Protective Effect Of Calcitriol On Hepatic Fibrosis Induced By Bile Duct Ligation In Mice And Its Mechanism

ObjectiveTo investigate the protective effect of calcitriol on liver fibrosis induced by common bile duct ligation(BDL)in the mice,and to explain its possible mechanism.MethodsForty-five male C57BL/6 mice aged 8 to 10 weeks were randomly divided into sham operation group,model group(BDL group)and treatment group(BDL+calcitriol group),with 15 mice in each group.The mice were anesthetized with 2% isoflurane and fixed on the operating table.During the operation,the body temperature was kept at 37℃ and the skin was prepared.The mice in sham operation group received the surgical line around the common bile duct without ligation,in BDL group and BDL+calcitriol group,the common bile ducts of the mice were double-ligated and severed,and the mice in BDL+calcitriol group were injected intraperitoneally with calcitriol(2.5μg·kg-1)3times a week after operation.The mice in sham operation group and BDL group were injected with an equal volume of saline.The administration was continued until 4 weeks after the operation.On the28 th day,blood was taken from the orbit and the liver of the mouse was taken.The activities of alanine aminotransferase(ALT),aspartate aminotransferase(AST)and hydroxyproline(Hyp)in serum of each group of mice were detected after operation,as well as total bile in liver tissue of each group of mice acid(TBA)and total bilirubin(TBIL)levels.The pathomorphology and fibrosis degrees of liver tissue were observed with hematoxylin and eosin(HE)and Sirius red staining.Western blotting method was used to detect the expression levels of phosphorylated extracellular signal-regulated protein kinase(p ERK),extracellular signal-regulated protein kinase(ERK),α-smooth muscle protein(α-SMA),transforming growth factor-β1(TGF-β1),type I collagen(Col Ι)and Desmin in liver tissues of the mice in various groups.Immunohistochemistry method was used to detect the expressions of α-SMA and TGF-β1 in liver tissue of the mice in various groups.ResultsIn the detection of serum enzymes,compared with sham operation group,the liver activities of ALT,AST and Hyp and the serum levels of TBA and TBIL in BDL group were significantly increased(P<0.01);compared with BDL group,the liver activities of ALT,AST and Hyp and the serum levels of TBA and TBIL in BDL+calcitriol group were significantly reduced(P<0.01).The HE staining results showed that the liver lobules of the mice in sham operation group were normal,the tissue structure was relatively complete,and no inflammatory cell infiltration was seen;the liver tissue of the mice in BDL group had inflammatory cell infiltration,and necrosis and collagen deposition were more obvious;compared with BDL group,the necrotic area of??liver tissue of the mice in BDL+calcitriol group was significantly reduced,and the inflammatory infiltration was improved.The Sirius red staining results showed that only a small amount of collagen deposition in the liver tissue of mice in sham operation group appeared around the central vein;the central vein and portal area of liver tissue of the mice in BDL group showed obvious collagen deposits;compared with BDL group,the collagen deposition in BDL+calcitriol group was significantly reduced.The Western blot results showed that compared with sham operation group,the expression levels of p ERK,ERK,α-SMA,TGF-β1,Col Ι and Desmin inliver tissue of the mice in BDL group were significantly increased(P<0.01);compared with BDL group,the expression levels of p ERK,ERK,α-SMA,TGF-β1,Col Ι and Desmin in liver tissue of the BDL+calcitriol group mice were significantly reduced(P<0.01).The immunohistochemical staining results showed that the expressions ofα-SMA and TGF-β1 proteins in liver tissue of the mice in BDL group were higher than those in sham operation group(P<0.01),and the expressions of α-SMA and TGF-β1 proteins in BDL+calcitriol group were significantly lower than those in BDL group(P<0.01).ConclusionsCalcitriol can reduce the activation of hepatic stellate cells(HSCs)by activating the vitamin D receptor(VDR),down-regulating ERK protein,reducing the expression of TGF-β1,and inhibiting the deposition of collagen between tissues,thereby alleviate BDL-induced liver fibrosis in the mice.