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AIM To clarify whether Agtr1 a methylation is involved in the development of nonalcoholic steatohepatitis(NASH)-related liver fibrosis in adult rats.METHODS A choline-deficient amino acid(CDAA) diet model was employed for methylation analysis of NASH-related liver fibrosis.Agtr1 a methylation levels were measured in the livers of CDAA- and control choline-sufficient amino acid(CSAA)-fed rats for 8 and 12 wk using quantitative methylation-specific PCR.Hepatic stellate cells(HSCs) were isolated by collagenase digestion of the liver,followed by centrifugation of the crude cell suspension through a density gradient.Agtr1 a methylation and its gene expression were also analyzed during the activation of HSCs.RESULTS The mean levels of Agtr1 a methylation in the livers of CDAA-fed rats(11.5% and 18.6% at 8 and 12 wk,respectively) tended to be higher(P = 0.06 and 0.09,respectively) than those in the livers of CSAA-fed rats(2.1% and 5.3% at 8 and 12 wk,respectively).Agtr1 a was not methylated at all in quiescent HSCs,but was clearly methylated in activated HSCs(13.8%,P < 0.01).Interestingly,although Agtr1 a was hypermethylated,the Agtr1 a m RNA level increased up to 2.2-fold(P < 0.05) in activated HSCs compared with that in quiescent HSCs,suggesting that Agtr1 a methylation did not silence its expression but instead had the potential to upregulate its expression.These findings indicate that Agtr1 a methylation and its upregulation of gene expression are associated with the development of NASH-related liver fibrosis.CONCLUSION This is the first study to show that DNA methylation is potential y involved in the regulation of a renin-angiotensin system-related gene expression during liver fibrosis.
AIM To clarify whether Agtr1 a methylation is involved in the development of nonalcoholic steatohepatitis (NASH) -latedlated liver fibrosis in adult rats. METHODS A choline-deficient amino acid (CDAA) diet model was employed for methylation analysis of NASH-related liver fibrosis. Agtr1 a methylation levels were measured in the livers of CDAA- and control choline-sufficient amino acid (CSAA) -fed rats for 8 and 12 wk using quantitative methylation-specific PCR. Hepatitis stellate cells (HSCs) were isolated by collagenase digestion of the liver, followed by centrifugation of the crude cell suspension through a density gradient. Agtr1 a methylation and its gene expression were also analyzed during the activation of HSCs.RESULTS The mean levels of Agtr1 a methylation in the livers of CDAA-fed rats (11.5% and 18.6% at 8 and 12 wk, respectively) tended to be higher (P = 0.06 and 0.09, respectively) than those in the livers of CSAA-fed rats (2.1% and 5.3% at 8 and 12 wk, respectively) a was not methylated a In all the quiescent HSCs, but was clearly methylated in activated HSCs (13.8%, P <0.01) .Interestingly, although Agtr1 a was hypermethylated, the Agtr1 am RNA level increased up to 2.2-fold with that in quiescent HSCs, suggesting that Agtr1 a methylation did not silence its expression but instead had the potential to upregulate its expression. These findings show that Agtr1 a methylation and its upregulation of gene expression are associated with the development of NASH-related liver fibrosis .CONCLUSION This is the first study to show that DNA methylation is potential y involved in the regulation of a renin-angiotensin system-related gene expression during liver fibrosis.