Association of low-density neutrophils with lung function and disease progression in adult cystic fibrosis.

BACKGROUND: Cystic fibrosis (CF) neutrophils fail to eradicate infection despite their massive recruitment into the lung. While studies mostly focus on pathogen clearance by normal density neutrophils in CF, the contribution of low-density neutrophil (LDNs) subpopulations to disease pathogenesis remains unclear. METHODS: LDNs were isolated from whole blood donations of clinically stable adult CF patients and from healthy donors. LDN proportion and immunophenotype was assessed by flow cytometry. Associations of LDNs with clinical parameters were determined. RESULTS: LDN proportion was increased in CF patients' circulation compared with healthy donors. LDNs are a heterogeneous population of both mature and immature cells in CF and in healthy individuals. Moreover, a higher proportion of mature LDN correlates with a gradual decline in lung function and repeated pulmonary exacerbations in CF patients. CONCLUSIONS: Collectively, our observations suggest that low-density neutrophils are linked to CF pathogenesis and underscore the potential clinical relevance of neutrophil subpopulations in CF.

Serum Analyte Profiles Associated With Crohn's Disease and Disease Location.

BACKGROUND: Crohn's disease (CD) can affect any segment of the digestive tract but is most often localized in the ileal, ileocolonic, and colorectal regions of the intestines. It is believed that the chronic inflammation in CD is a result of an imbalance between the epithelial barrier, the immune system, and the intestinal microbiota. The aim of the study was to identify circulating markers associated with CD and/or disease location in CD patients. METHODS: We tested 49 cytokines, chemokines, and growth factors in serum samples from 300 patients with CD and 300 controls. After quality control, analyte levels were tested for association with CD and disease location. RESULTS: We identified 13 analytes that were higher in CD patients relative to healthy controls and that remained significant after conservative Bonferroni correction (P < 0.0015). In particular, CXCL9, CXCL1, and interleukin IL-6 had the greatest effect and were highly significant (P < 5 × 10-7). We also identified 9 analytes that were associated with disease location, with VEGF, IL-12p70, and IL-6 being elevated in patients with colorectal disease (P < 3 × 10-4). CONCLUSIONS: Multiple serum analytes are elevated in CD. These implicate the involvement of multiple cell types from the immune, epithelial, and endothelial systems, suggesting that circulating analytes reflect the inflammatory processes that are ongoing within the gut. Moreover, the identification of distinct profiles according to disease location supports the existence of a biological difference between ileal and colonic CD, consistent with previous genetic and clinical observations.

The Potential Causes of Cystic Fibrosis-Related Diabetes.

Cystic fibrosis (CF) is a genetic disease caused by mutations in the cystic fibrosis transmembrane conductance regulator gene (CFTR). Cystic fibrosis-related diabetes (CFRD) is the most common comorbidity, affecting more than 50% of adult CF patients. Despite this high prevalence, the etiology of CFRD remains incompletely understood. Studies in young CF children show pancreatic islet disorganization, abnormal glucose tolerance, and delayed first-phase insulin secretion suggesting that islet dysfunction is an early feature of CF. Since insulin-producing pancreatic ß-cells express very low levels of CFTR, CFRD likely results from ß-cell extrinsic factors. In the vicinity of ß-cells, CFTR is expressed in both the exocrine pancreas and the immune system. In the exocrine pancreas, CFTR mutations lead to the obstruction of the pancreatic ductal canal, inflammation, and immune cell infiltration, ultimately causing the destruction of the exocrine pancreas and remodeling of islets. Both inflammation and ductal cells have a direct effect on insulin secretion and could participate in CFRD development. CFTR mutations are also associated with inflammatory responses and excessive cytokine production by various immune cells, which infiltrate the pancreas and exert a negative impact on insulin secretion, causing dysregulation of glucose homeostasis in CF adults. In addition, the function of macrophages in shaping pancreatic islet development may be impaired by CFTR mutations, further contributing to the pancreatic islet structural defects as well as impaired first-phase insulin secretion observed in very young children. This review discusses the different factors that may contribute to CFRD.

Humoral responses to the measles, mumps and rubella vaccine are impaired in Leigh Syndrome French Canadian patients.

