Proteome analysis of liver nonparenchymal cells from rats with alcoholic liver fibrosis

Objective The development of alcoholic liver fibrosis ( ALF) is a complex process involving both parenchymal and nonparenchymal cells. The knowledge about the proteome of liver nonparenchymal cells ( NPCs) in response to ethanol is limited. This paper aims to investigate the regulatory effect of alcohol on the protein expression in liver NPCs during liver fibrosis development and to provide new clues for understanding the molecular mechanism of liver fibrosis. Methods Rats were treated with ethanol by gastric administration to establish a liver fibrosis model. The pathological changes in the liver were evaluated by James staining. Liver NPCs were enriched by Percoll density gradient centrifugation, and proteins were extracted from NPCs by two-dimensional gel electrophoresis ( 2DE) and stained by Coomassie Brilliant Blue G-250. The differentially expressed proteins were identified by liquid chromatography-tandem mass spectrometry ( LC-MS / MS) . Some of the differentially expressed proteins were verified by real time RT-PCR and Western blot. The protein spots on 2DE gels were analyzed by two sample t-test. The RT-PCR results were statistically analyzed by Mann-Whitney test. Results A rat model of ALF was established. Among the NPCs purified by Percoll density gradient centrifugation, lymphocytes, Kupffer cells, and endothelial cells were enriched for 1. 5, 3. 2, and 3. 7 times, respectively. More than 800 protein spots were detected by 2DE, and there were 26 proteins with more than 2-fold increases or decreases in expression; 21 non-redundant proteins were identified by LC-MS /MS and real-time RT-PCR analysis of 7 of these proteins showed that the mRNA levels of ANXA3, CES3, ATPA and NDUFV2 were in accordance with the proteome analysis results. Conclusion This study detected and identified a group of differentially expressed proteins related to ALF. Our work might offer some new clues to understand the mechanism of ALF.