Omega-3 PUFA modulate lipogenesis, ER stress, and mitochondrial dysfunction markers in NASH - Proteomic and lipidomic insight.

BACKGROUND & AIMS: Currently there is no FDA-approved therapy for nonalcoholic steatohepatitis (NASH). Increased n-6/n-3 polyunsaturated fatty acids (PUFA) ratio can induce endoplasmic reticulum (ER) stress and mitochondrial dysfunction that characterize NASH. Our recent study with n-3 PUFA showed improvement in individual histologic parameters like steatosis, ballooning and lobular inflammation. We hypothesized that n-3 PUFA therapy mediated improvement in histologic parameters is modulated by lipidomic and proteomic changes. METHODS: We therefore evaluated hepatic proteomic and plasma lipidomic profiles before and after n-3 PUFA therapy in subjects with NASH. In a double-blind, randomized, placebo-controlled trial, patients with NASH received 6-month treatment with n-3 PUFA (0.945 g/day [64% alpha-linolenic (ALA), 21% eicosapentaenoic (EPA), and 16% docosahexaenoic (DHA) acids]). Paired liver biopsy and plasma collected before and after-n-3 PUFA therapy were assessed using mass spectrometry and gas chromatography for hepatic proteomics and plasma lipidomics. Data were matched to UniProt and LIPID MAPS database, respectively. Cytoscape software was used to analyze functional pathways. Twenty-seven NASH patients with paired liver histology and plasma before and after n-3 PUFA treatment were studied. RESULTS: Treatment with n-3 PUFA significantly increased ALA, EPA, and glycerophospholipids, and decreased arachidonic acid (p < 0.05 for all). Further, proteomic markers of cell matrix, lipid metabolism, ER stress and cellular respiratory pathways were also modulated. Interestingly, these alterations reflected functional changes highly suggestive of decreased cellular lipotoxicity potential; reduced ER proteasome degradation of proteins and induction of chaperones; and a shift in cell energy homeostasis towards mitochondrial beta-oxidation. CONCLUSION: Six-month treatment with omega-3 PUFAs significantly improved hepatic proteomic and plasma lipidomic markers of lipogenesis, endoplasmic reticulum stress and mitochondrial functions in patients with NASH.

Omega-3 polyunsaturated fatty acids in treating non-alcoholic steatohepatitis: A randomized, double-blind, placebo-controlled trial.

BACKGROUND: & aims: Few clinical trials have addressed the potential benefits of omega-3 polyunsaturated fatty acids (PUFAs) on non-alcoholic steatohepatitis (NASH). We evaluated the effects of supplementation with omega-3 PUFAs from flaxseed and fish oils in patients with biopsy-proven NASH. METHODS: Patients received three capsules daily, each containing 0.315 g of omega-3 PUFAs (64% alpha-linolenic [ALA], 16% eicosapentaenoic [EPA], and 21% docosahexaenoic [DHA] acids; n-3 group, n = 27) or mineral oil (placebo group, n = 23). Liver biopsies were evaluated histopathologically by the NASH activity score (NAS). Plasma levels of omega-3 PUFAs were assessed as a marker of intake at baseline and after 6 months of treatment. Secondary endpoints included changes in plasma biochemical markers of lipid metabolism, inflammation, and liver function at baseline and after 3 and 6 months of treatment. RESULTS: At baseline, NAS was comparable between the groups (p = 0.98). After intervention with omega-3 PUFAs, plasma ALA and EPA levels increased (p ≤ 0.05). However in the placebo group, we also observed increased EPA and DHA (p ≤ 0.05), suggesting an off-protocol intake of PUFAs. NAS improvement/stabilization was correlated with increased ALA in the n-3 group (p = 0.02) and with increased EPA (p = 0.04) and DHA (p = 0.05) in the placebo group. Triglycerides were reduced after 3 months in the n-3 group compared to baseline (p = 0.01). CONCLUSIONS: In NASH patients, the supplementation of omega-3 PUFA from flaxseed and fish oils significantly impacts on plasma lipid profile of patients with NASH. Plasma increase of these PUFAs was associated with better liver histology. (ID 01992809).

