Publications by authors named "Debapriya Basu"

20 Publications

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Lipolytic enzymes and free fatty acids at the endothelial interface.

Atherosclerosis 2021 Jul 28;329:1-8. Epub 2021 May 28.

Department of Medicine, Center for Human Nutrition, Washington University School of Medicine, Saint Louis, MO, USA. Electronic address:

Lipids released from circulating lipoproteins by intravascular action of lipoprotein lipase (LpL) reach parenchymal cells in tissues with a non-fenestrated endothelium by transfer through or around endothelial cells. The actions of LpL are controlled at multiple sites, its synthesis and release by myocytes and adipocytes, its transit and association with the endothelial cell luminal surface, and finally its activation and inhibition by a number of proteins and by its product non-esterified fatty acids. Multiple pathways mediate endothelial transit of lipids into muscle and adipose tissues. These include movement of fatty acids via the endothelial cell fatty acid transporter CD36 and movement of whole or partially LpL-hydrolyzed lipoproteins via other apical endothelial cell receptors such as SR-B1and Alk1. Lipids also likely change the barrier function of the endothelium and operation of the paracellular pathway around endothelial cells. This review summarizes in vitro and in vivo support for the key role of endothelial cells in delivery of lipids and highlights incompletely understood processes that are the focus of active investigation.
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http://dx.doi.org/10.1016/j.atherosclerosis.2021.05.018DOI Listing
July 2021

Endothelial Cell Receptors in Tissue Lipid Uptake and Metabolism.

Circ Res 2021 02 4;128(3):433-450. Epub 2021 Feb 4.

Division of Endocrinology, Diabetes and Metabolism, Department of Medicine, New York University Grossman School of Medicine (A.G.C., D.B., I.J.G.).

Lipid uptake and metabolism are central to the function of organs such as heart, skeletal muscle, and adipose tissue. Although most heart energy derives from fatty acids (FAs), excess lipid accumulation can cause cardiomyopathy. Similarly, high delivery of cholesterol can initiate coronary artery atherosclerosis. Hearts and arteries-unlike liver and adrenals-have nonfenestrated capillaries and lipid accumulation in both health and disease requires lipid movement from the circulation across the endothelial barrier. This review summarizes recent in vitro and in vivo findings on the importance of endothelial cell receptors and uptake pathways in regulating FAs and cholesterol uptake in normal physiology and cardiovascular disease. We highlight clinical and experimental data on the roles of ECs in lipid supply to tissues, heart, and arterial wall in particular, and how this affects organ metabolism and function. Models of FA uptake into ECs suggest that receptor-mediated uptake predominates at low FA concentrations, such as during fasting, whereas FA uptake during lipolysis of chylomicrons may involve paracellular movement. Similarly, in the setting of an intact arterial endothelial layer, recent and historic data support a role for receptor-mediated processes in the movement of lipoproteins into the subarterial space. We conclude with thoughts on the need to better understand endothelial lipid transfer for fuller comprehension of the pathophysiology of hyperlipidemia, and lipotoxic diseases such as some forms of cardiomyopathy and atherosclerosis.
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http://dx.doi.org/10.1161/CIRCRESAHA.120.318003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7959116PMC
February 2021

Hypertriglyceridemia and Atherosclerosis: Using Human Research to Guide Mechanistic Studies in Animal Models.

Front Endocrinol (Lausanne) 2020 6;11:504. Epub 2020 Aug 6.

Department of Medicine, University of Washington Medicine Diabetes Institute, University of Washington School of Medicine, Seattle, WA, United States.

