Reanalysis of mGWAS results and in vitro validation show that lactate dehydrogenase interacts with branched-chain amino acid metabolism.

The assignment of causative genes to noncoding variants identified in genome-wide association studies (GWASs) is challenging. We show how combination of knowledge from gene and pathway databases and chromatin interaction data leads to reinterpretation of published quantitative trait loci for blood metabolites. We describe a previously unidentified link between the rs2403254 locus, which is associated with the ratio of 3-methyl-2-oxobutanoate and alpha-hydroxyisovalerate levels, and the distal LDHA gene. We confirmed that lactate dehydrogenase can catalyze the conversion between these metabolites in vitro, suggesting that it has a role in branched-chain amino acid metabolism. Examining datasets from the ENCODE project we found evidence that the locus and LDHA promoter physically interact, showing that LDHA expression is likely under control of distal regulatory elements. Importantly, this discovery demonstrates that bioinformatic workflows for data integration can have a vital role in the interpretation of GWAS results.

Effects of low-stearate palm oil and high-stearate lard high-fat diets on rat liver lipid metabolism and glucose tolerance.

BACKGROUND: Excess consumption of energy-dense, high-fat Western diets contributes to the development of obesity and obesity-related disorders, such as fatty liver disease. However, not only the quantity but also the composition of dietary fat may play a role in the development of liver steatosis. The aim of this study was to determine the effects of low-stearate palm oil and high-stearate lard high-fat diets on in vivo liver lipid metabolism. METHODS: Wistar rats were fed with either normal chow (CON), a high-fat diet based on palm oil (HFP), or a high-fat diet based on lard (HFL). After 10 weeks of diet, magnetic resonance spectroscopy was applied for the in vivo determination of intrahepatocellular lipid content and the uptake and turnover of dietary fat after oral administration of (13)C-labeled lipids. Derangements in liver lipid metabolism were further assessed by measuring hepatic very-low density lipoprotein (VLDL) secretion and ex vivo respiratory capacity of liver mitochondria using fat-derived substrates. In addition, whole-body and hepatic glucose tolerance were determined with an intraperitoneal glucose tolerance test. RESULTS: Both high-fat diets induced liver lipid accumulation (p < 0.001), which was accompanied by a delayed uptake and/or slower turnover of dietary fat in the liver (p < 0.01), but without any change in VLDL secretion rates. Surprisingly, liver lipid content was higher in HFP than in HFL (p < 0.05), despite the increased fatty acid oxidative capacity in isolated liver mitochondria of HFP animals (p < 0.05). In contrast, while both high-fat diets induced whole-body glucose intolerance, only HFL impaired hepatic glucose tolerance. CONCLUSION: High-fat diets based on palm oil and lard similarly impair the handling of dietary lipids in the liver, but only the high-fat lard diet induces hepatic glucose intolerance.

FcRγ-chain deficiency reduces the development of diet-induced obesity.

OBJECTIVE: Pathogenic immunoglobulins are produced during the development of obesity and contribute to the development of insulin resistance (IR). However, the mechanisms by which these antibodies affect IR are largely unknown. This study investigated whether Fc-receptors contribute to the development of diet-induced obesity and IR by studying FcRγ(-/-) mice that lack the γ-subunit necessary for signaling and cell surface expression of FcγR and FcεRI. METHODS: FcRγ(-/-) and wild-type (WT) mice were fed a high-fat diet (HFD) to induce obesity. At 4 and 11 weeks, body weight and insulin sensitivity were measured, and adipose tissue (AT) inflammation was determined. Furthermore, intestinal triglyceride (TG) uptake and plasma TG clearance were determined, and gut microbiota composition was analyzed. RESULTS: FcRγ(-/-) mice gained less weight after 11 weeks of HFD. They had reduced adiposity, adipose tissue inflammation, and IR. Interestingly, FcRγ(-/-) mice had higher lean mass compared to WT mice, which was associated with increased energy expenditure. Intestinal TG absorption was increased whereas plasma TG clearance was not affected in FcRγ(-/-) mice. Gut microbial composition differed significantly and might therefore have added to the observed phenotype. CONCLUSIONS: FcRγ-chain deficiency reduces the development of diet-induced obesity, as well as associated AT inflammation and IR at 11 weeks of HFD.

