| 【Objective】In this study,the traditional Chinese prescription "yiguanjian" is adopted under the guidance of theory of kidney bone marrow,liver and kidney yin moisturizing organs,in order to maintain the stability of tissue structure.Taking bone marrow MSCs’ ability of inhibiting the activation of hepatic stellate cells as the starting point,observe the changes of FGF2-Dlk1 signaling pathway and its key protein factor in the liver tissue of rats with hepatic fibrosis,and investigate the effect of FGF2-DLK1 signaling pathway and its key protein factor on the bone marrow MSCs transferred from liver tissue,combined with the fluorescence tracing of bone marrow MSCs transferred into liver tissue,to promote the key link for reversal of bone marrow MSCs from tissue,cell and gene level Liver fibrosis,thus to provide scientific experimental basis for the clinical prevention and treatment of liver fibrosis.【Method】54 male SPF grade SD rats were randomly divided into normal group,model group.CCl4-induced rat liver fibrosis model was randomly divided into yiguanjian group,bone marrow MSCs group,model group,yiguanjian and bone marrow MSCs group and positive group after modeling for three days.Each rat in bone marrow MSCs group and yiguanjian and bone marrow MSCs group were injected with 1 mol bone marrow MSCs(concentration of 5×106mmol/L);The rats in bone marrow MSCs group and yiguanjian and bone marrow MSCs group were added the samedosage of 2 mg / 200 g traditional Chinese medicine yiguanjian for gaveve,1 time / d for 4 weeks.The positive group was fed with colchicine solution for 0.25 mg / kg,1time / D for 4 weeks.Normal group and model group were treated with the same dose of saline by the same means.Anesthesia and sampling at the abdominal aorta for the blood and separation of serum to test liver function.Each group of rats received the same part of the liver lobe for the observation of pathological sections,and take the same site of the liver of the other bone marrow stem cells and yiguanjian and bone marrow MSCs group for observation of bone marrow MSCs fluorescence tracing.The remaining liver tissue was used for WB to detect col-1,DLK-1 and Notch1 protein expression;Col1,col3 and DLK1 mRNA were detected by RT-PCR.And the expression of FGF2 protein was detected by ELLSA.【Result】Rat general situation: Compared with the normal group,the spirit,water,hair color and activity,etc of the rats in the model group had varying degrees of change.Compared with the model group,the general condition of the rats in the other groups had different degrees of improvement and improvement.Rats serum liver function test: compared with normal group,the liver tissue inflammation activity,difference obvious fibrosis degree of model group had significant difference with statistically significance(P < 0.05),suggesting the success of liver fibrosis in rats model building.Compared with the model group,the degree of fibrosis of yiguanjian group and bone marrow MSCs group were decreased,and compared with the model group,the fibrosis of yiguanjian group and bone marrow MSCs group waslower than that of the model group,which was statistically significant(P <0.05).It was suggested that both yiguanjian and bone marrow MSCs could improve the degree of inflammation and the degree of fibrosis in liver tissue ofliver fibrosis rats,and the improvement effect was the most obvious when the two cooperate with each other.Detection of Yin Deficiency Index in Rats: weight: the weight growth speed of Chinese traditional medicine group,the bone marrow MSCs group,yiguanjian and bone marrow MSCs group and positive group were significantly higher than that the model group(P < 0.05);Tongue surface dry humidity: The weight of the tongue fluid secretion of rats of yiguanjian and bone marrow MSCs group always was higher compared with model group(P < 0.05);Tongue surface temperature: tongue surface temperature of model was obviously increased group compared with normal group(P< 0.05);Compared with the model group,the MSCs group was significantly lower than that of the model group,but with no statistical differences;but yiguanjian and bone marrow MSCs group was obviously reduced compared with the model group(P<0.05);The blood flow velocity of the tongue: yiguanjian group was significantly reduced compared with the positive control group and the model group(P <0.05).The blood flow velocity of the tongue of yiguanjian and bone marrow MSCs group was much more reduced compared with the model group(P <0.05);The water consumption: the model group was not significantly different from that in the normal group;The water consumption of the rats of yiguanjian and bone marrow MSCs group was significantly lower than that of the model group(P <0.05).Detection of Collagen Area in Rat Liver Tissue: The collagen area in the liver tissue of the model group was significantly different from that of the normal group(P<0.05).Suggesting that liver fibrosis rat model was established successfully.Compared with the model group,the collagen area in the liver tissue of the positive control group,yiguanjian group,the bone marrow MSCs group and yiguanjian and bone marrow MSCs group was significantly lower than that of the model group(P<0.05).It is suggested that yiguanjian can effectively reduce the collagen area in the liver tissue of rats with hepatic fibrosis,thus improving the degree of fibrosis.Detection of yiguanjian transfer bone marrow MSCs to rat liver tissue: The results of yiguanjian and bone marrow MSCs group showed that its fluorescent area was significantly higher than those in MSCs group(P <0.05),which indicated that yiguanjian could promote bone marrow MSCs transfer to the liver tissue.Expression of col1,DLK1 and Notch1 in rat liver: Compared with the normal group,the expression of col1,DLk1 and Notch1 in the liver tissue of the model group was significantly higher than that of the normal group(P <0.05).Compared with the model group,the expression of col1,DLk1 and Notch1 in bone marrow MSCs group,yiguanjian and bone marrow MSCs group were significantly decreased(P <0.05).Expression of col1,col3 and DLK1 mRNA in Rat Liver Tissue:The expression of col1,col3 and DLK1 mRNA in liver tissue of model group was significantly higher than that of normal group(P <0.05).Compared with the model group,the expressions of col1,col3 and DLK1 mRNA were signi-ficantly decreased in MSCs group,yiguanjian and bone marrow MSCs group and yiguanjian,P <0.05.Expression of FGF-2 protein in rat liver tissue: Compared with the normal group,the expression of FGF2 in the liver tissue of the model group was increased,but P>0.05.Compared with the model group,the expression of FGF2 in the liver tissue of yiguanjian and bone marrow MSCs group was significantly higher than that of the model group(P <0.05).【Conclusion】Nourishing liver and kidney prescription yiguanjian can effectively promote bone marrow MSCs transfer to liver tissue metastasis,to improve liver fibrosis model of liver function and Yin deficiency syndrome,reversing its role in the developmentprocess.The mechanism of action may be related to the expression of FGF2-DLK1 signaling pathway,which can increase the expression of FGF2 protein and decrease the expression of DLK1 gene,so as to reverse the effect of hepatic fibrosis,which has a certain positive significance for the clinical application of fouling treatment of liver fiber. |
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