Human studies support a strong association between hypertriglyceridemia and atherosclerotic cardiovascular disease (CVD). However, whether a causal relationship exists between hypertriglyceridemia and increased CVD risk is still unclear. One plausible explanation for the difficulty establishing a clear causal role for hypertriglyceridemia in CVD risk is that lipolysis products of triglyceride-rich lipoproteins (TRLs), rather than the TRLs themselves, are the likely mediators of increased CVD risk. This hypothesis is supported by studies of rare mutations in humans resulting in impaired clearance of such lipolysis products (remnant lipoprotein particles; RLPs). Several animal models of hypertriglyceridemia support this hypothesis and have provided additional mechanistic understanding. Mice deficient in lipoprotein lipase (LPL), the major vascular enzyme responsible for TRL lipolysis and generation of RLPs, or its endothelial anchor GPIHBP1, are severely hypertriglyceridemic but develop only minimal atherosclerosis as compared with animal models deficient in apolipoprotein (APO) E, which is required to clear TRLs and RLPs. Likewise, animal models convincingly show that increased clearance of TRLs and RLPs by LPL activation (achieved by inhibition of APOC3, ANGPTL3, or ANGPTL4 action, or increased APOA5) results in protection from atherosclerosis. Mechanistic studies suggest that RLPs are more atherogenic than large TRLs because they more readily enter the artery wall, and because they are enriched in cholesterol relative to triglycerides, which promotes pro-atherogenic effects in lesional cells. Other mechanistic studies show that hepatic receptors (LDLR and LRP1) and APOE are critical for RLP clearance. Thus, studies in animal models have provided additional mechanistic insight and generally agree with the hypothesis that RLPs derived from TRLs are highly atherogenic whereas hypertriglyceridemia due to accumulation of very large TRLs in plasma is not markedly atherogenic in the absence of TRL lipolysis products.
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http://dx.doi.org/10.3389/fendo.2020.00504DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7423973PMC
June 2021

Regulation of lipoprotein lipase-mediated lipolysis of triglycerides.

Curr Opin Lipidol 2020 06;31(3):154-160

Division of Endocrinology, Diabetes and Metabolism, Department of Medicine, New York University Grossman School of Medicine, New York, New York, USA.

Purpose Of Review: To discuss the recent developments in structure, function and physiology of lipoprotein lipase (LpL) and the regulators of LpL, which are being targeted for therapy.

Recent Findings: Recent studies have revealed the long elusive crystal structure of LpL and its interaction with glycosylphosphatidylinositol anchored high-density lipoprotein binding protein 1 (GPIHBP1). New light has been shed on LpL being active as a monomer, which brings into questions previous thinking that LpL inhibitors like angiopoietin-like 4 (ANGPTL4) and stabilizers like LMF1 work on disrupting or maintaining LpL in dimer form. There is increasing pharmaceutical interest in developing targets to block LpL inhibitors like ANGPTL3. Other approaches to reducing circulating triglyceride levels have been using an apoC2 mimetic and reducing apoC3.

Summary: Lipolysis of triglyceride-rich lipoproteins by LpL is a central event in lipid metabolism, releasing fatty acids for uptake by tissues and generating low-density lipoprotein and expanding high-density lipoprotein. Recent mechanistic insights into the structure and function of LpL have added to our understanding of triglyceride metabolism. This has also led to heightened interest in targeting its posttranslational regulators, which can be the next generation of lipid-lowering agents used to prevent hypertriglyceridemic pancreatitis and, hopefully, cardiovascular disease.
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http://dx.doi.org/10.1097/MOL.0000000000000676DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7478854PMC
June 2020

A dual apolipoprotein C-II mimetic-apolipoprotein C-III antagonist peptide lowers plasma triglycerides.

Sci Transl Med 2020 01;12(528)

Lipoprotein Metabolism Laboratory, Translational Vascular Medicine Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA.