Increased PUFA Content and 5-Lipoxygenase Pathway Expression Are Associated with Subcutaneous Adipose Tissue Inflammation in Obese Women with Type 2 Diabetes.

Obese women with type 2 diabetes mellitus (T2DM) have more inflammation in their subcutaneous white adipose tissue (sWAT) than age-and-BMI similar obese women with normal glucose tolerance (NGT). We aimed to investigate whether WAT fatty acids and/or oxylipins are associated with the enhanced inflammatory state in WAT of the T2DM women. Fatty acid profiles were measured in both subcutaneous and visceral adipose tissue (vWAT) of 19 obese women with NGT and 16 age-and-BMI similar women with T2DM. Oxylipin levels were measured in sWAT of all women. Arachidonic acid (AA) and docosahexaenoic acid (DHA) percentages were higher in sWAT, but not vWAT of the T2DM women, and AA correlated positively to the gene expression of macrophage marker CD68. We found tendencies for higher oxylipin concentrations of the 5-LOX leukotrienes in sWAT of T2DM women. Gene expression of the 5-LOX leukotriene biosynthesis pathway was significantly higher in sWAT of T2DM women. In conclusion, AA and DHA content were higher in sWAT of T2DM women and AA correlated to the increased inflammatory state in sWAT. Increased AA content was accompanied by an upregulation of the 5-LOX pathway and seems to have led to an increase in the conversion of AA into proinflammatory leukotrienes in sWAT.

Chronic helminth infection and helminth-derived egg antigens promote adipose tissue M2 macrophages and improve insulin sensitivity in obese mice.

Chronic low-grade inflammation associated with obesity contributes to insulin resistance and type 2 diabetes. Helminth parasites are the strongest natural inducers of type 2 immune responses, and short-lived infection with rodent nematodes was reported to improve glucose tolerance in obese mice. Here, we investigated the effects of chronic infection (12 weeks) with Schistosoma mansoni, a helminth that infects millions of humans worldwide, on whole-body metabolic homeostasis and white adipose tissue (WAT) immune cell composition in high-fat diet-induced obese C57BL/6 male mice. Our data indicate that chronic helminth infection reduced body weight gain (-62%), fat mass gain (-89%), and adipocyte size; lowered whole-body insulin resistance (-23%) and glucose intolerance (-16%); and improved peripheral glucose uptake (+25%) and WAT insulin sensitivity. Analysis of immune cell composition by flow cytometry and quantitative PCR (qPCR) revealed that S. mansoni promoted strong increases in WAT eosinophils and alternatively activated (M2) macrophages. Importantly, injections with S. mansoni-soluble egg antigens (SEA) recapitulated the beneficial effect of parasite infection on whole-body metabolic homeostasis and induced type 2 immune responses in WAT and liver. Taken together, we provide novel data suggesting that chronic helminth infection and helminth-derived molecules protect against metabolic disorders by promoting a T helper 2 (Th2) response, eosinophilia, and WAT M2 polarization.

Salsalate activates brown adipose tissue in mice.

Salsalate improves glucose intolerance and dyslipidemia in type 2 diabetes patients, but the mechanism is still unknown. The aim of the current study was to unravel the molecular mechanisms involved in these beneficial metabolic effects of salsalate by treating mice with salsalate during and after development of high-fat diet-induced obesity. We found that salsalate attenuated and reversed high-fat diet-induced weight gain, in particular fat mass accumulation, improved glucose tolerance, and lowered plasma triglyceride levels. Mechanistically, salsalate selectively promoted the uptake of fatty acids from glycerol tri[(3)H]oleate-labeled lipoprotein-like emulsion particles by brown adipose tissue (BAT), decreased the intracellular lipid content in BAT, and increased rectal temperature, all pointing to more active BAT. The treatment of differentiated T37i brown adipocytes with salsalate increased uncoupled respiration. Moreover, salsalate upregulated Ucp1 expression and enhanced glycerol release, a dual effect that was abolished by the inhibition of cAMP-dependent protein kinase (PKA). In conclusion, salsalate activates BAT, presumably by directly activating brown adipocytes via the PKA pathway, suggesting a novel mechanism that may explain its beneficial metabolic effects in type 2 diabetes patients.

