DOI: 10.60816/ydzf-en15
title: Swim Training Affects on Muscle Lactate Metabolism, Nicotinamide Adenine Dinucleotides Concentration, and the Activity of NADH Shuttle Enzymes in a Mouse Model of Amyotrophic Lateral Sclerosis

description: <p><span style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif;font-size:18.6667px">In this study, we aim to verify whether swim training can improve lactate metabolism, NAD</span><span style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif">&#43;</span><span style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif;font-size:18.6667px"> and NADH levels, as well as modify the activity of glycolytic and NADH shuttle enzymes and monocarboxylate transporters (MCTs) in skeletal muscle of amyotrophic lateral sclerosis (ALS) mice. </span></p><p><span style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif;font-size:18.6667px">ALS mice (SOD1G93A) (</span><em style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif;font-size:18.6667px">n</em><span style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif;font-size:18.6667px"> &#61; 7 per group) were analyzed before the onset of ALS, at first disease symptoms (trained and untrained), and the last stage of disease (trained and untrained), and then compared with a wild-type (WT) group of mice. The blood lactate and the skeletal muscle concentration of lactate, NAD</span><span style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif">&#43;</span><span style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif;font-size:18.6667px"> and NADH, MCT1 and MCT4 protein levels, as well as lactate dehydrogenase (LDH) and malate dehydrogenase (MDH) activities in skeletal muscle were determined by fluorometric, Western blotting, liquid chromatography-MS3 spectrometry, and spectrometric methods. </span></p><p><span style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif;font-size:18.6667px">In the untrained terminal ALS group, there were decreased blood lactate levels (</span><em style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif;font-size:18.6667px">p</em><span style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif;font-size:18.6667px"> &lt; 0.001) and increased skeletal muscle lactate levels (</span><em style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif;font-size:18.6667px">p</em><span style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif;font-size:18.6667px"> &lt; 0.05) as compared with a WT group of mice. The amount of nicotinamide adenine dinucleotides in the ALS groups were also significantly reduced as well as LDH activity and the level of MCT1. Swim training increased lactate levels in the blood (</span><em style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif;font-size:18.6667px">p</em><span style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif;font-size:18.6667px"> &lt; 0.05 vs. ALS TERMINAL untrained). In addition, cytosolic MDH activity and the cMDH/LDH 2.1 ratio were significantly higher in trained vs. untrained mice (</span><em style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif;font-size:18.6667px">p</em><span style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif;font-size:18.6667px"> &lt; 0.05). </span></p><p><span style="color:rgb( 27 , 27 , 27 );font-family:&#39;cambria&#39; , &#39;cambria math&#39; , &#39;charter&#39; , &#39;times new roman&#39; , &#39;times&#39; , &#39;stixgeneral&#39; , serif;font-size:18.6667px">The data indicate significant dysfunction of lactate metabolism in ALS mice, associated with a reduction in muscle anaerobic metabolism and NADH transporting enzymes, as well as swim-induced compensation of energy demands in the ALS mice.</span></p>
description: <p><span style="font-size:12.04px">The obtained results were grouped for the following research groups: ALS mice (male) - ALS BEFORE (untrained ALS mice with no apparent disease symptoms), ALS ONSET (ALS mice with first disease symptoms), and untrained and trained ALS TERMINAL (untrained and trained ALS mice in the last stage of the disease) and wild-type (WT) B6SJL mice (male). </span><span style="font-size:0.86rem;letter-spacing:0px">The animals
in the ALS BEFORE group were euthanized at the 10th week of age. The ALS ONSET
mice were euthanized when the first disease symptoms were observed in untrained
animals (16th week of age). The ALS TERMINAL groups were euthanized when the
last stage of disease was observed in untrained animals (18th to 19th week of
life).</span></p><p><span style="font-size:12.04px">The summarized study results are derived from the following analyses:</span></p><p><span style="font-size:12.04px">a. Lactate concentration in plasma and skeletal muscle homogenates was measured fluorometrically (GloMax-Multi&#43;, Promega Corporation; Madison, WI, USA)</span></p><p><span style="font-size:12.04px">b. The total amount of nicotinamide adenine dinucleotide (NAD&#43; and NADH) levels were conducted according to an NAD&#43;/NADH Assay Kit (Abcam, ab176723, Cambridge, UK)</span></p><p><span style="font-size:12.04px">c. All enzyme activities were measured spectrophotometrically (Cecil CE9200, Cecil Instruments Limited, Cambridge, UK) in tibialis anterior muscles homogenates.</span></p><p><span style="font-size:12.04px">d. MCT1 and MCT4 protein levels, were measured using Western Blotting.Changes in protein levels were assessed using immunoreactive band densitometry. This was then normalized to the total amount of protein in the samples measured on the membranes after transfer using stain-free technology, which allows for avoiding mistakes related to housekeeping proteins. Finally, protein values ​​were expressed as A.U.</span></p><p><span style="font-size:12.04px">e. Proteomic analysis: The levels of 16 investigated proteins related to the glycolysis process (HXK1, HXK2, G6PI, K6PF, ALDOA, TPIS, GAPDH, PGK1, PGAM2, ENOA, ENOB, B0QZL1, KPYM, KPYM1, E9Q509, and LDHA) and 4 NADH shuttle enzymes (cASPAT, mASPAT, cMDH, and mMDH) were determined in ALS BEFORE, ALS TERMINAL, untrained and trained groups, using liquid chromatography-MS3 spectrometry (LC-MS/MS) at the Thermo Fisher Center for Multiplexed Proteomics (Department of Cell Biology, Harvard Medical School, Cambridge, MA, USA) Principal component analysis (PCA) enabled the linear transformation of the 15 variables into 2D or 3D space, simultaneously retaining the maximum information about the analyzed individual variables.The results were grouped as follows: TERMINAL untrained, TERMINAL trained, and BEFORE.</span></p><p><br /></p>