Exercise-induced increase in muscle insulin sensitivity in men is amplified when assessed using a meal test.

AIMS/HYPOTHESIS: Exercise has a profound effect on insulin sensitivity in skeletal muscle. The euglycaemic-hyperinsulinaemic clamp (EHC) is the gold standard for assessment of insulin sensitivity but it does not reflect the hyperglycaemia that occurs after eating a meal. In previous EHC investigations, it has been shown that the interstitial glucose concentration in muscle is decreased to a larger extent in previously exercised muscle than in rested muscle. This suggests that previously exercised muscle may increase its glucose uptake more than rested muscle if glucose supply is increased by hyperglycaemia. Therefore, we hypothesised that the exercise-induced increase in muscle insulin sensitivity would appear greater after eating a meal than previously observed with the EHC. METHODS: Ten recreationally active men performed dynamic one-legged knee extensor exercise for 1 h. Following this, both femoral veins and one femoral artery were cannulated. Subsequently, 4 h after exercise, a solid meal followed by two liquid meals were ingested over 1 h and glucose uptake in the two legs was measured for 3 h. Muscle biopsies from both legs were obtained before the meal test and 90 min after the meal test was initiated. Data obtained in previous studies using the EHC (n=106 participants from 13 EHC studies) were used for comparison with the meal-test data obtained in this study. RESULTS: Plasma glucose and insulin peaked 45 min after initiation of the meal test. Following the meal test, leg glucose uptake and glucose clearance increased twice as much in the exercised leg than in the rested leg; this difference is twice as big as that observed in previous investigations using EHCs. Glucose uptake in the rested leg plateaued after 15 min, alongside elevated muscle glucose 6-phosphate levels, suggestive of compromised muscle glucose metabolism. In contrast, glucose uptake in the exercised leg plateaued 45 min after initiation of the meal test and there were no signs of compromised glucose metabolism. Phosphorylation of the TBC1 domain family member 4 (TBC1D4; p-TBC1D4Ser704) and glycogen synthase activity were greater in the exercised leg compared with the rested leg. Muscle interstitial glucose concentration increased with ingestion of meals, although it was 16% lower in the exercised leg than in the rested leg. CONCLUSIONS/INTERPRETATION: Hyperglycaemia after meal ingestion results in larger differences in muscle glucose uptake between rested and exercised muscle than previously observed during EHCs. These findings indicate that the ability of exercise to increase insulin-stimulated muscle glucose uptake is even greater when evaluated with a meal test than has previously been shown with EHCs.

Pharmacological activation of PDC flux reverses lipid-induced inhibition of insulin action in muscle during recovery from exercise.

Insulin resistance is a risk factor for type 2 diabetes and exercise can improve insulin sensitivity. However, following exercise high circulating fatty acid (FA) levels might counteract this. We hypothesized that such inhibition would be reduced by forcibly increasing carbohydrate oxidation through pharmacological activation of the pyruvate dehydrogenase complex (PDC). Insulin-stimulated glucose uptake was examined with a cross-over design in healthy young men (n = 8) in a previously exercised and a rested leg during a hyperinsulinemiceuglycemic clamp five hours after one-legged exercise with: 1) infusion of saline, 2) infusion of intralipid imitating circulating FA levels during recovery from whole-body exercise, and 3) infusion of intralipid + oral PDC-activator, dichloroacetate (DCA). Intralipid infusion reduced insulin-stimulated glucose uptake by 19% in the previously exercised leg, which was not observed in the contralateral rested leg. Interestingly, this effect of intralipid in the exercised leg was abolished by DCA, which increased muscle PDC activity (130%) and flux (acetylcarnitine 130%) and decreased inhibitory phosphorylation of PDC on Ser293 (∼40%) and Ser300 (∼80%). Novel insight is provided into the regulatory interaction between glucose and lipid metabolism during exercise recovery. Coupling exercise and PDC flux activation upregulated the capacity for both glucose transport (exercise) and oxidation (DCA), which seems necessary to fully stimulate insulin-stimulated glucose uptake during recovery.

Transcellular Barriers to Glucose Delivery in the Body.