Recent genetic studies have established that hypertriglyceridemia (HTG) is causally related to cardiovascular disease, making it an active area for drug development. We describe a strategy for lowering triglycerides (TGs) with an apolipoprotein C-II (apoC-II) mimetic peptide called D6PV that activates lipoprotein lipase (LPL), the main plasma TG-hydrolyzing enzyme, and antagonizes the TG-raising effect of apoC-III. The design of D6PV was motivated by a combination of all-atom molecular dynamics simulation of apoC-II on the Anton 2 supercomputer, structural prediction programs, and biophysical techniques. Efficacy of D6PV was assessed ex vivo in human HTG plasma and was found to be more potent than full-length apoC-II in activating LPL. D6PV markedly lowered TG by more than 80% within a few hours in both apoC-II-deficient mice and h-transgenic (Tg) mice. In h-Tg mice, D6PV treatment reduced plasma apoC-III by 80% and apoB by 65%. Furthermore, low-density lipoprotein (LDL) cholesterol did not accumulate but rather was decreased by 10% when h-Tg mice lacking the LDL-receptor (h-Tg × ) were treated with the peptide. D6PV lowered TG by 50% in whole-body inducible knockout (i ) mice, confirming that it can also act independently of LPL. D6PV displayed good subcutaneous bioavailability of about 80% in nonhuman primates. Because it binds to high-density lipoproteins, which serve as a long-term reservoir, it also has an extended terminal half-life (42 to 50 hours) in nonhuman primates. In summary, D6PV decreases plasma TG by acting as a dual apoC-II mimetic and apoC-III antagonist, thereby demonstrating its potential as a treatment for HTG.
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http://dx.doi.org/10.1126/scitranslmed.aaw7905DOI Listing
January 2020

Incompatible crossmatch: First sign of a hemolytic transfusion reaction due to out-of-group platelet transfusion.

Asian J Transfus Sci 2019 Jan-Jun;13(1):57-59

Department of Gynaecological Surgery, Tata Medical Center, Kolkata, West Bengal, India.

Platelet (PLT) transfusion is undertaken in a variety of clinical settings with thrombocytopenia, with or without bleeding. Since PLTs are most often stored in donor plasma, group-specific PLT transfusions are preferred to out-of-group transfusions. PLTs adsorb ABO antigens over their surface from the plasma. In major ABO-incompatible PLT transfusions, anti-A/B from the patient plasma react with the ABO antigens on transfused PLTs and can potentially cause adverse reactions or PLT refractoriness. Transfusion of PLTs with major ABO incompatibility, though effective in preventing clinical bleeding, is associated with reduced posttransfusion PLT count increments. In minor incompatible PLT transfusion transfused, anti-A/B can cause hemolytic transfusion reaction (HTR) which is not always related to a high titer of anti-A/B in the donor. Although attempts are made to practice ABO identical PLT transfusion, most centers practice out-of-group random donor platelets (RDPs) as well as single-donorplatelets (SDP) transfusion. The limited PLT shelf life does not always permit ABO identical PLT transfusion. At our center, ABO-specific PLT transfusions are practiced where possible, and in case of minor ABO-incompatible transfusions, antibody titers are not done. Here, we report a case of HTR due to out-of-group SDP transfusion, detected in the laboratory after an incompatible red blood cell (RBC) crossmatch.
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http://dx.doi.org/10.4103/ajts.AJTS_36_17DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6580831PMC
July 2019

Mechanism of Increased LDL (Low-Density Lipoprotein) and Decreased Triglycerides With SGLT2 (Sodium-Glucose Cotransporter 2) Inhibition.

Arterioscler Thromb Vasc Biol 2018 09;38(9):2207-2216

From the Division of Endocrinology, Diabetes and Metabolism, New York University School of Medicine (D.B., L.-A.H., D.S., J.O., I.J.G.).

Objective- SGLT2 (sodium-glucose cotransporter 2) inhibition in humans leads to increased levels of LDL (low-density lipoprotein) cholesterol and decreased levels of plasma triglyceride. Recent studies, however, have shown this therapy to lower cardiovascular mortality. In this study, we aimed to determine how SGLT2 inhibition alters circulating lipoproteins. Approach and Results- We used a mouse model expressing human CETP (cholesteryl ester transfer protein) and human ApoB100 (apolipoprotein B100) to determine how SGLT2 inhibition alters plasma lipoprotein metabolism. The mice were fed a high-fat diet and then were made partially insulin deficient using streptozotocin. SGLT2 was inhibited using a specific antisense oligonucleotide or canagliflozin, a clinically available oral SGLT2 inhibitor. Inhibition of SGLT2 increased circulating levels of LDL cholesterol and reduced plasma triglyceride levels. SGLT2 inhibition was associated with increased LpL (lipoprotein lipase) activity in the postheparin plasma, decreased postprandial lipemia, and faster clearance of radiolabeled VLDL (very-LDL) from circulation. Additionally, SGLT2 inhibition delayed turnover of labeled LDL from circulation. Conclusions- Our studies in diabetic CETP-ApoB100 transgenic mice recapitulate many of the changes in circulating lipids found with SGLT2 inhibition therapy in humans and suggest that the increased LDL cholesterol found with this therapy is because of reduced clearance of LDL from the circulation and greater lipolysis of triglyceride-rich lipoproteins. Most prominent effects of SGLT2 inhibition in the current mouse model were seen with antisense oligonucleotides-mediated knockdown of SGLT2.
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http://dx.doi.org/10.1161/ATVBAHA.118.311339DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6207215PMC
September 2018