Hypocaloric high-protein diet improves clinical and biochemical markers in patients with nonalcoholic fatty liver disease (NAFLD).

OBJECTIVE: To investigate the role of hypocaloric highprotein diet, a prospective clinical study was conducted in NAFLD patients. RESEARCH METHODS AND PROCEDURES: Pre-versus post-interventional data were analyzed in 48 stable NAFLD patients (submitted to a hypocaloric high-protein diet during 75 days. Variables included anthropometrics (body mass index/ BMI and waist circumference/WC), whole-body and segmental bioimpedance analysis and biochemical tests. Diet compliance was assessed by interviews every two weeks. RESULTS: BMI, WC and body fat mass remained relatively stable (-1.3%, -1.8% and -2.5% respectively, no significance). HDL- cholesterol increased (P < 0.05) whereas total, LDL and VLDL cholesterol, triglycerides, aspartate aminotransferase/ AST, gamma glutamyltransferase/GGT, alkaline phosphatase/ AP, fasting blood glucose and glycated hemoglobin/ HbA1c decreased (P < 0.05). When patients were stratified according to increase (22/48, 45.8%) and decrease (21/48, 43.8%) of BMI, association between weight decrease and liver benefit could be elicited in such circumstances for ALT, AP and AST/ALT ratio. No change could be demonstrated in patients who gained weight. Multivariate assessment confirmed that waist circumference, ferritin, triacylglycerol, and markers of glucose homeostasis were the most relevant associated with liver enzymes. DISCUSSION: Ours results are consistent with the literature of calorie restriction in the management of NAFLD. Changes in lifestyle and weight loss are recommended for NAFLD patients. European guidelines also support this recommendation. CONCLUSION: This is the first study that demonstrated that a high protein, hypocaloric diet were associated with improvement of lipid profile, glucose homeostasis and liver enzymes in NAFLD independent on BMI decrease or body fat mass reduction.

Combination of N-acetylcysteine and metformin improves histological steatosis and fibrosis in patients with non-alcoholic steatohepatitis.

AIM: There is no proven medical therapy for the treatment of non-alcoholic steatohepatitis (NASH). Oxidative stress and insulin resistance are the mechanisms that seem to be mostly involved in its pathogenesis. The aim of our study was to evaluate the efficacy of N-acetylcysteine (NAC) in combination with metformin (MTF) in improving the aminotransferases and histological parameters (steatosis, inflammation, hepatocellular ballooning, and fibrosis) after 12 months of treatment. METHODS: Twenty consecutive patients (mean age 53 +/- 2 years [36-68] and body mass index [BMI] 29 [25-35]) with biopsy-proven NASH were enrolled in the study. NAC (1.2 g/day) and MTF (850-1000 mg/day) were given orally for 12 months. All patients underwent evaluation of serum aminotransferases, fasting lipid profile and serum glucose, anthropometric parameters, and nutritional status at 0 and 12 months. A low calorie diet was prescribed for all patients. RESULTS: Serum alanine aminotransferase, high-density lipoprotein, insulin, and glucose concentrations and thehomeostasis model assessment-insulin resistance (HOMA-IR) index were reduced significantly at the end of study (P < 0.05). The BMI declined, but without statistical significance. Aspartate aminotransferase, gamma-glutamyl transferase, alkaline phosphatase, cholesterol, and triglycerides levels were not altered with the treatment. Liver steatosis and fibrosis decreased (P < 0.05), but no improvement was noted in lobular inflammation or hepatocellular ballooning. The NASH activity score was significantly improved after treatment. CONCLUSION: Based on the biochemical and histological evidence in this pilot study, NAC in combination with MTF appears to ameliorate several aspects of NASH, including fibrosis. Further studies of this form of combination therapy are warranted to assess its potential efficacy.