Leigh Syndrome French Canadian (LSFC) is a rare autosomal recessive metabolic disorder characterized by severe lactic acidosis crises and early mortality. LSFC patients carry mutations in the Leucine Rich Pentatricopeptide Repeat Containing (LRPPRC) gene, which lead to defects in the respiratory chain complexes and mitochondrial dysfunction. Mitochondrial respiration modulates cellular metabolic activity, which impacts many cell types including the differentiation and function of immune cells. Hence, we postulated that, in addition to neurological and metabolic disorders, LSFC patients may show impaired immune activity. To gain insight into the quality of the immune response in LSFC patients, we examined the response to the measles, mumps and rubella (MMR) vaccine by measuring antibody titers to MMR in the plasma. In a cohort of eight LSFC patients, the response to the MMR vaccine was variable, with some individuals showing antibodies to all three viruses, while others had antibodies to two or fewer viruses. These results suggest that the mutations in the LRPPRC gene present in LSFC patients may affect the immune response to vaccines. Monitoring vaccine response in this fragile population should be considered to ensure full protection against pathogens.

Induced and spontaneous colitis mouse models reveal complex interactions between IL-10 and IL-12/IL-23 pathways.

Crohn's disease (CD) and ulcerative colitis (UC) are the two major forms of inflammatory bowel disease (IBD). These idiopathic and chronic diseases result from inflammation of the gastrointestinal tract and are mainly mediated by the immune system. Genome wide association studies link genes of the IL-12 and IL-23 biology to both CD and UC susceptibility. IL-12 and IL-23 cytokines share a functional subunit, p40, and their respective receptors also share a functional subunit, IL-12Rß1. However, clinical trials targeting p40, and thus inhibiting both IL-12 and IL-23 pathways, provided mitigated effects on IBD, suggesting context dependent effects for each cytokine. In addition to IL-12 and IL-23, genetic deficiencies in IL-10 also result in severe IBD pathology. We generated various mouse models to determine how IL-12 or IL-23 interacts with IL-10 in IBD pathology. Whereas defects in both IL-10 and IL-12R do not impact the severity of the Dextran Sulfate Sodium (DSS)-induced colitis, combined deficiencies in both IL-10 and IL-23R aggravate the disease. In contrast to DSS-induced colitis, defects in IL-12R and IL-23R both protect from the spontaneous colitis observed in IL10-/- mice. Together, these studies exemplify the complexity of genetic and environmental interactions for identifying biological pathways predictive of pathological inflammatory processes.

mTORC1 is required for expression of LRPPRC and cytochrome-c oxidase but not HIF-1α in Leigh syndrome French Canadian type patient fibroblasts.

Leigh syndrome French Canadian type (LSFC) is a mitochondrial disease caused by mutations in the leucine-rich pentatricopeptide repeat-containing (LRPPRC) gene leading to a reduction of cytochrome-c oxidase (COX) expression reaching 50% in skin fibroblasts. We have shown that under basal conditions, LSFC and control cells display similar ATP levels. We hypothesized that this occurs through upregulation of mechanistic target of rapamycin (mTOR)-mediated metabolic reprogramming. Our results showed that compared with controls, LSFC cells exhibited an upregulation of the mTOR complex 1 (mTORC1)/p70 ribosomal S6 kinase pathway and higher levels of hypoxia-inducible factor 1α (HIF-1α) and its downstream target pyruvate dehydrogenase kinase 1 (PDHK1), a regulator of mitochondrial pyruvate dehydrogenase 1 (PDH1). Consistent with these signaling alterations, LSFC cells displayed a 40-61% increase in [U-13C6]glucose contribution to pyruvate, lactate, and alanine formation, as well as higher levels of the phosphorylated and inactive form of PDH1-α. Interestingly, inhibition of mTOR with rapamycin did not alter HIF-1α or PDHK1 protein levels in LSFC fibroblasts. However, this treatment increased PDH1-α phosphorylation in control and LSFC cells and reduced ATP levels in control cells. Rapamycin also decreased LRPPRC expression by 41 and 11% in LSFC and control cells, respectively, and selectively reduced COX subunit IV expression in LSFC fibroblasts. Taken together, our data demonstrate the importance of mTORC1, independent of the HIF-1α/PDHK1 axis, in maintaining LRPPRC and COX expression in LSFC cells.

