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Impaired insulin signalling leads to decreased expression of GPIHBP1
Abstracts / Atherosclerosis 241 (2015) e32ee71
duodenal intralipid infusion. The lymphatic HDL was isolated; associated lipid, protein and miRNA composition analyzed. Jejunal explants from IR rats were treated with niacin or nicotinamide and mRNA expression was assessed. Results: In IR rats apo-AI lymphatic HDL secretion was reduced (47%), but associated TG content enriched (86%), compared to non-IR rats. Niacin was found to stimulate the secretion of lymph HDL (>60%) compared to nontreated IR rats. Niacin treatment normalized the TG content of lymphatic HDL particles. Niacin but not nicotinamide increased Ppara and Cpt1a mRNA, and annulled Tnf alpha mRNA in jejunal explants. Top three regulated miR's by niacin in the lymph HDL fraction were: rno-miR-223-3p, rno-miR-29c-3p and rno-miR93-5p. Analysis of these miR's (IPA software) showed PPAR signalling to be amongst the significantly predicted pathways. Conclusion: IR reduces the secretion of apoAI HDL into mesenteric lymph, whilst niacin may contribute to improving CVD risk by restoring the number and TG enrichment of intestinal apoAI particles. The altered miRNA profile in response to niacin treatment might be involved in exerting hypolipidemic effects of niacin.
EAS-0505. ARGININE-DIRECTED GLYCATION BY METHYLGLYOXAL IS A FACTOR DECREASING HDL PLASMA CONCENTRATION AND FUNCTIONALITY IN VIVO N. Rabbani, P.J. Thornalley. Warwick Medical School, University Hospitals Coventry and Warwickshire NHS Trust, Coventry, United Kingdom Aim: The aim of this study was to quantify the modification of HDL by the glucose-derived reactive metabolite, methylglyoxal, in healthy people and patients with type 2 diabetes (T2DM), characterise structural, functional and physiological consequences of the modification and predict their importance for risk of cardiovascular disease. Methods: Major fractions of HDL were isolated from healthy human subjects and patients with T2DM and fractions modified by methylglyoxal and related dicarbonyl metabolites quantified. HDL fractions were glycated by methylglyoxal to minimum extent in vitro and molecular, functional and physiological characteristics determined. A one compartment model of HDL plasma clearance was produced including formation and clearance of dicarbonyl modified HDL. Results: HDL modified by methylglyoxal and related dicarbonyl metabolites accountedfor 2.6% HDL and increased to 4.5% in patients with T2DM. Methylglyoxal modification in vitro induced re-structuring of the HDL particles, decreasing stability and plasma half-life in vivo. It occurred at sites of apolipoprotein A-1 linked to membrane fusion, intramolecular bonding and ligand binding. Kinetic modelling of methylglyoxal modification of HDL predicted a negative correlation of plasma HDL-C with methylglyoxal-modified HDL. This was validated clinically. It also predicted dicarbonyl modification produces 2 e 6% decrease in total plasma HDL and 5 -13% decrease in functional HDL clinically. Conclusions: Methylglyoxal modification of apolipoprotein A-1 may accelerate HDL degradation and impairs its functionality in vivo, likely contributing to increased risk of cardiovascular disease. Quantitation of methylglyoxal-modified HDL may improve cardiovascular disease risk models and therapeutics to decrease methylglyoxal may improve preventive treatment of cardiovascular disease.
