Effects of substitutions of glycine and asparagine for serine132 on activity and binding of human lipoprotein lipase to very low density lipoproteins

AbstractFor studying the role of Ser132 in the putative catalytic site of human lipoprotein lipase (LPL), mutant LPL cDNAs expressing LPLs with amino acid substitutions of Gly or Asn for Ser132 were obtained by site-directed mutagenesis, and were expressed in COS-1 cells. Considerable amounts of LPL enzyme protein mass were detected in the culture medium of COS-1 cells transfected with wild-type LPL, LPL-Gly132, or LPL-Asn132. LPL-Gly132 hydrolyzed Triton X-100-triolein and tributyrin as effectively as wild-type LPL, whereas LPL-Asn132 showed no activity. LPL-Asn132 bound to very low density lipoproteins as effectively as wild-type LPL

Serum Apolipoprotein M Levels are Correlated with Biomarkers of Coagulation

Background:Apolipoprotein M (ApoM) is bound to high-density lipoprotein (HDL) in plasma, and HDL has anticoagulation effects. However, the association between ApoM and biomarkers of coagulation was unclear. Therefore, we investigated relationships between ApoM and biomarkers of coagulation. Methods: Serum samples from 233 Japanese participants including with diabetes mellitus, hypertension, dyslipidemia, or healthy controls were analyzed. Serum ApoM levels were measured using Enzyme-Linked Immuno-Sorbent Assay(ELISA). Results:Analysis of all 233 participants showed that ApoM levels were positively correlated with age (r＝0.284, p＜0.001), total cholesterol (TC;r＝0.477, p＜0.001), HDL-cholesterol (HDL-C;r＝0.234, p＜0.001) and lowdensity lipoprotein cholesterol (LDL-C;r＝0.331, p＜0.001). Higher ApoM levels were correlated with shorter activated partial thromboplastin time(APTT;r＝－0.226,p＝0.001) and prothrombin time(PT,％;r＝0.326,p＜ 0.001). Separate analysis of the 115 healthy controls showed that ApoM levels were positively correlated with age, TC, HDL-C and LDL-C, and higher ApoM levels were correlated with shorter PT. Conclusion:Serum levels of ApoM may influence biomarkers of coagulation

Soluble LR11/SorLA represses thermogenesis in adipose tissue and correlates with BMI in humans.

Thermogenesis in brown adipose tissue (BAT) is an important component of energy expenditure in mammals. Recent studies have confirmed its presence and metabolic role in humans. Defining the physiological regulation of BAT is therefore of great importance for developing strategies to treat metabolic diseases. Here we show that the soluble form of the low-density lipoprotein receptor relative, LR11/SorLA (sLR11), suppresses thermogenesis in adipose tissue in a cell-autonomous manner. Mice lacking LR11 are protected from diet-induced obesity associated with an increased browning of white adipose tissue and hypermetabolism. Treatment of adipocytes with sLR11 inhibits thermogenesis via the bone morphogenetic protein/TGFβ signalling pathway and reduces Smad phosphorylation. In addition, sLR11 levels in humans are shown to positively correlate with body mass index and adiposity. Given the need for tight regulation of a tissue with a high capacity for energy wastage, we propose that LR11 plays an energy conserving role that is exaggerated in states of obesity.AW and AVP were supported by FP7 – BetaBAT, BBSRC (BB/J009865/1), the British Heart Foundation (PG/12/53/29714) and MDU MRC. MJ and HB were supported by Japan Health and Labour Sciences Research grant (H22-rinkensui-ippan-001) and Grants-in–aid for Scientific Research from Japanese Ministry of Education, Culture, Sports, Science and Technology (24390231 and 24790907). VP was supported by Wellcome Trust and the Cambridge Overseas Trust. JR was supported by Ministerio de Educación, through “Programa Nacional de Movilidad de Recursos Humanos del Plan Nacional de I-D+i 2008-2011 (Subprograma de Estancias de Movilidad en el Extranjero “José Castillejo” para jóvenes Doctores, ref: JC2011-0248). SV was supported by MRC. WJS was supported by the Austrian Science Fund (FWF P-20218 and P-20455). Animal work was performed at the MDU DMC Core facilities.This is the final version of the article. It first appeared from NPG via http://dx.doi.org/10.1038/ncomms995

Soluble LR11/SorLA represses thermogenesis in adipose tissue and correlates with BMI in humans

