Effects Of CREBH On Hepatic Fibrosis In Nonalcoholic Fatty Liver Disease

Objective: The aim of this study was to gain insight into the role of cyclic AMP response element binding protein H(CREBH)in nonalcoholic fatty liver disease(NAFLD)and explore the molecular mechanisms.Methods: Healthy 8-week male CREBH knock out(KO)and wild type(WT)mice were averagely divided into a methionine and choline deficiency(MCD)or high fat(HF)diet group and a chow diet(CD)group,respectively.Hepatic pathology,liver enzymes,metabolic parameters,hydroxyproline content,expression of key m RNAs and proteins of hepatic fibrosis in the livers of mice were determined after the 4-week treatment for the MCD model and 24-week treatment for the HF model.Results: 1)Characteristics of nonalcoholic steatohepatitis(NASH)-related liver fibrosis in KO-MCD/HF group were verified by hematoxylin and eosin staining,oil red staining and sirius red staining.In WT-MCD/HF group,inflammatory infiltration and lipid droplet formation were obvious while hepatic fibrosis was not observed.Immunohistochemical staining revealed that KO-MCD/HF group had increased αSMA-positive hepatic stellate cells in the liver.2)Liver injury,as determined by plasma ALT levels,aggravated significantly in KO-MCD/HF group compared to WT-MCD/HF group.The indicators of lipid metabolism and fasting blood glucose were abnormal to various degrees in KOMCD/HF group.Furthermore,hydroxyproline analysis was conducted and indicated progressive fibrosis in KO-HF group.3)RT-q PCR results of MCP-1,αSMA,DES,COL-1,TIMP-1,TGF-β1,TGF-β2 showed significantly higher level in KO-MCD/HF group compared with that in WT-MCD/HF group.There was also a significant difference of inflammation and liver fibrosis in KO-HF group as assessed by m RNAs of TNFα,CTGF and CCND1 compared with WT-HF group.MMP-9,FGF21 m RNA level decreased in KOMCD/HF group compared to WT-MCD/HF group.4)Protein levels of MCP-1,BAX,αSMA,COL-1,TGF-β1 and SMAD2/3 in KO-MCD/HF group significantly increased compared to WT-MCD/HF group.Besides,protein level of CCND1 was also upregulated in KO-HF group compared to WT-HF group.Conclusion: CREBH knock-out may activate the TGF-β1 signaling pathway directly or indirectly and lead to more severe inflammation and NASH-related liver fibrosis.CREBH could be a potentially antifibrotic strategy in progressive NAFLD.