A novel method for determining human <i>ex vivo</i> submaximal skeletal muscle mitochondrial function

ABSTRACT: Despite numerous studies, there is no consensus about whether mitochondrial function is altered with increased age. The novelty of the present study is the determination of mitochondrial function at submaximal activity rates, which is more physiologically relevant than the ex vivo functionality protocols used previously. Muscle biopsies were taken from 64 old or young male subjects (aged 60–70 or 20–30 years). Aged subjects were recruited as trained or untrained. Muscle biopsies were used for the isolation of mitochondria and subsequent measurements of DNA repair, anti-oxidant capacity and mitochondrial protein levels (complexes I–V). Mitochondrial function was determined by simultaneous measurement of oxygen consumption, membrane potential and hydrogen peroxide emission using pyruvate + malate (PM) or succinate + rotenone (SR) as substrates. Proton leak was lower in aged subjects when determined at the same membrane potential and was unaffected by training status. State 3 respiration was lower in aged untrained subjects. This effect, however, was alleviated in aged trained subjects. H(2)O(2) emission with PM was higher in aged subjects, and was exacerbated by training, although it was not changed when using SR. However, with a higher manganese superoxide dismuthase content, the trained aged subjects may actually have lower or similar mitochondrial superoxide emission compared to the untrained subjects. We conclude that ageing and the physical activity level in aged subjects are both related to changes in the intrinsic functionality of the mitochondrion in skeletal muscle. Both of these changes could be important factors in determining the metabolic health of the aged skeletal muscle cell. KEY POINTS: The present study utilized a novel method aiming to investigate mitochondrial function in human skeletal muscle at submaximal levels and at a predefined membrane potential. . The effect of age and training status was investigated using a cross-sectional design. . Ageing was found to be related to decreased leak regardless of training status. . Increased training status was associated with increased mitochondrial hydrogen peroxide emission.

Muscle-Saturated Bioactive Lipids Are Increased with Aging and Influenced by High-Intensity Interval Training

Ceramide and diacylglycerol are linked to insulin resistance in rodents, but in humans the data are inconsistent. Insulin resistance is frequently observed with aging, but the role of ceramide and diacylglycerol is not clarified. Training improves metabolic health and, therefore, we aimed to elucidate the influence of age and high-intensity interval training (HIIT) on ceramide and diacylglycerol content in muscle. Fourteen young (33 &plusmn; 1) and 22 older (63 &plusmn; 1) overweight to obese subjects performed 6 weeks HIIT three times a week. Maximal oxygen uptake and body composition were measured and muscle biopsies and fasting blood samples were obtained. Muscle ceramide and diacylglycerol were measured by gas-liquid chromatography and proteins in insulin signaling, lipid and glucose metabolism were measured by Western blotting. Content of ceramide and diacylglycerol total, saturated, C16:0 and C18:0 fatty acids and C18:1 ceramide were higher in older compared to young. HIIT reduced saturated and C18:0 ceramides, while the content of the proteins involved in glucose (GLUT4, glycogen synthase, hexokinase II, AKT) and lipid metabolism (adipose triglyceride lipase, fatty acid binding protein) were increased after HIIT. We demonstrate a higher content of saturated ceramide and diacylglycerol fatty acids in the muscle of older subjects compared to young. Moreover, the content of saturated ceramides was reduced and muscle glucose metabolism improved at protein level after HIIT. This study highlights an increased content of saturated ceramides in aging which could be speculated to influence insulin sensitivity

Preoperative β-cell function in patients with type 2 diabetes is important for the outcome of Roux-en-Y gastric bypass surgery

The majority of the patients with type 2 diabetes (T2DM) show remission after Roux-en-Y gastric bypass (RYGB). This is the result of increased postoperative insulin sensitivity and β-cell secretion. The aim of the present study was to elucidate the importance of the preoperative β-cell function in T2DM for the chance of remission after RYGB. Fifteen patients with and 18 without T2DM had 25 g oral (OGTT) and intravenous (IVGTT) glucose tolerance tests performed at inclusion, after a diet-induced weight loss, and 4 and 18 months after RYGB. Postoperative first phase insulin secretion rate (ISR) during the IVGTT and β-cell glucose sensitivity during the OGTT increased in T2DM. Postoperative insulin sensitivity and the disposition index (DI) markedly increased in both groups. By stratifying the T2DM into two groups according to highest (T2DM(high)) and lowest (T2DM(low)) baseline DI, a restoration of first phase ISR and β-cell glucose sensitivity were seen only in T2DM(high). Remission of type 2 diabetes was 71 and 38% in T2DM(high) and T2DM(low), respectively. Postoperative postprandial GLP-1 concentrations increased markedly, but did not differ between the groups. Our findings emphasize the importance of the preoperative of β-cell function for remission of diabetes after RYGB. KEY POINTS: Roux-en-Y gastric bypass surgery leads to remission of type 2 diabetes in the majority of patients suffering from the disease. . The gut hormone glucagon-like peptide-1 is believed to be of major importance for the remission process. . The present project demonstrates a marked difference in the chance of remission of type 2 diabetes in patients with low or high preoperative β-cell function in spite of a similar post-surgery increase in postprandial glucagon-like peptide-1 release. . Furthermore, post-surgery intravenous glucose administration, which does not stimulate release of glucagon-like peptide-1, leads to increased insulin secretion in the patients with the best preoperative β-cell function. . Together the present findings indicate that patients with type 2 diabetes with high preoperative β-cell function experience a glucagon-like peptide-1-independent increase in β-cell function after gastric bypass surgery.

Hepatic mitochondrial oxidative phosphorylation is normal in obese patients with and without type 2 diabetes

KEY POINTS: Hepatic insulin resistance in patients with obesity or type 2 diabetes has been suggested to result from hepatic mitochondrial dysfunction. High‐resolution respirometry (HRR) can be used to assess oxidative phosphorylation by measuring the mitochondrial oxygen consumption rate in the individual complexes of the mitochondria. By using HRR, the present study demonstrates no difference in hepatic mitochondrial oxidative phosphorylation among subjects with obesity with or without type 2 diabetes and non‐obese controls. Furthermore, the amount of mitochondria, assessed by the citrate synthase activity, is not different between the three groups. Together the present findings indicate that hepatic mitochondrial oxidative phosphorylation capacity is not impaired in patients with obesity or type 2 diabetes. ABSTRACT: Obese patients with type 2 diabetes (T2DM) and without type 2 diabetes (OB) are characterized by high hepatic lipid content and hepatic insulin resistance. This may be linked to impaired hepatic mitochondrial oxidative phosphorylation (OXPHOS) capacity. The aim of the present study was to investigate and compare hepatic mitochondrial OXPHOS capacity in T2DM, OB and non‐obese controls (CON). Seventeen obese patients (nine OB and eight T2DM) and six CON patients had perioperative liver biopsies taken. Samples were divided into three parts to measure (1) complex I, II and IV linked respiration, (2) citrate synthase (CS) activity and (3) lipid droplet (LD) size and area (% of total tissue area filled by LDs). State 3 respiration of complex I, II and IV and the CS activity did not differ in OB, T2DM and CON. LD size was significantly higher in T2DM compared with CON, and LD area tended (P = 0.10) to be higher in T2DM and OB compared with CON. The present findings indicate that hepatic OXPHOS capacity is not different in patients with markedly different weight and glycaemic control. Furthermore, the results do not support impaired hepatic mitochondrial respiratory capacity playing a major role in the development of obesity‐induced type 2 diabetes