Experimental Study On Human Umbilical Cord Blood Stem Cell Differentiating Into Hepatocyte Through Tonification Traditional Chinese Medicine Promotion

Background and Objective: There are plenty of experiments which have proved that human umbilical cord blood stem cell(HUCBSC )can differentiate into hepatocyte in certain condition. However, the achievement ratio of HUCBSC expansion in vitro is very low and the period of HUCBSC differentiation into hepatocyte is very long. Through preliminary experiments we have found that Astragalus mongholicus, angelica, and Lycium chinense Mill can speed up HUCBSC growth and differentiation. So we select some tonification traditional Chinese medicine to promote HUCBSC differentiation into hepatocyte to enhance the therapeutic efficacy of human umbilical cord blood stem cell transplantation( HUCBSCT) and establish a new therapeutic approach for severe hepatitis.Methods: 1. Separating and cultivating HUCBSC in vitro: HUCBMNC is separated from human umbilical cord blood. Hepatocyte growth factor( HGF) and Fibroblast growth factor (FGF) are added into the HUCBMNC culture medium or only FGF is added. The growth of the cells is observed and human AFP, ALB are detected by immunohistochemistry, RT-PCR.2. Preparation of traditional Chinese medicine: The injection was bought from market. We observe the growth of cells so that we can know the influence of different dosages and kinds of medicine and we can select the suitable dosage to be used in experiment.3. We picked up some kinds of medicine (Astragalus mongholicus, angelica, and Lycium chinense Mill) which are added into culture medium of HUCBSC. The expression of human AFP and ALB are detected by immunohistochemistry and RT-PCR.Result: 1. HUCBSC in culture medium added HGF and FGF are round, spherical and polygon shape besides fusiform cells. These cells are human AFP, ALB positive by immunohistochemistry and RT-PCR; however those cells in which only FGF is added showed fusiform shape mostly and human AFP, ALB detection is negative.2. The growth of the cells is observed and the results showed that suitable dosage medicine can speed up HUCBSC growth and differentiation, the high-dosage medicine is toxic to cells and the low-dosage has no obvious influence on cell growth.3. The result of the experiment of HUCBSC differentiation into hepatocyte showed that the group which traditional Chinese medicine were added expressed human AFP, ALB more than the control group cells (no medicine is added); moreover, the former needs less time to grow and differentiate.Conclusion: 1. HUCBSC can be passaged and expanded in vitro and remain its undifferentiation.2. HUCBSC can differentiate into hepato-like cell induced by certain cytokine and can express human AFP, ALB.3. Suitable dosage tonification traditional Chinese medicine can speed up HUCBSC proliferation and differentiation.4. Large dosage traditional Chinese medicine is toxic to HUCBSC.5. Suitable dosage tonification traditional Chinese medicine can speed up HUCBSC differentiation into hepatocyte.