Patellar tendon biomechanical and morphologic properties and their relationship to serum clinical variables in persons with prediabetes and type 2 diabetes.

Tendon biomechanical properties and fibril organization are altered in patients with diabetes compared to healthy individuals, yet few biomarkers have been associated with in vivo tendon properties. We investigated the relationships between in vivo imaging-based tendon properties, serum variables, and patient characteristics across healthy controls (n = 14, age: 45 ± 5 years, body mass index [BMI]: 24 ± 1, hemoglobin A1c [HbA1c]: 5.3 ± 0.1%), prediabetes (n = 14, age: 54 ± 5 years, BMI: 29 ± 2; HbA1c: 5.7 ± 0.1), and type 2 diabetes (n = 13, age: 55 ± 3 years, BMI: 33 ± 2, HbA1c: 6.7 ± 0.3). We used ultrasound speckle-tracking and measurements from magnetic resonance imaging (MRI) to estimate the patellar tendon in vivo tangent modulus. Analysis of plasma c-peptide, interleukin-1ß (IL-1ß), IL-6, IL-8, tumor necrosis factor-α (TNF-α), adiponectin, leptin, insulin-like growth factor 1 (IGF-1), and C-reactive protein (CRP) was completed. We built regression models incorporating statistically significant covariates and indicators for the clinically defined groups. We found that tendon cross-sectional area normalized to body weight (BWN CSA) and modulus were lower in patients with type 2 diabetes than in healthy controls (p < 0.05). Our regression analysis revealed that a model that included BMI, leptin, high-density lipoprotein (HDL), low-density lipoprotein (LDL), age, and group explained ~70% of the variability in BWN CSA (R2 = 0.70, p < 0.001). For modulus, including the main effects LDL, groups, HbA1c, age, BMI, cholesterol, IGF-1, c-peptide, leptin, and IL-6, accounted for ~54% of the variability in modulus (R2 = 0.54, p < 0.05). While BWN CSA and modulus were lower in those with diabetes, group was a poor predicter of tendon properties when considering the selected covariates. These data highlight the multifactorial nature of tendon changes with diabetes and suggest that blood variables could be reliable predictors of tendon properties.

Soluble RAGE and skeletal muscle tissue RAGE expression profiles in lean and obese young adults across differential aerobic exercise intensities.

Nearly 40% of Americans have obesity and are at increased risk for developing type 2 diabetes. Skeletal muscle is responsible for >80% of insulin-stimulated glucose uptake that is attenuated by the inflammatory milieu of obesity and augmented by aerobic exercise. The receptor for advanced glycation endproducts (RAGE) is an inflammatory receptor directly linking metabolic dysfunction with inflammation. Circulating soluble isoforms of RAGE (sRAGE) formed either by proteolytic cleavage (cRAGE) or alternative splicing (esRAGE) act as decoys for RAGE ligands, thereby counteracting RAGE-mediated inflammation. We aimed to determine if RAGE expression or alternative splicing of RAGE is altered by obesity in muscle, and whether acute aerobic exercise (AE) modifies RAGE and sRAGE. Young (20-34 yr) participants without [n = 17; body mass index (BMI): 22.6 ± 2.6 kg/m2] and with obesity (n = 7; BMI: 32.8 ± 2.9 kg/m2) performed acute aerobic exercise (AE) at 40%, 65%, or 80% of maximal aerobic capacity (V̇o2max; mL/kg/min) on separate visits. Blood was taken before and 30 min after each AE bout. Muscle biopsy samples were taken before, 30 min, and 3 h after the 80% V̇o2max AE bout. Individuals with obesity had higher total RAGE and esRAGE mRNA and RAGE protein (P < 0.0001). In addition, RAGE and esRAGE transcripts correlated to transcripts of the NF-κB subunit P65 (P < 0.05). There was no effect of AE on total RAGE or esRAGE transcripts, or RAGE protein (P > 0.05), and AE tended to decrease circulating sRAGE in particular at lower intensities of exercise. RAGE expression is exacerbated in skeletal muscle with obesity, which may contribute to muscle inflammation via NF-κB. Future work should investigate the consequences of increased skeletal muscle RAGE on the development of obesity-related metabolic dysfunction and potential mitigating strategies.NEW & NOTEWORTHY This study is the first to investigate the effects of aerobic exercise intensity on circulating sRAGE isoforms, muscle RAGE protein, and muscle RAGE splicing. sRAGE isoforms tended to diminish with exercise, although this effect was attenuated with increasing exercise intensity. Muscle RAGE protein and gene expression were unaffected by exercise. However, individuals with obesity displayed nearly twofold higher muscle RAGE protein and gene expression, which positively correlated with expression of the P65 subunit of NF-κB.

Neutralization of excessive CCL28 improves wound healing in diabetic mice.