Fibrin in blood samples interferes with blood grouping by automation.

Transfusion 2018 08;58(8):1831-1832

Department of Transfusion Medicine, Tata Medical Center, Kolkata, India.

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http://dx.doi.org/10.1111/trf.14593DOI Listing
August 2018

Endothelial cell CD36 optimizes tissue fatty acid uptake.

J Clin Invest 2018 10 26;128(10):4329-4342. Epub 2018 Jul 26.

Division of Endocrinology, Diabetes and Metabolism, New York University School of Medicine, New York, New York, USA.

Movement of circulating fatty acids (FAs) to parenchymal cells requires their transfer across the endothelial cell (EC) barrier. The multiligand receptor cluster of differentiation 36 (CD36) facilitates tissue FA uptake and is expressed in ECs and parenchymal cells such as myocytes and adipocytes. Whether tissue uptake of FAs is dependent on EC or parenchymal cell CD36, or both, is unknown. Using a cell-specific deletion approach, we show that EC, but not parenchymal cell, CD36 deletion increased fasting plasma FAs and postprandial triglycerides. EC-Cd36-KO mice had reduced uptake of radiolabeled long-chain FAs into heart, skeletal muscle, and brown adipose tissue; these uptake studies were replicated using [11C]palmitate PET scans. High-fat diet-fed EC-CD36-deficient mice had improved glucose tolerance and insulin sensitivity. Both EC and cardiomyocyte (CM) deletion of CD36 reduced heart lipid droplet accumulation after fasting, but CM deletion did not affect heart glucose or FA uptake. Expression in the heart of several genes modulating glucose metabolism and insulin action increased with EC-CD36 deletion but decreased with CM deletion. In conclusion, EC CD36 acts as a gatekeeper for parenchymal cell FA uptake, with important downstream effects on glucose utilization and insulin action.
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http://dx.doi.org/10.1172/JCI99315DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6159965PMC
October 2018

ABO, Rhesus, and Kell Antigens, Alleles, and Haplotypes in West Bengal, India.

Transfus Med Hemother 2018 Jan 20;45(1):62-66. Epub 2017 Oct 20.

Department of Immunohematology and Blood Transfusion, Medical College Hospital, Kolkata, India; Department of Transfusion Medicine, The Mission Hospital, Durgapur, India; NIH Clinical Center, National Institutes of Health, Bethesda MD, USA.

Background: Few studies have documented the blood group antigens in the population of eastern India. Frequencies of some common alleles and haplotypes were unknown. We describe phenotype, allele, and haplotype frequencies in the state of West Bengal, India.

Methods: We tested 1,528 blood donors at the Medical College Hospital, Kolkata. The common antigens of the ABO, Rhesus, and Kell blood group systems were determined by standard serologic methods in tubes. Allele and haplotype frequencies were calculated with an iterative method that yielded maximum-likelihood estimates under the assumption of a Hardy-Weinberg equilibrium.

Results: The prevalence of ABO antigens were B (34%), O (32%), A (25%), and AB (9%) with allele frequencies for O = 0.567, A = 0.189, and B = 0.244. The D antigen (RH1) was observed in 96.6% of the blood donors with haplotype frequencies, such as for CDe = 0.688809, cde = 0.16983 and CdE = 0.000654. The K antigen (K1) was observed in 12 donors (0.79%) with allele frequencies for K = 0.004 and k = 0.996. For the Bengali population living in the south of West Bengal, we established the frequencies of the major clinically relevant antigens in the ABO, Rhesus, and Kell blood group systems and derived estimates for the underlying ABO and KEL alleles and RH haplotypes. Such blood donor screening will improve the availability of compatible red cell units for transfusion. Our approach using widely available routine methods can readily be applied in other regions, where the sufficient supply of blood typed for the Rh and K antigens is lacking.
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http://dx.doi.org/10.1159/000475507DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5836283PMC
January 2018

Clinical and laboratory profile of anti-M.

