Liquid chromatography-mass spectrometry (LC-MS) is the method of choice for the untargeted profiling of biological samples. A multiplatform LC-MS-based approach is needed to screen polar metabolites and lipids comprehensively. Different mobile phase modifiers were tested to improve the electrospray ionization process during metabolomic and lipidomic profiling. For polar metabolites, hydrophilic interaction LC using a mobile phase with 10 mM ammonium formate/0.125% formic acid provided the best performance for amino acids, biogenic amines, sugars, nucleotides, acylcarnitines, and sugar phosphate, while reversed-phase LC (RPLC) with 0.1% formic acid outperformed for organic acids. For lipids, RPLC using a mobile phase with 10 mM ammonium formate or 10 mM ammonium formate with 0.1% formic acid permitted the high signal intensity of various lipid classes ionized in ESI(+) and robust retention times. For ESI(-), the mobile phase with 10 mM ammonium acetate with 0.1% acetic acid represented a reasonable compromise regarding the signal intensity of the detected lipids and the stability of retention times compared to 10 mM ammonium acetate alone or 0.02% acetic acid. Collectively, we show that untargeted methods should be evaluated not only on the total number of features but also based on common metabolites detected by a specific platform along with the long-term stability of retention times.

• Klíčová slova
• LC-MS, additives, lipidomics, liquid chromatography, mass spectrometry, metabolomics, mobile phase, modifiers, optimization,
• MeSH
• chromatografie kapalinová metody   MeSH
• formiáty MeSH
• hmotnostní spektrometrie s elektrosprejovou ionizací metody   MeSH
• kyselina octová MeSH
• lipidomika * MeSH
• metabolomika metody   MeSH
• tandemová hmotnostní spektrometrie * metody   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• ammonium acetate MeSH Prohlížeč
• formiáty MeSH
• formic acid MeSH Prohlížeč
• kyselina octová MeSH

Treatment with all-trans retinoic acid (ATRA), the carboxylic form of vitamin A, lowers body weight in rodents by promoting oxidative metabolism in multiple tissues including white and brown adipose tissues. We aimed to identify novel markers of the metabolic impact of ATRA through targeted blood metabolomics analyses, with a focus on acylcarnitines and amino acids. Blood was obtained from mice treated with a high ATRA dose (50 mg/kg body weight/day, subcutaneous injection) or placebo (controls) during the 4 days preceding collection. LC-MS/MS analyses with a focus on acylcarnitines and amino acids were conducted on plasma and PBMC. Main results showed that, relative to controls, ATRA-treated mice had in plasma: increased levels of carnitine, acetylcarnitine, and longer acylcarnitine species; decreased levels of citrulline, and increased global arginine bioavailability ratio for nitric oxide synthesis; increased levels of creatine, taurine and docosahexaenoic acid; and a decreased n-6/n-3 polyunsaturated fatty acids ratio. While some of these features likely reflect the stimulation of lipid mobilization and oxidation promoted by ATRA treatment systemically, other may also play a causal role underlying ATRA actions. The results connect ATRA to specific nutrition-modulated biochemical pathways, and suggest novel mechanisms of action of vitamin A-derived retinoic acid on metabolic health.

• Klíčová slova
• acylcarnitines, amino acids, retinoic acid, targeted metabolomics, vitamin A,
• MeSH
• aminokyseliny krev   MeSH
• biologické modely MeSH
• karnitin analogy a deriváty   krev   MeSH
• leukocyty mononukleární účinky léků   metabolismus   MeSH
• lipidy krev   MeSH
• metabolismus lipidů účinky léků   MeSH
• metabolom účinky léků   MeSH
• metabolomika * metody   MeSH
• myši MeSH
• oxidace-redukce účinky léků   MeSH
• stanovení celkové genové exprese MeSH
• tretinoin farmakologie   MeSH
• tuková tkáň MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• acylcarnitine MeSH Prohlížeč
• aminokyseliny MeSH
• karnitin MeSH
• lipidy MeSH
• tretinoin MeSH

