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Proteomics approach to investigate dynamic protein profile involved in high fat diet-induced fatty liver disease in rats.

ABSTRACT: Nonalcoholic fatty liver disease (NAFLD) is a disorder of the liver found worldwide. The molecular mechanisms underlying NAFLD initiation and progression, however, remain poorly understood. In this study, fluorescence difference gel electrophoresis (DIGE) combined with mass spectrometry was performed to profile the intracellular processes in the rat liver at the proteome level when rats were fed a high-fat diet for 8 weeks. Dynamic changes of 27 protein spots were observed. Among them, upregulation of 14 spots and downregulation of 13 spots were observed during the eight weeks of the high fat diet-induction period. These spots were analyzed by matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry (MALDI-TOF/TOF), and ultimately 24 proteins were identified with more than 95% confidence. Gene ontology (GO) annotation indicated that these proteins were implicated in the metabolism of carbohydrates, lipids, and amino acids. Four proteins were validated by western blot. Further functional studies on these dynamically changing proteins may lead to a better understanding of the mechanisms of high fat diet-induced fatty liver disease.

SUBMITTER: Huang B

PROVIDER: S-EPMC6831498 | biostudies-literature | 2019 Oct

REPOSITORIES: biostudies-literature

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Proteomics approach to investigate dynamic protein profile involved in high fat diet-induced fatty liver disease in rats.

Huang Baohua B Yao Yanling Y Li Yaping Y Yang Hua H Liu Huchen H Liu Heng H Li Dongming D Shu Wei W Chen Ming M

Journal of toxicologic pathology 20190319 4

Nonalcoholic fatty liver disease (NAFLD) is a disorder of the liver found worldwide. The molecular mechanisms underlying NAFLD initiation and progression, however, remain poorly understood. In this study, fluorescence difference gel electrophoresis (DIGE) combined with mass spectrometry was performed to profile the intracellular processes in the rat liver at the proteome level when rats were fed a high-fat diet for 8 weeks. Dynamic changes of 27 protein spots were observed. Among them, upregulat ...[more]

PMID: 31719749

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Project description:BackgroundThe "multiple-hit" hypothesis is currently the most widely accepted theory for non-alcoholic fatty liver disease (NAFLD) pathogenesis. The present study aimed to investigate the effects of the water extract of artichoke (WEA) on NAFLD and its underlying mechanism.MethodsRats were fed a high-fat diet (HFD) for 8 weeks to induce NAFLD and then treated with WEA at three doses (0.4, 0.8, and 1.6 g/kg body weight, BW) for 8 weeks. At the end of the intervention, serum biochemical parameters, hepatic antioxidant capacity, hepatic levels of pro-inflammatory cytokines, liver histopathology, hepatic inflammatory gene and lipid metabolism gene expression, and Akt and p-Akt (S473) protein levels were determined.ResultsThe body weight, liver weight, liver triglyceride (TG) and serum levels of TG, total cholesterol, low-density lipoprotein cholesterol, alanine aminotransferase, aspartate aminotransferase, glucose, and insulin were all significantly reduced in the WEA-treated groups (0.8 and 1.6 g/kg BW) compared with the HFD group (P < 0.01). A significant decrease in hepatic content of malondialdehyde (P < 0.01) and glutathione (P < 0.01), as well as a significant increase in liver superoxide dismutase activity (P < 0.01) were observed in WEA-treated groups (0.8 and 1.6 g/kg BW) compared to the HFD group. In addition, there was a marked decrease in the hepatic levels of pro-inflammatory cytokines (TNF-α, IL-1β, and IL-6) in the WEA-treated groups compared to the HFD group (P < 0.01). In line with these findings, the histopathology of the livers of rats treated with WEA (0.8 and 1.6 g/kg BW) showed a decrease in steatosis, ballooning, and lobular inflammation. Mechanistically, the reduced hepatic TG content might be related to the downregulation of lipogenic genes (SREBP1c, FASN, SCD1) and upregulation of lipolytic gene (PPARα), and the improved insulin signaling might be associated with the observed increase in antioxidant activity and reduction in inflammation in the WEA-treated groups.ConclusionThe hepatoprotective role of WEA in NAFLD may be attributed to its anti-steatotic, antioxidant, anti-inflammatory, and anti-insulin resistance effects.

