Effects of 1-Methylnicotinamide (MNA) on Exercise Capacity and Endothelial Response in Diabetic Mice

RESEARCH ARTICLE Effects of 1-Methylnicotinamide (MNA) on Exercise Capacity and Endothelial Response in Diabetic Mice Kamil Przyborowski1, Marta Wojewoda1, Barbara Sitek1, Agnieszka Zakrzewska1, Agnieszka Kij1,2, Krystyna Wandzel1, Jerzy Andrzej Zoladz3, Stefan Chlopicki1,4* 1 Jagiellonian Centre for Experimental Therapeutics (JCET), Jagiellonian University, Krakow, Poland, 2 Department of Pharmacokinetics and Physical Pharmacy, Jagiellonian University Medical College, Krakow, Poland, 3 Department of Muscle Physiology, Faculty of Rehabilitation, University School of Physical Education, Krakow, Poland, 4 Department of Experimental Pharmacology, Jagiellonian University Medical College, Krakow, Poland * Abstract OPEN ACCESS Citation: Przyborowski K, Wojewoda M, Sitek B, Zakrzewska A, Kij A, Wandzel K, et al. (2015) Effects of 1-Methylnicotinamide (MNA) on Exercise Capacity and Endothelial Response in Diabetic Mice. PLoS ONE 10(6): e0130908. doi:10.1371/journal. pone.0130908 Editor: Gustavo Batista Menezes, UFMG, BRAZIL Received: February 17, 2015 Accepted: May 25, 2015 Published: June 26, 2015 Copyright: © 2015 Przyborowski et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. 1-Methylnicotinamide (MNA), which was initially considered to be a biologically inactive endogenous metabolite of nicotinamide, has emerged as an anti-thrombotic and anti-inflammatory agent with the capacity to release prostacyclin (PGI2). In the present study, we characterized the effects of MNA on exercise capacity and the endothelial response to exercise in diabetic mice. Eight-week-old db/db mice were untreated or treated with MNA for 4 weeks (100 mg·kg-1), and their exercise capacity as well as NO- and PGI2-dependent response to endurance running were subsequently assessed. MNA treatment of db/db mice resulted in four-fold and three-fold elevation of urine concentrations of MNA and its metabolites (Met2PY + Met-4PY), respectively (P<0.01), but did not affect HbA1c concentration, fasting glucose concentration or lipid profile. However, insulin sensitivity was improved (P<0.01). In MNA-treated db/db mice, the time to fatigue for endurance exercise was significantly prolonged (P<0.05). Post-exercise Δ6-keto-PGF1α (difference between mean concentration in the sedentary and exercised groups) tended to increase, and post-exercise leukocytosis was substantially reduced in MNA-treated animals. In turn, the post-exercise fall in plasma concentration of nitrate was not affected by MNA. In conclusion, we demonstrated for the first time that MNA improves endurance exercise capacity in mice with diabetes, and may also decrease the cardiovascular risk of exercise. Data Availability Statement: All relevant data are within the paper. Funding: European Union from the resources of the European Regional Development Fund under the Innovative Economy Programme (grant coordinated by JCET-UJ, No. POIG.01.01.02-00-069/09). Prof. SC received the funding. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. Introduction 1-methylnicotinamide (MNA) is major product of nicotinamide (vit B3, PP) metabolism, and is generated by nicotinamide N-methyltransferase (NNMT) and then further converted into 1-methyl-2-pyridone-5-carboxamide (Met-2-PY) and 1-methyl-4-pyridone-5-carboxamide (Met-4-PY) [1, 2]. It has been reported that MNA is a biologically active compound, and PLOS ONE | DOI:10.1371/journal.pone.0130908 June 26, 2015 1 / 15 MNA, Endothelial Response and Exercise Capacity in Diabetic Mice experimental studies in in vivo animal models have demonstrated that the anti-thrombotic [3], anti-inflammatory [4] and gastroprotective [5] effects of MNA are mediated by a prostacyclin (PGI2)-dependent mechanism. Additionally, chronic administration of MNA in animal models of hypertriglyceridemia and diabetes resulted in improvement of nitric oxide (NO)-dependent endothelial function [6]. It is well known that PGI2 production is increased during exercise [7, 8] and PGI2 release from the vascular endothelium in response to exercise appears to be an important factor regulating exercise tolerance and exercise capacity [9]. Furthermore, Zoladz et al. [9] have suggested that impairment of the exercise-induced release of PGI2 may be responsible for the increased cardiovascular risk of vigorous exercise. Since it has been reported that diabetic patients have decreased ability to release PGI2 during exercise [10], and are characterized by higher cardiovascular risk during vigorous exercise [11] pharmacological stimulation of post-exercise PGI2 production may prove beneficial. NO is also involved in the regulation of exercise capacity, and NO generated by NO synthase is metabolized in the body to inorganic anions: nitrite (NO2-) and nitrate (NO3-) [12]. On the other hand, nitrite may be reduced back to NO by enzymatic and non-enzymatic pathways, particularly in acidic environments with low oxygen availability [12], which occurs during exercise [13]. It has been reported that single bout of strenuous physical exercise had no effect on plasma nitrate concentrations in humans [14]. However, others have demonstrated a small post-exercise increase in plasma nitrate concentrations [15] or increase in plasma nitrite concentrations [16]. Furthermore, exogenous nitrate and the subsequent increase in plasma nitrite concentrations was accompanied by enhanced exercise tolerance in humans [17]. Thus, enhanced NO bioavailability appears to enhance exercise capacity in humans. We previously showed that endogenous MNA was involved in the regulation of exercise capacity, since the NNMT-MNA pathway was activated by a single bout of strenuous exercise, with an elevated post-exercise plasma concentration of MNA [18]. Considering the pharmacological profile of MNA, including PGI2 release and improvement of NO-dependent function, one could speculate that MNA supplementation could improve exercise capacity in diabetics and therefore, could be considered as a protective agent against cardiovascular risk during physical activity. Accordingly, the aim of this work was to characterize the effects of MNA supplementation on exercise capacity and endothelial-, PGI2- and NO-dependent response to exercise in diabetic db/db mice. For this purpose, db/db mice were treated with MNA in drinking water (100 mg. kg-1) for 4 weeks, their exercise capacity during an endurance running test and post-exercise MNA, nitrite, nitrate and 6-keto-PGF1α concentrations were subsequently assessed. Materials and Methods Animals Male C57BL6/Jdb/db mice (henceforth referred to as db/db mice) purchased from Charles River Laboratories were housed w (...truncated)