Immunohematology 2017 Dec;33(4):165-169

Clinical Haematology, Tata Medical Center, Kolkata, India.

Conclusions: Anti-M is a frequently detected naturally occurring antibody that has been reported in various clinical settings and also in voluntary donors. We describe here the clinical and laboratory findings of 11 cases with anti-M detected at our center. This report is a retrospective study in which we reviewed our immunohematology laboratory records for cases involving anti-M. Both donor and patient data from a 28-month period (September 2014 to December 2016) were reviewed. During this period, 11 examples of anti-M were detected (8 patients, 1 voluntary whole blood donor, and 1 hematopoietic stem cell donor. Anti-M was also detected in one external quality assessment scheme sample received during this period. In conclusion, anti-M can be detected in various clinical settings. This antibody can be clinically significant; in the laboratory, it can present as a serologic problem such as an ABO group discrepancy or an incompatible crossmatch. After detection, management and course of action is determined by both the antibody characteristics and the clinical setting.
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December 2017

Novel Reversible Model of Atherosclerosis and Regression Using Oligonucleotide Regulation of the LDL Receptor.

Circ Res 2018 02 10;122(4):560-567. Epub 2018 Jan 10.

From the Department of Medicine, New York University Langone Health, New York (D.B., Y.H., L.-A.H., A.M., E.A.F., I.J.G.); Ionis Pharmaceuticals, Carlsbad, CA (A.E.M., M.J.G.); Division of Cardiology, Department of Medicine (T.W.), Division of Metabolism, Endocrinology and Nutrition, Department of Medicine, UW Diabetes Institute (S.B., K.E.B.), and Department of Pathology (K.E.B.), University of Washington, Seattle; and Department of Cardiology and Angiology I, Heart Center, Freiburg University, Germany (K.P., A.Z., F.W.).

Rationale: Animal models have been used to explore factors that regulate atherosclerosis. More recently, they have been used to study the factors that promote loss of macrophages and reduction in lesion size after lowering of plasma cholesterol levels. However, current animal models of atherosclerosis regression require challenging surgeries, time-consuming breeding strategies, and methods that block liver lipoprotein secretion.

Objective: We sought to develop a more direct or time-effective method to create and then reverse hypercholesterolemia and atherosclerosis via transient knockdown of the hepatic LDLR (low-density lipoprotein receptor) followed by its rapid restoration.

Methods And Results: We used antisense oligonucleotides directed to LDLR mRNA to create hypercholesterolemia in wild-type C57BL/6 mice fed an atherogenic diet. This led to the development of lesions in the aortic root, aortic arch, and brachiocephalic artery. Use of a sense oligonucleotide replicating the targeted sequence region of the LDLR mRNA rapidly reduced circulating cholesterol levels because of recovery of hepatic LDLR expression. This led to a decrease in macrophages within the aortic root plaques and brachiocephalic artery, that is, regression of inflammatory cell content, after a period of 2 to 3 weeks.

Conclusions: We have developed an inducible and reversible hepatic LDLR knockdown mouse model of atherosclerosis regression. Although cholesterol reduction decreased early en face lesions in the aortic arches, macrophage area was reduced in both early and late lesions within the aortic sinus after reversal of hypercholesterolemia. Our model circumvents many of the challenges associated with current mouse models of regression. The use of this technology will potentially expedite studies of atherosclerosis and regression without use of mice with genetic defects in lipid metabolism.
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http://dx.doi.org/10.1161/CIRCRESAHA.117.311361DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5815899PMC
February 2018

Neutrophil-derived S100 calcium-binding proteins A8/A9 promote reticulated thrombocytosis and atherogenesis in diabetes.

J Clin Invest 2017 Jun 15;127(6):2133-2147. Epub 2017 May 15.