Glucose is the universal fuel of most mammalian cells, and it is largely replenished through dietary intake. Glucose availability to tissues is paramount for the maintenance of homeostatic energetics and, hence, supply should match demand by the consuming organs. In its journey through the body, glucose encounters cellular barriers for transit at the levels of the absorbing intestinal epithelial wall, the renal epithelium mediating glucose reabsorption, and the tight capillary endothelia (especially in the brain). Glucose transiting through these cellular barriers must escape degradation to ensure optimal glucose delivery to the bloodstream or tissues. The liver, which stores glycogen and generates glucose de novo, must similarly be able to release it intact to the circulation. We present the most up-to-date knowledge on glucose handling by the gut, liver, brain endothelium, and kidney, and discuss underlying molecular mechanisms and open questions. Diseases associated with defects in glucose delivery and homeostasis are also briefly addressed. We propose that the universal problem of sparing glucose from catabolism in favor of translocation across the barriers posed by epithelia and endothelia is resolved through common mechanisms involving glucose transfer to the endoplasmic reticulum, from where glucose exits the cells via unconventional cellular mechanisms.

TNIK is a conserved regulator of glucose and lipid metabolism in obesity.

Obesity and type 2 diabetes (T2D) are growing health challenges with unmet treatment needs. Traf2- and NCK-interacting protein kinase (TNIK) is a recently identified obesity- and T2D-associated gene with unknown functions. We show that TNIK governs lipid and glucose homeostasis in Drosophila and mice. Loss of the Drosophila ortholog of TNIK, misshapen, altered the metabolite profiles and impaired de novo lipogenesis in high sugar-fed larvae. Tnik knockout mice exhibited hyperlocomotor activity and were protected against diet-induced fat expansion, insulin resistance, and hepatic steatosis. The improved lipid profile of Tnik knockout mice was accompanied by enhanced skeletal muscle and adipose tissue insulin-stimulated glucose uptake and glucose and lipid handling. Using the T2D Knowledge Portal and the UK Biobank, we observed associations of TNIK variants with blood glucose, HbA1c, body mass index, body fat percentage, and feeding behavior. These results define an untapped paradigm of TNIK-controlled glucose and lipid metabolism.

GDF15 increases insulin action in the liver and adipose tissue via a ß-adrenergic receptor-mediated mechanism.

Growth differentiation factor 15 (GDF15) induces weight loss and increases insulin action in obese rodents. Whether and how GDF15 improves insulin action without weight loss is unknown. Obese rats were treated with GDF15 and displayed increased insulin tolerance 5 h later. Lean and obese female and male mice were treated with GDF15 on days 1, 3, and 5 without weight loss and displayed increased insulin sensitivity during a euglycemic hyperinsulinemic clamp on day 6 due to enhanced suppression of endogenous glucose production and increased glucose uptake in WAT and BAT. GDF15 also reduced glucagon levels during clamp independently of the GFRAL receptor. The insulin-sensitizing effect of GDF15 was completely abrogated in GFRAL KO mice and also by treatment with the ß-adrenergic antagonist propranolol and in ß1,ß2-adrenergic receptor KO mice. GDF15 activation of the GFRAL receptor increases ß-adrenergic signaling, in turn, improving insulin action in the liver and white and brown adipose tissue.

The Rho guanine dissociation inhibitor α inhibits skeletal muscle Rac1 activity and insulin action.

The molecular events governing skeletal muscle glucose uptake have pharmacological potential for managing insulin resistance in conditions such as obesity, diabetes, and cancer. With no current pharmacological treatments to target skeletal muscle insulin sensitivity, there is an unmet need to identify the molecular mechanisms that control insulin sensitivity in skeletal muscle. Here, the Rho guanine dissociation inhibitor α (RhoGDIα) is identified as a point of control in the regulation of insulin sensitivity. In skeletal muscle cells, RhoGDIα interacted with, and thereby inhibited, the Rho GTPase Rac1. In response to insulin, RhoGDIα was phosphorylated at S101 and Rac1 dissociated from RhoGDIα to facilitate skeletal muscle GLUT4 translocation. Accordingly, siRNA-mediated RhoGDIα depletion increased Rac1 activity and elevated GLUT4 translocation. Consistent with RhoGDIα's inhibitory effect, rAAV-mediated RhoGDIα overexpression in mouse muscle decreased insulin-stimulated glucose uptake and was detrimental to whole-body glucose tolerance. Aligning with RhoGDIα's negative role in insulin sensitivity, RhoGDIα protein content was elevated in skeletal muscle from insulin-resistant patients with type 2 diabetes. These data identify RhoGDIα as a clinically relevant controller of skeletal muscle insulin sensitivity and whole-body glucose homeostasis, mechanistically by modulating Rac1 activity.

