Ceramide analog C2-cer induces a loss in insulin sensitivity in muscle cells through the salvage/recycling pathway.

Ceramides have been shown to play a major role in the onset of skeletal muscle insulin resistance and therefore in the prevalence of type 2 diabetes. However, many of the studies involved in the discovery of deleterious ceramide actions used a nonphysiological, cell-permeable, short-chain ceramide analog, the C2-ceramide (C2-cer). In the present study, we determined how C2-cer promotes insulin resistance in muscle cells. We demonstrate that C2-cer enters the salvage/recycling pathway and becomes deacylated, yielding sphingosine, re-acylation of which depends on the availability of long chain fatty acids provided by the lipogenesis pathway in muscle cells. Importantly, we show these salvaged ceramides are actually responsible for the inhibition of insulin signaling induced by C2-cer. Interestingly, we also show that the exogenous and endogenous monounsaturated fatty acid oleate prevents C2-cer to be recycled into endogenous ceramide species in a diacylglycerol O-acyltransferase 1-dependent mechanism, which forces free fatty acid metabolism towards triacylglyceride production. Altogether, the study highlights for the first time that C2-cer induces a loss in insulin sensitivity through the salvage/recycling pathway in muscle cells. This study also validates C2-cer as a convenient tool to decipher mechanisms by which long-chain ceramides mediate insulin resistance in muscle cells and suggests that in addition to the de novo ceramide synthesis, recycling of ceramide could contribute to muscle insulin resistance observed in obesity and type 2 diabetes.

Fish oil minimises feed intake and improves insulin sensitivity in Zucker fa/fa rats.

Long-chain n-3 PUFA (LC n-3 PUFA) prevent, in rodents, insulin resistance (IR) induced by a high-fat and/or fructose diet but not IR induced by glucocorticoids. In humans, contrasting effects have also been reported. We investigated their effects on insulin sensitivity, feed intake (FI) and body weight gain in genetically insulin resistant male obese (fa/fa) Zucker (ZO) rats during the development of obesity. ZO rats were fed a diet supplemented with 7 % fish oil (FO) + 1 % corn oil (CO) (wt/wt) (ZOFO), while the control group was fed a diet containing 8 % fat from CO (wt/wt) (ZOCO). Male lean Zucker (ZL) rats fed either FO (ZLFO) or CO (ZLCO) diet were used as controls. FO was a marine-derived TAG oil containing EPA 90 mg/g + DHA 430 mg/g. During an oral glucose tolerance test, glucose tolerance remained unaltered by FO while insulin response was reduced in ZOFO only. Liver insulin sensitivity (euglycaemic-hyperinsulinaemic clamp + 2 deoxyglucose) was improved in ZOFO rats, linked to changes in phosphoenolpyruvate carboxykinase expression, activity and glucose-6-phosphatase activity. FI in response to intra-carotid insulin/glucose infusion was decreased similarly in ZOFO and ZOCO. Hypothalamic ceramides levels were lower in ZOFO than in ZOCO. Our study demonstrates that LC n-3 PUFA can minimise weight gain, possibly by alleviating hypothalamic lipotoxicity, and liver IR in genetically obese Zucker rats.

Circulating Sphingolipids and Glucose Homeostasis: An Update.

Sphingolipids are a family of lipid molecules produced through different pathways in mammals. Sphingolipids are structural components of membranes, but in response to obesity, they are implicated in the regulation of various cellular processes, including inflammation, apoptosis, cell proliferation, autophagy, and insulin resistance which favors dysregulation of glucose metabolism. Of all sphingolipids, two species, ceramides and sphingosine-1-phosphate (S1P), are also found abundantly secreted into the bloodstream and associated with lipoproteins or extracellular vesicles. Plasma concentrations of these sphingolipids can be altered upon metabolic disorders and could serve as predictive biomarkers of these diseases. Recent important advances suggest that circulating sphingolipids not only serve as biomarkers but could also serve as mediators in the dysregulation of glucose homeostasis. In this review, advances of molecular mechanisms involved in the regulation of ceramides and S1P association to lipoproteins or extracellular vesicles and how they could alter glucose metabolism are discussed.

Stearoyl CoA desaturase is a gatekeeper that protects human beta cells against lipotoxicity and maintains their identity.