Haematopoiesis and Leukocyte Biology, Baker Heart and Diabetes Institute, Melbourne, Victoria, Australia.

Platelets play a critical role in atherogenesis and thrombosis-mediated myocardial ischemia, processes that are accelerated in diabetes. Whether hyperglycemia promotes platelet production and whether enhanced platelet production contributes to enhanced atherothrombosis remains unknown. Here we found that in response to hyperglycemia, neutrophil-derived S100 calcium-binding proteins A8/A9 (S100A8/A9) interact with the receptor for advanced glycation end products (RAGE) on hepatic Kupffer cells, resulting in increased production of IL-6, a pleiotropic cytokine that is implicated in inflammatory thrombocytosis. IL-6 acts on hepatocytes to enhance the production of thrombopoietin, which in turn interacts with its cognate receptor c-MPL on megakaryocytes and bone marrow progenitor cells to promote their expansion and proliferation, resulting in reticulated thrombocytosis. Lowering blood glucose using a sodium-glucose cotransporter 2 inhibitor (dapagliflozin), depleting neutrophils or Kupffer cells, or inhibiting S100A8/A9 binding to RAGE (using paquinimod), all reduced diabetes-induced thrombocytosis. Inhibiting S100A8/A9 also decreased atherogenesis in diabetic mice. Finally, we found that patients with type 2 diabetes have reticulated thrombocytosis that correlates with glycated hemoglobin as well as increased plasma S100A8/A9 levels. These studies provide insights into the mechanisms that regulate platelet production and may aid in the development of strategies to improve on current antiplatelet therapies and to reduce cardiovascular disease risk in diabetes.
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http://dx.doi.org/10.1172/JCI92450DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5451242PMC
June 2017

ApoC-III inhibits clearance of triglyceride-rich lipoproteins through LDL family receptors.

J Clin Invest 2016 08 11;126(8):2855-66. Epub 2016 Jul 11.

Hypertriglyceridemia is an independent risk factor for cardiovascular disease, and plasma triglycerides (TGs) correlate strongly with plasma apolipoprotein C-III (ApoC-III) levels. Antisense oligonucleotides (ASOs) for ApoC-III reduce plasma TGs in primates and mice, but the underlying mechanism of action remains controversial. We determined that a murine-specific ApoC-III-targeting ASO reduces fasting TG levels through a mechanism that is dependent on low-density lipoprotein receptors (LDLRs) and LDLR-related protein 1 (LRP1). ApoC-III ASO treatment lowered plasma TGs in mice lacking lipoprotein lipase (LPL), hepatic heparan sulfate proteoglycan (HSPG) receptors, LDLR, or LRP1 and in animals with combined deletion of the genes encoding HSPG receptors and LDLRs or LRP1. However, the ApoC-III ASO did not lower TG levels in mice lacking both LDLR and LRP1. LDLR and LRP1 were also required for ApoC-III ASO-induced reduction of plasma TGs in mice fed a high-fat diet, in postprandial clearance studies, and when ApoC-III-rich or ApoC-III-depleted lipoproteins were injected into mice. ASO reduction of ApoC-III had no effect on VLDL secretion, heparin-induced TG reduction, or uptake of lipids into heart and skeletal muscle. Our data indicate that ApoC-III inhibits turnover of TG-rich lipoproteins primarily through a hepatic clearance mechanism mediated by the LDLR/LRP1 axis.
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http://dx.doi.org/10.1172/JCI86610DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4966320PMC
August 2016

Hepatic S1P deficiency lowers plasma cholesterol levels in apoB-containing lipoproteins when LDLR function is compromised.

Nutr Metab (Lond) 2015 20;12:35. Epub 2015 Oct 20.

Department of Cell Biology, State University of New York, Downstate Medical Center, Brooklyn, NY 11203 USA.

Background: Site-1 protease (S1P) is the key enzyme required for activation of the sterol regulatory element binding proteins (SREBPs) that govern lipid synthesis. While S1P has been speculated to influence plasma apoB-containing lipoprotein (Blp) metabolism, there has been little investigative work. LDL receptor (LDLR) is the major receptor for clearing plasma LDL cholesterol (LDL-c). Proprotein convertase subtilisin kexin type 9 (PCSK9) modulates LDL-c through post-translational degradation of the LDLR.