Prolonged niacin treatment leads to increased adipose tissue PUFA synthesis and anti-inflammatory lipid and oxylipin plasma profile.

Prolonged niacin treatment elicits beneficial effects on the plasma lipid and lipoprotein profile that is associated with a protective CVD risk profile. Acute niacin treatment inhibits nonesterified fatty acid release from adipocytes and stimulates prostaglandin release from skin Langerhans cells, but the acute effects diminish upon prolonged treatment, while the beneficial effects remain. To gain insight in the prolonged effects of niacin on lipid metabolism in adipocytes, we used a mouse model with a human-like lipoprotein metabolism and drug response [female APOE*3-Leiden.CETP (apoE3 Leiden cholesteryl ester transfer protein) mice] treated with and without niacin for 15 weeks. The gene expression profile of gonadal white adipose tissue (gWAT) from niacin-treated mice showed an upregulation of the "biosynthesis of unsaturated fatty acids" pathway, which was corroborated by quantitative PCR and analysis of the FA ratios in gWAT. Also, adipocytes from niacin-treated mice secreted more of the PUFA DHA ex vivo. This resulted in an increased DHA/arachidonic acid (AA) ratio in the adipocyte FA secretion profile and in plasma of niacin-treated mice. Interestingly, the DHA metabolite 19,20-dihydroxy docosapentaenoic acid (19,20-diHDPA) was increased in plasma of niacin-treated mice. Both an increased DHA/AA ratio and increased 19,20-diHDPA are indicative for an anti-inflammatory profile and may indirectly contribute to the atheroprotective lipid and lipoprotein profile associated with prolonged niacin treatment.

Long-term niacin treatment induces insulin resistance and adrenergic responsiveness in adipocytes by adaptive downregulation of phosphodiesterase 3B.

The lipid-lowering effect of niacin has been attributed to the inhibition of cAMP production in adipocytes, thereby inhibiting intracellular lipolysis and release of nonesterified fatty acids (NEFA) to the circulation. However, long-term niacin treatment leads to a normalization of plasma NEFA levels and induces insulin resistance, for which the underlying mechanisms are poorly understood. The current study addressed the effects of long-term niacin treatment on insulin-mediated inhibition of adipocyte lipolysis and focused on the regulation of cAMP levels. APOE*3-Leiden.CETP transgenic mice treated with niacin for 15 wk were subjected to an insulin tolerance test and showed whole body insulin resistance. Similarly, adipocytes isolated from niacin-treated mice were insulin resistant and, interestingly, exhibited an increased response to cAMP stimulation by 8Br-cAMP, ß1- and ß2-adrenergic stimulation. Gene expression analysis of the insulin and ß-adrenergic pathways in adipose tissue indicated that all genes were downregulated, including the gene encoding the cAMP-degrading enzyme phosphodiesterase 3B (PDE3B). In line with this, we showed that insulin induced a lower PDE3B response in adipocytes isolated from niacin-treated mice. Inhibiting PDE3B with cilostazol increased lipolytic responsiveness to cAMP stimulation in adipocytes. These data show that long-term niacin treatment leads to a downregulation of PDE3B in adipocytes, which could explain part of the observed insulin resistance and the increased responsiveness to cAMP stimulation.

Niacin Reduces Atherosclerosis Development in APOE*3Leiden.CETP Mice Mainly by Reducing NonHDL-Cholesterol.