AIMS/HYPOTHESIS: During the onset of type 2 diabetes, excessive dietary intake of saturated NEFA and fructose lead to impaired insulin production and secretion by insulin-producing pancreatic beta cells. The majority of data on the deleterious effects of lipids on functional beta cell mass were obtained either in vivo in rodent models or in vitro using rodent islets and beta cell lines. Translating data from rodent to human beta cells remains challenging. Here, we used the human beta cell line EndoC-ßH1 and analysed its sensitivity to a lipotoxic and glucolipotoxic (high palmitate with or without high glucose) insult, as a way to model human beta cells in a type 2 diabetes environment. METHODS: EndoC-ßH1 cells were exposed to palmitate after knockdown of genes related to saturated NEFA metabolism. We analysed whether and how palmitate induces apoptosis, stress and inflammation and modulates beta cell identity. RESULTS: EndoC-ßH1 cells were insensitive to the deleterious effects of saturated NEFA (palmitate and stearate) unless stearoyl CoA desaturase (SCD) was silenced. SCD was abundantly expressed in EndoC-ßH1 cells, as well as in human islets and human induced pluripotent stem cell-derived beta cells. SCD silencing induced markers of inflammation and endoplasmic reticulum stress and also IAPP mRNA. Treatment with the SCD products oleate or palmitoleate reversed inflammation and endoplasmic reticulum stress. Upon SCD knockdown, palmitate induced expression of dedifferentiation markers such as SOX9, MYC and HES1. Interestingly, SCD knockdown by itself disrupted beta cell identity with a decrease in mature beta cell markers INS, MAFA and SLC30A8 and decreased insulin content and glucose-stimulated insulin secretion. CONCLUSIONS/INTERPRETATION: The present study delineates an important role for SCD in the protection against lipotoxicity and in the maintenance of human beta cell identity. DATA AVAILABILITY: Microarray data and all experimental details that support the findings of this study have been deposited in in the GEO database with the GSE130208 accession code.

Sphingolipid Metabolism: New Insight into Ceramide-Induced Lipotoxicity in Muscle Cells.

Insulin-resistance is a characteristic feature of type 2 diabetes (T2D) and plays a major role in the pathogenesis of this disease. Skeletal muscles are quantitatively the biggest glucose users in response to insulin and are considered as main targets in development of insulin-resistance. It is now clear that circulating fatty acids (FA), which are highly increased in T2D, play a major role in the development of muscle insulin-resistance. In healthy individuals, excess FA are stored as lipid droplets in adipocytes. In situations like obesity and T2D, FA from lipolysis and food are in excess and eventually accumulate in peripheral tissues. High plasma concentrations of FA are generally associated with increased risk of developing diabetes. Indeed, ectopic fat accumulation is associated with insulin-resistance; this is called lipotoxicity. However, FA themselves are not involved in insulin-resistance, but rather some of their metabolic derivatives, such as ceramides. Ceramides, which are synthetized de novo from saturated FA like palmitate, have been demonstrated to play a critical role in the deterioration of insulin sensitivity in muscle cells. This review describes the latest progress involving ceramides as major players in the development of muscle insulin-resistance through the targeting of selective actors of the insulin signaling pathway.

Protective role of the ELOVL2/docosahexaenoic acid axis in glucolipotoxicity-induced apoptosis in rodent beta cells and human islets.