Protecting the heart through MK2 modulation, toward a role in diabetic cardiomyopathy and lipid metabolism.

Various signaling pathways have been identified in the heart as important players during development, physiological adaptation or pathological processes. This includes the MAPK families, particularly p38MAPK, which is involved in several key cellular processes, including differentiation, proliferation, apoptosis, inflammation, metabolism and survival. Disrupted p38MAPK signaling has been associated with several diseases, including cardiovascular diseases (CVD) as well as diabetes and its related complications. Despite efforts to translate this knowledge into therapeutic avenues, p38 inhibitors have failed in clinical trials due to adverse effects. Inhibition of MK2, a downstream target of p38, appears to be a promising alternative strategy. Targeting MK2 activity may avoid the adverse effects linked to p38 inhibition, while maintaining its beneficial effects. MK2 was first considered as a therapeutic target in inflammatory diseases such as rheumatoid polyarthritis. A growing body of evidence now supports a key role of MK2 signaling in the pathogenesis of CVD, particularly ischemia/reperfusion injury, hypertrophy, and hypertension and that its inhibition or inactivation is associated with improved heart and vascular functions. More recently, MK2 was shown to be a potential player in diabetes and related complications, particularly in liver and heart, and perturbations in calcium handling and lipid metabolism. In this review, we will discuss recent advances in our knowledge of the role of MK2 in p38MAPK-mediated signaling and the benefits of its loss of function in CVD and diabetes, with an emphasis on the roles of MK2 in calcium handling and lipid metabolism. This article is part of a Special issue entitled Cardiac adaptations to obesity, diabetes and insulin resistance, edited by Professors Jan F.C. Glatz, Jason R.B. Dyck and Christine Des Rosiers.

MK2 Deletion in Mice Prevents Diabetes-Induced Perturbations in Lipid Metabolism and Cardiac Dysfunction.

Heart disease remains a major complication of diabetes, and the identification of new therapeutic targets is essential. This study investigates the role of the protein kinase MK2, a p38 mitogen-activated protein kinase downstream target, in the development of diabetes-induced cardiomyopathy. Diabetes was induced in control (MK2(+/+)) and MK2-null (MK2(-/-)) mice using repeated injections of a low dose of streptozotocin (STZ). This protocol generated in MK2(+/+) mice a model of diabetes characterized by a 50% decrease in plasma insulin, hyperglycemia, and insulin resistance (IR), as well as major contractile dysfunction, which was associated with alterations in proteins involved in calcium handling. While MK2(-/-)-STZ mice remained hyperglycemic, they showed improved IR and none of the cardiac functional or molecular alterations. Further analyses highlighted marked lipid perturbations in MK2(+/+)-STZ mice, which encompass increased 1) circulating levels of free fatty acid, ketone bodies, and long-chain acylcarnitines and 2) cardiac triglyceride accumulation and ex vivo palmitate ß-oxidation. MK2(-/-)-STZ mice were also protected against all these diabetes-induced lipid alterations. Our results demonstrate the benefits of MK2 deletion on diabetes-induced cardiac molecular and lipid metabolic changes, as well as contractile dysfunction. As a result, MK2 represents a new potential therapeutic target to prevent diabetes-induced cardiac dysfunction.

Mitochondrial vulnerability and increased susceptibility to nutrient-induced cytotoxicity in fibroblasts from leigh syndrome French canadian patients.

