| Placenta of pregnant women as a source of amniotic tissue waste after childbirth, wide sources added quickly and without ethical restrictions in recent years, the research focus. Amnion are fetal side of the placenta wrapped in a layer of translucent membrane, its surface is not nerves, blood vessels, muscles and other tissues and lymph, separable from amniotic epithelial cells and amniotic mesenchymal stem cells. Where human placental amnion-derived mesenchymal stem cells (human amnion mesenchymal cells, hAMSCs) from extraembryonic mesoderm primitive streak stage, not only the expression of adult stem cell properties of mesenchymal stem cell properties also expressed partial embryonic stem cell properties.In addition, hAMSCs under certain conditions also have inwardly, the outer layer of the three embryonic differentiation potential, the seed cells are ideal for transplantation and tissue engineering. This study human amniotic mesenchymal stem cells isolated from in vitro culture and induced to differentiate between human amniotic mesenchymal stem cells in mice tracing and locating repair liver injury model in mice, human amniotic mesenchymal stem cell transplantation promote liver damage repair three-part study conducted to explore the role and mechanism to provide experimental data for further clinical application between mesenchymal stem cell transplantation in the treatment of liver disease.To observe the therapeutic effect of hAMSCs transplantation labeled with 5,6,2-carboxy fluorescein diacetate succinimidy ester (CFSE) on mice model with liver damage induced by carbon tetrachloride. The amniotic mesenchymal stem cells were obtained by using two-step digestion with trypsin and collagenase type I, then the flow cytometry and immunofluorescent staining techniques were adopted to identify the surface molecular markers and embryonic stem cell characteristics. Hepatic damage model groups were established through carbon tetrachloride and oil inducing,20 μl/g doses of the mixture were given to mice by abdominal injection. Mouse models of hepatic damage were randomly divided into two groups. The control group was injected with the same volume of PBS via the tail vein; the treatment group was injected with human amnion mesenchymal stem cells suspension labeled with CFSE, and the number of the stem cells was 1×106. At 4 weeks after transplantation, animals were sacrificed. Liver tissues were obtained for the histological observation and the eyeball blood was collected to detecte the liver function indicators. The human amniotic mesenchymal stem cells obtained by using two-step digestion with trypsin and collagenase type I were high-purity. At one week after transplantation, the immunofluorescent staining of frozen slice showed that human amnion mesenchymal stem cell colonization could be seen in the mice liver tissues of the cell transplantation group. The labeled human amnion mesenchymal stem cell showed strong green fluorescence. At 4 weeks after cell transplantation, compared with the model group, levels of serum aspartate aminotransferase, alanine aminotransferase in the cell transplantation group were significantly decreased, while the total albumin level was increased significantly, the liver cell inflammatory necrosis, steatosis and liver fibrosis were improved significantly. After 4 weeks, the immunofluorescent staining of frozen slice result indicated that the expressions of human serum albumin could be observed in the mice liver tissue of the cell transplantation group, but no expression could be seen in the model group. It is visible that human amnion mesenchymal stem cells can improve liver function and pathological damage of liver damage mice in a certain extent.The findings may provide useful experimental datas for cell posited transplantation with treatment liver disease.HAMSCs are multipotential adult stem cells. hAMSCs with the ability of self-proliferation and multipotential differentiation have some rescue effect for lesion tissues, but the repair mechanism of hAMSCs in liver injury remains largely unknown.This study aimed to investigate hepatocyte growth factor(HGF),silent information regulator 1(SIRT-1), a-smooth muscle actin(a-SMA), kinase inhibition protein 1(P27kip1) in liver injury rats induced by carbon tetrachloride(CCl4) after hAMSCs transplantation, and the purpose of this study is to explore the mechanism of action of hAMSCs transplanted to mice with liver fibrosis. The mesenchymal stem cells were isolated from human amniotic membrane with trypsin-collagenase, and the phenotype of hAMSCs was identified by immunofluorescence and flow cytometry. Model group were established by intraperitoneal injection of 10% CCl4 olive solution at 20Ll/g. Then 0.2 ml hAMSCs containing 1×106 cells was injected into mice in the treatment group via the tail vein. Before and after hAMSCs transplantation, Western blot and RT-PCR analysis was performed to determine the protein levels and mRNA levels of HGF, SIRT-1,α-SMA P27kipl in mice liver tissue taken from the model group and the treatment group.. Real time RT-PCR analysis showed that the mRNA levels of HGF and SIRT-1 in treatment group were higher than in model group (P<0.05). However, the mRNA levels of a-SMA and p27kipl in treatment group were lower than in model group (P<0.05). And Western blot analysis showed that the protein levels of HGF and SIRT-1 in treatment group were significantly higher than in model group (P<0.01). However, the protein levels of a-SMA and p27kipl in treatment group were significantly lower than in model group (P<0.01). The results suggest that the transplantation of hAMSCs not only can improve liver fibrosis induced by CCl4 but also can inhibit the apoptosis of hepatocytes. Besides, it can promote the mitosis of hepatocytes and had some rescue effect for liver injury. The a-SMA HGF SIRT-1 p27kipl may be key genes in liver regeneration and they can provide experimental basis for the exploration of signal pathway in liver regeneration and the repair mechanism in liver injury.In summary, hAMSCs can be isolated and cultured in vitro differentiation potential of having three germ layers, after marking hAMSCs homing able to migrate to the damaged liver tissue, and can be involved in the regulation of genes related to liver cells effectively alleviate the degree of liver damage, for the treatment of acute liver provide new experimental models and clinical aspects based on damage, cell homing and repair mechanisms.These investigations eould offer original experimental model and evidence about efficient cure of MSCs to liver disorders such as acute injury, serve a new source of cells for cell therapy of hepatic diseases. |
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