Insulin resistance in patients with cancer: a systematic review and meta-analysis.

BACKGROUND: Insulin resistance is a critical cause of metabolic dysfunctions. Metabolic dysfunction is common in patients with cancer and is associated with higher cancer recurrence rates and reduced overall survival. Yet, insulin resistance is rarely considered in the clinic and thus it is uncertain how frequently this condition occurs in patients with cancer. METHODS: To address this knowledge gap, we performed a systematic review and a meta-analysis guided by the Preferred Items for Systematic Review and Meta-Analyses (PRISMA) statement. We included studies assessing insulin resistance in patients with various cancer diagnoses, using the gold-standard hyperinsulinemic-euglycemic clamp method. Studies eligible for inclusion were as follows: (1) included cancer patients older than 18 years of age; (2) included an age-matched control group consisting of individuals without cancer or other types of neoplasms; (3) measured insulin sensitivity using the hyperinsulinemic-euglycemic clamp method. We searched the databases MEDLINE, Embase, and Cochrane Central Register of Controlled Trials for articles published from database inception through March 2023 with no language restriction, supplemented by backward and forward citation searching. Bias was assessed using funnel plot. FINDINGS: Fifteen studies satisfied the criteria. The mean insulin-stimulated rate of glucose disposal (Rd) was 7.5 mg/kg/min in control subjects (n = 154), and 4.7 mg/kg/min in patients with a cancer diagnosis (n = 187). Thus, the Rd mean difference was -2.61 mg/kg/min [95% confidence interval, -3.04; -2.19], p<.01). Heterogeneity among the included studies was insignificant (p=.24). INTERPRETATION: These findings suggest that patients with a cancer diagnosis are markedly insulin resistant. As metabolic dysfunction in patients with cancer associates with increased recurrence and reduced overall survival, future studies should address if ameliorating insulin resistance in this population can improve these outcomes thereby improving patient care.Key pointsMetabolic dysfunction increases cancer recurrence rates and reduces survival for patients with cancer.Insulin resistance is a critical cause of metabolic dysfunctions.To date, no comprehensive compilation of research investigating insulin resistance in cancer patients has been produced.In this meta-analysis, we found that patients with various cancers were markedly insulin-resistant.

Insulin Sensitization Following a Single Exercise Bout Is Uncoupled to Glycogen in Human Skeletal Muscle: A Meta-analysis of 13 Single-Center Human Studies.

Exercise profoundly influences glycemic control by enhancing muscle insulin sensitivity, thus promoting glucometabolic health. While prior glycogen breakdown so far has been deemed integral for muscle insulin sensitivity to be potentiated by exercise, the mechanisms underlying this phenomenon remain enigmatic. We have combined original data from 13 of our studies that investigated insulin action in skeletal muscle either under rested conditions or following a bout of one-legged knee extensor exercise in healthy young male individuals (n = 106). Insulin-stimulated glucose uptake was potentiated and occurred substantially faster in the prior contracted muscles. In this otherwise homogenous group of individuals, a remarkable biological diversity in the glucometabolic responses to insulin is apparent both in skeletal muscle and at the whole-body level. In contrast to the prevailing concept, our analyses reveal that insulin-stimulated muscle glucose uptake and the potentiation thereof by exercise are not associated with muscle glycogen synthase activity, muscle glycogen content, or degree of glycogen utilization during the preceding exercise bout. Our data further suggest that the phenomenon of improved insulin sensitivity in prior contracted muscle is not regulated in a homeostatic feedback manner from glycogen. Instead, we put forward the idea that this phenomenon is regulated by cellular allostatic mechanisms that elevate the muscle glycogen storage set point and enhance insulin sensitivity to promote the uptake of glucose toward faster glycogen resynthesis without development of glucose overload/toxicity or feedback inhibition.

Phosphoproteomics of three exercise modalities identifies canonical signaling and C18ORF25 as an AMPK substrate regulating skeletal muscle function.

