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Contains fulltext : 282360.pdf (Publisher’s version ) (Open Access), Triglycerides are carried in the bloodstream as part of very low-density lipoproteins (VLDLs) and chylomicrons, which represent the triglyceride-rich lipoproteins. Triglyceride-rich lipoproteins and their remnants contribute to atherosclerosis, possibly by carrying remnant cholesterol and/or by exerting a proinflammatory effect on macrophages. Nevertheless, little is known about how macrophages process triglyceride-rich lipoproteins. Here, using VLDL-sized triglyceride-rich emulsion particles, we aimed to study the mechanism by which VLDL triglycerides are taken up, processed, and stored in macrophages. Our results show that macrophage uptake of VLDL-sized emulsion particles is dependent on lipoprotein lipase (LPL) and requires the lipoprotein-binding C-terminal domain but not the catalytic N-terminal domain of LPL. Subsequent internalization of VLDL-sized emulsion particles by macrophages is carried out by caveolae-mediated endocytosis, followed by triglyceride hydrolysis catalyzed by lysosomal acid lipase. It is shown that STARD3 is required for the transfer of lysosomal fatty acids to the ER for subsequent storage as triglycerides, while NPC1 likely is involved in promoting the extracellular efflux of fatty acids from lysosomes. Our data provide novel insights into how macrophages process VLDL triglycerides and suggest that macrophages have the remarkable capacity to excrete part of the internalized triglycerides as fatty acids.

Contains fulltext : 189879.pdf (publisher's version ) (Open Access)

Contains fulltext : 197966.pdf (publisher's version ) (Open Access)

Contains fulltext : 174075.pdf (publisher's version ) (Open Access), AIMS/HYPOTHESIS: Obesity induces macrophages to drive inflammation in adipose tissue, a crucial step towards the development of type 2 diabetes. The tricarboxylic acid (TCA) cycle intermediate succinate is released from cells under metabolic stress and has recently emerged as a metabolic signal induced by proinflammatory stimuli. We therefore investigated whether succinate receptor 1 (SUCNR1) could play a role in the development of adipose tissue inflammation and type 2 diabetes. METHODS: Succinate levels were determined in human plasma samples from individuals with type 2 diabetes and non-diabetic participants. Succinate release from adipose tissue explants was studied. Sucnr1 -/- and wild-type (WT) littermate mice were fed a high-fat diet (HFD) or low-fat diet (LFD) for 16 weeks. Serum metabolic variables, adipose tissue inflammation, macrophage migration and glucose tolerance were determined. RESULTS: We show that hypoxia and hyperglycaemia independently drive the release of succinate from mouse adipose tissue (17-fold and up to 18-fold, respectively) and that plasma levels of succinate were higher in participants with type 2 diabetes compared with non-diabetic individuals (+53%; p < 0.01). Sucnr1 -/- mice had significantly reduced numbers of macrophages (0.56 +/- 0.07 vs 0.92 +/- 0.15 F4/80 cells/adipocytes, p < 0.05) and crown-like structures (0.06 +/- 0.02 vs 0.14 +/- 0.02, CLS/adipocytes p < 0.01) in adipose tissue and significantly improved glucose tolerance (p < 0.001) compared with WT mice fed an HFD, despite similarly increased body weights. Consistently, macrophages from Sucnr1 -/- mice showed reduced chemotaxis towards medium collected from apoptotic and hypoxic adipocytes (-59%; p < 0.05). CONCLUSIONS/INTERPRETATION: Our results reveal that activation of SUCNR1 in macrophages is important for both infiltration and inflammation of adipose tissue in obesity, and suggest that SUCNR1 is a promising therapeutic target in obesity-induced type 2 diabetes. DAT

Contains fulltext : 170968.pdf (publisher's version ) (Closed access), Microbial stimuli such as lipopolysaccharide (LPS) induce robust metabolic rewiring in immune cells known as the Warburg effect. It is unknown whether this increase in glycolysis and decrease in oxidative phosphorylation (OXPHOS) is a general characteristic of monocytes that have encountered a pathogen. Using CD14+ monocytes from healthy donors, we demonstrated that most microbial stimuli increased glycolysis, but that only stimulation of Toll-like receptor (TLR) 4 with LPS led to a decrease in OXPHOS. Instead, activation of other TLRs, such as TLR2 activation by Pam3CysSK4 (P3C), increased oxygen consumption and mitochondrial enzyme activity. Transcriptome and metabolome analysis of monocytes stimulated with P3C versus LPS confirmed the divergent metabolic responses between both stimuli, and revealed significant differences in the tricarboxylic acid cycle, OXPHOS and lipid metabolism pathways following stimulation of monocytes with P3C versus LPS. At a functional level, pharmacological inhibition of complex I of the mitochondrial electron transport chain diminished cytokine production and phagocytosis in P3C- but not LPS-stimulated monocytes. Thus, unlike LPS, complex microbial stimuli and the TLR2 ligand P3C induce a specific pattern of metabolic rewiring that involves upregulation of both glycolysis and OXPHOS, which enables activation of host defence mechanisms such as cytokine production and phagocytosis.

