Effect Of Human UPA Gene-modified Bone Marrow-derived Mesenchymal Stem Cells On The Wnt Signaling Pathway In Rats With Liver Fibrosis

Objective To explore the therapeutic effect of human urokinase-type plasminogen activator(uPA)gene-modified bone marrow derived mesenchymal stem cells(BMSCs)transplantation on liver fibrosis rats included by carbon tetrachloride(CCl4),and to further investigate the effect of uPA gene-modified BMSCs on Wnt signaling pathway.Methods BMSCs,isolated and cultured in vitro,were transfected with adenoviral vector expressing human uPA(AduPA),and then the AduPA-BMSCs were transplanted into rats with liver fibrosis.Sprague-Dawley(SD)rats with 6-8 weeks old(n=40)were randomly divided into four groups.To induce liver fibrosis,the three groups(n=30)were injected subcutaneously with 40%CCl4 every three days for eight weeks.At the end of the fourth week,the four groups were respectively divided into uPA-BMSCs group(injected via the tail vein with 2×106 AduPA-transfected BMSCs,n=10),BMSCs group(injected via the tail vein with 2×106 untransfected BMSCs,n=10),model group(injected via the tail vein with an equal volume of normal saline,n=10)and normal control group(injected via the tail vein with an equal volume of normal saline,n= 10).All rats were sacrificed after 8 weeks,the serum levels of liver function and liver fibrosis were detected,histological pathology change of liver tissues were observed.The uPA expression in liver tissues was analysed by western blot.Meanwhile,reverse transcription-quantitative polymerase chain reaction(RT-qPCR)and western blot were used to examined mRNA and protein expressions of β-catenin,Wnt4 and Wnt5a in liver tissues.Results The serum levels of liver function and liver fibrosis(HA 105.71±15.21.LN 143.82±5.99.PCIII 27.14±5.92)were significantly ameliorated in the uPA-BMSCs group compared with BMSCs group and model group(P<0.05 for all).Meanwhile,HE and Masson staining showed that the area of fibrosis was obviously reduced in the uPA-BMSCs group compared with the BMSCs group(8.31%±1.21%versus 12.38%±2.27%for uPA-BMSCs group and BMSCs group,P<0.05)and model group(8.31%±1.21%versus 16.69%±1.30%for uPA-BMSCs group and model group,P<0.05).Contrasted with other groups,the uPA protein expression in liver tissues was up-regulated distinctly in the uPA-BMSCs group,whereas the mRNA as well as the protein expression of β-catenin,Wnt4 and Wnt5a in liver tissues was down-regulated in the uPA-BMSCs group(P<0.05 for all).Conclusion Transplantation of uPA gene-modified BMSCs ameliorates liver function and reduces collagen accumulation effectively in liver fibrosis rats included by CCl4,thus suppress liver fibrosis.Moreover,the rusult shows that this may be a more favorable therapeutic method than transplantation of BMSCs.Furthermore,uPA gene-modified BMSCs transplantation exerts an anti-fibrosis effect mainly by up-regulating the uPA expression,which enhances the ability of degradation for ECM.Meanwhile,it suggests that the effect may be partly due to the inhibition of Wnt signaling pathway.