Metformin counters the insulin-induced suppression of fatty acid oxidation and stimulation of triacylglycerol storage in rodent skeletal muscle

The present study examined the acute effects of metformin on fatty acid (FA) metabolism in oxidative soleus (SOL) and glycolytic epitrochlearis (EPT) rodent muscle. SOL and EPT were incubated for either 30 or 180 min in the absence or presence of 2 mM metformin and with or without insulin (10 mU/ml). Metformin did not alter basal FA metabolism but countered the effects of insulin on FA oxidation and incorporation into triacylglyerol (TAG). Specifically, metformin prevented the insulin-induced suppression of FA oxidation in SOL but did not alter FA incorporation into lipid pools. In contrast, in EPT metformin blunted the incorporation of FA into TAG when insulin was present but did not alter FA oxidation. In SOL, metformin resulted in a 50% increase in AMP-activated protein kinase α2 activity and prevented the insulin-induced increase in malonyl-CoA content. In both fiber types, basal and insulin-stimulated glucose oxidation were not significantly altered by metformin. All effects were similar regardless of whether they were measured after 30 or 180 min. Because increased muscle lipid storage and impaired FA oxidation have been associated with insulin resistance in this tissue, the ability of metformin to reverse these abnormalities in muscle FA metabolism may be a part of the mechanism by which metformin improves glucose clearance and insulin sensitivity. The present data also suggest that increased glucose clearance is not due to its enhanced subsequent oxidation. Additional studies are warranted to determine whether chronic metformin treatment has similar effects on muscle FA metabolism.

Interleukin-6 and tumor necrosis factor-α are not increased in patients with type 2 diabetes: Evidence that plasma interleukin-6 is related to fat mass and not insulin responsiveness

Aims/hypothesis. Our aim was to examine the possible direct relationship of interleukin-6 and TNFα with insulin sensitivity in humans. Methods. We carried out two series of euglycaemic-hyperinsulinaemic clamp experiments. In the first (CLAMP1), skeletal muscle mRNA expression and plasma concentrations of IL-6 and TNFα were examined in patients with Type 2 diabetes (n=6), subjects matched for age (n=6), and young healthy (n=11) control subjects during a 120-min supra-physiological hyperinsulinaemic (40 mU·m -2·min-1) euglycaemic clamp. In the second series of experiments (CLAMP2), patients with Type 2 diabetes (n=6) and subjects matched for age (n=7) were studied during a 240-min high-physiological hyperinsulinaemic (7 mU·m-2·min-1) euglycaemic clamp, during which arterial and venous (femoral and subclavian) blood samples were measured for IL-6 and TNFα flux. Results. In both experiments the glucose infusion rate in the patients was markedly lower than that in the other groups. In CLAMP1, basal skeletal muscle IL-6 and TNFα mRNA were the same in all groups. They were not affected by insulin and they were not related to the glucose infusion rate. In CLAMP2, neither cytokine was released from the arm or leg during insulin stimulation in either group. In both experiments plasma concentrations of these cytokines were similar in the patients and in the control subjects, although in CLAMP1 the young healthy control group had lower (p<0.05) plasma IL-6 concentrations. Using data from all subjects, a strong positive correlation (r=0.85; p<0.00001) was observed between basal plasma IL-6 and BMI. Conversely, a negative relationship (r=-0.345; p<0.05) was found between basal plasma TNFα and BMI, although this was not significant when corrected for BMI. When corrected for BMI, no relationship was observed between either basal plasma IL-6 or TNFα and GIR. Conclusions/interpretation. These data show that the increased circulating IL-6 concentrations seen in patients with Type 2 diabetes are strongly related to fat mass and not insulin responsiveness, and suggest that neither IL-6 nor TNFα are indicative of insulin resistance.

