We have previously identified flavin-containing monooxygenase 5 (FMO5) as a regulator of metabolic aging. The aim of the present study was to investigate the role of FMO5 in glucose homeostasis and the impact of diet and gut flora on the phenotype of mice in which the Fmo5 gene has been disrupted (Fmo5−/− mice). In comparison with wild-type (WT) counterparts, Fmo5−/− mice are resistant to age-related changes in glucose homeostasis and maintain the higher glucose tolerance and insulin sensitivity characteristic of young animals. When fed a high-fat diet, they are protected against weight gain and reduction of insulin sensitivity. The phenotype of Fmo5−/− mice is independent of diet and the gut microbiome and is determined solely by the host genotype. Fmo5−/− mice have metabolic characteristics similar to those of germ-free mice, indicating that FMO5 plays a role in sensing or responding to gut bacteria. In WT mice, FMO5 is present in the mucosal epithelium of the gastrointestinal tract where it is induced in response to a high-fat diet. In comparison with WT mice, Fmo5−/− mice have fewer colonic goblet cells, and they differ in the production of the colonic hormone resistin-like molecule β. Fmo5−/− mice have lower concentrations of tumor necrosis factor α in plasma and of complement component 3 in epididymal white adipose tissue, indicative of improved inflammatory tone. Our results implicate FMO5 as a regulator of body weight and of glucose disposal and insulin sensitivity and, thus, identify FMO5 as a potential novel therapeutic target for obesity and insulin resistance.
Intracellular Na elevation in the heart is a hallmark of pathologies where both acute and chronic metabolic remodelling occurs. Here, we assess whether acute (75 μM ouabain 100 nM blebbistatin) or chronic myocardial Na i load (PLM 3SA mouse) are causally linked to metabolic remodelling and whether the failing heart shares a common Na-mediated metabolic 'fingerprint'. Control (PLM WT), transgenic (PLM 3SA), ouabain-treated and hypertrophied Langendorff-perfused mouse hearts are studied by 23 Na, 31 P, 13 C NMR followed by 1 H-NMR metabolomic profiling. Elevated Na i leads to common adaptive metabolic alterations preceding energetic impairment: a switch from fatty acid to carbohydrate metabolism and changes in steady-state metabolite concentrations (glycolytic, anaplerotic, Krebs cycle intermediates). Inhibition of mitochondrial Na/Ca exchanger by CGP37157 ameliorates the metabolic changes. In silico modelling indicates altered metabolic fluxes (Krebs cycle, fatty acid, carbohydrate, amino acid metabolism). Prevention of Na i overload or inhibition of Na/ Ca mito may be a new approach to ameliorate metabolic dysregulation in heart failure.
Mice over-expressing the creatine transporter have elevated myocardial creatine levels [Cr] and are protected against ischaemia/reperfusion injury via improved energy reserve. However, mice with very high [Cr] develop cardiac hypertrophy and dysfunction. To investigate these contrasting effects, we applied a non-biased hypothesis-generating approach to quantify global protein and metabolite changes in the LV of mice stratified for [Cr] levels: wildtype, moderately elevated, and high [Cr] (65–85; 100–135; 160–250 nmol/mg protein, respectively). Male mice received an echocardiogram at 7 weeks of age with tissue harvested at 8 weeks. RV was used for [Cr] quantification by HPLC to select LV tissue for subsequent analysis. Two-dimensional difference in-gel electrophoresis identified differentially expressed proteins, which were manually picked and trypsin digested for nano-LC–MS/MS. Principal component analysis (PCA) showed efficient group separation (ANOVA P ≤ 0.05) and peptide sequences were identified by mouse database (UniProt 201203) using Mascot. A total of 27 unique proteins were found to be differentially expressed between normal and high [Cr], with proteins showing [Cr]-dependent differential expression, chosen for confirmation, e.g. α-crystallin B, a heat shock protein implicated in cardio-protection and myozenin-2, which could contribute to the hypertrophic phenotype. Nuclear magnetic resonance (¹H-NMR at 700 MHz) identified multiple strong correlations between [Cr] and key cardiac metabolites. For example, positive correlations with α-glucose (r² = 0.45; P = 0.002), acetyl-carnitine (r² = 0.50; P = 0.001), glutamine (r² = 0.59; P = 0.0002); and negative correlations with taurine (r² = 0.74; P < 0.0001), fumarate (r² = 0.45; P = 0.003), aspartate (r² = 0.59; P = 0.0002), alanine (r² = 0.66; P < 0.0001) and phosphocholine (r² = 0.60; P = 0.0002). These findings suggest wide-ranging and hitherto unexpected adaptations in substrate utilisation and energy metabolism with a general pattern of impaired energy generating pathways in mice with very high creatine levels.Electronic supplementary materialThe online version of this article (doi:10.1007/s00726-016-2236-x) contains supplementary material, which is available to authorized users.
BACKGROUND AND PURPOSEIn drug research using the rat Langendorff heart preparation, it is possible to study left ventricular (LV) contractility using an intraventricular balloon (IVB), and arrhythmogenesis during coronary ligation-induced regional ischaemia. Assessing both concurrently would halve animal requirements. We aimed to test the validity of this approach. EXPERIMENTAL APPROACHThe electrocardiogram (ECG) and LV function (IVB) were recorded during regional ischaemia of different extents in a randomized and blinded study. KEY RESULTSIVB-induced proarrhythmia was anticipated, but in hearts with an ischaemic zone (IZ) made deliberately small, an inflated IVB reduced ischaemia-induced ventricular fibrillation (VF) incidence as a trend. Repeating studies in hearts with large IZs revealed the effect to be significant. There were no changes in QT interval or other variables that might explain the effect. Insertion of an IVB that was minimally inflated had no effect on any variable compared with 'no IVB' controls. The antiarrhythmic effect of verapamil (a positive control drug) was unaffected by IVB inflation. Removal of an inflated (but not a non-inflated) IVB caused a release of lactate commensurate with reperfusion of an endocardial/subendocardial layer of IVB-induced ischaemia. This was confirmed by intracellular 31 phosphorus ( 31 P) nuclear magnetic resonance (NMR) spectroscopy. CONCLUSIONS AND IMPLICATIONSIVB inflation does not inhibit VF suppression by a standard drug, but it has profound antiarrhythmic effects of its own, likely to be due to inflation-induced localized ischaemia. This means rhythm and contractility cannot be assessed concurrently by this approach, with implications for drug discovery and safety assessment.
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