Obesity is associated with insulin resistance and impaired glucose tolerance, which represent characteristic features of the metabolic syndrome. Development of obesity is also linked to changes in fatty acid and amino acid metabolism observed in animal models of obesity as well as in humans. The aim of this study was to explore whether plasma metabolome, namely the levels of various acylcarnitines and amino acids, could serve as a biomarker of propensity to obesity and impaired glucose metabolism. Taking advantage of a high phenotypic variation in diet-induced obesity in C57BL/6J mice, 12-week-old male and female mice (n = 155) were fed a high-fat diet (lipids ~32 wt%) for a period of 10 weeks, while body weight gain (BWG) and changes in insulin sensitivity (ΔHOMA-IR) were assessed. Plasma samples were collected before (week 4) and after (week 22) high-fat feeding. Both univariate and multivariate statistical analyses were then used to examine the relationships between plasma metabolome and selected phenotypes including BWG and ΔHOMA-IR. Partial least squares-discrimination analysis was able to distinguish between animals selected either for their low or high BWG (or ΔHOMA-IR) in male but not female mice. Among the metabolites that differentiated male mice with low and high BWG, and which also belonged to the major discriminating metabolites when analyzed in plasma collected before and after high-fat feeding, were amino acids Tyr and Orn, as well as acylcarnitines C16-DC and C18:1-OH. In general, the separation of groups selected for their low or high ΔHOMA-IR was less evident and the outcomes of a corresponding multivariate analysis were much weaker than in case of BWG. Thus, our results document that plasma acylcarnitines and amino acids could serve as a gender-specific complex biomarker of propensity to obesity, however with a limited predictive value in case of the associated impairment of insulin sensitivity.

• MeSH
• aminokyseliny krev   MeSH
• analýza rozptylu MeSH
• biologické markery MeSH
• dieta s vysokým obsahem tuků škodlivé účinky   MeSH
• fenotyp MeSH
• glukózový toleranční test MeSH
• inzulinová rezistence MeSH
• karnitin analogy a deriváty   krev   MeSH
• krevní glukóza MeSH
• metabolom MeSH
• metabolomika metody   MeSH
• modely nemocí na zvířatech MeSH
• myši MeSH
• obezita krev   diagnóza   etiologie   MeSH
• porucha glukózové tolerance MeSH
• prognóza MeSH
• shluková analýza MeSH
• tendenční skóre MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• acylcarnitine MeSH Prohlížeč
• aminokyseliny MeSH
• biologické markery MeSH
• karnitin MeSH
• krevní glukóza MeSH

Insulin resistance, the key defect in type 2 diabetes (T2D), is associated with a low capacity to adapt fuel oxidation to fuel availability, i.e., metabolic inflexibility. This, in turn, contributes to a further damage of insulin signaling. Effectiveness of T2D treatment depends in large part on the improvement of insulin sensitivity and metabolic adaptability of the muscle, the main site of whole-body glucose utilization. We have shown previously in mice fed an obesogenic high-fat diet that a combined use of n-3 long-chain polyunsaturated fatty acids (n-3 LC-PUFA) and thiazolidinediones (TZDs), anti-diabetic drugs, preserved metabolic health and synergistically improved muscle insulin sensitivity. We investigated here whether n-3 LC-PUFA could elicit additive beneficial effects on metabolic flexibility when combined with a TZD drug rosiglitazone. Adult male C57BL/6N mice were fed an obesogenic corn oil-based high-fat diet (cHF) for 8 weeks, or randomly assigned to various interventions: cHF with n-3 LC-PUFA concentrate replacing 15% of dietary lipids (cHF+F), cHF with 10 mg rosiglitazone/kg diet (cHF+ROSI), cHF+F+ROSI, or chow-fed. Indirect calorimetry demonstrated superior preservation of metabolic flexibility to carbohydrates in response to the combined intervention. Metabolomic and gene expression analyses in the muscle suggested distinct and complementary effects of the interventions, with n-3 LC-PUFA supporting complete oxidation of fatty acids in mitochondria and the combination with n-3 LC-PUFA and rosiglitazone augmenting insulin sensitivity by the modulation of branched-chain amino acid metabolism. These beneficial metabolic effects were associated with the activation of the switch between glycolytic and oxidative muscle fibers, especially in the cHF+F+ROSI mice. Our results further support the idea that the combined use of n-3 LC-PUFA and TZDs could improve the efficacy of the therapy of obese and diabetic patients.