Introduction: Chronic, non-healing skin wounds such as diabetic foot ulcers (DFUs) are common in patients with type 2 diabetes mellitus (T2DM) and often result in limb amputation and even death. However, mechanisms by which T2DM and inflammation negatively impact skin wound healing remains poorly understood. Here we investigate a mechanism by which an excessive level of chemokine CCL28, through its receptor CCR10, impairs wound healing in patients and mice with T2DM. Methods & Results: Firstly, a higher level of CCL28 was observed in skin and plasma in both patients with T2DM, and in obesity-induced type 2 diabetic db/db mice. Compared with WT mice, adipose tissue from db/db mice released 50% more CCL28, as well as 2- to 3-fold more IL-1ß, IL-6, and TNF-α, and less VEGF, as determined by ELISA measurements. Secondly, overexpression of CCL28 with adenovirus (Adv-CCL28) caused elevation of proinflammatory cytokines as well as CCR10 expression and also reduced eNOS expression in the dorsal skin of WT mice as compared with control Adv. Thirdly, topical application of neutralizing anti-CCL28 Ab dose-dependently accelerated wound closure and eNOS expression, and decreased IL-6 level, with an optimal dose of 1 µg/wound. In addition, mRNA levels of eNOS and anti-inflammatory cytokine IL-4 were increased as shown by real-time RT-PCR. The interaction between eNOS and CCR10 was significantly reduced in diabetic mouse wounds following application of the optimal dose of anti-CCL28 Ab, and eNOS expression increased. Finally, enhanced VEGF production and increased subdermal vessel density as indicated by CD31 immunostaining were also observed with anti-CCL28 Ab. Discussion: Taken together, topical application of neutralizing anti-CCL28 Ab improved dorsal skin wound healing by reducing CCR10 activation and inflammation in part by preventing eNOS downregulation, increasing VEGF production, and restoring angiogenesis. These results indicate anti-CCL28 Ab has significant potential as a therapeutic strategy for treatment of chronic non-healing diabetic skin wounds such as DFUs.

Advanced Glycation End Products and Inflammatory Cytokine Profiles in Maintenance Hemodialysis Patients After the Ingestion of a Protein-Dense Meal.

OBJECTIVE: The goal of this investigation was to evaluate circulating and skeletal muscle inflammatory biomarkers between maintenance hemodialysis (MHD) and demographic-matched control subjects (CON) before and after ingestion of a protein-rich meal. DESIGN AND METHODS: CON (n = 8; 50 ± 2 years; 31 ± 1 kg/m2) and MHD patients (n = 8; 56 ± 5 years; 32 ± 2 kg/m2) underwent a basal blood draw and muscle biopsy and serial blood draws after the ingestion of a mixed meal on a nondialysis day. Plasma advanced glycation end products (AGEs) and markers of oxidation were assessed via liquid chromatography-tandem mass spectrometry before and after the meal (+240 min). Circulating inflammatory cytokines and soluble receptors for AGE (sRAGE) isoforms (endogenous secretory RAGEs and cleaved RAGEs) were determined before and after the meal (+240 min). Basal muscle was probed for inflammatory cytokines and protein expression of related signaling components (RAGE, Toll-like receptor 4, oligosaccharyltransferase subunit 48, TIR-domain-containing adapter-inducing interferon-ß, total IκBα, and pIκBα). RESULTS: Basal circulating AGEs were 7- to 343-fold higher (P < .001) in MHD than those in CON, but only MG-H1 increased in CON after the meal (P < .001). There was a group effect (MHD > CON) for total sRAGEs (P = .02) and endogenous secretory RAGEs (P < .001) and a trend for cleaved RAGEs (P=.09), with no meal effect. In addition, there was a group effect (MHD < CON; P < .05) for circulating fractalkine, interleukin (IL)10, IL17A, and IL1ß and a trend (P < .10) for IL6 and macrophage inflammatory protein 1 alpha, whereas tumor necrosis factor alpha was higher in MHD (P < .001). In muscle, Toll-like receptor 4 (P = .03), TIR-domain-containing adapter-inducing interferon-ß (P = .002), and oligosaccharyltransferase subunit 48 (P = .02) expression was lower in MHD than that in CON, whereas IL6 was higher (P = .01) and IL8 (P = .08) tended to be higher in MHD. CONCLUSION: Overall, MHD exhibited an exaggerated, circulating, and skeletal muscle inflammatory biomarker environment, and the meal did not appreciably affect the inflammatory status.

Inhibition of CCL28/CCR10-Mediated eNOS Downregulation Improves Skin Wound Healing in the Obesity-Induced Mouse Model of Type 2 Diabetes.