Exercise induces signaling networks to improve muscle function and confer health benefits. To identify divergent and common signaling networks during and after different exercise modalities, we performed a phosphoproteomic analysis of human skeletal muscle from a cross-over intervention of endurance, sprint, and resistance exercise. This identified 5,486 phosphosites regulated during or after at least one type of exercise modality and only 420 core phosphosites common to all exercise. One of these core phosphosites was S67 on the uncharacterized protein C18ORF25, which we validated as an AMPK substrate. Mice lacking C18ORF25 have reduced skeletal muscle fiber size, exercise capacity, and muscle contractile function, and this was associated with reduced phosphorylation of contractile and Ca2+ handling proteins. Expression of C18ORF25 S66/67D phospho-mimetic reversed the decreased muscle force production. This work defines the divergent and canonical exercise phosphoproteome across different modalities and identifies C18ORF25 as a regulator of exercise signaling and muscle function.

An exercise-inducible metabolite that suppresses feeding and obesity.

Exercise confers protection against obesity, type 2 diabetes and other cardiometabolic diseases1-5. However, the molecular and cellular mechanisms that mediate the metabolic benefits of physical activity remain unclear6. Here we show that exercise stimulates the production of N-lactoyl-phenylalanine (Lac-Phe), a blood-borne signalling metabolite that suppresses feeding and obesity. The biosynthesis of Lac-Phe from lactate and phenylalanine occurs in CNDP2+ cells, including macrophages, monocytes and other immune and epithelial cells localized to diverse organs. In diet-induced obese mice, pharmacological-mediated increases in Lac-Phe reduces food intake without affecting movement or energy expenditure. Chronic administration of Lac-Phe decreases adiposity and body weight and improves glucose homeostasis. Conversely, genetic ablation of Lac-Phe biosynthesis in mice increases food intake and obesity following exercise training. Last, large activity-inducible increases in circulating Lac-Phe are also observed in humans and racehorses, establishing this metabolite as a molecular effector associated with physical activity across multiple activity modalities and mammalian species. These data define a conserved exercise-inducible metabolite that controls food intake and influences systemic energy balance.

Clenbuterol exerts antidiabetic activity through metabolic reprogramming of skeletal muscle cells.

Activation of the sympathetic nervous system causes pronounced metabolic changes that are mediated by multiple adrenergic receptor subtypes. Systemic treatment with ß2-adrenergic receptor agonists results in multiple beneficial metabolic effects, including improved glucose homeostasis. To elucidate the underlying cellular and molecular mechanisms, we chronically treated wild-type mice and several newly developed mutant mouse strains with clenbuterol, a selective ß2-adrenergic receptor agonist. Clenbuterol administration caused pronounced improvements in glucose homeostasis and prevented the metabolic deficits in mouse models of ß-cell dysfunction and insulin resistance. Studies with skeletal muscle-specific mutant mice demonstrated that these metabolic improvements required activation of skeletal muscle ß2-adrenergic receptors and the stimulatory G protein, Gs. Unbiased transcriptomic and metabolomic analyses showed that chronic ß2-adrenergic receptor stimulation caused metabolic reprogramming of skeletal muscle characterized by enhanced glucose utilization. These findings strongly suggest that agents targeting skeletal muscle metabolism by modulating ß2-adrenergic receptor-dependent signaling pathways may prove beneficial as antidiabetic drugs.

Exercise increases phosphorylation of the putative mTORC2 activity readout NDRG1 in human skeletal muscle.