Project description:Bioactive proanthocyanidins have been reported to have several beneficial effects on health in relation to metabolic syndrome, type 2 diabetes, and cardiovascular disease. We studied the effect of grape seed proanthocyanidin extract (GSPE) in rats fed a high fat diet (HFD). This is the first study of the effects of flavonoids on the liver proteome of rats suffering from metabolic syndrome. Three groups of rats were fed over a period of 13 weeks either a chow diet (control), an HFD, or a high fat diet supplemented for the last 10 days with GSPE (HFD + GSPE). The liver proteome was fractionated, using a Triton X-114-based two-phase separation, into soluble and membrane protein fractions so that total proteome coverage was considerably improved. The data from isobaric tag for relative and absolute quantitation (iTRAQ)-based nano-LC-MS/MS analysis revealed 90 proteins with a significant (p < 0.05) minimal expression difference of 20% due to metabolic syndrome (HFD versus control) and 75 proteins due to GSPE treatment (HFD + GSPE versus HFD). The same animals have previously been studied (Quesada, H., del Bas, J. M., Pajuelo, D., Díaz, S., Fernandez-Larrea, J., Pinent, M., Arola, L., Salvadó, M. J., and Bladé, C. (2009) Grape seed proanthocyanidins correct dyslipidemia associated with a high-fat diet in rats and repress genes controlling lipogenesis and VLDL assembling in liver. Int. J. Obes. 33, 1007-1012), and GSPE was shown to correct dyslipidemia observed in HFD-fed rats probably through the repression of hepatic lipogenesis. Our data corroborate those findings with an extensive list of proteins describing the induction of hepatic glycogenesis, glycolysis, and fatty acid and triglyceride synthesis in HFD, whereas the opposite pattern was observed to a large extent in GSPE-treated animals. GSPE was shown to have a wider effect than previously thought, and putative targets of GSPE involved in the reversal of the symptoms of metabolic syndrome were revealed. Some of these novel candidate proteins such as GFPT1, CD36, PLAA (phospholipase A(2)-activating protein), METTL7B, SLC30A1, several G signaling proteins, and the sulfide-metabolizing ETHE1 and SQRDL (sulfide-quinone reductase-like) might be considered as drug targets for the treatment of metabolic syndrome.

Project description:BackgroundAlthough metabolic associated fatty liver disease (MAFLD) is the most common chronic liver disease worldwide, the exact molecular mechanism of MAFLD progression remains unknown. In the present study, Tandem Mass Tag-labeled quantitative proteomic technology was used to elucidate the protein expression patterns of liver tissues in the progression of MAFLD, providing new potential therapeutic targets of it.MethodsFive 6-week-old male C57BL/6 mice were fed with high fat diet (HFD) for 22 weeks to establish the MAFLD mouse models. Five C57BL/6 mice of the same age were fed with normal diet (ND) and taken as controls. Mice serum were sampled for biochemical tests, and livers were isolated for histopathological examinations. Six mouse liver samples (three from each group) were performed for proteomic analysis. Differentially expressed proteins were defined using fold change of > 1.5 or < 0.67 and p value < 0.05 as thresholds. Bioinformatic analysis was used to identify the hub proteins. Real-Time Quantitative Polymerase Chain Reaction (RT-qPCR), Gene Expression Omnibus dataset, western blotting and immunohistochemistry were used to validate the expression of identified hub proteins.ResultsAfter 22 weeks on HFD diet, all mice developed MAFLD demonstrated by histopathological examination. Mouse body weights, liver weights, serum alanine transaminase and aspartate transaminase levels were significantly higher in the HFD group than ND group. Proteomics technology identified 4915 proteins in the mouse livers, among which 71 proteins were differentially expressed. Kyoto Encyclopedia of Genes and Genomes pathway analysis showed that majority of the differentially expressed proteins were involved in the peroxisome and peroxisome proliferator-activated receptor signaling pathway, as well as biosynthesis of unsaturated fatty acids. Protein-protein interaction analysis showed that these differentially expressed proteins interacted with each other and formed a complex network. Ten hub proteins were identified and validated using RT-qPCR. Five of these proteins were validated in the Gene Expression Omnibus dataset. Finally, Enoyl-CoA hydratase and 3-hydroxyacyl CoA dehydrogenase protein was validated in mouse liver tissue samples using western blotting and immunohistochemistry.ConclusionOur data showed that lipid metabolism-related pathways are closely associated with the development of MAFLD. The identified hub proteins might be novel targets for treating MAFLD.

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Proteomics approach to investigate dynamic protein profile involved in high fat diet-induced fatty liver disease in rats.

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Proteomics approach to investigate dynamic protein profile involved in high fat diet-induced fatty liver disease in rats.

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