Chronic, nonhealing skin wounds, such as diabetic foot ulcers (DFUs), are common in patients with type 2 diabetes. Here, we investigated the role of chemokine (C-C motif) ligand 28 (CCL28) and its receptor C-C chemokine receptor type 10 (CCR10) in downregulation of endothelial nitric (NO) oxide synthase (eNOS) in association with delayed skin wound healing in the db/db mouse model of type 2 diabetes. We observed reduced eNOS expression and elevated CCL28/CCR10 levels in dorsal skin of db/db mice and subdermal leg biopsy specimens from human subjects with type 2 diabetes. Further interrogation revealed that overexpression of CCR10 reduced eNOS expression, NO bioavailability, and tube formation of human dermal microvascular endothelial cells (HDMVECs) in vitro, which was recapitulated in mouse dorsal skin. In addition, incubation of HDMVECs with CCL28 led to internalization of the CCR10/eNOS complex and colocalization with lysosome-associated membrane protein 1. Finally, topical application of myristoylated CCR10 binding domain 7 amino acid (Myr-CBD7) peptide prevented CCR10-eNOS interaction and subsequent eNOS downregulation, enhanced eNOS/NO levels, eNOS/VEGF-R2+ microvessel density, and blood perfusion, reduced inflammatory cytokine levels, and importantly, decreased wound healing time in db/db mice. Thus, endothelial cell CCR10 activation in genetically obese mice with type 2 diabetes promotes eNOS depletion and endothelial dysfunction, and targeted disruption of CCR10/eNOS interaction improves wound healing.

Oral vitamin C restores endothelial function during acute inflammation in young and older adults.

Oxidative stress has been linked to reductions in vascular function during acute inflammation in young adults; however, the effect of acute inflammation on vascular function with aging is inconclusive. The aim of this study was to determine if oral antioxidant administration eliminates vascular dysfunction during acute inflammation in young and older adults. Brachial flow-mediated dilation (FMD) and carotid-femoral pulse wave velocity (PWV) were measured in nine young (3 male, 24 ± 4 yrs, 26.2 ± 4.9 kg/m2 ) and 16 older (13 male, 64 ± 5 yrs, 25.8 ± 3.2 kg/m2 ) adults before and 2-h after oral consumption of 2 g of vitamin C. The vitamin C protocol was completed at rest and 24 h after acute inflammation was induced via the typhoid vaccine. Venous blood samples were taken to measure markers of inflammation and vitamin C. Both interleukin-6 (Δ+0.7 ± 1.8 pg/ml) and C-reactive protein (Δ+1.9 ± 3.1 mg/L) were increased at 24 h following the vaccine (p < 0.01). There was no change in FMD or PWV following vitamin C administration at rest (p > 0.05). FMD was lower in all groups during acute inflammation (Δ-1.4 ± 1.9%, p < 0.01), with no changes in PWV (Δ-0.0 ± 0.9 m/s, p > 0.05). Vitamin C restored FMD back to initial values in young and older adults during acute inflammation (Δ+1.0 ± 1.8%, p < 0.01) with no change in inflammatory markers or PWV (p > 0.05). In conclusion, oral vitamin C restored endothelial function during acute inflammation in young and older adults, with no effect on aortic stiffness. The effect of vitamin C on endothelial function did not appear to be due to reductions in inflammatory markers. The exact mechanisms should be further investigated.

Depletion of Caveolin-1 in Type 2 Diabetes Model Induces Alzheimer's Disease Pathology Precursors.

Type 2 diabetes mellitus (T2DM) is a risk factor for the development of late-onset Alzheimer's disease (AD). However, the mechanism underlying the development of late-onset AD is largely unknown. Here we show that levels of the endothelial-enriched protein caveolin-1 (Cav-1) are reduced in the brains of T2DM patients compared with healthy aging, and inversely correlated with levels of ß-amyloid (Aß). Depletion of Cav-1 is recapitulated in the brains of db/db (Leprdb ) diabetic mice and corresponds with recognition memory deficits as well as the upregulation of amyloid precursor protein (APP), BACE-1, a trending increase in ß-amyloid Aß42/40 ratio and hyperphosphorylated tau (p-tau) species. Importantly, we show that restoration of Cav-1 levels in the brains of male db/db mice using adenovirus overexpressing Cav-1 (AAV-Cav-1) rescues learning and memory deficits and reduces pathology (i.e., APP, BACE-1 and p-tau levels). Knocking down Cav-1 using shRNA in HEK cells expressing the familial AD-linked APPswe mutant variant upregulates APP, APP carboxyl terminal fragments, and Aß levels. In turn, rescue of Cav-1 levels restores APP metabolism. Together, these results suggest that Cav-1 regulates APP metabolism, and that depletion of Cav-1 in T2DM promotes the amyloidogenic processing of APP and hyperphosphorylation of tau. This may suggest that depletion of Cav-1 in T2DM underlies, at least in part, the development of AD and imply that restoration of Cav-1 may be a therapeutic target for diabetic-associated sporadic AD.SIGNIFICANCE STATEMENT More than 95% of the Alzheimer's patients have the sporadic late-onset form (LOAD). The cause for late-onset Alzheimer's disease is unknown. Patients with Type 2 diabetes mellitus have considerably higher incidence of cognitive decline and AD compared with the general population, suggesting a common mechanism. Here we show that the expression of caveolin-1 (Cav-1) is reduced in the brain in Type 2 diabetes mellitus. In turn, reduced Cav-1 levels induce AD-associated neuropathology and learning and memory deficits. Restoration of Cav-1 levels rescues these deficits. This study unravels signals underlying LOAD and suggests that restoration of Cav-1 may be an effective therapeutic target.

