Study On The Mechanism Of Transplantation Of Differently Activated Macrophages For Murine Liver Fibrosist Treatment

【Background】Liver fibrosis following various chronic liver disesases might cause liver cirrhosis, orprimary carcinoma of liver, which seriously threatens people’s health. There are very feweffective treatment for end-stage liver cirrhosis except liver transplantation. However,fibrotic liver can return to near-normal architecture under effective treatments.Therefore, itis an urgent need to develop alternative therapies for liver fibrosis. Now, cell therapy, anemerging therapy for liver fibrosis, may be have promising clinical utility.Liver fibrosis is a wound-healing response. Kinds of chronic liver diseases, causedegeneration, necrosis and apoptosis of hepatocytes, acompanied by exudation of variousinflammatory cells. Interaction of many kinds of cells in the liver, leads to the activation ofhepatic stellate cells (HSCs), which can synthesize and secrete ECM. Once the balance ofsecretion and the degradation of ECM was broken, many ECM will be accumulated. Thenthe liver fibrosis forms. Macrophage plays an important role in this process. Macrophages are essential cells constituting innate immune system, which are central inthe pathogenesis of chronic liver injury and fibrosis. On the one hand, activatedmacrophages secrete a large number of cytokines, which activate HSCs and causedeposition of collagen; on the other hand, matrix metallopeptidases（MMPs） secreted bymacrophages, degrade the collagen fiber and induce the apoptosis of HSCs, whichalleviate liver fibrosis. These reveal that hepatic macrophages are remarkably theheterogeneous population that fulfill diverse functions in liver fibrosis. According to theirfunctions and phenotypes, macrophages can be subdivided into two groups: classicallyactivated macrophage (M1) and alternatively activated macrophage (M2). They havedifferent effect on liver fibeosis, which M1macrophage plays an antifibrotic role and M2macrophage acts as pro-fibrotic cells. A recent research suggests that macrophage celltherapy could improve clinically relevant parameters in experimental liver fibrosis. Butthere is no report about differently activated macrophages for therapy for murine liverfibrosis. In our study, we perform several experiments to explore this question.【Aims】1.To investigate the different effect of transplatation of differently activatedmacrophages for murine liver fibrosis.2.To explore the mechanism of differently activated macrophages for murine liverfibrosis treatment.【Methods】1.Establish CCl4induced liver fibrosis model in mouse.2.Establish cultural method of bone marrow derived-macrophages, and activate themby different cytokines.3.Track macrophages labeled by PEI-SPIO/Cy3-miRNA.4.Detect the fibrosis level with Sirius Red staining, hydroxyproline(Hyp) andimmunohistochemistry of Desmin.5.Detect the level of the gene expression of MMPs and chemokine (C-C motif)ligand(CCL) in differently activated macrophages, with RT-PCR, FACS and gelatinzymography assay. 6.Detect the number of macrophages infiltrating into liver, the level of geneexpression of MMPs in liver, the quantity of Hyp in urine.【Results】1. Liver fibrosis model is established; Cultural method of bone marrowderived-macrophages is successful; Labeled BMMs are detected in the Fibrotic Liver.2. Transplantation of M1and M0macrophages improves murine liver fibrosis, andM1macrophages are more effective than M0.3. Compared to M0and M2, M1macrophages have the highest-level gene expressionof MMPs, CCL2and CCL3.4. Compared to PBS, M0and M2, there are the highest-level gene expression ofCCL2, CCL3and MMPs in liver, more macrophages infiltrating into liver, and more Hypin urine of M1mice.【Conclusions】1.Transplantation of M1macrophages for liver fibrosis treatmenat is the mosteffective.2.The highest-level gene expression of MMPs in M1macrophages, and the secretionof MMPS from the macrophages infiltrating into liver, cause the degradation of the ECMin the fibrotic liver.