In mice, exercise is suggested to activate the mechanistic target of rapamycin complex 2 (mTORC2) in skeletal muscle, and mTORC2 is required for normal muscle glucose uptake during exercise. Whether this translates to human skeletal muscle and what signaling pathways facilitate the exercise-induced mTORC2 activation is unknown. We herein tested the hypothesis that exercise increases mTORC2 activity in human skeletal muscle and investigated if ß2-adrenergic receptor (AR) activation mediates exercise-induced mTORC2 activation. We examined several mTORC2 activity readouts (p-NDRG1 Thr346, p-Akt Ser473, p-mTOR S2481, and p-Akt Thr450) in human skeletal muscle biopsies after uphill walking or cycling exercise. In mouse muscles, we assessed mTORC2 activity readouts following acute activation of muscle ß2-adrenergic or GS signaling and during in vivo and ex vivo muscle contractions. Exercise increased phosphorylation of NDRG1 Thr346 in human soleus, gastrocnemius, and vastus lateralis muscle, without changing p-Akt Ser473, p-Akt Thr450, and p-mTOR Ser2481. In mouse muscle, stimulation of ß2-adrenergic or GS signaling and ex vivo contractions failed to increase p-NDRG1 Thr346, whereas in vivo contractions were sufficient to induce p-NDRG1 Thr346. In conclusion, the mTORC2 activity readout p-NDRG1 Thr346 is a novel exercise-responsive signaling protein in human skeletal muscle. Notably, contraction-induced p-NDRG1 Thr346 appears to require a systemic factor. Unlike exercise, and in contrast to published data obtained in cultured muscles cells, stimulation of ß2-adrenergic signaling is not sufficient to trigger NDRG1 phosphorylation in mature mouse skeletal muscle.NEW & NOTEWORTHY The mTORC2 readout p-NDRG Thr346 is a novel exercise-responsive protein in human skeletal muscle. ß2-AR and GS signaling are not sufficient to induce mTORC2 signaling in adult muscle. In vivo, but not ex vivo, contraction induced p-NDRG Thr346, which indicates requirement of a systemic factor for exercise-induced mTORC2 activation.

Personalized phosphoproteomics identifies functional signaling.

Protein phosphorylation dynamically integrates environmental and cellular information to control biological processes. Identifying functional phosphorylation amongst the thousands of phosphosites regulated by a perturbation at a global scale is a major challenge. Here we introduce 'personalized phosphoproteomics', a combination of experimental and computational analyses to link signaling with biological function by utilizing human phenotypic variance. We measure individual subject phosphoproteome responses to interventions with corresponding phenotypes measured in parallel. Applying this approach to investigate how exercise potentiates insulin signaling in human skeletal muscle, we identify both known and previously unidentified phosphosites on proteins involved in glucose metabolism. This includes a cooperative relationship between mTOR and AMPK whereby the former directly phosphorylates the latter on S377, for which we find a role in metabolic regulation. These results establish personalized phosphoproteomics as a general approach for investigating the signal transduction underlying complex biology.

GDF15 in Appetite and Exercise: Essential Player or Coincidental Bystander?

Growth differentiation factor 15 (GDF15) has recently moved to the forefront of metabolism research. When administered pharmacologically, GDF15 reduces food intake and lowers body weight via the hindbrain-situated receptor GFRAL (glial cell-derived neurotrophic factor family receptor alpha-like). Endogenous GDF15 is a ubiquitous cellular stress signal that can be produced and secreted by a variety of cell types. Circulating levels are elevated in a series of disease states, but also in response to exogenous agents such as metformin, colchicine, AICAR, and cisplatin. Recently, exercise has emerged as a relevant intervention to interrogate GDF15 physiology. Prolonged endurance exercise increases circulating GDF15 to levels otherwise associated with certain pathological states and in response to metformin treatment. The jury is still out on whether GDF15 is a functional "exerkine" mediating organ-to-brain crosstalk or whether it is a coincidental bystander. In this review, we discuss the putative physiological implication of exercise-induced GDF15, focusing on the potential impact on appetite and metabolism.

Interactions between insulin and exercise.

The interaction between insulin and exercise is an example of balancing and modifying the effects of two opposing metabolic regulatory forces under varying conditions. While insulin is secreted after food intake and is the primary hormone increasing glucose storage as glycogen and fatty acid storage as triglycerides, exercise is a condition where fuel stores need to be mobilized and oxidized. Thus, during physical activity the fuel storage effects of insulin need to be suppressed. This is done primarily by inhibiting insulin secretion during exercise as well as activating local and systemic fuel mobilizing processes. In contrast, following exercise there is a need for refilling the fuel depots mobilized during exercise, particularly the glycogen stores in muscle. This process is facilitated by an increase in insulin sensitivity of the muscles previously engaged in physical activity which directs glucose to glycogen resynthesis. In physically trained individuals, insulin sensitivity is also higher than in untrained individuals due to adaptations in the vasculature, skeletal muscle and adipose tissue. In this paper, we review the interactions between insulin and exercise during and after exercise, as well as the effects of regular exercise training on insulin action.

The many actions of insulin in skeletal muscle, the paramount tissue determining glycemia.

