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Beta cell failure is one of the most important features of type 2 diabetes mellitus (T2DM). High‐density lipoprotein (HDL) has been proposed to improve β‐cell function. However, the mechanisms involved in this process are still poorly understood. The aim of this study was to investigate the contribution of sphingosine‐1‐phosphate (S1P) in the impact of HDL treatment on insulin secretion by pancreatic β‐cells and to determine its mechanisms. Primary cultures of β‐cells isolated from rat were treated with or without HDL in the presence or absence of S1P pathway inhibitors and insulin secretion response was analyzed. The S1P content of HDL (HDL‐S1P) isolated from T2DM patients was analyzed and correlated to the HDL‐induced insulin secretion. The expression of genes involved in the biosynthesis of the insulin was also evaluated. HDL as well as S1P treatment enhanced glucose‐stimulated insulin secretion (GSIS). In HDL isolated from T2DM patients, while HDL‐S1P was strongly correlated to its pro‐secretory capacity (r = 0.633, p = 0.005), HDL‐cholesterol and apolipoprotein AI levels were not. HDL‐induced GSIS was blocked by the S1P1/3 antagonist but not by the S1P2 antagonist, and was also accompanied by increased intracellular S1P in β‐cells. We also observed that HDL improved GSIS without significant changes in expression levels of insulin biosynthesis genes. Our present study highlights the importance HDL‐S1P in GSIS in T2DM patients and demonstrates that HDL induces insulin secretion by a process involving both intra‐ and extra‐cellular sources of S1P independently of an effect on insulin biosynthesis genes., HDL improves insulin secretion by beta cells S1P plays a role in HDL‐induced insulin secretion HDL increases intracellular S1P content.

International audience; AIM: The worldwide increase in obesity and type 2 diabetes (T2D) represents a major health challenge. Chronically altered lipids induced by obesity further promote the development of T2D, and the accumulation of toxic lipid metabolites in serum and peripheral organs may contribute to the diabetic phenotype. METHODS: To better understand the complex metabolic pattern of lean and obese T2D and non-T2D individuals, we analysed the lipid profile of human serum, skeletal muscle and visceral adipose tissue of two cohorts by systematic mass spectrometry-based lipid analysis. RESULTS: Lipid homeostasis was strongly altered in a disease- and tissue-specific manner, allowing us to define T2D signatures associated with obesity from those that were obesity independent. Lipid changes encompassed lyso-, diacyl- and ether-phospholipids. Moreover, strong changes in sphingolipids included cytotoxic 1-deoxyceramide accumulation in a disease-specific manner in serum and visceral adipose tissue. The high amounts of non-canonical 1-deoxyceramide present in human adipose tissue most likely come from cell-autonomous synthesis because 1-deoxyceramide production increased upon differentiation to adipocytes in mouse cell culture experiments. CONCLUSION: Taken together, the observed lipidome changes in obesity and T2D will facilitate the identification of T2D patient subgroups and represent an important step towards personalized medicine in diabetes.

Context Hyperparathyroidism is associated with hypercalcemia and the excess of parathyroid hormone secretion; however, the alterations in molecular pattern of functional genes during parathyroid tumorigenesis have not been unraveled. We aimed at establishing transcriptional patterns of normal and pathological parathyroid glands (PGs) in sporadic primary (HPT1) and secondary hyperparathyroidism (HPT2). Objective To evaluate dynamic alterations in molecular patterns as a function of the type of PG pathology, a comparative transcript analysis was conducted in subgroups of healthy samples, sporadic HPT1 adenoma and hyperplasia, and HPT2. Design Normal, adenomatous, HPT1, and HPT2 hyperplastic PG formalin-fixed paraffin-embedded samples were subjected to NanoString analysis. In silico microRNA (miRNA) analyses and messenger RNA–miRNA network in PG pathologies were conducted. Individual messenger RNA and miRNA levels were assessed in snap-frozen PG samples. Results The expression levels of c-MET, MYC, TIMP1, and clock genes NFIL3 and PER1 were significantly altered in HPT1 adenoma compared with normal PG tissue when assessed by NanoString and quantitative reverse transcription polymerase chain reaction. RET was affected in HPT1 hyperplasia, whereas CaSR and VDR transcripts were downregulated in HPT2 hyperplastic PG tissue. CDH1, c-MET, MYC, and CaSR were altered in adenoma compared with hyperplasia. Correlation analyses suggest that c-MET, MYC, and NFIL3 exhibit collective expression level changes associated with HPT1 adenoma development. miRNAs, predicted in silico to target these genes, did not exhibit a clear tendency upon experimental validation. Conclusions The presented gene expression analysis provides a differential molecular characterization of PG adenoma and hyperplasia pathologies, advancing our understanding of their etiology.