Item does not contain fulltext, BACKGROUND & AIMS: Peroxisome proliferator-activated receptor alpha (PPARalpha) is a key regulator of hepatic fat oxidation that serves as an energy source during starvation. Vanin-1 has been described as a putative PPARalpha target gene in liver, but its function in hepatic lipid metabolism is unknown. METHODS: We investigated the regulation of vanin-1, and total vanin activity, by PPARalpha in mice and humans. Furthermore, the function of vanin-1 in the development of hepatic steatosis in response to starvation was examined in Vnn1 deficient mice, and in rats treated with an inhibitor of vanin activity. RESULTS: Liver microarray analyses reveals that Vnn1 is the most prominently regulated gene after modulation of PPARalpha activity. In addition, activation of mouse PPARalpha regulates hepatic- and plasma vanin activity. In humans, consistent with regulation by PPARalpha, plasma vanin activity increases in all subjects after prolonged fasting, as well as after treatment with the PPARalpha agonist fenofibrate. In mice, absence of vanin-1 exacerbates the fasting-induced increase in hepatic triglyceride levels. Similarly, inhibition of vanin activity in rats induces accumulation of hepatic triglycerides upon fasting. Microarray analysis reveal that the absence of vanin-1 associates with gene sets involved in liver steatosis, and reduces pathways involved in oxidative stress and inflammation. CONCLUSIONS: We show that hepatic vanin-1 is under extremely sensitive regulation by PPARalpha and that plasma vanin activity could serve as a readout of changes in PPARalpha activity in human subjects. In addition, our data propose a role for vanin-1 in regulation of hepatic TG levels during fasting.

BACKGROUND & AIMS: Peroxisome proliferator-activated receptor alpha (PPARalpha) is a key regulator of hepatic fat oxidation that serves as an energy source during starvation. Vanin-1 has been described as a putative PPARalpha target gene in liver, but its function in hepatic lipid metabolism is unknown. METHODS: We investigated the regulation of vanin-1, and total vanin activity, by PPARalpha in mice and humans. Furthermore, the function of vanin-1 in the development of hepatic steatosis in response to starvation was examined in Vnn1 deficient mice, and in rats treated with an inhibitor of vanin activity. RESULTS: Liver microarray analyses reveals that Vnn1 is the most prominently regulated gene after modulation of PPARalpha activity. In addition, activation of mouse PPARalpha regulates hepatic- and plasma vanin activity. In humans, consistent with regulation by PPARalpha, plasma vanin activity increases in all subjects after prolonged fasting, as well as after treatment with the PPARalpha agonist fenofibrate. In mice, absence of vanin-1 exacerbates the fasting-induced increase in hepatic triglyceride levels. Similarly, inhibition of vanin activity in rats induces accumulation of hepatic triglycerides upon fasting. Microarray analysis reveal that the absence of vanin-1 associates with gene sets involved in liver steatosis, and reduces pathways involved in oxidative stress and inflammation. CONCLUSIONS: We show that hepatic vanin-1 is under extremely sensitive regulation by PPARalpha and that plasma vanin activity could serve as a readout of changes in PPARalpha activity in human subjects. In addition, our data propose a role for vanin-1 in regulation of hepatic TG levels during fasting.

Contains fulltext : 118107.pdf (Publisher’s version ) (Open Access), Caspase-1 is known to activate the proinflammatory cytokines IL-1beta and IL-18. Additionally, it can cleave other substrates, including proteins involved in metabolism. Recently, we showed that caspase-1 deficiency in mice strongly reduces high-fat diet-induced weight gain, at least partly caused by an increased energy production. Increased feces secretion by caspase-1-deficient mice suggests that lipid malabsorption possibly further reduces adipose tissue mass. In this study we investigated whether caspase-1 plays a role in triglyceride-(TG)-rich lipoprotein metabolism using caspase-1-deficient and wild-type mice. Caspase-1 deficiency reduced the postprandial TG response to an oral lipid load, whereas TG-derived fatty acid (FA) uptake by peripheral tissues was not affected, demonstrated by unaltered kinetics of [(3)H]TG-labeled very low-density lipoprotein (VLDL)-like emulsion particles. An oral gavage of [(3)H]TG-containing olive oil revealed that caspase-1 deficiency reduced TG absorption and subsequent uptake of TG-derived FA in liver, muscle, and adipose tissue. Similarly, despite an elevated hepatic TG content, caspase-1 deficiency reduced hepatic VLDL-TG production. Intestinal and hepatic gene expression analysis revealed that caspase-1 deficiency did not affect FA oxidation or FA uptake but rather reduced intracellular FA transport, thereby limiting lipid availability for the assembly and secretion of TG-rich lipoproteins. The current study reveals a novel function for caspase-1, or caspase-1-cleaved substrates, in controlling intestinal TG absorption and hepatic TG secretion.

