Deficiency of electron transport chain in human skeletal muscle mitochondria in type 2 diabetes mellitus and obesity

Abstract

Insulin resistance in skeletal muscle in obesity and T2DM is associated with reduced muscle oxidative capacity, reduced expression in nuclear genes responsible for oxidative metabolism, and reduced activity of mitochondrial electron transport chain. The presented study was undertaken to analyze mitochondrial content and mitochondrial enzyme profile in skeletal muscle of sedentary lean individuals and to compare that with our previous data on obese or obese T2DM group. Frozen skeletal muscle biopsies obtained from lean volunteers were used to estimate cardiolipin content, mtDNA (markers of mitochondrial mass), NADH oxidase activity of mitochondrial electron transport chain (ETC), and activity of citrate synthase and beta-hydroxyacyl-CoA dehydrogenase (beta-HAD), key enzymes of TCA cycle and beta-oxidation pathway, respectively. Frozen biopsies collected from obese or T2DM individuals in our previous studies were used to estimate activity of beta-HAD. The obtained data were complemented by data from our previous studies and statistically analyzed to compare mitochondrial content and mitochondrial enzyme profile in lean, obese, or T2DM cohort. The total activity of NADH oxidase was reduced significantly in obese or T2DM subjects. The cardiolipin content for lean or obese group was similar, and although for T2DM group cardiolipin showed a tendency to decline, it was statistically insignificant. The total activity of citrate synthase for lean and T2DM group was similar; however, it was increased significantly in the obese group. Activity of beta-HAD and mtDNA content was similar for all three groups. We conclude that the total activity of NADH oxidase in biopsy for lean group is significantly higher than corresponding activity for obese or T2DM cohort. The specific activity of NADH oxidase (per mg cardiolipin) and NADH oxidase/citrate synthase and NADH oxidase/beta-HAD ratios are reduced two- to threefold in both T2DM and obesity.

Fig. 1.

The markers of mitochondrial mass and enzyme activity in skeletal muscle biopsy from type 2 diabetes mellitus (T2DM), obese, or healthy lean sedentary individuals. Data represent means ± SE. The total content of cardiolipin (A), citrate synthase enzymatic activity (B), and rotenone-sensitive NADH oxidase enzymatic activity (C) in biopsy were normalized to the creatine kinase activity. D: mitochondrial DNA (mtDNA) presented as a relative copy number of mtDNA/diploid nuclear genome. Biopsies were analyzed before (Pre-Int) and after intervention (Post-Int). P values are for mitochondrial parameters Pre-Int vs. after Post-Int. CK, creatine kinase.

Fig. 2.

The ratios between activity of mitochondrial electron transport chain and cardiolipin, β-oxidation, and TCA cycle in skeletal muscle biopsy from T2DM, obese, or healthy lean sedentary individuals Pre-Int and Post-Int. Activity of β-hydroxyacyl-CoA dehydrogenase (HAD) in skeletal muscle biopsy from T2DM, obese, or healthy lean sedentary individuals Pre-Int and Post-Int (C). Total NADH oxidase activity in biopsy was normalized to total cardiolipin (CL) content (A), citrate synthase (CS; B), or β-HAD activity (D). Data represent means ± SE. NADH oxidase/CL, NADH oxidase/CS, or NADH oxidase/HAD for T2DM vs. lean and for obese vs. lean are statistically significant before and after interventions. *P < 0.05, lean vs. obese or T2DM (baseline); #P < 0.05, lean vs. obese or T2DM (Post-Int).

Fig. 3.

The ratios between CL, TCA cycle, and activity of mitochondrial electron transport chain in skeletal muscle biopsy from obese individuals before and after completion of weight loss program only. The rotenone-sensitive NADH oxidase enzymatic activity was measured Pre-Int and Post-Int. Total NADH oxidase activity in biopsy wass normalized to total CL content or to total CS activity. Data represent means ± SE.

Fig. 4.

Potential effects of the disbalance between mitochondrial electron transport, TCA cycle, and β-oxidation on skeletal muscle metabolomics in obesity and type 2 diabetes (scheme). In the state of decreased contractile activity and excessive caloric intake, the suppressed ability of mitochondrial electron transport chain to oxidize NADH accompanied by excessive activity of CS in TCA cycle or HAD in β-oxidation pathway can lead to abnormally high steady-state concentration of intramitochondrial and cytoplasmic NADH. The increased concentration of NADH can impair substrate oxidation of 2 key enzymes that control oxidation of glucose and fatty acids, pyruvate dehydrogenase complex (PDC) and HAD, respectively. Thus the defect in mitochondrial electron transport chain can lead to general slowdown of basal metabolism in skeletal muscle, and perhaps it can result in decreased rate of ATP generation during contractile activity. Inhibition of PDC can divert more pyruvate to oxaloacetate and to citrate that can leak from mitochondria to cytoplasm. Excessive cytoplasmic citrate can be a precursor for acetyl-CoA and acyl-CoAs. Accumulation of array of metabolite byproducts and the products of incomplete oxidation of fatty acids and glucose in skeletal muscle of individuals adopting sedentary lifestyle and overeating could be a major cause of insulin resistance in skeletal muscle.