Background: The worldwide increase in obesity and type 2 diabetes (T2D) represents a major health challenge. Chronically altered lipids induced by obesity further promote the development of T2D, and the accumulation of toxic lipid metabolites in serum and peripheral organs may contribute to the diabetic phenotype. Methods: To better understand the complex metabolic pattern of lean and obese T2D and non-T2D individuals, we analysed the lipid profile of human serum, skeletal muscle and visceral adipose tissue of two cohorts by systematic mass spectrometry-based lipid analysis. Findings: Lipid homeostasis was strongly altered in a disease and tissue-specific manner, allowing us to define T2D signatures associated with obesity from those that were obesity-independent. Lipid changes encompassed lyso-, diacyl- and ether-phospholipids. Moreover, strong changes in sphingolipids included cytotoxic 1-deoxyceramide accumulation in a disease-specific manner in serum and visceral adipose tissue. Increased production of non-canonical 1-deoxyceramides in human adipose tissue was confirmed in adipocytes cell culture experiments. Interpretation: Taken together, the observed lipidome changes in obesity and T2D will facilitate the identification of T2D patient subgroups and represent an important step towards personalized medicine in diabetes. Funding Statement: This study was supported by the SNSF grants 31003A_166700/1 and 310030-184708, the Vontobel Foundation, the Olga Mayenfisch Foundation, the Novartis Foundation for Medical-Biological Research, the Fondation Ernst et Lucie Schmidheiny, the Jubilaumsstiftung Swiss Life Foundation, and the Fondation pour la Recherche sur le Cancer (C.D.); by SNSF grant CRSII3_154405 (H.R., C.D., E.L.); SNSF grant CRSII3_160741 (S.A.B., J.P, E.T.D); SNSF grant 310030B_166686 (H.R.), the NCCR Chemical Biology funded by the SNSF (H.R.), by the Bo & Kerstin Hjelt diabetes Foundation (U.L.-M.), and by the Young Independent Investigator Grant SGED/SSED (F. S.). Declaration of Interests: The authors declare no competing interests. Ethics Approval Statement: The study had ethics committee approval (CER11-015). The study conformed to the Declaration of Helsinki and the experimental protocol (‘DIOMEDE’) was approved by the Ethical Committee SUD EST IV (Agreement 12/111) and performed according to the French legislation (Huriet’s law).

PurposeHigh density lipoprotein (HDL) protects against myocardial infarction via mechanisms that remain unclear. STAT3 (signal transducer and activator of transcription 3) plays a key role in HDL-induced cardioprotection. In the heart, microRNAs (miRNAs) are involved in ischemia reperfusion injury. We therefore investigated whether the cardioprotective effect of HDL modulates miRNAs as a downstream target of STAT3 activation.MethodsSTAT3 cardiomyocyte deficient mice (STAT3-KO) and wildtype littermates (STAT3-WT) were submitted to left coronary ligature and reperfused (IR) with or without injection of HDL. Infarct size (IS) was determined and cardiac miRNA expression was evaluated after reperfusion in sham, IR and IR+HDL hearts by microarray analysis. In vitro, neonatal rat ventricular cardiomyocytes were submitted to hypoxia with or without HDL incubation. Cell viability and miRNA expression were analysed.ResultsIn vivo, HDL reduced IS from 40.5±4.3% to 24.4±2.1% (pConclusionsOur study, performed both in vivo and in vitro, delineates a novel cardioprotective signalling pathway activated by HDL, involving STAT3-mediated decrease of miR-34b and miR-337 expression.

Background: Chronic kidney disease (CKD) exacerbates the risk of death due to cardiovascular disease (CVD). Modifications to blood lipid metabolism which manifest as increases in circulating triglycerides and reductions in high-density lipoprotein (HDL) cholesterol are thought to contribute to increased risk. In CKD patients, higher HDL cholesterol levels were not associated with reduced mortality risk. Recent research has revealed numerous mechanisms by which HDL could favourably influence CVD risk. In this study, we compared plasma levels of sphingosine-1-phosphate (S1P), HDL-associated S1P (HDL-S1P) and HDL-mediated protection against oxidative stress between CKD and control patients. Methods: High-density lipoprotein was individually isolated from 20 CKD patients and 20 controls. Plasma S1P, apolipoprotein M (apoM) concentrations, HDL-S1P content and the capacity of HDL to protect cardiomyocytes against doxorubicin-induced oxidative stress in vitro were measured. Results: Chronic kidney disease patients showed a typical profile with significant reductions in plasma HDL cholesterol and albumin and an increase in triglycerides and pro-inflammatory cytokines (TNF-alpha and IL-6). Unexpectedly, HDL-S1P content (P = .001) and HDL cardioprotective capacity (P = .034) were increased significantly in CKD patients. Linear regression analysis of which factors could influence HDL-S1P content showed an independent, negative and positive association with plasma albumin and apoM levels, respectively. Discussion: The novel and unexpected observation in this study is that uremic HDL is more effective than control HDL for protecting cardiomyocytes against oxidative stress. It is explained by its higher S1P content which we previously demonstrated to be the determinant of HDL-mediated cardioprotective capacity. Interestingly, lower concentrations of albumin in CKD are associated with higher HDL-S1P. (Less)

