Induced differentiation of bone marrow stem cells in transplanted rat liver / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the differentiation of bone marrow stem cells in transplanted livers and its impact on the long-term survival of rats with orthotopic liver transplantation.</p><p><b>METHODS</b>Twenty-four female recipient rats with orthotopic liver transplantation were randomized into blank-control group, D-hanks solution group, bone marrow stem cells group with postoperative infusion of stem cells, and the pathological changes of the liver grafts and survival time of the rats were observed. The differentiation of the bone marrow stem cells were assessed 60 days after transplantation using in situ hybridization histochemistry for Sry gene and alpha-fetoprotein (AFP) immunohistochemistry.</p><p><b>RESULTS</b>In rats with postoperative infusion of bone marrow stem cells through the portal vein, the median long-term graft survival time exceeded 180 days, significantly longer than that in the other two groups (P<0.05), and no obvious evidence of acute rejection was observed with positive Sry expression and AFP expression.</p><p><b>CONCLUSION</b>Infusion of bone marrow stem cells through the portal vein following liver transplantation may alleviate acute graft rejection and promote long-term liver graft survival and AFP expression.</p>

Human bone marrow multipotent adult progenitor cells differentiate into hepatocyte-like cells with hepatocyte growth factor plus fibroblast growth factor-4 in vitro / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the possibility of the human bone marrow multipotent adult progenitor cells (hMAPCs) to differentiate into hepatocytes with hepatocyte growth factor (HGF)/ fibroblast growth factor-4 (FGF-4) in vitro.</p><p><b>METHODS</b>(1) Obtaining the hMAPCs. Bone marrow was obtained from volunteers and then centrifuged through density gradient centrifugation methods. The collected mononuclear cells were cultured through adheret culture to get mesenchymal stem cells (MSCs). The hMAPCs were obtained through collecting and isolating the MSCs by magnetic activated cell sorting (MACS) through depletion selection by use of CD45 and GlyA microbeads. (2) Differentiation of the hMAPCs with HGF+FGF-4. Group A: HGF (20 ng/ml) + FGF-4 (10 ng/ml) induced hMAPCs; group B (positive control group): L-02 human hepatocytes(cell lines); and group C (negative control group): the undifferentiated hMAPCs. (3) The expressions of albumin (Alb), alpha fetoprotein (AFP), cytokeratin-18 (CK-18), and cytokeratin-19 (CK-19) were detected with immunocytochemistry to identify the characteristics of the differentiated cells at different times and the ratio of the positive cells was determined. (4) ALB, AFP, CK-18, and CK-19 expressions of the differentiated cells were detected by RT-PCR assay to investigate the mRNA transcriptions of characteristic hepatic proteins. (5) Alb expressions of the differentiated cells at different times were detected by Western blot on the 21st and 35th days.</p><p><b>RESULTS</b>(1) The results of immunocytochemistry. The staining of Alb, CK18 were essentially positive in group A. As an early marker of immature hepatocytes, AFP staining was positive on the 7th day but negative in later differentiating periods in group A. (2) The results of RT-PCR. On the 7th day, the differentiated hMAPCs expressed AFP mRNA but were negative in later differentiating periods. On the contrary, the mRNA of Alb and CK-18 were positive at all times. (3) The results of Western blot assay. Alb protein was positive on the 21st day and 35th day.</p><p><b>CONCLUSIONS</b>Under some definite inducing conditions hMAPCs can differentiate into hepatocyte-like cells. They may serve as a potential cell source for liver engineering.</p>

Protecting effects of p44/42 MAPK signal transduction pathway on hepatocytes in ischemic preconditioning / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the significance of PKC and p44/42 mitogen-activated protein kinase (MAPK) signal transduction in ischemic preconditioning (IP).</p><p><b>METHODS</b>Through liver cell IP models, PKC inhibitor and MEK inhibitor were utilized to analyze the phosphorylation level of p44/42 MAPK and cell viability was also observed. Rat liver IP models were established which were treated with various drugs. Then the phosphorylation level of p44/42 MAPK in vivo and serum alanine aminotransferase (ALT), aspartate aminotransferase (AST) concentrations were detected. And cellular structures were observed under light microscopy.</p><p><b>RESULTS</b>Similar results were obtained in vivo and in vitro IP models. Compared with the ischemia reperfusion (IR) group in vivo, the phosphorylation level of p44/42 MAPK was obviously increased in IP treated rats (q = 27.217, P < 0.01), and the cellular structure injured slightly. The concentrations of serum ALT and AST in IP group were significantly lower than those in IR group (281.0 U/L +/-35.6 U/L vs 762.8 U/L +/-130.5 U/L and 407.7 U/L +/-73.7 U/L vs 820.9 U/L +/-111.3 U/L, P < 0.01). However, opposite changes were found in PKC and MEK inhibited groups, when compared to IP group. The phosphorylation level of p44/42 MAPK was obviously decreased, the liver tissues injured evidently, and the concentrations of serum ALT and AST (645.61 U/L +/-90.4 U/L, 678.6 U/L +/-136.5U/L and 466.2 U/L +/-82.8 U/L, 732.9 U/L +/-91.1 U/L, respectively) were significantly greater than those in IP group.</p><p><b>CONCLUSION</b>These results suggest that p44/42 MAPK pathway plays a vital role in the protection of hepatocytes in ischemic preconditioning.</p>