Experimental Hyperglycemia Alters Circulating Concentrations and Renal Clearance of Oxidative and Advanced Glycation End Products in Healthy Obese Humans.

The purpose of this investigation was to evaluate the effects of experimental hyperglycemia on oxidative damage (OX), advanced glycation end products (AGEs), and the receptor for AGEs (RAGE) through an in vivo approach. Obese subjects (n = 10; 31.2 ± 1.2 kg·m-2; 56 ± 3 years) underwent 24 h of hyperglycemic clamp (+5.4 mM above basal), where plasma at basal and after 2 h and 24 h of hyperglycemic challenge were assayed for OX (methionine sulfoxide, MetSO, and aminoadipic acid, AAA) and AGE-free adducts (Ne-carboxymethyllysine, CML; Ne-carboxyethyllysine, CEL; glyoxal hydroimidazolone-1, GH-1; methylglyoxal hydroimidazolone-1, MG-H1; and 3-deoxyglucosone hydroimidazolone, 3DG-H) via liquid chromatography⁻tandem mass spectrometry (LC⁻MS/MS). Urine was also analyzed at basal and after 24 h for OX and AGE-free adducts and plasma soluble RAGE (sRAGE) isoforms (endogenous secretory RAGE, esRAGE, and cleaved RAGE, cRAGE), and inflammatory markers were determined via enzyme-linked immunosorbent assay (ELISA). Skeletal muscle tissue collected via biopsy was probed at basal, 2 h, and 24 h for RAGE and OST48 protein expression. Plasma MetSO, AAA, CEL, MG-H1, and G-H1 decreased (-18% to -47%; p < 0.05), while CML increased (72% at 24 h; p < 0.05) and 3DG-H remained unchanged (p > 0.05) with the hyperglycemic challenge. Renal clearance of MetSO, AAA, and G-H1 increased (599% to 1077%; p < 0.05), CML decreased (-30%; p < 0.05), and 3DG-H, CEL, and MG-H1 remained unchanged (p > 0.05). Fractional excretion of MetSO, AAA, CEL, G-H1, and MG-H1 increased (5.8% to 532%; p < 0.05) and CML and 3DG-H remained unchanged (p > 0.05). Muscle RAGE and OST48 expression, plasma sRAGE, IL-1ß, IL-1Ra, and TNFα remained unchanged (p > 0.05), while IL-6 increased (159% vs. basal; p > 0.05). These findings suggest that individuals who are obese but otherwise healthy have the capacity to prevent accumulation of OX and AGEs during metabolic stress by increasing fractional excretion and renal clearance.

Reciprocal regulation of eNOS and caveolin-1 functions in endothelial cells.

We hypothesized that the maintenance of vascular homeostasis is critically dependent on the expression and reciprocal regulation of caveolin-1 (Cav-1) and endothelial nitric oxide synthase (eNOS) in endothelial cells (ECs). Skeletal muscle biopsies from subjects with type 2 diabetes showed 50% less Cav-1 and eNOS than those from lean healthy controls. The Cav-1:eNOS expression ratio was 200:1 in primary culture human ECs. Cav-1 small interfering RNA (siRNA) reduced eNOS protein and gene expression in association with a twofold increase in eNOS phosphorylation and nitrate production per molecule of eNOS, which was reversed in cells overexpressing Adv-Cav-1-GFP. Upon addition of the Ca2+ ionophore A23187 to activate eNOS, we observed eNOS Ser1177 phosphorylation, its translocation to ß-catenin-positive cell-cell junctions, and increased colocalization of eNOS and Cav-1 within 5 min. We also observed Cav-1 S-nitrosylation and destabilization of Cav-1 oligomers in cells treated with A23187 as well as insulin or albumin, and this could be blocked by L-NAME, PP2, or eNOS siRNA. Finally, caveola-mediated endocytosis of albumin or insulin was reduced by Cav-1 or eNOS siRNA, and the effect of Cav-1 siRNA was rescued by Adv-Cav-1-GFP. Thus, Cav-1 stabilizes eNOS expression and regulates its activity, whereas eNOS-derived NO promotes caveola-mediated endocytosis.

Nox2 contributes to hyperinsulinemia-induced redox imbalance and impaired vascular function.