Mutations in LRPPRC are responsible for the French Canadian variant of Leigh Syndrome (LSFC), a severe disorder characterized biochemically by a tissue-specific deficiency of cytochrome c oxidase (COX) and clinically by the occurrence of severe and deadly acidotic crises. Factors that precipitate these crises remain unclear. To better understand the physiopathology and identify potential treatments, we performed a comprehensive analysis of mitochondrial function in LSFC and control fibroblasts. Furthermore, we have used this cell-based model to screen for conditions that promote premature cell death in LSFC cells and test the protective effect of ten interventions targeting well-defined aspects of mitochondrial function. We show that, despite maintaining normal ATP levels, LSFC fibroblasts present several mitochondrial functional abnormalities under normal baseline conditions, which likely impair their capacity to respond to stress. This includes mitochondrial network fragmentation, impaired oxidative phosphorylation capacity, lower membrane potential, increased sensitivity to Ca2+-induced permeability transition, but no changes in reactive oxygen species production. We also show that LSFC fibroblasts display enhanced susceptibility to cell death when exposed to palmitate, an effect that is potentiated by high lactate, while high glucose or acidosis alone or in combination were neutral. Furthermore, we demonstrate that compounds that are known to promote flux through the electron transport chain independent of phosphorylation (methylene blue, dinitrophenol), or modulate fatty acid (L-carnitine) or Krebs cycle metabolism (propionate) are protective, while antioxidants (idebenone, N-acetyl cysteine, resveratrol) exacerbate palmitate plus lactate-induced cell death. Collectively, beyond highlighting multiple alterations in mitochondrial function and increased susceptibility to nutrient-induced cytotoxicity in LSFC fibroblasts, these results raise questions about the nature of the diets, particularly excess fat intake, as well as on the use of antioxidants in patients with LSFC and, possibly, other COX defects.

Impact of sex on insulin secretion in cystic fibrosis.

CONTEXT: Cystic fibrosis-related diabetes is primarily due to a defect in insulin secretion. Women with cystic fibrosis (CF) are at higher risk of developing CF-related diabetes. OBJECTIVE: The objective of the study was to examine sex differences in insulin and glucose homeostasis. We hypothesized that in CF, women would display lower insulin secretion than men. DESIGN: This was a study based on an ongoing observational CF cohort with a mean follow-up of 19.9 ± 5.2 months. SETTING: The study was conducted at the CF clinic of the Centre Hospitalier de l'Université de Montréal (Québec, Canada). PATIENTS: From 230 adults with CF (123 men, 107 women) of similar age and functional pulmonary status, 104 retested after the follow-up. Age-matched healthy individuals (25 men, 19 women) were included in the study. INTERVENTIONS: Participants underwent a 2-hour oral glucose tolerance test with 30-minute interval sample measurements. MAIN OUTCOME MEASURE: Plasma insulin and glucose levels were measured. RESULTS: Women with CF had higher overall insulin secretion as compared with men with CF (P ≤ .05) but similar to healthy women (P = .606). Men with CF had lower overall insulin secretion than healthy men (P = .020) and higher insulin sensitivity (P = .009) than women with CF. PATIENTS with CF displayed higher overall glucose excursions than healthy patients. Sex-related differences were still observed in the CF cohort after follow-up. CONCLUSIONS: Surprisingly, in CF, adult women presented higher insulin secretion than adult men at a comparable level with what is observed in healthy individuals. Potential implications of this sex dimorphism in CF remain to be established.

The Visceral Adiposity Index: Relationship with cardiometabolic risk factors in obese and overweight postmenopausal women--a MONET group study.

A recent study suggested visceral adipose index (VAI) as an indicator of adipose tissue distribution and function associated with cardiometabolic risk. We aim to examine the association between VAI and visceral adipose tissue (VAT), insulin sensitivity, and a large panel of associated cardiometabolic risk factors, and to determine if changes in VAI after weight loss intervention will reflect changes in VAT. We performed a secondary analysis using the data of 99 overweight and postmenopausal women that completed a 6-month weight loss program (Montreal Ottawa New Emerging Team Study). VAI was calculated according to the equation by Amato et al. (2010; Diabetes Care, 33(4):920-922). At baseline, VAI was associated with VAT (r = 0.284, p < 0.01) but not with subcutaneous adipose tissue (SAT) while body mass index (BMI) and waist circumference (WC) were significantly related to both. BMI and WC demonstrated significantly stronger predictive value of VAT accumulation (area under the curve = 0.84 and 0.86, respectively) than VAI (area under the curve = 0.61; p < 0.01). However, VAT, BMI, WC, and VAI were similarly related to fasting insulin and glucose disposal rates. After a 6-month weight loss program, VAI decreased significantly and similarly in both intervention groups (p < 0.01). In addition, the percentage of change in VAI showed the significantly weakest correlation (r = 0.25) with the percentage of change in VAT than BMI (r = 0.56; p < 0.01 for r comparisons) and was not a significant predictor of interindividual percentage of change in VAT while BMI accounted for 33.7%. VAI is a weak indicator of VAT function and did not predict changes in VAT after weight loss. Furthermore, this index was not superior to BMI or WC. However, VAI is a good indicator of metabolic syndrome.