Thermogenesis in brown adipose tissue (BAT) is an important component of energy expenditure in mammals. Recent studies have confirmed its presence and metabolic role in humans. Defining the physiological regulation of BAT is therefore of great importance for developing strategies to treat metabolic diseases. Here we show that the soluble form of the low-density lipoprotein receptor relative, LR11/SorLA (sLR11), suppresses thermogenesis in adipose tissue in a cell-autonomous manner. Mice lacking LR11 are protected from diet-induced obesity associated with an increased browning of white adipose tissue and hypermetabolism. Treatment of adipocytes with sLR11 inhibits thermogenesis via the bone morphogenetic protein/TGFb signalling pathway and reduces Smad phosphorylation. In addition, sLR11 levels in humans are shown to positively correlate with body mass index and adiposity. Given the need for tight regulation of a tissue with a high capacity for energy wastage, we propose that LR11 plays an energy conserving role that is exaggerated in states of obesity

An ELISA for quantifying GPIHBP1 autoantibodies and making a diagnosis of the GPIHBP1 autoantibody syndrome

BackgroundAutoantibodies against GPIHBP1, the endothelial cell transporter for lipoprotein lipase (LPL), cause severe hypertriglyceridemia ("GPIHBP1 autoantibody syndrome"). Affected patients have low serum GPIHBP1 and LPL levels. We report the development of a sensitive and specific ELISA, suitable for routine clinical use, to detect GPIHBP1 autoantibodies in serum and plasma.MethodsSerum and plasma samples were added to wells of an ELISA plate that had been coated with recombinant human GPIHBP1. GPIHBP1 autoantibodies bound to GPIHBP1 were detected with an HRP-labeled antibody against human immunoglobulin. Sensitivity, specificity, and reproducibility of the ELISA was evaluated with plasma or serum samples from patients with the GPIHBP1 autoantibody syndrome.ResultsA solid-phase ELISA to detect and quantify GPIHBP1 autoantibodies in human plasma and serum was developed. Spiking recombinant human GPIHBP1 into the samples reduced the ability of the ELISA to detect GPIHBP1 autoantibodies. The ELISA is reproducible and sensitive; it can detect GPIHBP1 autoantibodies in samples diluted by >1000-fold.ConclusionWe have developed a sensitive and specific ELISA for detecting GPIHBP1 autoantibodies in human serum and plasma; this assay will make it possible to rapidly diagnose the GPIHBP1 autoantibody syndrome

An enzyme-linked immunosorbent assay for measuring GPIHBP1 levels in human plasma or serum

BackgroundGlycosylphosphatidylinositol-anchored high-density lipoprotein-binding protein 1 (GPIHBP1), a glycosylphosphatidylinositol (GPI)-anchored protein of capillary endothelial cells, transports lipoprotein lipase to the capillary lumen and is essential for the lipolytic processing of triglyceride-rich lipoproteins.ObjectiveBecause some GPI-anchored proteins have been detected in plasma, we tested whether GPIHBP1 is present in human blood and whether GPIHBP1 deficiency or a history of cardiovascular disease affected GPIHBP1 circulating levels.MethodsWe developed 2 monoclonal antibodies against GPIHBP1 and used the antibodies to establish a sandwich enzyme-linked immunosorbent assay (ELISA) to measure GPIHBP1 levels in human blood.ResultsThe GPIHBP1 ELISA was linear in the 8 to 500 pg/mL range and allowed the quantification of GPIHBP1 in serum and in pre- and post-heparin plasma (including lipemic samples). GPIHBP1 was undetectable in the plasma of subjects with null mutations in GPIHBP1. Serum GPIHBP1 median levels were 849 pg/mL (range: 740-1014) in healthy volunteers (n = 28) and 1087 pg/mL (range: 877-1371) in patients with a history of cardiovascular or metabolic disease (n = 415). There was an extremely small inverse correlation between GPIHBP1 and triglyceride levels (r = 0.109; P < .0275). GPIHBP1 levels tended to be slightly higher in patients who had a major cardiovascular event after revascularization.ConclusionWe developed an ELISA for quantifying GPIHBP1 in human blood. This assay will be useful to identify patients with GPIHBP1 deficiency and patients with GPIHBP1 autoantibodies. The potential of plasma GPIHBP1 as a biomarker for metabolic or cardiovascular disease is yet questionable but needs additional testing