EAS-0555. IMPAIRED INSULIN SIGNALLING LEADS TO DECREASED EXPRESSION OF GPIHBP1 R.P. Surendran 1, M. Clemente-Postigo 2, R.S. Kootte 3, M.M. van Eijk 4, L. Garrido-Sanchez 2, F. Cardona 2, F. Tinahones 2, M. Nieuwdorp 3, G. Dallinga-Thie 1. 1 Experimental Vascular Medicine, Academic Medical Center University of Amsterdam, Amsterdam, The Netherlands; 2 Unidad de n Clínica Endocrinología y Nutricio n, Instituto de Investigacio n Gestio laga (IBIMA) Complejo Hospitalario de Ma laga, Malaga, Biom edica de Ma Spain; 3 Vascular Medicine, Academic Medical Center University of
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Amsterdam, Amsterdam, The Netherlands; 4 Medical Biochemistry, Academic Medical Center University of Amsterdam, Amsterdam, The Netherlands Hypertriglyceridemia (HTG) is a major risk factor for cardiovascular diseases in patients with T2D, which may be due to impaired TG lipolysis and/ or impaired hepatic remnant clearance. GPI-anchored-HDL binding protein 1(GPIHBP1) anchors LPL at the cell surface and enables TG lipolysis. Therefore, the aim was to determine the effect of insulin on GPIHBP1 expression and the impact of insulin resistance leads to impaired regulation of GPIHBP1. To study the effect of insulin on GPIHBP1, human microvascular endothelial cells were treated with 100nM of insulin. On stimulation with insulin, GPIHBP1mRNA level were significantly (p<0.05) decreased. Additionally, immunofluorescence image showed decreased expression of GPIHBP1 protein in the insulin stimulated cells. To study the effect of insulin resistance, GPIHBP1mRNA and protein expression were analyzed in adipose tissue depots in a severe insulin resistant mouse model (Leprdb/db). Interestingly, GPIHBP1 mRNA expression was significantly increased (P<0.05) whereas the protein expression showed a trend towards a decrease compared to its control (Leprdb/m) (P¼0.11). Next, GPIHBP1 mRNA and protein expression in visceral adipose tissue biopsy of 216 subjects with a BMI range of 18 to 75kg/m2 was measured. GPIHBP1 mRNA showed no correlation to any study parameters whereas, in a subset of 45 subjects with a BMI range of 18 to 45kg/m2 GPIHBP1 protein levels were significantly negatively correlated with insulin(r¼-0.49,P¼0.001) and HOMA-IR(r¼ 0.46,P¼0.002). GPIHBP1 protein expression is decreased in insulin resistance and reveals a novel insight in the post-translational regulatory machinery of GPIHBP1. Further studies will be focused upon identification of the specific mediators responsible for the observed effects.
EAS-0822. INSULIN DEPENDENT TRIGLYCERIDE-RICH LIPOPROTEINS UPTAKE INTO BROWN ADIPOSE TISSUE N. Mangels 1, M. Heine 1, K. Gottschling 1, R. Reimer 2, J. Heeren 3. 1 Department of Biochemistry and Molecular Biology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; 2 Department of Electron Microscopy and Micro Technology, Heinrich-Pette Institute, Martinistrasse 52, 20246, Hamburg, Germany; 3 Department of Biochemistry and Molecular Biology, University Medical Center HamburgEppendorf Hamburg, Hamburg, Germany Objective: Catabolic, cold-activated brown adipose tissue (BAT) burns triglycerides stored in lipid droplets for heat production. Consequently, endogenous lipid stores need to be replenished by anabolic processes. The canonical lipid uptake pathway involves the hydrolysis of triglyceride-rich lipoproteins (TRL) by active lipoprotein lipase (LPL) and the subsequent fatty acid uptake by active adipocytes. In addition to fatty acid uptake, there is evidence for a whole TRL particle internalization into active BAT. The aim of this study was to investigate the catabolic and anabolic processes in lipid handling of activated BAT. Methods: BAT activity was stimulated by cold exposure or CL316,243 (CL) treatment in wild-type and transgenic mice. The uptake processes were investigated by metabolic turnover studies, intravital microscopy and by electron microscopy after the injection of nanoparticle-labelled TRL. Results: Increased BAT activity