Hypertriglyceridemia is associated with insulin levels in adult cystic fibrosis patients.

BACKGROUND: Recent studies have identified hypertriglyceridemic cystic fibrosis patients (CF-TG). However, whether hypertriglyceridemia is associated with an altered metabolic profile remains unknown. OBJECTIVE: To characterize CF-TG and determine whether triglycerides (TG) levels are associated with metabolic alterations. METHODS: 210 adult CF subjects from the Montreal Cystic Fibrosis Cohort without known diabetes were included in the analysis. All subjects underwent an OGTT to assess glucose tolerance, insulin secretion (insulin AUC) and insulin sensitivity (Stumvoll index). Fasting lipid profiles, pulmonary function (%FEV1) and BMI were determined. Hypertriglyceridemia (TG>1.7mmol/L) was observed in 20 CF patients. These subjects were matched for age, sex and glucose tolerance category with 20 CF patients (CF-normal-TG) and 20 healthy controls that had TG levels below 1.7mmol/L. Pearson correlations were performed in the complete study sample (n=210) to examine the associations between TG levels and other parameters. RESULTS: The prevalence of hypertriglyceridemia was 9.5%. Compared to CF-normal-TG, CF-TG subjects displayed significantly higher %FEV1, insulin AUC (AUC0-120, AUC0-30, AUC30-120), cholesterol levels and a higher ratio of total cholesterol to HDL-cholesterol. Pearson analysis demonstrated that TG levels were associated with BMI, %FEV1, fasting insulin, insulin AUC0-120 and AUC30-120, Stumvoll index, cholesterol levels and the ratio of total cholesterol to HDL-cholesterol. All these correlations remained significant after correction for BMI except %FEV1. CONCLUSION: TG levels are associated with a mild alteration of the metabolic profile. Whether these changes will increase the long-term risk of CF patients in developing cardiometabolic complications remains to be investigated.

LDL-cholesterol and insulin are independently associated with body mass index in adult cystic fibrosis patients.

BACKGROUND: The median life expectancy of cystic fibrosis (CF) patients has increased dramatically over the last few years and we now observe a subset of patients with a body mass index (BMI) exceeding 25 kg/m(2). The aim of this study was to characterize these individuals and to identify factors associated with higher BMI. METHODS: This is a cross sectional study including 187 adult CF subjects. Percent predicted forced expiratory volume in 1s (%FEV(1)), blood lipid profiles as well as fasting glucose and insulin levels were evaluated. Subjects also had an oral glucose tolerance test (OGTT) and the area under the curve (AUC) for glucose and insulin was calculated. CF subjects were then stratified according to the following BMI categories: underweight: BMI≤18.5 kg/m(2); normal weight: 18.5 kg/m(2)

Fiber intake predicts ghrelin levels in overweight and obese postmenopausal women.

BACKGROUND: Ghrelin levels are decreased upon food intake, but the impact of specific diet-derived macronutrients on its regulation remains unclear. In addition, because of ghrelin's association with body weight regulation, it is important to understand the mechanisms regulating its levels in obese individuals. OBJECTIVE: To examine the effect of specific macronutrients on ghrelin levels in overweight and obese postmenopausal women. METHODS: Thirty-five subjects underwent a euglycemic/hyperinsulinemic clamp (EHC) to examine glucose disposal and total ghrelin (TotG) and acylated ghrelin (AG) levels. Macronutrient intake was evaluated with a 3-day food questionnaire. RESULTS: Under fasting conditions, positive associations were observed between fiber intake and TotG and AG levels. Fasting AG also correlated positively with the intake of total energy, as well as monounsaturated and polyunsaturated lipids. Importantly, fiber consumption explained up to 26 and 23% of the variation in TotG and AG respectively. During the EHC, TotG levels were significantly reduced at all times, while AG was decreased at 60 min only. TotG area under the curve (AUC) values were positively associated with fiber and polyunsaturated lipid intake, while AG AUC values correlated positively with fiber, total energy, carbohydrate, and lipid intake. Interestingly, fiber intake explained up to 21% of the variation in TotG AUC, while total energy intake predicted up to 21% of the variation in the AG AUC. CONCLUSION: The present study suggests that fiber intake is an important regulator of ghrelin levels both in fasting and in hyperinsulinemic conditions. Overall, these results reinforce the importance of the intimate association between eating habits and gastrointestinal hormonal regulation.

