantly.The co-staining of GRN with CD68 or CDllb in liver tissue of CLD patients was significantly higher than that of normal people.The co-staining of GRN with F4/80 and CD11b in liver tissue of fibrotic mice increased significantly.Compared with normal people,CD68 and CD11b immunohistochemistry showed that the number of CD68+and CD11b+cells in liver tissue slices of patients with hepatic fibrosis significantly increased.The number of F4/80+and CDllb+cells in the liver fibrosis mouse model was also significantly higher than that in the blank control group.Compared with wild type mice,the activation of HSCs by monocytes-derived macrophages in GRN-/-mice was significantly reduced.Conclusions:GRN expression significantly increased during liver fibrosis,which is closely related to hepatic inflammation and fibrosis.After antagonizing GRN,HSCs activation and hepatic fibrosis were alleviated.The activation of HSCs by GRN is related to phosphorylation of ERK1/2 and STAT3.GRN mainly comes from monocytes-derived macrophages.Monocyte-derived macrophages can promote HSCs activation by secreting GRN and aggravate liver fibrosis.Part ? Curcumin reduces Ly6Chi monocyte infiltration to protect against liver fibrosis by inhibiting Kupffer cells activation to reduce chemokines secretionObjectives:Curcumin has been reported to have anti-fibrotic effect.However,the anti-fibrotic mechanism of curcumin for liver fibrosis remains obscure.In the presenting study,we aimed to investigate whether curcumin reduce chemokines secretion by inhibiting kupffer cells(KCs)activation to decrease Ly6Chi monocyte infiltration in the treatment of liver fibrosis.Methods:Liver fibrosis was induced by intraperitoneal carbon tetrachloride(CCl4)-injection in mice.Mice in curcumin group received curcumin treatment by gavage.Results:Pretreatment with curcumin significantly protected mice from liver inflammation and fibrosis.Compared to CCl4 group,mice in the curcumin group showed significantly less intrahepatic infiltration of Ly6Chi monocytes,but no difference of other leucocyte subtypes.Moreover,curcumin significantly reduced Ly6Chi monocytes associated pro-inflammatory and pro-fibrogenic cytokines,which was in line with the decreased numbers of intrahepatic Ly6Chi monocytes.Further study found that curcumin is able to decrease KCs activation and monocyte chemokines,which explains why curcumin can reduce Ly6Chi monocytes infiltration during liver fibrosis.In vitro,we discovered that curcumin prevents the polarization of macrophages toward Mlandreduces monocyte chemokines secretion,which is involved with ERK1/2 and p38 pathways.Conclusions:Taken together,for the first time,we verified that curcumin can reduce chemokines secretion by inhibiting KCs activation to decrease Ly6Chi monocyte infiltration in the treatment of liver fibrosis.These results suggested that curcumin maybe considered a promising candidate in the prevention andtreatment of liver fibrosis.