GSE136821
Combined Treatment with L-Carnitine and Nicotinamide Riboside Improves Hepatic Metabolism and Attenuates Obesity and Liver Steatosis
Abstract: Obesity characterized by adiposity and ectopic fat accumulation is associated with the development of non-alcoholic fatty liver disease (NAFLD). Treatments that stimulate lipid utilization may prevent the development of obesity and comorbidities. This study evaluated the potential anti-obesogenic hepatoprotective effects of combined treatment with L-carnitine and nicotinamide riboside, i.e., components that can enhance fatty acid transfer across the inner mitochondrial membrane and increase nicotinamide adenine nucleotide (NAD+) levels, which are necessary for ß-oxidation and the TCA cycle, respectively. Ldlr -/-.Leiden mice were treated with high-fat diet (HFD) supplemented with L-carnitine (LC; 0.4% w/w), nicotinamide riboside (NR; 0.3% w/w) or both (COMBI) for 21 weeks. L-carnitine plasma levels were reduced by HFD and normalized by LC. NR supplementation raised its plasma metabolite levels demonstrating effective delivery. Although food intake and ambulatory activity were comparable in all groups, COMBI treatment significantly attenuated HFD-induced body weight gain, fat mass gain (-17%) and hepatic steatosis (-22%). Also, NR and COMBI reduced hepatic 4-hydroxynonenal adducts. Upstream-regulator gene analysis demonstrated that COMBI reversed detrimental effects of HFD on liver metabolism pathways and associated regulators, e.g., ACOX, SCAP, SREBF, PPARGC1B, and INSR. Combination treatment with LC and NR exerts protective effects on metabolic pathways and constitutes a new approach to attenuate HFD-induced obesity and NAFLD. Overall design: Experimental design  The mice were supplied with food and tap water ad libitum and mice were fed standard lab chow (Ssniff R/M diet V1530, Uden, The Netherlands) until they were 12–15 weeks old. Subsequently, 74 mice were divided into five experimental groups. The healthy reference group continued to receive a chow diet (n = 13) and the high-fat diet control group (HFD) received a NAFLD inducing diet (45 kcal% fat from lard, 35 kcal% from carbohydrates, primarily sucrose and 20% kcal from protein; D12451, Research Diets, New Brunswick, NJ, USA) as described previously [17] (n = 16). L-carnitine (0.4% w/w, Sigma-Aldrich, Zwijndrecht, The Netherlands; LC group; n = 15), nicotinamide riboside (0.3% w/w, NovAlix, Cedex, France; NR group; n = 15) and the combination of both (COMBI group, n = 15) were mixed through the HFD and the respective groups were treated with these diets for 21 weeks.   Gene Expression and Pathway Analysis: Total-RNA isolation kit (Bio-connect, Huissen, The Netherlands) was used to extract RNA from snap-frozen livers. The RNA concentration was measured spectrophotometrically using a NanoDrop 1000 (Isogen Life Science, De Meern, The Netherlands). To assess the quality of the isolated RNA, the 2100 Bioanalyzer (Agilent Technologies, Amstelveen, The Netherlands) was used. Next, messenger RNA (mRNA) library was produced for next generation sequencing following the manufacturer's protocol NEBNext Ultra Directional RNA Library Prep Kit for Illumina (New England Biolabs, Ipswich, MA, USA). The quality and yield of the PCR products were consistent with the expected size distribution (300–800 base pairs). Clustering and DNA sequencing were performed by GenomeScan BV (Leiden, The Netherlands) using a NextSeq 500 sequencer (Illumina, San Diego, CA, USA). The sequences were directly aligned with the mouse reference genome (GRCm38p4) using the Start 2.5 algorithm with default settings.