Change in plasma acylation stimulating protein during euglycaemic-hyperinsulinaemic clamp in overweight and obese postmenopausal women: a MONET study.

OBJECTIVE: Acylation-stimulating protein (ASP) has been shown to positively stimulate fatty acid esterification and glucose uptake in adipocytes. In vitro studies demonstrate that insulin stimulates ASP secretion from adipocytes. Individuals with obesity and/or metabolic disturbances (insulin resistance and type 2 diabetes) have increased plasma ASP. DESIGN: The present study was designed to evaluate whether ASP levels are influenced by the metabolic profiles of overweight and obese postmenopausal women during a euglycaemic/hyperinsulinaemic clamp (EHC). Patients The study population consisted of 76 overweight and obese sedentary postmenopausal women. MEASUREMENTS: We evaluated insulin sensitivity, plasma ASP levels, body composition including visceral adipose tissue area, blood lipid profiles, liver enzymes, peak aerobic capacity, resting metabolic rate (RMR) and total energy expenditure (TEE). RESULTS: We observed wide interindividual variations of ASP levels during the EHC. Therefore, subjects were divided into three groups based on ASP changes. Negative ASP Responders (NAR; n = 24) showed a -20% or greater decrease in ASP levels while Positive ASP Responders (PAR; n = 42) displayed ASP fluctuations superior to +20%. Ten subjects had little or no ASP change and were considered as Zero ASP responders (ZAR). PAR women displayed a worse metabolic profile than NAR women, including higher BMI, visceral adipose tissue, fasting insulin levels, lean body mass, and alanine aminotransferase (ALT), a marker of impaired liver function. After adjustment for BMI, only ALT remained significantly different, while lean body mass (P = 0.08) and visceral adipose tissue (P = 0.07) remained marginally higher. Correlation analysis of all subjects demonstrated that fasting ASP levels correlated positively with albumin and VO(2 peak) and this association remained significant after adjustments for the effect of BMI. In addition, the percentage maximal change in ASP levels during the EHC was positively associated with BMI, lean body mass, visceral adipose tissue, fasting insulin, HOMA, TEE, RMR, ALT and AST. CONCLUSION: Overall these results suggest that an elevated ASP response during the EHC is associated with metabolic disturbances in overweight and obese postmenopausal women.

Tissue- and fibre-specific modifications of insulin-signalling molecules in cardiac and skeletal muscle of diabetic rats.

1. Levels of insulin-signalling molecules are altered in streptozotocin (STZ)-induced diabetes, a model of Type 1 diabetes. However, the tissue-specific regulation of these changes and the effect of insulin supplementation on signalling molecule protein levels have not been well characterized. 2. In the present study, we evaluated the level of proximal insulin-signalling intermediates in the heart and in red and white gastrocnemius muscles of 2 week diabetic rats and diabetic rats supplemented with insulin. 3. Diabetes augmented levels of the insulin receptor and the p85 regulatory subunit of phosphatidylinositol 3-kinase in the red gastrocnemius, but not in the white gastrocnemius or the heart. Furthermore, diabetes reduced insulin receptor substrate-1 levels in both the red and white gastrocnemius, but not in the heart. Examination of the levels and basal activities of distal insulin-signalling intermediates (protein kinase B (PKB)/Akt, extracellular signal-regulated kinase (ERK) 1/2, p38 mitogen-activated protein kinase (MAPK)) also failed to reveal a specific pattern in these changes. Thus, diabetes reduced basal ERK1/2 and PKB/Akt phosphorylation in the heart and white gastrocnemius, respectively, whereas it augmented basal p38 MAPK activity in the red gastrocnemius. Insulin supplementation normalized the levels and activities of some but not all proteins. 4. In conclusion, the results of the present study demonstrate that adaptation to STZ-induced diabetes varies among skeletal muscle fibre types and the heart, emphasizing the complex tissue-specific responses to diabetes.

