Experimental Study On Promoting Liver Injury Repair By Mesenchymal Stem Cells

Stem cells usually mean a cell population, who has the proliferative ability and differentiative potentiality at the same time. These cells can differentiate into kinds of functional cells in specific conditions, and sometimes they even form different tissues and organs. A kind of the stem cells is mesenchymal stem cells (MSCs). Among all the stem cells scientists are engaging in, MSCs may be an ideal candidate for seed cells of tissue engineering because of their multidifferentiation potentiality, simple availability, powerful proliferative ability and easy culture and enlarging. Nowadays, there still exist several problems that restrict the success of allotypic liver transplantation, for example, deficient liver donars, extensive charges for operation and sustaining, high risks, and especially the liver failure resulting from immunological repelling. Therefore researchers hope to resolve these difficulties using transplantation of MSCs. However, there is a great controversial challenge about whether MSCs can differentiate into hepatocytes. In addition, MSCs are usually cultured in vitro and transplanted into liver through portal vein, splenic vein or direct liver injection. The MSCs transplanted in this way usually have many changed biological behavior on account of differences between inner and culturing environments. So it is a hot field to research how to avail MSCs effectively and safely.In previous study we found that Bone marrow mononuclearcells(include MSCs and HSCs) which transplanted into the mice who suffered from 10 Gyγ-ray irradiation of 60Co through caudal vein could achieve and implanted in many tissue such as bone marrow,intestine,liver,spleen, skeletal muscle,brain and so on. In this research we used GFP-labeld Bone marrow mononuclearcells to establish bone marrow chimeric mice model which can easily observe the differentiation of Bone marrow mononuclearcells in vivo.To identify whether MSCs can differentiate into hepatocytes in vivo, we we transplanted MSCs into injuried liver directly. Green fluorescence protein (GFP) gene is a kind of labeling gene, the protein expressed by which can emit bright green fluorescence when it is exposed to specific band of stimulating light. Hence the differentiation of transplanted MSCs in vivo can be easily observed when we use the MSCs of GFP transgene mice.According to the reports that stem cells can be recruited to participate regeneration of cardiac infarction, we speculate the possibility to recruit MSCs to promote liver injury. We cultured and identified MSCs from G-CSF mobilized peripheral blood of rats, and observed the changes of mobilization related signal molecules such as SDF-1 after liver injury to explore the mechanisms of how MSCs in peripheral blood migrate to injuryed liver, we slso detectd their ability to differentiate into hepatocytes. The main results and conclusions are as follows.1. Bone marrow chimeric model established using MSCs transplantation could effectively promote the recovery of peripheral blood and bone marrow, therefore, helped to improve survival rate. The transplanted mice had still been alive for 16 months after irradiation. Our research group found GFP labeled cells could implanted in kinds of tissues, which provide an easy basis for observation of their behavior in vivo.2. In bone marrow chimeric mice and local implanted model we observed transplanted cells in liver. They had similar form with peripheral hepatocytes. The immunological fluorescence indicated these cells expressed GFP and albumin at the same time, suggesting Bone marrow mononuclearcells and MSCs could differentiate into functional hepatocytes.3. In chimeric mice GFP positive cells were also observed in liver as fibroblast-like and endothelial-like cells, which could be hepato-oval cells and fibroblasts or endothelial cells in sinus, suggesting there could exist several ways in which MSCs participate repairing process.4. MSCs colonies of peripheral blood were significantly higher after mobilization by 20μg/Kg G-CSF than that of 10μg/Kg. According to the form and stem cell markers we identified the cultured cells had no differences with the bone marrow MSCs. This indicates G-CSF can help to mobilize MSCs and increase their number in peripheral blood.5. The mRNA and protein leveles of SDF-1 in injuried liver tissue had increased in 5 days after injury. SDF-1 is believed to have important association with stem cell mobilization, and we speculate the increased SDF-1 in injuried liver has an ability to attract MSCs in peripheral blood to participate injury repairing. Howevr, the specific mechanisms still need to be further explored.6. Both HGF+FGF-4 and homogenate of liver tissue could induce MSCs cultured from peripheral blood to differentiate into hepatocyte-like cells. This suggests MSCs in peripheral blood have the potentiality to differentiate into hepatocytes, and they can participate the repairing process.