Effect Of Knock-out Nrf2on CYP2A5Expression In Non-alcoholic Fatty Liver In Mouse

With the more possibility of the fat and energy input, the incidence rate of non-alcoholic fattyliver disease(NAFLD) has been increasing in recently years.If not to find and treat it in time, it willget exacerbated and may lead to liver tumor.“two hit” hypothesis, is believed to be the mechanismof NAFLD, but actually which is not all clear. And we are lack of the effective medicine for it.Nuclear factor erythroid2-related factor2(Nrf2) were found in vairous cells, which functionhad been explored generally by home and abraod. Nrf2played a role in protecting oxidativestress,inflammation and apoptosis inliver by inducing antioxidase andⅡ phase detoxifyingenzymes.CYP2A5which belonged to Ⅰphase metabolic enzyme, was one of liver microsomalmixed-function oxidases. It had an important role in drug biotransformation and some diseasesdevelopment. The research had showed that, when hepatic injury was caused by toxic substanceand tissue damage directly or indirectly, the expression of CYP2A5was up regulation in livercytoplasm, which neighboring the area of inflammation and hepatic cirrhosis. Nrf2deficiency willlead to down regulation of CYP2A5. There were rare report about the relationship between Nrf2and CYP2A5in the NAFLD.In order to research the relationship of the up-regulation CYP2A5and Nrf2expression inNAFLD, we established an animal model of NAFLD induced by high fatty diet with wild type ICRmice and Nrf2knock out mice, respectively, under large preliminary experiment.We exploried therelationship of CYP2A5and Nrf2in NAFLD animal model including wild type ICR mice and Nrf2knock out mice by using immunohistochemistry.The groups included WT-HFD、KO-HFD(Wildtype ICR mice and Nrf2knock out mice were induced by high fatty diet,respectively.) andWT-ND、 KO-ND(Wild type ICR mice and Nrf2knock out mice were fed by normaldiet,respectively.). The mice were killed at4,6and8week. The results of the modeling wereevaluated by blood serum biochemical indicators, liver index and liver histopathology. Comparingthe disease processes of WT-HFD and KO-HFD, we analyzed the role of Nrf2in NAFLD. Theexpression of Nrf2and CYP2A5in mice liver were tested by immunohistochemistry. Thefollowing results were obtained in this experiment:（1）After fed High-fatty-diet8weeks, the serum biochemical indicators and lipid content ofWT-HFD and KO-HFD changed apparently. Among the ALT,AST,GGT,T-Chol,LDL and GLU allaraised up,but TG content decreased.Comparing WT-ND and KO-ND, respectively, ALT,AST andT-Chol in WT-HFD and KO-HFD araised up with very significant difference. At4th,6th,8th week,ALT,AST,T-Chol and GLU were higher in KO-HFD than in WT-HFD, with significant difference. This showed that Nrf2palyed a role in NAFLD processes. At8th week, ALT and T-Chol weresignificantly higher than at4th week in KO-HFD. At8th week,T-Chol were significantly higherthan at4th and6th week in WT-HFD.（2） The histopathology observation showed that,from4to8weeks, there were apparenthepatocytes steatosis in WT-HFD and KO-HFD,comparing WT-ND and KO-ND, respectively.Hepatocytes steatosis were mainly around the central veins, with lots of bullule and large bullulemixed type steatosis and also inflammatory cell infiltrating in WT-HFD and KO-HFD.At6th and8th week, hepatocytes steatosis and inflammatory cell infiltrating were more serious in KO-HFDthan in WT-HFD.At8th week, there were more lipochondria in KO-HFD than in WT-HFD,but notsignificant difference.This suggested that Nrf2played a role in NAFLD pathogenesis. And Nrf2probably decreased the NAFLD processes or contributed to the lipid metabolism. There were morehepatocytes steatosis at8th week than at4th and6th in WT-HFD,with very significantdifference,but not significant difference between4th and6th.From4th to8th week, hepatocytessteatosis were very significant difference in KO-HFD. From4th to8th, inflammatory cellinfiltrating were significant difference in WT-HFD.At8th week, there were higher inflammatorycell infiltrating than at6th week in KO-HFD with significant difference,and very significantdifference between8th,6th with4th.（3）Comparing WT-ND mice,the expression of CYP2A5and Nrf2were very significantdifference.From4th to8th, the expression of Nrf2and CYP2A5all araised up in WT-HFD withvery significant difference and significant difference,respectively. There was a positive correlationbetween Nrf2and CYP2A5.（4）From4th to8th week, the expression of CYP2A5in WT-ND were much more inKO-ND,but not significant difference.From4th to8th week, the expression of CYP2A5inWT-HFD were much more in KO-HFD,with very significant difference.The expression of Nrf2were not detected in Nrf2knock out mice.This suggested that suppression of Nrf2resulted indecreasing the expression of CYP2A5.And Nrf2played a role in regulating CYP2A5inNAFLD.Comparing KO-ND,the expression of CYP2A5araised up in KO-HFD,but not significant.This suggested that there were other transcription factors in regulationg expression of CYP2A5besides Nrf2.Under the successful establishment of NAFLD models, we had tested the expression of Nrf2and CYP2A5in WT-HFD and KO-HFD. From the research results, we may learn the relationshipbetween Nrf2and CYP2A5in NAFLD, and the influential factors including accumulation of fattyacids, steatosis and inflammation. Thereby we investigated the role of Nrf2and CYP2A5on themechanism of NAFLD. This may explored a new way for preparing new drugs.