AIMS/HYPOTHESIS: Dietary n-3 polyunsaturated fatty acids, especially docosahexaenoic acid (DHA), are known to influence glucose homeostasis. We recently showed that Elovl2 expression in beta cells, which regulates synthesis of endogenous DHA, was associated with glucose tolerance and played a key role in insulin secretion. The present study aimed to examine the role of the very long chain fatty acid elongase 2 (ELOVL2)/DHA axis on the adverse effects of palmitate with high glucose, a condition defined as glucolipotoxicity, on beta cells. METHODS: We detected ELOVL2 in INS-1 beta cells and mouse and human islets using quantitative PCR and western blotting. Downregulation and adenoviral overexpression of Elovl2 was carried out in beta cells. Ceramide and diacylglycerol levels were determined by radio-enzymatic assay and lipidomics. Apoptosis was quantified using caspase-3 assays and poly (ADP-ribose) polymerase cleavage. Palmitate oxidation and esterification were determined by [U-14C]palmitate labelling. RESULTS: We found that glucolipotoxicity decreased ELOVL2 content in rodent and human beta cells. Downregulation of ELOVL2 drastically potentiated beta cell apoptosis induced by glucolipotoxicity, whereas adenoviral Elovl2 overexpression and supplementation with DHA partially inhibited glucolipotoxicity-induced cell death in rodent and human beta cells. Inhibition of beta cell apoptosis by the ELOVL2/DHA axis was associated with a decrease in ceramide accumulation. However, the ELOVL2/DHA axis was unable to directly alter ceramide synthesis or metabolism. By contrast, DHA increased palmitate oxidation but did not affect its esterification. Pharmacological inhibition of AMP-activated protein kinase and etomoxir, an inhibitor of carnitine palmitoyltransferase 1 (CPT1), the rate-limiting enzyme in fatty acid ß-oxidation, attenuated the protective effect of the ELOVL2/DHA axis during glucolipotoxicity. Downregulation of CPT1 also counteracted the anti-apoptotic action of the ELOVL2/DHA axis. By contrast, a mutated active form of Cpt1 inhibited glucolipotoxicity-induced beta cell apoptosis when ELOVL2 was downregulated. CONCLUSIONS/INTERPRETATION: Our results identify ELOVL2 as a critical pro-survival enzyme for preventing beta cell death and dysfunction induced by glucolipotoxicity, notably by favouring palmitate oxidation in mitochondria through a CPT1-dependent mechanism.

Lipoprotein lipase in hypothalamus is a key regulator of body weight gain and glucose homeostasis in mice.

AIMS/HYPOTHESIS: Regulation of energy balance involves the participation of many factors, including nutrients, among which are circulating lipids, acting as peripheral signals informing the central nervous system of the energy status of the organism. It has been shown that neuronal lipoprotein lipase (LPL) participates in the control of energy balance by hydrolysing lipid particles enriched in triacylglycerols. Here, we tested the hypothesis that LPL in the mediobasal hypothalamus (MBH), a well-known nucleus implicated in the regulation of metabolic homeostasis, could also contribute to the regulation of body weight and glucose homeostasis. METHODS: We injected an adeno-associated virus (AAV) expressing Cre-green fluorescent protein into the MBH of Lpl-floxed mice (and wild-type mice) to specifically decrease LPL activity in the MBH. In parallel, we injected an AAV overexpressing Lpl into the MBH of wild-type mice. We then studied energy homeostasis and hypothalamic ceramide content. RESULTS: The partial deletion of Lpl in the MBH in mice led to an increase in body weight compared with controls (37.72 ± 0.7 g vs 28.46 ± 0.12, p < 0.001) associated with a decrease in locomotor activity. These mice developed hyperinsulinaemia and glucose intolerance. This phenotype also displayed reduced expression of Cers1 in the hypothalamus as well as decreased concentration of several C18 species of ceramides and a 3-fold decrease in total ceramide intensity. Conversely, overexpression of Lpl specifically in the MBH induced a decrease in body weight. CONCLUSIONS/INTERPRETATION: Our study shows that LPL in the MBH is an important regulator of body weight and glucose homeostasis.

The suppression of hepatic glucose production improves metabolism and insulin sensitivity in subcutaneous adipose tissue in mice.

AIMS/HYPOTHESIS: Despite the strong correlation between non-alcoholic fatty liver disease and insulin resistance, hepatic steatosis is associated with greater whole-body insulin sensitivity in several models. We previously reported that the inhibition of hepatic glucose production (HGP) protects against the development of obesity and diabetes despite severe steatosis, thanks to the secretion of specific hepatokines such as fibroblast growth factor 21 (FGF21) and angiopoietin-related growth factor. In this work, we focused on adipose tissue to assess whether liver metabolic fluxes might, by interorgan communication, control insulin signalling in lean animals. METHODS: Insulin signalling was studied in the adipose tissue of mice lacking the catalytic subunit of glucose 6-phosphatase, the key enzyme in endogenous glucose production, in the liver (L-G6pc -/- mice). Morphological and metabolic changes in the adipose tissues were characterised by histological analyses, gene expression and protein content. RESULTS: Mice lacking HGP exhibited improved insulin sensitivity of the phosphoinositide 3-kinase/Akt pathway in the subcutaneous adipose tissue associated with a browning of adipocytes. The suppression of HGP increased FGF21 levels in lean animals, and increased FGF21 was responsible for the metabolic changes in the subcutaneous adipose tissue but not for its greater insulin sensitivity. The latter might be linked to an increase in the ratio of monounsaturated to saturated fatty acids released by the liver. CONCLUSIONS: Our work provides evidence that HGP controls subcutaneous adipose tissue browning and insulin sensitivity through two pathways: the release of beneficial hepatokines and changes in hepatic fatty acids profile.