Disassociation of muscle triglyceride content and insulin sensitivity after exercise training in patients with Type 2 diabetes

Aim/hypothesis. We determined the effect of exercise training on insulin sensitivity and muscle lipids (triglyceride [TGm] and long-chain fatty acyl CoA [LCACoA] concentration) in patients with Type 2 diabetes. Methods. Seven patients with Type 2 diabetes and six healthy control subjects who were matched for age, BMI, % body fat and VO2peak participated in a 3 days per week training program for 8 weeks. Insulin sensitivity was determined pre- and post-training during a 120 min euglycaemic- hyperinsulinaemic clamp and muscle biopsies were obtained before and after each clamp. Oxidative enzyme activities [citrate synthase (CS), β-hydroxy-acyl- CoA (β-HAD)] and TGm were determined from basal muscle samples pre- and post training, while total LCACoA content was measured in samples obtained before and after insulin-stimulation, pre- and post training. Results. The training-induced increase in VO2peak (∼20%, p<0.01) was similar in both groups. Compared with control subjects, insulin sensitivity was lower in the diabetic patients before and after training (∼60%; p<0.05), but was increased to the same extent in both groups with training (∼30%; p<0.01). TGm was increased in patients with Type 2 diabetes (170%; p<0.05) before, but was normalized to levels observed in control subjects after training. Basal LCACoA content was similar between groups and was unaltered by training. Insulin-stimulation had no detectable effect on LCACoA content. CS and β-HAD activity were increased to the same extent in both groups in response to training (p<0.001). Conclusion/interpretation. We conclude that the enhanced insulin sensitivity observed after short-term exercise training was associated with a marked decrease in TGm content in patients with Type 2 diabetes. However, despite the normalization of TGm to levels observed in healthy individuals, insulin resistance was not completely reversed in the diabetic patients.

Postexercise muscle glycogen resynthesis in obese insulin-resistant Zucker rats

We determined the effect of an acute bout of swimming (8 × 30 min) followed by either carbohydrate administration (0.5 mg/g glucose ip and ad libitum access to chow; CHO) or fasting (Fast) on postexercise glycogen resynthesis in soleus muscle and liver from female lean (ZL) and obese insulin-resistant (ZO) Zucker rats. Resting soleus muscle glycogen concentration ([glycogen]) was similar between genotypes and was reduced by 73 (ZL) and 63% (ZO) after exercise (P < 0.05). Liver [glycogen] at rest was greater in ZO than ZL (334 ± 31 vs. 247 ± 16 μmol/g wet wt; P < 0.01) and fell by 44 and 94% after exercise (P < 0.05). The fractional activity of glycogen synthase (active/total) increased immediately after exercise (from 0.22 ± 0.05 and 0.32 ± 0.04 to 0.63 ± 0.08 vs. 0.57 ± 0.05; P < 0.01 for ZL and ZO rats, respectively) and remained elevated above resting values after 30 min of recovery. During this time, muscle [glycogen] in ZO increased 68% with CHO (P < 0.05) but did not change in Fast. Muscle [glycogen] was unchanged in ZL from postexercise values after both treatments. After 6 h recovery, GLUT-4 protein concentration was increased above resting levels by a similar extent for both genotypes in both fasted (∼45%) and CHO-supplemented (∼115%) rats. Accordingly, during this time CHO refeeding resulted in supercompensation in both genotypes (68% vs. 44% for ZL and ZO). With CHO, liver [glycogen] was restored to resting levels in ZL but remained at postexercise values for ZO after both treatments. We conclude that the increased glucose availability with carbohydrate refeeding after glycogen-depleting exercise resulted in glycogen supercompensation, even in the face of muscle insulin-resistance.

Mitochondrial dysfunction has divergent, cell type-dependent effects on insulin action

The contribution of mitochondrial dysfunction to insulin resistance is a contentious issue in metabolic research. Recent evidence implicates mitochondrial dysfunction as contributing to multiple forms of insulin resistance. However, some models of mitochondrial dysfunction fail to induce insulin resistance, suggesting greater complexity describes mitochondrial regulation of insulin action. We report that mitochondrial dysfunction is not necessary for cellular models of insulin resistance. However, impairment of mitochondrial function is sufficient for insulin resistance in a cell type-dependent manner, with impaired mitochondrial function inducing insulin resistance in adipocytes, but having no effect, or insulin sensitising effects in hepatocytes. The mechanism of mitochondrial impairment was important in determining the impact on insulin action, but was independent of mitochondrial ROS production. These data can account for opposing findings on this issue and highlight the complexity of mitochondrial regulation of cell type-specific insulin action, which is not described by current reductionist paradigms.

