| Objective:Hepatocellular carcinoma(HCC)is one of the most common malignant tumors,the majority of patients have a history of hepatic fibrosis and cirrhosis,cirrhosis is the late stage of hepatic fibrosis,a considerable number of cirrhosis eventually develop into liver cancer.Diethylnitrosamine(DEN)is a strong chemical carcinogens,which has an obvious affinity for the liver.The process of inducing hepatocarcinoma has gone through the hepatit,hepatic fibrosis,cirrhosis and hepatocarcinoma,which is very similar to the formation of human liver cancer.To establish the rat model of liver cancer by oral diethylnitrosamine discontinuously.Epithelial mesenchymal transition(EMT)refers to the dynamic biological process of epithelial cells transformed into mesenchymal cells,its main feature is that the expression of the epithelial marker,such as E-cadherin,is decreased,and the expression of the interstitial cell marker is over expressed.EMT plays an important role in embryonic development,inflammation,organ fibrosis and tumorigenesis.As a novel gene regulator,MicroRNA(miRNA)has been found to play an important role in tumor formation.It has been found that miRNA can regulate EMT.MiR-122 show sustained expression during liver development,abnormal expression of which may lead to a series of liver diseases.The study found that the down-regulation of miR-122 expression is associated with the occurrence of liver cancer,it may play the same role as a tumor suppressor gene.To observe the dynamic changes of miR-122 during malignant transformation of hepatocytes in rats,and to study the significance of miR-122 in the occurrence of liver cancer,by analyzing the correlation between miR-122 and TGF-?1?E-cadherin.Methods:1.Experimental animal: Clean grade male SD rats,weight 180~200g.2.Model: Fifty-five SD rats were randomly divided into the experimental group(40 rats)and the control group(15 rats).Rats of the experiment group were given 0.01% DEN solution in water for 6 weeks,followed by free drinking water for 2 weeks,and then continued drink 0.01% DEN solution in water for another 6 weeks.And the rats of control group were received free drinking water conventionally.3.Both experiment group and control group were sacrificed at the end of 4th,8th,12 th and 16 th week respectively.We got two groups of rats boby weight and liver weight,and observed the change of liver morphology and pathology.The liver mass index was calculated.4.The expression of miR-122 and TGF-?1 in each group were detected by qRT-PCR.5.The expression of TGF-?1 and E-cadherin in each group were detected by Westeren-blot.6.Statistical analysis: use SPSS17.0 to analysis.Result:1.Hepatit was showed on 4th week,hepatic fibrosis-cirrhosis was showed on 8th week,hepatocellular atypical hyperplasia was showed on 12 th week and hepatocellular carcinoma was showed on 16 th week in the experiment group.The carcinogenesis rate was 100%(15/15)at 16 weeks with an overall mortality was 17.5%(7/40)in the experiment group.There was no abnormal death in the control group.2.The weight of rats in the control group increased linearly,and the weight of the experimental group increased slowly.The weight of the experimental group was significantly lower than the control group at various periods in the liver(P<0.05).The liver mass index of the experimental group in each period was higher than the control group(P<0.05).3.qRT-PCR analysis indicated that miR-122 decline gradually in the experiment group,and there is a significant difference compared to the control group in each period(P <0.05);there is no significant difference between 4 weeks and 8 weeks in the experiment group(P=0.109>0.05).The expression of TGF-?1 mRNA increase gradually in the experimental group,and there is a significant difference compared to the control group in each period(P <0.05),except for 4 weeks(P=0.178>0.05);there is no significant difference between 4 weeks and 8 weeks in the experiment group(P=0.233>0.05).4.Western-blot analysis indicated that the expression of TGF-?1 increase gradually in the experimental group,and there is a significant difference compared to the control group in each period(P<0.05),except for 4 weeks(P =0.054>0.05);there is no significant difference between 4 weeks and 8 weeks in the experiment group(P=0.155 > 0.05).The expression of E-cadherin decline gradually in the experimental group,and there is a significant difference compared to the control group in each period(P <0.05),except for 4 weeks(P=0.310>0.05).5.Pearson correlation analysis: It show negative correlation between the miR-122 and TGF-?1 mRNA expression(r=-0.842,P<0.001);It show positive correlation between the miR-122 and E-cadherin protein expression(r=0.808,P<0.001);It show negative correlation between the TGF-?1 and E-cadherin protein expression(r=-0.820,P<0.001).Conclusion:1.Induced liver cancer model is established by oral DEN,which has the advantages of simple operation,increase the rate of induced hepatocarcinoma and decrease the mortality of rats.2.The expression of miR-122 decline gradually in the process of malignant transformation of hepatocytes in rats,and miR-122 may be closely associated with the occurrence of HCC.3.MiR-122 participate in regulating the process of EMT by TGF-?1,inhibiting its expression then inhibiting the cancer formation. |
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