Role of palmitate-induced sphingoid base-1-phosphate biosynthesis in INS-1 ß-cell survival.

Sphingoid base-1-phosphates represent a very low portion of the sphingolipid pool but are potent bioactive lipids in mammals. This study was undertaken to determine whether these lipids are produced in palmitate-treated pancreatic ß cells and what role they play in palmitate-induced ß cell apoptosis. Our lipidomic analysis revealed that palmitate at low and high glucose supplementation increased (dihydro)sphingosine-1-phosphate levels in INS-1 ß cells. This increase was associated with an increase in sphingosine kinase 1 (SphK1) mRNA and protein levels. Over-expression of SphK1 in INS-1 cells potentiated palmitate-induced accumulation of dihydrosphingosine-1-phosphate. N,N-dimethyl-sphingosine, a potent inhibitor of SphK, potentiated ß-cell apoptosis induced by palmitate whereas over-expression of SphK1 significantly reduced apoptosis induced by palmitate with high glucose. Endoplasmic reticulum (ER)-targeted SphK1 also partially inhibited apoptosis induced by palmitate. Inhibition of INS-1 apoptosis by over-expressed SphK1 was independent of sphingosine-1-phosphate receptors but was associated with a decreased formation of pro-apoptotic ceramides induced by gluco-lipotoxicity. Moreover, over-expression of SphK1 counteracted the defect in the ER-to-Golgi transport of proteins that contribute to the ceramide-dependent ER stress observed during gluco-lipotoxicity. In conclusion, our results suggest that activation of palmitate-induced SphK1-mediated sphingoid base-1-phosphate formation in the ER of ß cells plays a protective role against palmitate-induced ceramide-dependent apoptotic ß cell death.

Brain lipid sensing and the neural control of energy balance.

The central nervous system continuously detects circulating concentrations of lipids such as fatty acids and troglycerides. Once information has been detected, the central nervous system can in turn participate in the control of energy balance and blood sugar levels and in particular regulate the secretion and action of insulin. Neurons capable of detecting circulating lipid variations are located in the hypothalamus and in other regions such as the nucleus accumbens, the striatum or the hippocampus. An excess of lipids will have deleterious effects and may induce central lipotoxicity, in particular following local production of ceramides and the appearance of neuroinflammation which may lead to metabolic diseases such as obesity and type 2 diabetes.

Sphingosine-1-phosphate signalling in the heart: exploring emerging perspectives in cardiopathology.

Cardiometabolic disorders contribute to the global burden of cardiovascular diseases. Emerging sphingolipid metabolites like sphingosine-1-phosphate (S1P) and its receptors, S1PRs, present a dynamic signalling axis significantly impacting cardiac homeostasis. S1P's intricate mechanisms extend to its transportation in the bloodstream by two specific carriers: high-density lipoprotein particles and albumin. This intricate transport system ensures the accessibility of S1P to distant target tissues, influencing several physiological processes critical for cardiovascular health. This review delves into the diverse functions of S1P and S1PRs in both physiological and pathophysiological conditions of the heart. Emphasis is placed on their diverse roles in modulating cardiac health, spanning from cardiac contractility, angiogenesis, inflammation, atherosclerosis and myocardial infarction. The intricate interplays involving S1P and its receptors are analysed concerning different cardiac cell types, shedding light on their respective roles in different heart diseases. We also review the therapeutic applications of targeting S1P/S1PRs in cardiac diseases, considering existing drugs like Fingolimod, as well as the prospects and challenges in developing novel therapies that selectively modulate S1PRs.

Role of Sphingosine Kinase 1 in Glucolipotoxicity-Induced Early Activation of Autophagy in INS-1 Pancreatic ß Cells.