Association of acylated ghrelin profiles with chronic inflammatory markers in overweight and obese postmenopausal women: a MONET study.

OBJECTIVE: Recent reports have suggested that the existence of associations between hormonal dysregulation and chronic upregulation of inflammatory markers, which may cause obesity-related disturbances. Thus, we examined whether acylated ghrelin (AcylG) and total ghrelin (TotG) levels could be associated with the following inflammatory markers: C-reactive protein (CRP), tumor necrosis factor alpha (TNF-alpha), and soluble TNF receptor 1 (sTNF-R1). DESIGN: Cross-sectional study consisting of 50 overweight and obese postmenopausal women. METHODS: AcylG and TotG levels were assessed at 0, 60, 160, 170, and 180 min of the euglycemic/hyperinsulinemic clamp (EHC). We evaluated insulin sensitivity, body composition, and blood lipid profiles as well as fasting concentrations of CRP, TNF-alpha, and sTNF-R1. RESULTS: In fasting conditions, sTNF-R1 was negatively correlated with AcylG (r = -0.48, P < 0.001) levels. In addition, AcylG/TotG was associated negatively with sTNF-R1 (r = -0.44, P = 0.002) and positively with TNF-alpha (r = 0.38, P = 0.009) values. During the EHC, TotG (at all time points) and AcylG (at 60 and 160 min) values were significantly decreased from fasting concentrations. AcylG maximal reduction and area under the curve (AUC) values were correlated to sTNF-R1 (r = -0.35, P = 0.02 and r = -0.34, P = 0.02, respectively). Meanwhile, the AcylG/TotG AUC ratio was associated negatively with sTNF-R1 (r = -0.29, P < 0.05) and positively with TNF-alpha (r = 0.36, P = 0.02). Following adjustments for total adiposity, sTNF-R1 remained correlated with fasting and maximal reduction AcylG values. Similarly, AcylG/TotG ratios remained significantly correlated with sTNF-R1 and TNF-alpha. Importantly, 23% of the variation in sTNF-R1 was independently predicted by fasting AcylG. CONCLUSION: These results are the first to suggest that both fasting and EHC-induced AcylG profiles are correlated with fasting values of sTNF-R1, a component of the TNF-alpha system. Thus, AcylG may act, at least in part, as one mediator of chronic inflammatory activity in human obesity.

Regulation of glycogen concentration and glycogen synthase activity in skeletal muscle of insulin-resistant rats.

The aim of this study was to investigate the effect of insulin resistance on glycogen concentration and glycogen synthase activity in the red and white gastrocnemius muscles and to determine whether the inverse relationship existing between glycogen concentration and enzyme activity is maintained in insulin resistant state. These questions were addressed using 3 models that induce various degrees of insulin resistance: sucrose feeding, dexamethasone administration, and a combination of both treatments (dex+sucrose). Sucrose feeding raised triglyceride levels without affecting plasma glucose or insulin concentrations whereas dexamethasone and dex+sucrose provoked severe hyperinsulinemia, hyperglycemia and hypertriglyceridemia. Sucrose feeding did not alter muscle glycogen concentration but provoked a small reduction in the glycogen synthase activity ratio (-/+ glucose-6-phosphate) in red but not in white gastrocnemius. Dexamethasone administration augmented glycogen concentration and reduced glycogen synthase activity ratio in both muscle fiber types. In contrast, dex+sucrose animals showed decreased muscle glycogen concentration compared to dexamethasone group, leading to levels similar to those of control animals. This was associated with lower glycogen synthase activity compared to control animals leading to levels comparable to those of dexamethasone-treated animals. Thus, in dex+sucrose animals, the inverse relationship observed between glycogen levels and glycogen synthase activity was not maintained, suggesting that factors other than the glycogen concentration modulate the enzyme's activity. In conclusion, while insulin resistance was associated with a reduced glycogen synthase activity ratio, we found no correlation between muscle glycogen concentration and insulin resistance. Furthermore, our results suggest that sucrose treatment may modulate dexamethasone action in skeletal muscle.