Further investigation of the psychometric properties of the Insulin Treatment Appraisal Scale among insulin-using and non-insulin-using adults with type 2 diabetes : results from Diabetes MILES-Australia

Negative attitudes towards insulin are commonly reported by people with type 2 diabetes mellitus (T2DM) and can act as a barrier to timely insulin initiation. The Insulin Treatment Appraisal Scale (ITAS) is a widely used 20-item measure of attitudes towards insulin. While designed for completion by both insulin using and non-insulin using adults with T2DM, its psychometric properties have not been investigated separately for these groups. Furthermore, the total score is routinely reported in preference to the published two-factor structure (negative/positive appraisals). Further psychometric validation of the ITAS is required to examine its properties.

The CDP-Ethanolamine Pathway Regulates Skeletal Muscle Diacylglycerol Content and Mitochondrial Biogenesis without Altering Insulin Sensitivity.

Accumulation of diacylglycerol (DG) in muscle is thought to cause insulin resistance. DG is a precursor for phospholipids, thus phospholipid synthesis could be involved in regulating muscle DG. Little is known about the interaction between phospholipid and DG in muscle; therefore, we examined whether disrupting muscle phospholipid synthesis, specifically phosphatidylethanolamine (PtdEtn), would influence muscle DG content and insulin sensitivity. Muscle PtdEtn synthesis was disrupted by deleting CTP:phosphoethanolamine cytidylyltransferase (ECT), the rate-limiting enzyme in the CDP-ethanolamine pathway, a major route for PtdEtn production. While PtdEtn was reduced in muscle-specific ECT knockout mice, intramyocellular and membrane-associated DG was markedly increased. Importantly, however, this was not associated with insulin resistance. Unexpectedly, mitochondrial biogenesis and muscle oxidative capacity were increased in muscle-specific ECT knockout mice and were accompanied by enhanced exercise performance. These findings highlight the importance of the CDP-ethanolamine pathway in regulating muscle DG content and challenge the DG-induced insulin resistance hypothesis.

Ascorbic acid supplementation improves skeletal muscle oxidative stress and insulin sensitivity in people with type 2 diabetes: findings of a randomized controlled study

AIM/HYPOTHESIS: Skeletal muscle insulin resistance and oxidative stress are characteristic metabolic disturbances in people with type 2 diabetes. Studies in insulin resistant rodents show an improvement in skeletal muscle insulin sensitivity and oxidative stress following antioxidant supplementation. We therefore investigated the potential ameliorative effects of antioxidant ascorbic acid (AA) supplementation on skeletal muscle insulin sensitivity and oxidative stress in people with type 2 diabetes. METHODS: Participants with stable glucose control commenced a randomized cross-over study involving four months of AA (2×500mg/day) or placebo supplementation. Insulin sensitivity was assessed using a hyperinsulinaemic, euglycaemic clamp coupled with infusion of 6,6-D2 glucose. Muscle biopsies were measured for AA concentration and oxidative stress markers that included basal measures (2',7'-dichlorofluorescin [DCFH] oxidation, ratio of reduced-to-oxidized glutathione [GSH/GSSG] and F2-Isoprostanes) and insulin-stimulated measures (DCFH oxidation). Antioxidant concentrations, citrate synthase activity and protein abundances of sodium-dependent vitamin C transporter 2 (SVCT2), total Akt and phosphorylated Akt (ser473) were also measured in muscle samples. RESULTS: AA supplementation significantly increased insulin-mediated glucose disposal (delta rate of glucose disappearance; ∆Rd) (p=0.009), peripheral insulin-sensitivity index (p=0.046), skeletal muscle AA concentration (p=0.017) and muscle SVCT2 protein expression (p=0.008); but significantly decreased skeletal muscle DCFH oxidation during hyperinsulinaemia (p=0.007) when compared with placebo. Total superoxide dismutase activity was also lower following AA supplementation when compared with placebo (p=0.006). Basal oxidative stress markers, citrate synthase activity, endogenous glucose production, HbA1C and muscle Akt expression were not significantly altered by AA supplementation. CONCLUSIONS/INTERPRETATION: In summary, oral AA supplementation ameliorates skeletal muscle oxidative stress during hyperinsulinaemia and improves insulin-mediated glucose disposal in people with type 2 diabetes. Findings implicate AA supplementation as a potentially inexpensive, convenient, and effective adjunct therapy in the treatment of insulin resistance in people with type 2 diabetes.