Insulin-producing pancreatic ß cells play a crucial role in the regulation of glucose homeostasis, and their failure is a key event for diabetes development. Prolonged exposure to palmitate in the presence of elevated glucose levels, termed gluco-lipotoxicity, is known to induce ß cell apoptosis. Autophagy has been proposed to be regulated by gluco-lipotoxicity in order to favor ß cell survival. However, the role of palmitate metabolism in gluco-lipotoxcity-induced autophagy is presently unknown. We therefore treated INS-1 cells for 6 and 24 h with palmitate in the presence of low and high glucose concentrations and then monitored autophagy. Gluco-lipotoxicity induces accumulation of LC3-II levels in INS-1 at 6 h which returns to basal levels at 24 h. Using the RFP-GFP-LC3 probe, gluco-lipotoxicity increased both autophagosomes and autolysosmes structures, reflecting early stimulation of an autophagy flux. Triacsin C, a potent inhibitor of the long fatty acid acetyl-coA synthase, completely prevents LC3-II formation and recruitment to autophagosomes, suggesting that autophagic response requires palmitate metabolism. In contrast, etomoxir and bromo-palmitate, inhibitors of fatty acid mitochondrial ß-oxidation, are unable to prevent gluco-lipotoxicity-induced LC3-II accumulation and recruitment to autophagosomes. Moreover, bromo-palmitate and etomoxir potentiate palmitate autophagic response. Even if gluco-lipotoxicity raised ceramide levels in INS-1 cells, ceramide synthase 4 overexpression does not potentiate LC3-II accumulation. Gluco-lipotoxicity also still stimulates an autophagic flux in the presence of an ER stress repressor. Finally, selective inhibition of sphingosine kinase 1 (SphK1) activity precludes gluco-lipotoxicity to induce LC3-II accumulation. Moreover, SphK1 overexpression potentiates autophagic flux induced by gluco-lipotxicity. Altogether, our results indicate that early activation of autophagy by gluco-lipotoxicity is mediated by SphK1, which plays a protective role in ß cells.

Interesterified palm oil promotes insulin resistance and altered insulin secretion and signaling in Swiss mice.

Interesterified fats have been used to replace trans-fat in ultra-processed foods. However, their metabolic effects are not completely understood. Hence, this study aimed to investigate the effects related to glucose homeostasis in response to interesterified palm oil or refined palm oil intake. Four-week-old male Swiss mice were randomly divided into four experimental groups and fed the following diets for 8 weeks: a normocaloric and normolipidic diet containing refined palm oil (PO group) or interesterified palm oil (IPO group); a hypercaloric and high-fat diet containing refined PO (POHF group) or interesterified PO (IPOHF group). Metabolic parameters related to body mass, adiposity and food consumption showed no significant differences. As for glucose homeostasis parameters, interesterified palm oil diets (IPO and IPOHF) resulted in higher glucose intolerance than unmodified palm oil diets (PO and POHF). Euglycemic-hyperinsulinemic clamp assessment showed a higher endogenous glucose production in the IPO group compared with the PO group. Moreover, the IPO group showed significantly lower p-AKT protein content (in the muscle and liver tissues) when compared with the PO group. Analysis of glucose-stimulated static insulin secretion (11.1 mmol/L glucose) in isolated pancreatic islets showed a higher insulin secretion in animals fed interesterified fat diets (IPO and IPOHF) than in those fed with palm oil (PO and POHF). Interesterified palm oil, including in normolipidic diets, can impair insulin signaling in peripheral tissues and increase insulin secretion by ß-cells, characterizing insulin resistance in mice.

Hepatic deletion of serine palmitoyl transferase 2 impairs ceramide/sphingomyelin balance, bile acids homeostasis and leads to liver damage in mice.

Ceramides (Cer) have been shown as lipotoxic inducers, which disturb numerous cell-signaling pathways, leading to metabolic disorders such as type 2 diabetes. In this study, we aimed to determine the role of de novo hepatic ceramide synthesis in energy and liver homeostasis in mice. We generated mice lacking serine palmitoyltransferase 2 (Sptlc2), the rate limiting enzyme of ceramide de novo synthesis, in liver under albumin promoter. Liver function, glucose homeostasis, bile acid (BA) metabolism and hepatic sphingolipids content were assessed using metabolic tests and LC-MS. Despite lower expression of hepatic Sptlc2, we observed an increased concentration of hepatic Cer, associated with a 10-fold increase in neutral sphingomyelinase 2 (nSMase2) expression, and a decreased sphingomyelin content in the liver. Sptlc2ΔLiv mice were protected against obesity induced by high fat diet and displayed a defect in lipid absorption. In addition, an important increase in tauro-muricholic acid was associated with a downregulation of the nuclear BA receptor FXR target genes. Sptlc2 deficiency also enhanced glucose tolerance and attenuated hepatic glucose production, while the latter effect was dampened in presence of nSMase2 inhibitor. Finally, Sptlc2 disruption promoted apoptosis, inflammation and progressive development of hepatic fibrosis, worsening with age. Our data suggest a compensatory mechanism to regulate hepatic ceramides content from sphingomyelin hydrolysis, with deleterious impact on liver homeostasis. In addition, our results show the involvement of hepatic sphingolipid modulation in BA metabolism and hepatic glucose production in an insulin-independent manner, which highlight the still under-researched role of ceramides in many metabolic functions.