• MeSH
• dieta s vysokým obsahem tuků škodlivé účinky   MeSH
• glykolýza účinky léků   MeSH
• kosterní svalová vlákna účinky léků   metabolismus   MeSH
• kosterní svaly účinky léků   metabolismus   MeSH
• metabolomika MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• obezita etiologie   metabolismus   MeSH
• omega-3 mastné kyseliny farmakologie   MeSH
• oxidace-redukce účinky léků   MeSH
• regulace genové exprese účinky léků   MeSH
• rosiglitazon MeSH
• synergismus léků MeSH
• thiazolidindiony farmakologie   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• omega-3 mastné kyseliny MeSH
• rosiglitazon MeSH
• thiazolidindiony MeSH

Combining pharmacological treatments and life style interventions is necessary for effective therapy of major diseases associated with obesity, which are clustered in the metabolic syndrome. Acting via multiple mechanisms, combination treatments may reduce dose requirements and, therefore, lower the risk of adverse side effects, which are usually associated with long-term pharmacological interventions. Our previous study in mice fed high-fat diet indicated additivity in preservation of insulin sensitivity and in amelioration of major metabolic syndrome phenotypes by the combination treatment using n-3 long-chain polyunsaturated fatty acids (n-3 LC-PUFA) and rosiglitazone, i.e. an anti-diabetic drug of the thiazolidinedione (TZD) family. We investigated here whether pioglitazone, a TZD-drug in clinical use, could elicit the additive beneficial effects when combined with n-3 LC-PUFA. Adult male mice (C57BL/6N) were fed an obesogenic corn oil-based high-fat diet (cHF) for 8 weeks, or randomly assigned to various dietary treatments (i) cHF+F, cHF with n-3 LC-PUFA concentrate replacing 15% of dietary lipids; (ii) cHF+ROSI, cHF with 10 mg rosiglitazone/kg diet; (iii) cHF+F+ROSI; (iv) cHF+PIO, cHF with 50 mg pioglitazone/kg diet; and (v) cHF+F+PIO, or chow-fed. Plasma concentrations of 163 metabolites were evaluated using a targeted metabolomics approach. Both TZDs preserved glucose homeostasis and normal plasma lipid levels while inducing adiponectin, with pioglitazone showing better effectiveness. The beneficial effects of TZDs were further augmented by the combination treatments. cHF+F+ROSI but not cHF+F+PIO counteracted development of obesity, in correlation with inducibility of fatty acid β-oxidation, as revealed by the metabolomic analysis. By contrast, only cHF+F+PIO eliminated hepatic steatosis and this treatment also reversed insulin resistance in dietary obese mice. Our results reveal differential effects of rosiglitazone and pioglitazone, unmasked in the combination treatment with n-3 LC-PUFA, and support the notion that n-3 LC-PUFA could be used as add-on treatment to TZDs in order to improve diabetic patient's therapy.

• MeSH
• dietní tuky aplikace a dávkování   MeSH
• homeostáza MeSH
• hypoglykemika aplikace a dávkování   MeSH
• krevní glukóza analýza   MeSH
• lipidy krev   MeSH
• metabolomika MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• obezita prevence a kontrola   MeSH
• omega-3 mastné kyseliny aplikace a dávkování   MeSH
• pioglitazon MeSH
• rosiglitazon MeSH
• thiazolidindiony aplikace a dávkování   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• dietní tuky MeSH
• hypoglykemika MeSH
• krevní glukóza MeSH
• lipidy MeSH
• omega-3 mastné kyseliny MeSH
• pioglitazon MeSH
• rosiglitazon MeSH
• thiazolidindiony MeSH