The impact of insulin therapy and attitudes towards insulin intensification among adults with type 2 diabetes: A qualitative study

BACKGROUND: As type 2 diabetes (T2DM) is a progressive chronic condition, regular clinical review and treatment intensification are critical for prevention of long-term complications. Our aim was to explore the personal impact of insulin therapy, both positive and negative consequences, and attitudes towards future insulin intensification. METHODS: Twenty face-to-face interviews were conducted, and transcripts were analysed using thematic inductive analysis. Eligible participants were adults with T2DM, using insulin injections for <4years. Participants were mostly men (n=13, 65%), (median (range)) aged 65 (43-76) years, living with T2DM for 11.5 (2-27) years. RESULTS: Five themes emerged regarding the consequences (positive and negative) of insulin therapy, including: physical impact, personal control, emotional well-being, freedom/flexibility, (concerns about) others' reactions. Increased inconvenience and the perceived seriousness of using fast-acting insulin were both reported as barriers to future insulin intensification, despite most participants being receptive to the idea of administering additional injections. CONCLUSIONS: Positive and negative experiences of insulin therapy were reported by adults with T2DM and most were receptive to insulin intensification despite reported barriers. These findings may inform clinical interactions with people with T2DM and interventions to promote receptiveness to insulin initiation and intensification.

Elevated hypothalamic beacon gene expression in Psammomys obesus prone to develop obesity and type 2 diabetes

Objective: To investigate hypothalamic beacon gene expression at various developmental stages in genetically selected diabetes-resistant and diabetes-prone Psammomys obesus. In addition, effects of dietary energy composition on beacon gene expression were investigated in diabetes-prone P. obesus. Methods: Hypothalamic beacon gene expression was measured using TaqmanÔ fluorogenic PCR in 4-, 8- and 16-week-old animals from each genetically selected line. Results: Expression of beacon was elevated in the diabetes-prone compared with diabetes-resistant P. obesus at 4 weeks of age despite no difference in body weight between the groups. At 8 weeks of age, hypothalamic beacon gene expression was elevated in diabetes-prone animals fed a high-energy diet, and was correlated with serum insulin concentration. Conclusion: P. obesus with a genetic predisposition for the development of obesity and type 2 diabetes have elevated hypothalamic beacon gene expression at an early age. Overexpression of beacon may contribute to the development of obesity and insulin resistance in these animals.

The mitochondrial rhomboid protease PSARL is a new candidate gene for type 2 diabetes

Aims/hypothesis This study aimed to identify genes that are expressed in skeletal muscle, encode proteins with functional significance in mitochondria, and are associated with type 2 diabetes.
Methods We screened for differentially expressed genes in skeletal muscle of Psammomys obesus (Israeli sand rats), and prioritised these on the basis of genomic localisation and bioinformatics analysis for proteins with likely mitochondrial functions.
Results We identified a mitochondrial intramembrane protease, known as presenilins-associated rhomboid-like protein (PSARL) that is associated with insulin resistance and type 2 diabetes. Expression of PSARL was reduced in skeletal muscle of diabetic Psammomys obesus, and restored after exercise training to successfully treat the diabetes. PSARL gene expression in human skeletal muscle was correlated with insulin sensitivity as assessed by glucose disposal during a hyperinsulinaemic–euglycaemic clamp. In 1,031 human subjects, an amino acid substitution (Leu262Val) in PSARL was associated with increased plasma insulin concentration, a key risk factor for diabetes. Furthermore, this variant interacted strongly with age to affect insulin levels, accounting for 5% of the variation in plasma insulin in elderly subjects.
Conclusions/interpretation Variation in PSARL sequence and/or expression may be an important new risk factor for type 2 diabetes and other components of the metabolic syndrome.