OBJECTIVE: Niacin potently lowers triglycerides, mildly decreases LDL-cholesterol, and largely increases HDL-cholesterol. Despite evidence for an atheroprotective effect of niacin from previous small clinical studies, the large outcome trials, AIM-HIGH and HPS2-THRIVE did not reveal additional beneficial effects of niacin (alone or in combination with laropiprant) on top of statin treatment. We aimed to address this apparent discrepancy by investigating the effects of niacin without and with simvastatin on atherosclerosis development and determine the underlying mechanisms, in APOE*3Leiden.CETP mice, a model for familial dysbetalipoproteinemia (FD). APPROACH AND RESULTS: Mice were fed a western-type diet containing cholesterol without or with niacin (120 mg/kg/day), simvastatin (36 mg/kg/day) or their combination for 18 weeks. Similarly as in FD patients, niacin reduced total cholesterol by -39% and triglycerides by -50%, (both P<0.001). Simvastatin and the combination reduced total cholesterol (-30%; -55%, P<0.001) where the combination revealed a greater reduction compared to simvastatin (-36%, P<0.001). Niacin decreased total cholesterol and triglycerides primarily by increasing VLDL clearance. Niacin increased HDL-cholesterol (+28%, P<0.01) and mildly increased reverse cholesterol transport. All treatments reduced monocyte adhesion to the endothelium (-46%; -47%, P<0.01; -53%, P<0.001), atherosclerotic lesion area (-78%; -49%, P<0.01; -87%, P<0.001) and severity. Compared to simvastatin, the combination increased plaque stability index [(SMC+collagen)/macrophages] (3-fold, P<0.01). Niacin and the combination reduced T cells in the aortic root (-71%, P<0.01; -81%, P<0.001). Lesion area was strongly predicted by nonHDL-cholesterol (R(2) = 0.69, P<0.001) and to a much lesser extent by HDL-cholesterol (R(2) = 0.20, P<0.001). CONCLUSION: Niacin decreases atherosclerosis development mainly by reducing nonHDL-cholesterol with modest HDL-cholesterol-raising and additional anti-inflammatory effects. The additive effect of niacin on top of simvastatin is mostly dependent on its nonHDL-cholesterol-lowering capacities. These data suggest that clinical beneficial effects of niacin are largely dependent on its ability to lower LDL-cholesterol on top of concomitant lipid-lowering therapy.

Apolipoprotein A5 deficiency aggravates high-fat diet-induced obesity due to impaired central regulation of food intake.

Mutations in apolipoprotein A5 (APOA5) have been associated with hypertriglyceridemia in humans and mice. This has been attributed to a stimulating role for APOA5 in lipoprotein lipase-mediated triglyceride hydrolysis and hepatic clearance of lipoprotein remnant particles. However, because of the low APOA5 plasma abundance, we investigated an additional signaling role for APOA5 in high-fat diet (HFD)-induced obesity. Wild-type (WT) and Apoa5(-/-) mice fed a chow diet showed no difference in body weight or 24-h food intake (Apoa5(-/-), 4.5±0.6 g; WT, 4.2±0.5 g), while Apoa5(-/-) mice fed an HFD ate more in 24 h (Apoa5(-/-), 2.8±0.4 g; WT, 2.5±0.3 g, P<0.05) and became more obese than WT mice. Also, intravenous injection of APOA5-loaded VLDL-like particles lowered food intake (VLDL control, 0.26±0.04 g; VLDL+APOA5, 0.11±0.07 g, P<0.01). In addition, the HFD-induced hyperphagia of Apoa5(-/-) mice was prevented by adenovirus-mediated hepatic overexpression of APOA5. Finally, intracerebroventricular injection of APOA5 reduced food intake compared to injection of the same mouse with artificial cerebral spinal fluid (0.40±0.11 g; APOA5, 0.23±0.08 g, P<0.01). These data indicate that the increased HFD-induced obesity of Apoa5(-/-) mice as compared to WT mice is at least partly explained by hyperphagia and that APOA5 plays a role in the central regulation of food intake.

Protein-induced geometric constraints and charge transfer in bacteriochlorophyll-histidine complexes in LH2.

Bacteriochlorophyll-histidine complexes are ubiquitous in nature and are essential structural motifs supporting the conversion of solar energy into chemically useful compounds in a wide range of photosynthesis processes. A systematic density functional theory study of the NMR chemical shifts for histidine and for bacteriochlorophyll-a-histidine complexes in the light-harvesting complex II (LH2) is performed using the BLYP functional in combination with the 6-311++G(d,p) basis set. The computed chemical shift patterns are consistent with available experimental data for positive and neutral(tau) (N(tau) protonated) crystalline histidines. The results for the bacteriochlorophyll-a-histidine complexes in LH2 provide evidence that the protein environment is stabilizing the histidine close to the Mg ion, thereby inducing a large charge transfer of approximately 0.5 electronic equivalent. Due to this protein-induced geometric constraint, the Mg-coordinated histidine in LH2 appears to be in a frustrated state very different from the formal neutral(pi) (N(pi) protonated) form. This finding could be important for the understanding of basic functional mechanisms involved in tuning the electronic properties and exciton coupling in LH2.