Hepatic Fibrosis In Schistosomiasis Japonica Reversed By Macrophages Transfected With LV-mmp13

Background Our previous studies have shown that dense granule protein 15 of type II Toxoplasma gondii(GRA15_?)can induce macrophage polarization to M1;furthermore,the constructed lentivirus vector containing gra15_?(LV-gra15_?)can transform macrophage into M1(LV-gra15_?-M1)in vivo and vitro,which prevented liver fibrosis of schistosomiasis significantly.We found that LV-gra15_?-M1 secreted matrix metalloproteinase 13?MMP13?which is an important molecule for dissolving collagen protein.In this study,we firstly constructed lentivirus vector containing mmp-13?LV-mmp13?,then transfected it into macrophage in vitro?LV-mmp13-M?.Finally,we detected the activation state of LV-mmp13-M and infused it into mouse with liver fibrosis of schistosomiasis to understand the effect on and mechanism in anti-fibrosis.Methods LV-mmp13 and LV-gra15 IIwere first constructed,then transfected to RAW264.7.The stable expressing strain was chosen for further study.The Ly6 C were determined by flow cytometry?FCM?to estimate the expression of Ly6 C.After 6 weeks of Schistosoma japonicum cercariae challenge to female Balb/c mice,the mice were randomly divided into four groups: PBS,LV-blank-M,LV-gra15?-Mand LV-mmp13-M group,and the PBS and corresponding cell was infused into the mice through the tail by intravenous high pressure injection,respectively.All mice were anesthetized and sacrificed at 8 weeks post challenge.Serum was taken to detect ALT,hyaluronic acid?HA?content by ELISA.Liver tissue was taken for hydroxyproline?HYP?content detection,H&E,Masson and immunohistochemistry staining,as well as q RT-PCR and western-blotting analysis.Meanwhile,liver macrophages were extracted to detect the expression of LV-mmp13.In vitro,the RAW264.7,LV-blank-M,LV-gra15?-Mand LV-mmp13-M was co-cultured with mouse fibroblast cell line JS1,respectively,then JS1 apoptosis was detected by FCM.MMP13,Col? and CCL2 in cellular supernatant was assayedrespectively by ELISA.Result Stably expressing LV-mmp13-M and LV-gra15_?-M was successfully constructed,respectively,LV-gra15_?-M had a M1 phenotype.LV-mmp13-M showed low expression of Ly6 C compared with LV-gra15_?-M.In the LV-mmp13-M group,blood HA,liver HYP,Col?,liver granuloma size and fibrosis degree was significantly improved,respectively,which indicated that LV-mmp13-M could effectively reverse liver fibrosis.Further studies showed that LV-mmp13-M entered liver tissue smoothly and secreted a large amount of MMP13 protein,which promoted collagen dissolution.In addition,CCL2 and some other cytokinessecreted from LV-mmp13-M also play a role in inducing apoptosis of liver fibroblasts and inhibiting liver fibrosis.This study also suggests that LV-gra15_?-M no apparent therapeutical effect on liver fibrosis.Conclusion LV-mmp13-M can reverse liver fibrosis in schistosomiasis japonicum by secreting MMP13,CCL2 and some other cytokines to dissolve collagen fibers and promote the apoptosis of liver fibroblasts.