Legumes have the unique ability to host nitrogen-fixing Rhizobium bacteria as symbiosomes inside root nodule cells. To get insight into this key process, which forms the heart of the endosymbiosis, we isolated specific cells/tissues at different stages of symbiosome formation from nodules of the model legume Medicago truncatula using laser-capture microdissection. Next, we determined their associated expression profiles using Affymetrix Medicago GeneChips. Cells were collected from the nodule infection zone divided into a distal (where symbiosome formation and division occur) and proximal region (where symbiosomes are mainly differentiating), as well as infected cells from the fixation zone containing mature nitrogen fixing symbiosomes. As non-infected cells/tissue we included nodule meristem cells and uninfected cells from the fixation zone. Here, we present a comprehensive gene expression map of an indeterminate Medicago nodule and selected genes that show specific enriched expression in the different cells or tissues. Validation of the obtained expression profiles, by comparison to published gene expression profiles and experimental verification, indicates that the data can be used as digital “in situ”. This digital “in situ” offers a genome-wide insight into genes specifically associated with subsequent stages of symbiosome and nodule cell development, and can serve to guide future functional studies.

Item does not contain fulltext, Oncoprotein-induced transcript 1 (Oit1) was previously identified as a dietary fat-induced gene in the small intestine of C57Bl/6J mice. In this study, we further characterized Oit1 and its human ortholog family with sequence similarity 3, member D (Fam3D), on the messenger RNA as well as the protein level. Oit1 and Fam3D were found to be predominantly expressed in the gastrointestinal tract of mice and humans, respectively. Dietary fat induced a clear and acute up-regulation of Oit1, especially in the jejunum, whereas fasting led to a reduced gene expression in the small intestine. Regarding protein expression, we found a remarkable pattern of Oit1 along the longitudinal axis of the intestine, a predominant villus-restricted expression in the proximal small intestine and a more pronounced crypt expression in the distal parts of the intestine. Using transfection experiments, we confirmed secretion of the Oit1 protein, as was predicted by a signal peptide sequence. Detection of Oit1 and Fam3D in plasma samples indicated that both proteins are secreted to the basolateral site of enterocytes. Moreover, in human plasma samples, we also found an effect of nutritional status on Fam3D levels, with a postprandial elevation and a reduction after fasting. In conclusion, Oit1 and Fam3D are gut-derived proteins that are expressed and secreted in a nutritional status-dependent manner.

Contains fulltext : 97039.pdf (publisher's version ) (Closed access), Inflammation plays a key role in the pathogenesis of obesity. Chronic overfeeding leads to macrophage infiltration in the adipose tissue, resulting in proinflammatory cytokine production. Both microbial and endogenous danger signals trigger assembly of the intracellular innate immune sensor Nlrp3, resulting in caspase-1 activation and production of proinflammatory cytokines IL-1beta and IL-18. Here, we showed that mice deficient in Nlrp3, apoptosis-associated speck-like protein, and caspase-1 were resistant to the development of high-fat diet-induced obesity, which correlated with protection from obesity-induced insulin resistance. Furthermore, hepatic triglyceride content, adipocyte size, and macrophage infiltration in adipose tissue were all reduced in mice deficient in inflammasome components. Monocyte chemoattractant protein (MCP)-1 is a key molecule that mediates macrophage infiltration. Indeed, defective inflammasome activation was associated with reduced MCP-1 production in adipose tissue. Furthermore, plasma leptin and resistin that affect energy use and insulin sensitivity were also changed by inflammasome-deficiency. Detailed metabolic and molecular phenotyping demonstrated that the inflammasome controls energy expenditure and adipogenic gene expression during chronic overfeeding. These findings reveal a critical function of the inflammasome in obesity and insulin resistance, and suggest inhibition of the inflammasome as a potential therapeutic strategy.