Methods: A hepatic-specific knockdown (KD) of S1P was achieved using floxed S1P mouse models (S1P(f/f) and LDLR(-/-)S1P(f/f)) and hepatic expression of Cre recombinase. Lipids were measured in total plasma and size fractionated plasma using colorimetric assays. Realtime polymerase chain reaction, western blotting and ELISA were used to determine hepatic expression of key genes/protein. Plasmid mediated overexpression and siRNA mediated knockdown of genes were performed in mouse primary hepatocytes to determine the mechanistic basis of PCSK9 gene regulation.

Results: A hepatic-specific KD of S1P resulted in a 45 % and 38 % reduction in plasma total cholesterol and triglyceride levels, respectively. Hepatic S1P KD had a minimal effect on plasma Blp cholesterol (Blp-c) in S1P(f/f) mice, despite significantly reducing VLDL secretion. Notably, hepatic S1P KD decreased the LDL receptor (LDLR) mRNA expression by 50 %. However, the reduction in LDLR protein levels was less than that of mRNA expression, especially under fed conditions. Further assessment of hepatic S1P deficiency revealed that it increased LDLR protein stability in vivo. Mechanistically, hepatic S1P KD was shown to decrease the liver and plasma levels of the protein proprotein convertase subtilisin/kexin type 9 (PCSK9), which degrades LDLR protein. This effect was more prominent in the fed condition and sufficient to account for the discordance in LDLR mRNA and protein levels. Furthermore, hepatic S1P was shown to regulate PCSK9 expression through activation of the SREBPs. In the LDLR(-/-) background, hepatic S1P KD significantly reduced Blp-c levels.

Conclusion: Hepatic S1P is a physiological modulator of plasma Blp metabolism through its regulation of LDLR and PCSK9. Hepatic S1P is a valid target for lowering plasma Blp-c levels in the situation where LDLR function is compromised.
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http://dx.doi.org/10.1186/s12986-015-0031-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4613744PMC
October 2015

Hepatic overexpression of the prodomain of furin lessens progression of atherosclerosis and reduces vascular remodeling in response to injury.

Atherosclerosis 2014 Sep 1;236(1):121-30. Epub 2014 Jul 1.

Department of Cell Biology, State University of New York, Downstate Medical Center, Brooklyn, NY 11203, USA. Electronic address:

Objective: Atherosclerosis is a complex disease, involving elevated LDL-c, lipid accumulation in the blood vessel wall, foam cell formation and vascular dysfunction. Lowering plasma LDL-c is the cornerstone of current management of cardiovascular disease. However, new approaches which reduce plasma LDL-c and lessen the pathological vascular remodeling occurring in the disease should also have therapeutic value. Previously, we found that overexpression of profurin, the 83-amino acid prodomain of the proprotein convertase furin, lowered plasma HDL levels in wild-type mice. The question that remained was whether it had effects on apolipoprotein B (ApoB)-containing lipoproteins.

Methods: Adenovirus mediated overexpression of hepatic profurin in Ldlr(-/-)mice and wild-type mice were used to evaluate effects of profurin on ApoB-containing lipoproteins, atherosclerosis and vascular remodeling.

Results: Hepatic profurin overexpression resulted in a significant reduction in atherosclerotic lesion development in Ldlr(-/-)mice and a robust reduction in plasma LDL-c. Metabolic studies revealed lower secretion of ApoB and triglycerides in VLDL particles. Mechanistic studies showed that in the presence of profurin, hepatic ApoB, mainly ApoB100, was degraded by proteasomes. There was no effect on ApoB mRNA expression. Importantly, short-term hepatic profurin overexpression did not result in hepatic lipid accumulation. Blood vessel wall thickening caused by either wire-induced femoral artery injury or common carotid artery ligation was reduced. Profurin expression inhibited proliferation and migration in vascular smooth muscle cells in vitro.