Purpose: High-density lipoproteins (HDL) are atheroprotective. New evidence shows that HDL has widespread actions, including protection against cardiac ischemia reperfusion injury (IRI). This cardioprotective role has been attributed to several constituents of the HDL particle, including apolipoprotein A1 (apoA1) the major protein constituent and sphingosine-1-phosphate (S1P). However, the exact mechanisms of HDL-induced cardioprotection are still unknown and are under intensive investigation. A particular unresolved question remains the specific role of the HDL receptor, Scavenger Receptor BI (SR-BI), in the heart. The latter mediates many of the intracellular effects of HDL. The aim of this study was to evaluate the specific role of SR-BI in the protective impact of HDL on the heart. Methods and Results: The effects of HDL on the heart was evaluated in vitro using rat cultured neonatal cardiomyocytes and ex vivo in mice using the isolated heart Langendorff model of IRI (global no flow ischemia 35min, reperfusion 60min). In this model, HDL (400μg/mL) was injected during the first 7min of reperfusion and infarct size was assessed by triphenyltetrazolium chloride (TTC) staining. The specific role of SR-BI was investigated in vitro using specific siRNA facilitated knockdown and ex vivo using SR-BI knockout (SR-BI KO) mice. We confirmed that SR-BI is expressed in the cardiomyocytes and that HDL binds specifically to these cells. Treatment of cardiomyocytes with HDL induced the activation of several prosurvival signalling proteins, including Akt, STAT3 and ERK1/2 and protected the cells against oxidative stress induced by doxorubicin. Although specific binding of HDL was significantly reduced in SR-BI knockdown cardiomyocytes, activation of the prosurvival pathways was not affected. Similarly, SR-BI knockdown did not reduce the in vitro protective influence of HDL against oxidative stress induced by doxorubicin. Ex vivo, IRI induced an infarct size of 17.3±0.6% in isolated hearts of wildtype mice. HDL treatment during the first phase of reperfusion significantly reduced the infarct size by approximately 35% (p

Objective— The dyslipidemia of type 2 diabetes mellitus has multiple etiologies and impairs lipoprotein functionality, thereby increasing risk for cardiovascular disease. High-density lipoproteins (HDLs) have several beneficial effects, notably protecting the heart from myocardial ischemia. We hypothesized that glycation of HDL could compromise this cardioprotective effect. Approach and Results— We used in vitro (cardiomyocytes) and ex vivo (whole heart) models subjected to oxidative stress together with HDL isolated from diabetic patients and nondiabetic HDL glycated in vitro (methylglyoxal). Diabetic and in vitro glycated HDL were less effective ( P P P P P Conclusions— Our data demonstrate that glycation can reduce the S1P content of HDL, leading to increased cardiomyocyte cell death because of less effective activation of intracellular survival pathways. It has important implications for the functionality of HDL in diabetes mellitus because HDL-S1P has several beneficial effects on the vasculature.

Objective— To examine the contribution of the scavenger receptor (SR) BI to the mechanism by which high-density lipoprotein (HDL) acquires paraoxonase-1 (PON1). Methods and Results— Serum PON1 activity contributes to the antioxidant capacity of HDLs and is suggested to be an independent risk factor for atherosclerosis. The association of PON1 with HDL is a major determinant of its serum activity levels. PON1 secretion was studied in stably transfected Chinese hamster ovary and HepG2 models. Complementary analyses were performed in transgenic models. Modulation of SR-BI expression, by SR-BI small and interfering RNA knockdown and pharmacologically, correlated with significant changes ( P Conclusion— The present study identifies SR-BI as a major determinant of the capacity of HDL to acquire PON1. It reinforces the concept of the receptor as a docking molecule, allowing communication between HDL and the cell, and extends the importance of SR-BI to HDL metabolism and function.

Statins act by reducing hepatic cholesterol synthesis, thus stimulating uptake of serum cholesterol. Statin therapy modulates a number of genes involved in hepatic cholesterol homeostasis. These have rarely been analyzed simultaneously in the same experimental setting, with virtually no studies of primary human hepatocytes. This study analyzed the efficacy of rosuvastatin in the coordinated regulation of a number of genes implicated in cholesterol metabolism in primary human hepatocytes. Expression of five cholesterol-related genes were significantly upregulated, notably the Niemann-Pick C1 like 1 protein, for whom functional studies have been essentially limited to the intestine. Two genes were significantly downregulated, including sterol recognition element binding protein-1 gene that is implicated in control of hepatic lipogenesis. The results show the coordinated regulation of several genes implicated in hepatic cholesterol homeostasis and suggest therapeutic targets that could complement that clinical action of statins.