Insulin resistance promotes vascular endothelial dysfunction and subsequent development of cardiovascular disease. Previously we found that skeletal muscle arteriolar flow-induced dilation (FID) was reduced following a hyperinsulinemic clamp in healthy adults. Therefore, we hypothesized that hyperinsulinemia, a hallmark of insulin resistance, contributes to microvascular endothelial cell dysfunction via inducing oxidative stress that is mediated by NADPH oxidase (Nox) system. We examined the effect of insulin, at levels that are comparable with human hyperinsulinemia on 1) FID of isolated arterioles from human skeletal muscle tissue in the presence and absence of Nox inhibitors and 2) human adipose microvascular endothelial cell (HAMECs) expression of nitric oxide (NO), endothelial NO synthase (eNOS), and Nox-mediated oxidative stress. In six lean healthy participants (mean age 25.5±1.6 y, BMI 21.8±0.9), reactive oxygen species (ROS) were increased while NO and arteriolar FID were reduced following 60min of ex vivo insulin incubation. These changes were reversed after co-incubation with the Nox isoform 2 (Nox2) inhibitor, VAS2870. In HAMECs, insulin-induced time-dependent increases in Nox2 expression and P47phox phosphorylation were echoed by elevations of superoxide production. In contrast, phosphorylation of eNOS and expression of superoxide dismutase (SOD2 and SOD3) isoforms showed a biphasic response with an increased expression at earlier time points followed by a steep reduction phase. Insulin induced eNOS uncoupling that was synchronized with a drop of NO and a surge of ROS production. These effects were reversed by Tempol (SOD mimetic), Tetrahydrobiopterin (BH4; eNOS cofactor), and VAS2870. Finally, insulin induced nitrotyrosine formation which was reversed by inhibiting NO or superoxide generation. In conclusions, hyperinsulinemia may reduce FID via inducing Nox2-mediated superoxide production in microvascular endothelial cells which reduce the availability of NO and enhances peroxynitrite formation. Therefore, the Nox2 pathway should be considered as a target for the prevention of oxidative stress-associated endothelial dysfunction during hyperinsulinemia.

Short-term exercise reduces markers of hepatocyte apoptosis in nonalcoholic fatty liver disease.

Increased hepatocyte apoptosis is a hallmark of nonalcoholic fatty liver disease (NAFLD) and contributes to the profibrogenic state responsible for the progression to nonalcoholic steatohepatitis (NASH). Strategies aimed at reducing apoptosis may result in better outcomes for individuals with NAFLD. We therefore examined the effect of a short-term exercise program on markers of apoptosis-plasma cytokeratin 18 (CK18) fragments, alanine aminotransferase (ALT), aspartate aminotransferase (AST), soluble Fas (sFas), and sFas ligand (sFasL)-in 13 obese individuals with NAFLD [body mass index 35.2 ± 1.2 kg/m(2), >5% intrahepatic lipid (IHL) assessed by (1)H-MR spectroscopy]. Exercise consisted of treadmill walking for 60 min/day on 7 consecutive days at ∼85% of maximal heart rate. Additionally, subjects underwent an oral glucose tolerance test and a maximal oxygen consumption (Vo(2max)) test before and after the exercise intervention. The Matsuda index was used to assess insulin sensitivity. We observed significant decreases in CK18 fragments (558.4 ± 106.8 vs. 323.4 ± 72.5 U/l, P < 0.01) and ALT (30.2 ± 5.1 vs. 24.3 ± 4.8 U/l, P < 0.05), and an increase in whole body fat oxidation (49.3 ± 6.1 vs. 69.4 ± 7.1 mg/min, P < 0.05), while decreases in circulating sFasL approached statistical significance (66.5 ± 6.0 vs. 63.0 ± 5.7 pg/ml, P = 0.06), as did the relationship between percent change in circulating CK18 fragments and ALT (r = 0.55, P = 0.05). We also observed a significant correlation between changes in fat oxidation and circulating sFasL (rho = -0.65, P < 0.05). There was no change in IHL following the intervention (18.2 ± 2.5 vs. 17.5 ± 2.1%, NS). We conclude that short-term exercise reduces a circulatory marker of hepatocyte apoptosis in obese individuals with NAFLD and propose that changes in the proapoptotic environment may be mediated through improved insulin sensitivity and increased oxidative capacity.

A low-glycemic index diet and exercise intervention reduces TNF(alpha) in isolated mononuclear cells of older, obese adults.

Low-glycemic index diets and exercise independently improve glucose tolerance and reduce diabetes risk. However, the combined effect of a low-glycemic index diet and exercise on inflammation and glucose metabolism is not known. Therefore, we randomized 28 insulin-resistant adults (age: 66 ± 1 y; BMI: 34.2 ± 0.7 kg · m(-2)) to a 12-wk, low (LGI = 40) or high- (HGI = 80) glycemic index diet plus aerobic exercise (5 d · wk(-1), 60 min · d(-1), 80-85% heart rate(max)) intervention. All food and fluids were provided during the study. Inflammation was assessed from cytokine (TNFα and IL-6) secretion using peripheral blood mononuclear cells (MNC) stimulated overnight with LPS. Glycemic response was determined following ingestion of a 75-g glucose solution. Fasting blood samples were collected for additional cytokine [TNFα, IL-6, and monocyte chemoattractant protein 1 (MCP-1)] analysis. Both interventions decreased BMI (P < 0.001), fasting plasma glucose (P = 0.01), and insulin (P = 0.02). The glycemic response was reduced only in the LGI group (P = 0.04). Plasma and MNC-derived TNFα secretion were reduced in the LGI group (P = 0.02) but increased in the HGI group (P = 0.02). Secretion of IL-6 from MNC and plasma IL-6 and MCP-1 concentrations were reduced in the LGI group. The change in MNC-derived TNFα (r = 0.43; P = 0.04) and plasma MCP-1 (r = 0.44; P = 0.04) correlated with decreases in the glycemic response. These data highlight the importance of diet composition in the treatment and prevention of inflammation and hyperglycemia. A low-glycemic index diet has antiinflammatory and antidiabetogenic effects when combined with exercise in older, obese prediabetics.