Conclusion: These results indicate that a profurin-based therapy has the potential to treat atherosclerosis by improving metabolic lipid profiles and reducing both atherosclerotic lesion development and pathological vascular remodeling.
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http://dx.doi.org/10.1016/j.atherosclerosis.2014.06.015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7518654PMC
September 2014

Measurement of the phospholipase activity of endothelial lipase in mouse plasma.

J Lipid Res 2013 Jan 26;54(1):282-9. Epub 2012 Oct 26.

Department of Cell Biology, State University of New York Downstate Medical Center, Brooklyn, NY 11203, USA.

Endothelial lipase (EL) is a major negative regulator of plasma HDL levels in mice, rabbits, and most probably, humans. Although this regulatory function is critically dependent on EL's hydrolysis of HDL phospholipids, as yet there is no phospholipase assay specific for EL in plasma. We developed such an assay for the mouse enzyme using a commercially available phospholipid-like fluorescent substrate in combination with an EL neutralizing antibody. The specificity of the assay was established using EL knockout mice and its utility demonstrated by detection of an increase in plasma EL phospholipase activity following exposure of wild-type mice to lipopolysaccharide. The assay revealed that murine pre-heparin plasma does not contain measurable EL activity, indicating that the hydrolysis of HDL phospholipids by EL in vivo likely occurs on the cell surface.
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http://dx.doi.org/10.1194/jlr.D031112DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3520535PMC
January 2013

Proteolytic processing of angiopoietin-like protein 4 by proprotein convertases modulates its inhibitory effects on lipoprotein lipase activity.

J Biol Chem 2011 May 12;286(18):15747-56. Epub 2011 Mar 12.

Department of Cell Biology, State University of New York Downstate Medical Center, Brooklyn, New York 11203, USA.

Angiopoietin-like protein 4 (ANGPTL4) has been associated with a variety of diseases. It is known as an endogenous inhibitor of lipoprotein lipase (LPL), and it modulates lipid deposition and energy homeostasis. ANGPTL4 is cleaved by unidentified protease(s), and the biological importance of this cleavage event is not fully understood with respect to its inhibitory effect on LPL activity. Here, we show that ANGPTL4 appears on the cell surface as the full-length form, where it can be released by heparin treatment in culture and in vivo. ANGPTL4 protein is then proteolytically cleaved into several forms by proprotein convertases (PCs). Several PCs, including furin, PC5/6, paired basic amino acid-cleaving enzyme 4, and PC7, are able to cleave human ANGPTL4 at a consensus site. PC-specific inhibitors block the processing of ANGPTL4. Blockage of ANGPTL4 cleavage reduces its inhibitory effects on LPL activity and decreases its ability to raise plasma triglyceride levels. In summary, the cleavage of ANGPTL4 by these PCs modulates its inhibitory effect on LPL activity.
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http://dx.doi.org/10.1074/jbc.M110.217638DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3091183PMC
May 2011

Determination of lipoprotein lipase activity using a novel fluorescent lipase assay.

J Lipid Res 2011 Apr 26;52(4):826-32. Epub 2011 Jan 26.

Department of Cell Biology, State University of New York Downstate Medical Center, Brooklyn, NY 11203, USA.

A novel, real-time, homogeneous fluorogenic lipoprotein lipase (LPL) assay was developed using a commercially available substrate, the EnzChek lipase substrate, which is solubilized in Zwittergent. The triglyceride analog substrate does not fluoresce, owing to apposition of fluorescent and fluorescent quenching groups at the sn-1 and sn-2 positions, respectively, fluorescence becoming unquenched upon release of the sn-1 BODIPY FA derivative following hydrolysis. Increase in fluorescence intensity at 37°C was proportional to LPL concentration. The assay was more sensitive than a similar assay using 1,2-O-dilauryl-rac-glycero-3-glutaric acid-(6-methylresorufin ester) and was validated in biological samples, including determination of LPL-specific activity in postheparin mouse plasma. The simplicity and reproducibility of the assay make it ideal for in vitro, high-throughput screening for inhibitors and activators of LPL, thus expediting discovery of drugs of potential clinical value.
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http://dx.doi.org/10.1194/jlr.D010744DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3284171PMC
April 2011
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