Ceramide synthase 4 and de novo production of ceramides with specific N-acyl chain lengths are involved in glucolipotoxicity-induced apoptosis of INS-1 ß-cells.

Pancreatic ß-cell apoptosis induced by palmitate requires high glucose concentrations. Ceramides have been suggested to be important mediators of glucolipotoxicity-induced ß-cell apoptosis. In INS-1 ß-cells, 0.4 mM palmitate with 5 mM glucose increased the levels of dihydrosphingosine and dihydroceramides, two lipid intermediates in the de novo biosynthesis of ceramides, without inducing apoptosis. Increasing glucose concentrations to 30 mM amplified palmitate-induced accumulation of dihydrosphingosine and the formation of (dihydro)ceramides. Of note, glucolipotoxicity specifically induced the formation of C(18:0), C(22:0) and C(24:1) (dihydro)ceramide molecular species, which was associated with the up-regulation of CerS4 (ceramide synthase 4) levels. Fumonisin-B1, a ceramide synthase inhibitor, partially blocked apoptosis induced by glucolipotoxicity. In contrast, apoptosis was potentiated in the presence of D,L-threo-1-phenyl-2-palmitoylamino-3-morpholinopropan-1-ol, an inhibitor of glucosylceramide synthase. Moreover, overexpression of CerS4 amplified ceramide production and apoptosis induced by palmitate with 30 mM glucose, whereas down-regulation of CerS4 by siRNA (short interfering RNA) reduced apoptosis. CerS4 also potentiates ceramide accumulation and apoptosis induced by another saturated fatty acid: stearate. Collectively, our results suggest that glucolipotoxicity induces ß-cell apoptosis through a dual mechanism involving de novo ceramide biosynthesis and the formation of ceramides with specific N-acyl chain lengths rather than an overall increase in ceramide content.

Serum ceramides could predict durable diabetes remission following gastric bypass surgery.

Morbid obesity is a major risk factor for the development of type 2 diabetes (T2D). One strategy to both lose weight and counteract T2D is bariatric surgery (RYGB). In a study published in this issue of Med, Poss et al. revealed that circulating ceramides could predict the durability of T2D remission independently of weight loss following RYGB.

Homocysteine Metabolism Pathway Is Involved in the Control of Glucose Homeostasis: A Cystathionine Beta Synthase Deficiency Study in Mouse.

Cystathionine beta synthase (CBS) catalyzes the first step of the transsulfuration pathway from homocysteine to cystathionine, and its deficiency leads to hyperhomocysteinemia (HHcy) in humans and rodents. To date, scarce information is available about the HHcy effect on insulin secretion, and the link between CBS activity and the setting of type 2 diabetes is still unknown. We aimed to decipher the consequences of an inborn defect in CBS on glucose homeostasis in mice. We used a mouse model heterozygous for CBS (CBS+/-) that presented a mild HHcy. Other groups were supplemented with methionine in drinking water to increase the mild to intermediate HHcy, and were submitted to a high-fat diet (HFD). We measured the food intake, body weight gain, body composition, glucose homeostasis, plasma homocysteine level, and CBS activity. We evidenced a defect in the stimulated insulin secretion in CBS+/- mice with mild and intermediate HHcy, while mice with intermediate HHcy under HFD presented an improvement in insulin sensitivity that compensated for the decreased insulin secretion and permitted them to maintain a glucose tolerance similar to the CBS+/+ mice. Islets isolated from CBS+/- mice maintained their ability to respond to the elevated glucose levels, and we showed that a lower parasympathetic tone could, at least in part, be responsible for the insulin secretion defect. Our results emphasize the important role of Hcy metabolic enzymes in insulin secretion and overall glucose homeostasis.

cAMP-secretion coupling is impaired in diabetic GK/Par rat ß-cells: a defect counteracted by GLP-1.