Influence of acetaminophen and ibuprofen on skeletal muscle adaptations to resistance exercise in older adults.

Evidence suggests that consumption of over-the-counter cyclooxygenase (COX) inhibitors may interfere with the positive effects that resistance exercise training has on reversing sarcopenia in older adults. This study examined the influence of acetaminophen or ibuprofen consumption on muscle mass and strength during 12 wk of knee extensor progressive resistance exercise training in older adults. Thirty-six individuals were randomly assigned to one of three groups and consumed the COX-inhibiting drugs in double-blind placebo-controlled fashion: placebo (67 ± 2 yr; n = 12), acetaminophen (64 ± 1 yr; n = 11; 4 g/day), and ibuprofen (64 ± 1 yr; n = 13; 1.2 g/day). Compliance with the resistance training program (100%) and drug consumption (via digital video observation, 94%), and resistance training intensity were similar (P > 0.05) for all three groups. Drug consumption unexpectedly increased muscle volume (acetaminophen: 109 ± 14 cm(3), 12.5%; ibuprofen: 84 ± 10 cm(3), 10.9%) and muscle strength (acetaminophen: 19 ± 2 kg; ibuprofen: 19 ± 2 kg) to a greater extent (P < 0.05) than placebo (muscle volume: 69 ± 12 cm(3), 8.6%; muscle strength: 15 ± 2 kg), when controlling for initial muscle size and strength. Follow-up analysis of muscle biopsies taken from the vastus lateralis before and after training showed muscle protein content, muscle water content, and myosin heavy chain distribution were not influenced (P > 0.05) by drug consumption. Similarly, muscle content of the two known enzymes potentially targeted by the drugs, COX-1 and -2, was not influenced (P > 0.05) by drug consumption, although resistance training did result in a drug-independent increase in COX-1 (32 ± 8%; P < 0.05). Drug consumption did not influence the size of the nonresistance-trained hamstring muscles (P > 0.05). Over-the-counter doses of acetaminophen or ibuprofen, when consumed in combination with resistance training, do not inhibit and appear to enhance muscle hypertrophy and strength gains in older adults. The present findings coupled with previous short-term exercise studies provide convincing evidence that the COX pathway(s) are involved in the regulation of muscle protein turnover and muscle mass in humans.

A low-glycemic index diet combined with exercise reduces insulin resistance, postprandial hyperinsulinemia, and glucose-dependent insulinotropic polypeptide responses in obese, prediabetic humans.

BACKGROUND: The optimal lifestyle intervention that reverses diabetes risk factors is not known. OBJECTIVE: We examined the effect of a low-glycemic index (GI) diet and exercise intervention on glucose metabolism and insulin secretion in obese, prediabetic individuals. DESIGN: Twenty-two participants [mean ± SEM age: 66 ± 1 y; body mass index (in kg/m(2)): 34.4 ± 0.8] underwent a 12-wk exercise-training intervention (1 h/d for 5 d/wk at ≈ 85% of maximum heart rate) while randomly assigned to receive either a low-GI diet (LoGIX; 40 ± 0.3 units) or a high-GI diet (HiGIX; 80 ± 0.6 units). Body composition (measured by using dual-energy X-ray absorptiometry and computed tomography), insulin sensitivity (measured with a hyperinsulinemic euglycemic clamp with [6,6-(2)H(2)]-glucose), and oral glucose-induced insulin and incretin hormone secretion were examined. RESULTS: Both groups lost equal amounts of body weight (-8.8 ± 0.9%) and adiposity and showed similar improvements in peripheral tissue (+76.2 ± 14.9%) and hepatic insulin sensitivity (+27.1 ± 7.1%) (all P < 0.05). However, oral glucose-induced insulin secretion was reduced only in the LoGIX group (6.59 ± 0.86 nmol in the prestudy compared with 4.70 ± 0.67 nmol in the poststudy, P < 0.05), which was a change related to the suppressed postprandial response of glucose-dependent insulinotropic polypeptide. When corrected for changes in ß cell glucose exposure, changes in insulin secretion were attenuated in the LoGIX group but became significantly elevated in the HiGIX group. CONCLUSIONS: Although lifestyle-induced weight loss improves insulin resistance in prediabetic individuals, postprandial hyperinsulinemia is reduced only when a low-GI diet is consumed. In contrast, a high-GI diet impairs pancreatic ß cell and intestinal K cell function despite significant weight loss. These findings highlight the important role of the gut in mediating the effects of a low-GI diet on type 2 diabetes risk reduction.