is accompanied by enhanced insulin secretion. CL treatment stimulated anabolic processes in BAT via insulindependent phosphorylation of Akt kinase. This process is dependent on CD36 expression and associated with facilitated whole particle uptake which was visualized by electron microscopy. Inhibition of insulin secretion using the potassium channel agonist diazoxide during activation abolished TRL uptake into BAT. Conclusion: Our data show that cold promotes catabolic as well as anabolic processes in BAT whereas insulin orchestrates metabolic pathways that controls lipoprotein handling for the replenishment of endogenous energy stores. Impaired lipoprotein processing mediated by BAT
duodenal intralipid infusion. The lymphatic HDL was isolated; associated lipid, protein and miRNA composition analyzed. Jejunal explants from IR rats were treated with niacin or nicotinamide and mRNA expression was assessed. Results: In IR rats apo-AI lymphatic HDL secretion was reduced (47%), but associated TG content enriched (86%), compared to non-IR rats. Niacin was found to stimulate the secretion of lymph HDL (>60%) compared to nontreated IR rats. Niacin treatment normalized the TG content of lymphatic HDL particles. Niacin but not nicotinamide increased Ppara and Cpt1a mRNA, and annulled Tnf alpha mRNA in jejunal explants. Top three regulated miR's by niacin in the lymph HDL fraction were: rno-miR-223-3p, rno-miR-29c-3p and rno-miR93-5p. Analysis of these miR's (IPA software) showed PPAR signalling to be amongst the significantly predicted pathways. Conclusion: IR reduces the secretion of apoAI HDL into mesenteric lymph, whilst niacin may contribute to improving CVD risk by restoring the number and TG enrichment of intestinal apoAI particles. The altered miRNA profile in response to niacin treatment might be involved in exerting hypolipidemic effects of niacin.
EAS-0505. ARGININE-DIRECTED GLYCATION BY METHYLGLYOXAL IS A FACTOR DECREASING HDL PLASMA CONCENTRATION AND FUNCTIONALITY IN VIVO N. Rabbani, P.J. Thornalley. Warwick Medical School, University Hospitals Coventry and Warwickshire NHS Trust, Coventry, United Kingdom Aim: The aim of this study was to quantify the modification of HDL by the glucose-derived reactive metabolite, methylglyoxal, in healthy people and patients with type 2 diabetes (T2DM), characterise structural, functional and physiological consequences of the modification and predict their importance for risk of cardiovascular disease. Methods: Major fractions of HDL were isolated from healthy human subjects and patients with T2DM and fractions modified by methylglyoxal and related dicarbonyl metabolites quantified. HDL fractions were glycated by methylglyoxal to minimum extent in vitro and molecular, functional and physiological characteristics determined. A one compartment model of HDL plasma clearance was produced including formation and clearance of dicarbonyl modified HDL. Results: HDL modified by methylglyoxal and related dicarbonyl metabolites accountedfor 2.6% HDL and increased to 4.5% in patients with T2DM. Methylglyoxal modification in vitro induced re-structuring of the HDL particles, decreasing stability and plasma half-life in vivo. It occurred at sites of apolipoprotein A-1 linked to membrane fusion, intramolecular bonding and ligand binding. Kinetic modelling of methylglyoxal modification of HDL predicted a negative correlation of plasma HDL-C with methylglyoxal-modified HDL. This was validated clinically. It also predicted dicarbonyl modification produces 2 e 6% decrease in total plasma HDL and 5 -13% decrease in functional HDL clinically. Conclusions: Methylglyoxal modification of apolipoprotein A-1 may accelerate HDL degradation and impairs its functionality in vivo, likely contributing to increased risk of cardiovascular disease. Quantitation of methylglyoxal-modified HDL may improve cardiovascular disease risk models and therapeutics to decrease methylglyoxal may improve preventive treatment of cardiovascular disease.