cAMP-raising agents with glucagon-like peptide-1 (GLP-1) as the first in class, exhibit multiple actions that are beneficial for the treatment of type 2 diabetic (T2D) patients, including improvement of glucose-induced insulin secretion (GIIS). To gain additional insight into the role of cAMP in the disturbed stimulus-secretion coupling within the diabetic ß-cell, we examined more thoroughly the relationship between changes in islet cAMP concentration and insulin release in the GK/Par rat model of T2D. Basal cAMP content in GK/Par islets was significantly higher, whereas their basal insulin release was not significantly different from that of Wistar (W) islets. Even in the presence of IBMX or GLP-1, their insulin release did not significantly change despite further enhanced cAMP accumulation in both cases. The high basal cAMP level most likely reflects an increased cAMP generation in GK/Par compared with W islets since 1) forskolin dose-dependently induced an exaggerated cAMP accumulation; 2) adenylyl cyclase (AC)2, AC3, and G(s)α proteins were overexpressed; 3) IBMX-activated cAMP accumulation was less efficient and PDE-3B and PDE-1C mRNA were decreased. Moreover, the GK/Par insulin release apparatus appears less sensitive to cAMP, since GK/Par islets released less insulin at submaximal cAMP levels and required five times more cAMP to reach a maximal secretion rate no longer different from W. GLP-1 was able to reactivate GK/Par insulin secretion so that GIIS became indistinguishable from that of W. The exaggerated cAMP production is instrumental, since GLP-1-induced GIIS reactivation was lost in the presence the AC blocker 2',5'-dideoxyadenosine. This GLP-1 effect takes place in the absence of any improvement of the [Ca(2+)](i) response and correlates with activation of the cAMP-dependent PKA-dependent pathway.

Role of hypothalamic de novo ceramides synthesis in obesity and associated metabolic disorders.

BACKGROUND: Sphingolipid-mediated signalling pathways are described as important players in the normal functioning of neurons and nonneuronal cells in the central nervous system (CNS). SCOPE OF REVIEW: This review aims to show role of de novo ceramide synthesis in the CNS in controling key physiological processes, including food intake, energy expenditure, and thermogenesis. The corollary is a condition that leads to a dysfunction in ceramide metabolism in these central regions that can have major consequences on the physiological regulation of energy balance. MAJOR CONCLUSIONS: Excessive hypothalamic de novo ceramide synthesis has been shown to result in the establishment of central insulin resistance, endoplasmic reticulum stress, and inflammation. Additionally, excessive hypothalamic de novo ceramide synthesis has also been associated with changes in the activity of the autonomic nervous system. Such dysregulation of hypothalamic de novo ceramide synthesis forms the key starting point for the initiation of pathophysiological conditions such as obesity - which may or may not be associated with type 2 diabetes.

Ovarian insufficiency impairs glucose-stimulated insulin secretion through activation of hypothalamic de novo ceramide synthesis.

Oestrogens regulate body weight through their action on hypothalamus to modulate food intake and energy expenditure. Hypothalamic de novo ceramide synthesis plays a central role on obesity induced by oestrogen deficiency. Depletion in oestrogens is also known to be associated with glucose intolerance, which favours type 2 diabetes (T2D). However, the implication of hypothalamic ceramide in the regulation of glucose homeostasis by oestrogen is unknown. Here, we studied glucose homeostasis and insulin secretion in ovariectomized (OVX) female rats. OVX induces body weight gain associated with a hypothalamic inflammation and impaired glucose homeostasis. Genetic blockade of ceramide synthesis in the ventromedial nucleus of the hypothalamus (VMH) reverses hypothalamic inflammation and partly restored glucose tolerance induced by OVX. Furthermore, glucose-stimulated insulin secretion (GSIS) is increased in OVX rats due to a raise of insulin secretion second phase, a characteristic of early stage of T2D. In contrast, GSIS from isolated islets of OVX rats is totally blunted. Inhibition of ceramide synthesis in the VMH restores GSIS from isolated OVX islets and represses the second phase of insulin secretion. Stimulation of oestrogen receptor α (ERα) by oestradiol (E2) down-regulates ceramide synthesis in hypothalamic neuronal GT1-7 cells but no in microglial SIM-A9 cells. In contrast, genetic inactivation of ERα in VMH upregulates ceramide synthesis. These results indicate that hypothalamic neuronal de novo ceramide synthesis triggers the OVX-dependent impairment of glucose homeostasis which is partly mediated by a dysregulation of GSIS.