The Expression And Significance Of Autophagy-related Protein And P-MLC In Rats With Nonalcoholic Fatty Liver Fibrosis

Objective（1） To investigate characterization of rat with non-alcoholic fatty liver fibrosis inducedby injecting CCl₄subcutaneously in smaller dose plus feeding high fat diet（HFD） dailyat the same time.（2） To explore the expression and significance of autophagy markermicrotubule-associated proteins-light chain3（LC3） and lysosome-associatedmembrance protein LAMP-2A in this model.（3） To explore the expression and significance of P-MLC in this model.Method（1）70SD rats were randomly divided into the normal control group （group N）, CCl₄group （group M1） and HFD plus smaller dose CCl₄group （group M2）, Group N ratsand group M1rats were fed normal diet，and treated with subcutaneous injection ofarachis oil （3ml/kg of body weight） and40%CCl₄（3ml/kg of body weight） twice aweek respectively， group M2rats were fed HFD and treated with subcutaneousinjection of40%CCl₄（2ml/kg of body weight） twice a week.The rats were killed on the56th day and70th day. Liver function test、serumTG、TC were measured.Histologicalanalysis was carried out to assess the grade of hepatic steatosis and fibrosis.（2） The expression of P-MLC and LC3were examined by immunohistochemicalstaining. The content of LAMP-2A in the liver tissue was measured by ELISA.Result （1） Compared with group N, serum GLO,ALT,AST levels, grade of hepatic steatosis andfibrosis were significantly increased（P<0.05, P<0.01）, but A/G ratio was decreasedin group M1and M2both on the56th day and70th day（P<0.05, P<0.01）. SerumALT,AST levels in group M2were lower than that in group M1, but grade of hepaticsteatosis in group M2was significantly higher than that in group M1On the56th day（P<0.05）.SerumALB,A/G ratio in group M2were lower than that in group M1（P<0.05）,but serum ALT,AST levels were much higher than that in group M1on the70thday（P<0.01）.The degree of fibrosis in group M2were significantly higher than that ingroup M1both on the56th day and70th day（P<0.05）.Serum TC in group M2wassignificantly higher than that in group N and M1both on the56th day and70th day（P<0.05, P<0.01）. Serum TG in group M1and M2were both lower than that ingroup N on the70th day（P<0.01）.（2） The expression level of hepatic LC3in group M1and group M2were significantlyhigher than that in group N at the same time,but group M2was much weaker than groupM1（P<0.05, P<0.01）.Moreover,the expression level of hepatic LC3in group M1andgroup M2on the70th day was much weaker than that on the56th day（P<0.01）.（3） The content of LAMP-2A in the liver tissue homogenate in group M1and M2werehigher than that in group N（P<0.05, P<0.01）,but that in group M2was lower than thatin group M1（P<0.01） on the56th day and on the70th day. Moreover,in the samegroup,the content of LAMP-2A in the liver tissue homogenate on the70th day wasmuch weaker than that on the56th day（P<0.05, P<0.01）.（4） The expression level of hepatic P-MLC in group M1was significantly higher thanthat in group N（P<0.05, P<0.01）,but lower than that in group M2（P<0.05, P<0.01）on the56th day and on the70th day.Moreover,in the same model group,theexpression level of hepatic P-MLC on the70th day was much higher than that on the56th day（P<0.01）. Conclusion（1） Injecting CCl₄subcutaneously in smaller dose plus feeding high fat diet daily cannot only make satisfied model of rat liver fibrosis,but also can shorten the experimentaltime.（2） Hepatic LC3expression decreased along with the development of liver fibrosis.Itconcluded that hepatic autophagy decreased in fibrosis progression of nonalcholic fattyliver disease, autophagy may play a protective role in liver fibrosis.（3） The content of LAMP-2A in liver tissue was decreased along with the developmentof liver fibrosis,and was consistent with the decreased expression level of hepaticLC3,which suggested decrease of LAMP-2A may be one of cause resulting in thedecreased hepatic autophagy in nonalcholic fatty liver fibrosis.（4） P-MLC expression increased along with the development of liver fibrosis.Theincreased expression of P-MLC in liver may play an important role in fibrosisprogression of nonalcholic fatty liver disease.（5） HFD can accelerate the progression of nonalcholic fatty liver fibrosis by CCl₄,which could be related to its aggravating decrease of hepatic LC3and LAMP-2A andincerase of hepatic P-MLC in nonalcholic fatty liver fibrosis induced by CCl₄,...