Improved pancreatic beta-cell function in type 2 diabetic patients after lifestyle-induced weight loss is related to glucose-dependent insulinotropic polypeptide.

OBJECTIVE: Restoration of insulin secretion is critical for the treatment of type 2 diabetes. Exercise and diet can alter glucose-induced insulin responses, but whether this is due to changes in beta-cell function per se is not clear. The mechanisms by which lifestyle intervention may modify insulin secretion in type 2 diabetes have also not been examined but may involve the incretin axis. RESEARCH DESIGN AND METHODS: Twenty-nine older, obese (aged 65 +/- 1 years; BMI 33.6 +/- 1.0 kg/m(2)) subjects, including individuals with newly diagnosed type 2 diabetes (obese-type 2 diabetic) and individuals with normal glucose tolerance (obese-NGT), underwent 3 months of nutritional counseling and exercise training. beta-Cell function (oral glucose-induced insulin secretion corrected for insulin resistance assessed by hyperinsulinemic-euglycemic clamps) and the role of glucose-dependent insulinotropic polypeptide (GIP) were examined. RESULTS: After exercise and diet-induced weight loss (-5.0 +/- 0.7 kg), oral glucose-induced insulin secretion was increased in the obese-type 2 diabetic group and decreased in the obese-NGT group (both P < 0.05). When corrected for alterations in insulin resistance, the change in insulin secretion remained significant only in the obese-type 2 diabetic group (1.23 +/- 0.26 vs. 2.04 +/- 0.46 arbitrary units; P < 0.01). Changes in insulin secretion were directly related to the GIP responses to oral glucose (r = 0.64, P = 0.005), which were augmented in the obese-type 2 diabetic group and only moderately suppressed in the obese-NGT group. CONCLUSIONS: After lifestyle-induced weight loss, improvements in oral glucose-induced insulin secretion in older, obese, nondiabetic subjects seem to be largely dependent on improved insulin sensitivity. However, in older obese diabetic patients, improved insulin secretion is a consequence of elevated beta-cell function. We demonstrate for the first time that changes in insulin secretion after lifestyle intervention may be mediated via alterations in GIP secretion from intestinal K-cells.

Free fatty acid-induced hepatic insulin resistance is attenuated following lifestyle intervention in obese individuals with impaired glucose tolerance.

OBJECTIVE: The objective of the study was to examine the effects of an exercise/diet lifestyle intervention on free fatty acid (FFA)-induced hepatic insulin resistance in obese humans. RESEARCH DESIGN AND METHODS: Obese men and women (n = 23) with impaired glucose tolerance were randomly assigned to either exercise training with a eucaloric (EU; approximately 1800 kcal; n = 11) or hypocaloric (HYPO; approximately 1300 kcal; n = 12) diet for 12 wk. Hepatic glucose production (HGP; milligrams per kilogram fat-free mass(-1) per minute(-1)) and hepatic insulin resistance were determined using a two-stage sequential hyperinsulinemic (40 mU/m(2) . min(-1)) euglycemic (5.0 mm) clamp with [3-(3)H]glucose. Measures were obtained at basal, during insulin infusion (INS; 120 min), and insulin plus intralipid/heparin infusion (INS/FFA; 300 min). RESULTS: At baseline, basal HGP was similar between groups; hyperinsulinemia alone did not completely suppress HGP, whereas INS/FFA exhibited less suppression than INS (EU, 4.6 +/- 0.8, 2.0 +/- 0.5, and 2.6 +/- 0.4; HYPO, 3.8 +/- 0.5, 1.2 +/- 0.3, and 2.3 +/- 0.4, respectively). After the intervention the HYPO group lost more body weight (P < 0.05) and fat mass (P < 0.05). However, both lifestyle interventions reduced hepatic insulin resistance during basal (P = 0.005) and INS (P = 0.001) conditions, and insulin-mediated suppression of HGP during INS was equally improved in both groups (EU: -42 +/- 22%; HYPO: -50 +/- 20%, before vs. after, P = 0.02). In contrast, the ability of insulin to overcome FFA-induced hepatic insulin resistance and HGP was improved only in the HYPO group (EU: -15 +/- 24% vs. HYPO: -58 +/- 19%, P = 0.02). CONCLUSIONS: Both lifestyle interventions are effective in reducing hepatic insulin resistance under basal and hyperinsulinemic conditions. However, the reversal of FFA-induced hepatic insulin resistance is best achieved with a combined exercise/caloric-restriction intervention.

Muscle proteins during 60-day bedrest in women: impact of exercise or nutrition.