EAS-0555. IMPAIRED INSULIN SIGNALLING LEADS TO DECREASED EXPRESSION OF GPIHBP1 R.P. Surendran 1, M. Clemente-Postigo 2, R.S. Kootte 3, M.M. van Eijk 4, L. Garrido-Sanchez 2, F. Cardona 2, F. Tinahones 2, M. Nieuwdorp 3, G. Dallinga-Thie 1. 1 Experimental Vascular Medicine, Academic Medical Center University of Amsterdam, Amsterdam, The Netherlands; 2 Unidad de n Clínica Endocrinología y Nutricio n, Instituto de Investigacio n Gestio laga (IBIMA) Complejo Hospitalario de Ma laga, Malaga, Biom edica de Ma Spain; 3 Vascular Medicine, Academic Medical Center University of
e33
Amsterdam, Amsterdam, The Netherlands; 4 Medical Biochemistry, Academic Medical Center University of Amsterdam, Amsterdam, The Netherlands Hypertriglyceridemia (HTG) is a major risk factor for cardiovascular diseases in patients with T2D, which may be due to impaired TG lipolysis and/ or impaired hepatic remnant clearance. GPI-anchored-HDL binding protein 1(GPIHBP1) anchors LPL at the cell surface and enables TG lipolysis. Therefore, the aim was to determine the effect of insulin on GPIHBP1 expression and the impact of insulin resistance leads to impaired regulation of GPIHBP1. To study the effect of insulin on GPIHBP1, human microvascular endothelial cells were treated with 100nM of insulin. On stimulation with insulin, GPIHBP1mRNA level were significantly (p<0.05) decreased. Additionally, immunofluorescence image showed decreased expression of GPIHBP1 protein in the insulin stimulated cells. To study the effect of insulin resistance, GPIHBP1mRNA and protein expression were analyzed in adipose tissue depots in a severe insulin resistant mouse model (Leprdb/db). Interestingly, GPIHBP1 mRNA expression was significantly increased (P<0.05) whereas the protein expression showed a trend towards a decrease compared to its control (Leprdb/m) (P¼0.11). Next, GPIHBP1 mRNA and protein expression in visceral adipose tissue biopsy of 216 subjects with a BMI range of 18 to 75kg/m2 was measured. GPIHBP1 mRNA showed no correlation to any study parameters whereas, in a subset of 45 subjects with a BMI range of 18 to 45kg/m2 GPIHBP1 protein levels were significantly negatively correlated with insulin(r¼-0.49,P¼0.001) and HOMA-IR(r¼ 0.46,P¼0.002). GPIHBP1 protein expression is decreased in insulin resistance and reveals a novel insight in the post-translational regulatory machinery of GPIHBP1. Further studies will be focused upon identification of the specific mediators responsible for the observed effects.
EAS-0822. INSULIN DEPENDENT TRIGLYCERIDE-RICH LIPOPROTEINS UPTAKE INTO BROWN ADIPOSE TISSUE N. Mangels 1, M. Heine 1, K. Gottschling 1, R. Reimer 2, J. Heeren 3. 1 Department of Biochemistry and Molecular Biology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; 2 Department of Electron Microscopy and Micro Technology, Heinrich-Pette Institute, Martinistrasse 52, 20246, Hamburg, Germany; 3 Department of Biochemistry and Molecular Biology, University Medical Center HamburgEppendorf Hamburg, Hamburg, Germany Objective: Catabolic, cold-activated brown adipose tissue (BAT) burns triglycerides stored in lipid droplets for heat production. Consequently, endogenous lipid stores need to be replenished by anabolic processes. The canonical lipid uptake pathway involves the hydrolysis of triglyceride-rich lipoproteins (TRL) by active lipoprotein lipase (LPL) and the subsequent fatty acid uptake by active adipocytes. In addition to fatty acid uptake, there is evidence for a whole TRL particle internalization into active BAT. The aim of this study was to investigate the catabolic and anabolic processes in lipid handling of activated BAT. Methods: BAT activity was stimulated by cold exposure or CL316,243 (CL) treatment in wild-type and transgenic mice. The uptake processes were investigated by metabolic turnover studies, intravital microscopy and by electron microscopy after the injection of nanoparticle-labelled TRL. Results: Increased BAT activity is accompanied by enhanced insulin secretion. CL treatment stimulated anabolic processes in BAT via insulindependent phosphorylation of Akt kinase. This process is dependent on CD36 expression and associated with facilitated whole particle uptake which was visualized by electron microscopy. Inhibition of insulin secretion using the potassium channel agonist diazoxide during activation abolished TRL uptake into BAT. Conclusion: Our data show that cold promotes catabolic as well as anabolic processes in BAT whereas insulin orchestrates metabolic pathways that controls lipoprotein handling for the replenishment of endogenous energy stores. Impaired lipoprotein processing mediated by BAT