Ketone bodies alter dinitrophenol-induced glucose uptake through AMPK inhibition and oxidative stress generation in adult cardiomyocytes.

In aerobic conditions, the heart preferentially oxidizes fatty acids. However, during metabolic stress, glucose becomes the major energy source, and enhanced glucose uptake has a protective effect on heart function and cardiomyocyte survival. Thus abnormal regulation of glucose uptake may contribute to the development of cardiac disease in diabetics. Ketone bodies are often elevated in poorly controlled diabetics and are associated with increased cellular oxidative stress. Thus we sought to determine the effect of the ketone body beta-hydroxybutyrate (OHB) on cardiac glucose uptake during metabolic stress. We used 2,4-dinitrophenol (DNP), an uncoupler of the mitochondrial oxidative chain, to mimic hypoxia in cardiomyocytes. Our data demonstrated that chronic exposure to OHB provoked a concentration-dependent decrease of DNP action, resulting in 56% inhibition of DNP-mediated glucose uptake at 5 mM OHB. This was paralleled by a diminution of DNP-mediated AMP-activated protein kinase (AMPK) and p38 MAPK phosphorylation. Chronic exposure to OHB also increased reactive oxygen species (ROS) production by 1.9-fold compared with control cells. To further understand the role of ROS in OHB action, cardiomyocytes were incubated with H(2)O(2). Our results demonstrated that this treatment diminished DNP-induced glucose uptake without altering activation of the AMPK/p38 MAPK signaling pathway. Incubation with the antioxidant N-acetylcysteine partially restored DNP-mediated glucose but not AMPK/p38 MAPK activation. In conclusion, these results suggest that ketone bodies, through inhibition of the AMPK/p38 MAPK signaling pathway and ROS overproduction, regulate DNP action and thus cardiac glucose uptake. Altered glucose uptake in hyperketonemic states during metabolic stress may contribute to diabetic cardiomyopathy.

Chronic exposure to ketone bodies impairs glucose uptake in adult cardiomyocytes in response to insulin but not vanadate: the role of PI3-K.

There is a strong positive correlation between insulin resistance and cardiac diseases. We have already shown that chronic exposure to the ketone body beta-hydroxybutyrate (OHB) decreases insulin-mediated activation of protein kinase B (PKB) and glucose uptake in cardiomyocytes. To gain further insights into the mechanism underlying ketone body-induced insulin resistance, we examined whether OHB alters activation of the insulin-signaling cascade and whether the insulinomimetic agent vanadate could bypass insulin resistance and stimulate glucose uptake in these cells. Cardiomyocytes were incubated with 5 mM OHB, 50 microM vanadate or both for 16 h before the measurement of glucose uptake or the activation of insulin-signaling molecules. While chronic exposure to OHB did not alter insulin- or vanadate-mediated activation of the insulin receptor, it suppressed insulin receptor substrate-1 (IRS-1) tyrosine phosphorylation in response to both agonists. Furthermore, this treatment decreased by 54 and 36% the phosphorylation of the p85 regulatory subunit of phosphatidylinositol 3-kinase (PI3-K) and PKB in response to insulin, whereas it did not alter vanadate-mediated activation of these enzymes. Although insulin did not significantly stimulate p38MAPK phosphorylation, vanadate increased it by 3.8-fold. Furthermore, chronic exposure to OHB potentiated vanadate's action, resulting in a 250% increase in enzyme activation compared to control cells. Though OHB induced a 2.1-fold increase of basal ERK1/2 phosphorylation, inhibition of this enzyme with the MEK inhibitor PD98059 demonstrated that ERK1/2 did not participate in OHB-induced insulin resistance. In conclusion, ketone bodies promote insulin resistance probably through decreased activation of the PI3-K/PKB signaling cascade. Furthermore, vanadate can bypass insulin resistance and stimulate glucose uptake in OHB-treated cardiomyocytes.