We recently observed that free fatty acids impair the stimulation of glucose transport into cardiomyocytes in response to either insulin or metabolic stress. In vivo, fatty acids for the myocardium are mostly obtained from triglyceride-rich lipoproteins (chylomicrons and Very Low-Density Lipoproteins). We therefore determined whether exposure of cardiac myocytes to VLDL resulted in impaired basal and stimulated glucose transport. Primary adult rat cardiac myocytes were chronically exposed to VLDL before glucose uptake was measured in response to insulin or metabolic stress, provoked by the mitochondrial ATP synthase inhibitor oligomycin. Exposure of cardiac myocytes to VLDL reduced both insulin-and oligomycin-stimulated glucose uptake. The reduction of glucose uptake was associated with a moderately reduced tyrosine phosphorylation of the insulin receptor. No reduction of the phosphorylation of the downstream effectors of insulin signaling Akt and AS160 was however observed. Similarly only a modest reduction of the activating phosphorylation of the AMP-activated kinase (AMPK) was observed in response to oligomycin. Similar to our previous observations with free fatty acids, inhibition of fatty acid oxidation restored oligomycin-stimulated glucose uptake. In conclusions, VLDL-derived fatty acids impair stimulated glucose transport in cardiac myocytes by a mechanism that seems to be mediated by a fatty acid oxidation intermediate. Thus, in the clinical context of the metabolic syndrome high VLDL may contribute to enhancement of ischemic injury by reduction of metabolic stress-stimulated glucose uptake.

OBJECTIVES The objective was to examine the hypothesis that modifications to paraoxonase-1 specific activity (SP, activity per unit mass peptide) could contribute to serum paraoxonase-1 status, a determinant of the clinical efficacy of the enzyme. METHODS Enzyme activities and concentrations were determined in a large population (n=912) of patients and controls. SP were subsequently examined as a function of paraoxonase-1 gene polymorphisms, plasma lipids and lipoproteins, and physiological and pathophysiological parameters. RESULTS Pathophysiological parameters (diabetes, metabolic syndrome, smoking, aging) did not promote variations in paraoxonase-1 SP, whilst coronary disease lowered SP (P

BACKGROUND AND AIMS: Reduced high density lipoproteins (HDL) and increased oxidative stress are features of type 2 diabetes. Myeloperoxidase is an oxidative enzyme partly associated with HDL and causing HDL dysfunction. It is an independent risk factor for cardiovascular disease. Paraoxonase-1 is an HDL-associated enzyme that protects against cardiovascular disease and is reduced in diabetes. The present study examined if serum myeloperoxidase was (i) increased in type 2 diabetes, (ii) correlated with paraoxonase-1 activity. METHODS AND RESULTS: The study was based on cross-sectional analyses of serum myeloperoxidase and paraoxonase-1 in type 2 diabetic patients and non-diabetic participants, with and without cardiovascular disease. Serum myeloperoxidase concentrations were not increased in type 2 diabetic patients without cardiovascular disease compared to non-diabetic controls. They were significantly higher in type 2 patients and non-diabetic patients with angiographically confirmed coronary disease. HDL-associated myeloperoxidase was correlated with serum myeloperoxidase (r=0.80, p

Polymorphisms of the gene for the antioxidant enzyme, paraoxonase-1 (PON1), have been identified as risk factors for coronary disease (CHD), notably in diabetic patients. The polymorphisms have also been linked with other diabetic complications. The present study analyzed glucose metabolism as a function of PON1 polymorphisms in young healthy nondiabetic men from families with premature CHD and matched controls. The L55M PON1 polymorphism was independently associated with the glucose response to an oral glucose tolerance test. LL homozygotes had significantly impaired glucose disposal (P = 0.0007) compared with (LM+MM) genotypes. It was particularly marked for subjects from high CHD risk families and differentiated them from matched controls (P = 0.049). The area under the glucose curve (P = 0.0036) and the time to peak glucose value (P = 0.026) were significantly higher in the LL carriers, whereas the insulin response was slower (P = 0.013). Insulin resistance did not differ between L55M genotypes. There was a trend for reduced pancreatic beta-cell function as measured by glucose-induced insulin secretion (LL vs. LM vs. MM, 20.26 vs. 23.74 vs. 25.60; P = 0.077). The frequency of the L55 allele decreased significantly (P = 0.028) across regions defining a north-south European axis. No significant differences for the glucose response or case-control populations were observed as a function of the PON1 Q192R polymorphism. The study demonstrates an association between PON1 gene polymorphisms and glucose metabolism. The L55M-glucose interaction differentiated offspring of high CHD risk families, suggesting that it may be of particular relevance for vascular disease and possibly other diabetic complications.