The Study Of Transplant Mesenchymal Stem Cells To Liver Cirrhosis Rats Via Different Transplantation Path

Objective:To investigate the therapeutic effect of BMSCs on liver fibrosis induced by CCl4 and explore the best migration path.Methods:1. MSCs were isolated,expanded and purified by density gradient centrifugation combined with adherent culture.The growth curves of different passages MSCs were drawn by MTT.2.60 SD rats weighing about 200-250g, inject 40% CCl4 solution 0.3ml/100g,take 10% alcohol as the sole drinking water, to establish rat liver cirrhosis model.3. The model rats were randomly divided into control group, tail vein transplantion groups, partial liver transplant group, spleen and partial transplant group, with 10 rats each group.4. MSCs which received from the healthy SD rats marked with DAPI,were injected to every experimental group by 1×106 doses,control group do abdominal surgery only.5. Evaluation the survival state of the transplantation group and control group after transplanted 4 weeks later,observe the general changes in the liver,do hepatic function tests (ALB.ALT.TBIL.),do pathological examination, observe under fluorescence microscope,to understand the integration situation in the liver of BM-MSCs.Results:1. The isolation,culture and label of BMSCs. BMSCs were obtained by the combining method of density gradient centrifugation and adherent wall method, showing fibroblast-like, growing broomy. Primary culture needs 10 days. After passage and multiple-exchange fluid, suspension cells and fibroblasts cells were removed. BMSCs form a more homogeneous. Mass culture needs 3-4 days. BMSCs labeled by DAPI show blue under fluorescence microscope. Mark rate was 97%.2. The making of liver cirrhotic model. The model rats is activeless, apathetic, eating less, but the overall tolerance of rats is well. During the experiment,five rats died,who were more weighted than other individuals, probably duing to excessive injection of CCl4.The pathlogical examination results of the five randomly killed rats showed the formation of ascites, liver surface roughness, extensive formation of pseudo-leaves and re-segregation of fibers, accompanied by a clear thin regeneration of liver nodules.3. The effection of BMSCs after transplantation. Liver function significantly improved (P <0.05) in all experimental groups, compared with control group.The liver function of partial liver BMSCs transplantation group was more pronounced improved compared with other expermiment groups (P<0.05).There is no significant difference in the improvement of liver function between portal vein graft group, spleen injection transplantation group, tail vein injection transplantation group.Conclusions:1.The combining method of density gradient centrifugation and adherent wall culture access to the higher purity of BMSCs.2. DAPI is a good tracer of BMSCs.3.CCL4 combined with alcoholic can be done to shorten the time of the cirrhosis liver model and improve the success rate of making model.4. MSCs transplantation can improve liver cell synthesis,metabolism,improving the biochemical characteristics of rats blood and liver histological structure.5.Transplantation of BMSCs by the injection of local liver is the best way to improve the function of liver.