As the principal tissue for insulin-stimulated glucose disposal, skeletal muscle is a primary driver of whole-body glycemic control. Skeletal muscle also uniquely responds to muscle contraction or exercise with increased sensitivity to subsequent insulin stimulation. Insulin's dominating control of glucose metabolism is orchestrated by complex and highly regulated signaling cascades that elicit diverse and unique effects on skeletal muscle. We discuss the discoveries that have led to our current understanding of how insulin promotes glucose uptake in muscle. We also touch upon insulin access to muscle, and insulin signaling toward glycogen, lipid, and protein metabolism. We draw from human and rodent studies in vivo, isolated muscle preparations, and muscle cell cultures to home in on the molecular, biophysical, and structural elements mediating these responses. Finally, we offer some perspective on molecular defects that potentially underlie the failure of muscle to take up glucose efficiently during obesity and type 2 diabetes.

Pharmacological but not physiological GDF15 suppresses feeding and the motivation to exercise.

Growing evidence supports that pharmacological application of growth differentiation factor 15 (GDF15) suppresses appetite but also promotes sickness-like behaviors in rodents via GDNF family receptor α-like (GFRAL)-dependent mechanisms. Conversely, the endogenous regulation of GDF15 and its physiological effects on energy homeostasis and behavior remain elusive. Here we show, in four independent human studies that prolonged endurance exercise increases circulating GDF15 to levels otherwise only observed in pathophysiological conditions. This exercise-induced increase can be recapitulated in mice and is accompanied by increased Gdf15 expression in the liver, skeletal muscle, and heart muscle. However, whereas pharmacological GDF15 inhibits appetite and suppresses voluntary running activity via GFRAL, the physiological induction of GDF15 by exercise does not. In summary, exercise-induced circulating GDF15 correlates with the duration of endurance exercise. Yet, higher GDF15 levels after exercise are not sufficient to evoke canonical pharmacological GDF15 effects on appetite or responsible for diminishing exercise motivation.

Small Amounts of Dietary Medium-Chain Fatty Acids Protect Against Insulin Resistance During Caloric Excess in Humans.

Medium-chain fatty acids (MCFAs) have in rodents been shown to have protective effects on glucose homeostasis during high-fat overfeeding. In this study, we investigated whether dietary MCFAs protect against insulin resistance induced by a hypercaloric high-fat diet in humans. Healthy, lean men ingested a eucaloric control diet and a 3-day hypercaloric high-fat diet (increase of 75% in energy, 81-83% energy [E%] from fat) in randomized order. For one group (n = 8), the high-fat diet was enriched with saturated long-chain FAs (LCSFA-HFD), while the other group (n = 9) ingested a matched diet, but with ∼30 g (5E%) saturated MCFAs (MCSFA-HFD) in substitution for a corresponding fraction of the saturated long-chain fatty acids (LCFAs). A hyperinsulinemic-euglycemic clamp with femoral arteriovenous balance and glucose tracer was applied after the control and hypercaloric diets. In LCSFA-HFD, whole-body insulin sensitivity and peripheral insulin-stimulated glucose disposal were reduced. These impairments were prevented in MCSFA-HFD, accompanied by increased basal fatty acid oxidation, maintained glucose metabolic flexibility, increased nonoxidative glucose disposal related to lower starting glycogen content, and increased glycogen synthase activity, together with increased muscle lactate production. In conclusion, substitution of a small amount of dietary LCFAs with MCFAs rescues insulin action in conditions of lipid-induced energy excess.

Is GLUT4 translocation the answer to exercise-stimulated muscle glucose uptake?

Exercise in humans increases muscle glucose uptake up to 100-fold compared with rest. The magnitude of increase depends on exercise intensity and duration. Although knockout of glucose transporter type 4 (GLUT4) convincingly has shown that GLUT4 is necessary for exercise to increase muscle glucose uptake, studies only show an approximate twofold increase in GLUT4 translocation to the muscle cell membrane when transitioning from rest to exercise. Therefore, there is a big discrepancy between the increase in glucose uptake and GLUT4 translocation. It is suggested that either the methods for measurements of GLUT4 translocation in muscle grossly underestimate the real translocation of GLUT4 or, alternatively, GLUT4 intrinsic activity increases in muscle during exercise, perhaps due to increased muscle temperature and/or mechanical effects during contraction/relaxation cycles.