Almost no data exist regarding skeletal muscle responses to real or simulated spaceflight in women. We determined the impact of 60-day bedrest (BR, n=8), 60-day bedrest with exercise-training (BRE, n=8), and 60-day bedrest with a leucine-enriched, high-protein diet (BRN, n=8) on muscle protein composition. Vastus lateralis and soleus muscle biopsies were analyzed for global protein fractions (mixed, sarcoplasmic, myofibrillar) and force-specific proteins (myosin, actin, collagen). Concentrations (micrograms per milligram muscle wet weight) of these proteins were maintained (P>0.05) in BR, despite large changes in quadriceps (-21%) and triceps surae (-29%) volume. Neither countermeasure influenced muscle protein content in either muscle (P>0.05), despite exacerbation (BRN) or prevention (BRE) of atrophy. Pre-bedrest comparisons showed less myofibrillar protein in the soleus (-16%, P<0.05), primarily due to less myosin (-12%, P<0.05) and more collagen (234%, P<0.05) than the vastus lateralis. Muscle protein composition is tightly regulated in lower limb muscles of women, despite the most extreme weightlessness-induced atrophy reported in humans. In contrast, men who underwent prolonged unloading were unable to proportionally regulate atrophy of the soleus. These findings have implications for astronauts and clinical conditions of sarcopenia regarding the maintenance of muscle function and prevention of frailty.

Plasma ceramides are elevated in obese subjects with type 2 diabetes and correlate with the severity of insulin resistance.

OBJECTIVE: To quantitate plasma ceramide subspecies concentrations in obese subjects with type 2 diabetes and relate these plasma levels to the severity of insulin resistance. Ceramides are a putative mediator of insulin resistance and lipotoxicity, and accumulation of ceramides within tissues in obese and diabetic subjects has been well described. RESEARCH DESIGN AND METHODS: We analyzed fasting plasma ceramide subspecies by quantitative tandem mass spectrometry in 13 obese type 2 diabetic patients and 14 lean healthy control subjects. Results were related to insulin sensitivity measured with the hyperinsulinemic-euglycemic clamp technique and with plasma tumor necrosis factor-alpha (TNF-alpha) levels, a marker of inflammation. Ceramide species (C18:1, 18:0, 20:0, 24:1, and 24:0) were quantified using electrospray ionization tandem mass spectrometry after separation with high-performance liquid chromatography. RESULTS: Insulin sensitivity (mg x kg(-1) x min(-1)) was lower in type 2 diabetic patients (4.90 +/- 0.3) versus control subjects (9.6 +/- 0.4) (P < 0.0001). Type 2 diabetic subjects had higher (P < 0.05) concentrations of C18:0, C20:0, C24:1, and total ceramide. Insulin sensitivity was inversely correlated with C18:0, C20:0, C24:1, C24:0, and total ceramide (all P < 0.01). Plasma TNF-alpha concentration was increased (P < 0.05) in type 2 diabetic subjects and correlated with increased C18:1 and C18:0 ceramide subspecies. CONCLUSIONS: Plasma ceramide levels are elevated in type 2 diabetic subjects and may contribute to insulin resistance through activation of inflammatory mediators, such as TNF-alpha.

Collagen, cross-linking, and advanced glycation end products in aging human skeletal muscle.

We examined intramuscular endomysial collagen, cross-linking, and advanced glycation end products, as well as the general and contractile protein concentration of 20 young (25 +/- 3 yr) and 22 old (78 +/- 6 yr, range: 70-93 yr) sedentary men and women to better understand the underlying basis of changes in skeletal muscle mass and function that occur with aging. The old individuals had an impaired ability (increased time) (P < 0.05) to climb stairs (80%), rise from a chair (56%), and walk (44%), as well as lower (P < 0.05) quadriceps muscle volume (-29%), muscle strength (-35%), muscle power (-48%), and strength (-17%) and power (-33%) normalized to muscle size. Vastus lateralis muscle biopsies revealed that intramuscular endomysial collagen (young: 9.6 +/- 1.1, old: 10.2 +/- 1.2 microg/mg muscle wet wt) and collagen cross-linking (hydroxylysylpyridinoline) (young: 395 +/- 65, old: 351 +/- 45 mmol hydroxylysylpyridinoline/mol collagen) were unchanged (P > 0.05) with aging. The advanced glycation end product, pentosidine, was increased (P < 0.05) by approximately 200% (young: 5.2 +/- 1.3, old: 15.9 +/- 4.5 mmol pentosidine/mol collagen) with aging. While myofibrillar protein concentration was lower (-5%, P < 0.05), the concentration of the main contractile proteins myosin and actin were unchanged (P > 0.05) with aging. These data suggest that the synthesis and degradation of proteins responsible for the generation (myosin and actin) and transfer (collagen and pyridinoline cross-links) of muscle force are tightly regulated in aging muscle. Glycation-related cross-linking of intramuscular connective tissue may contribute to altered muscle force transmission and muscle function with healthy aging.