Dysfunctional HDL Research Articles

Abstract 434: LDL Apheresis Significantly Alters HDL Cargo and Function in Familial Hypercholesterolemia Subjects

In addition to cholesterol, HDL transports a wide-variety of cargo including vitamins, nucleic acids, and a diverse set of proteins. Outside of the reverse cholesterol transport pathway, HDL has alternative functions that contribute to its anti-atherogenicity, including anti-inflammatory, anti-oxidant, and signaling capacities. Recently, we found that HDL transports and delivers functional microRNAs to recipient cells, which likely confers HDL’s ability to suppress adhesion molecule expression in endothelial cells. As such, changes to HDL’s cargo impact many of these alternative functions, and thus its protective capacity. HDL dysfunction has been identified among patients with familial hypercholesterolemia (FH), an inherited disease due to mutations in the LDL receptor and associated with severe elevations in LDL-cholesterol (LDL-C) levels, which may necessitate LDL apheresis (LA) in management. Here we demonstrate that LA significantly alters HDL’s miRNA and protein signatures. We believe that these changes may have profound consequences on HDL’s protective capacity. Using density-gradient ultracentrifugation, we found 31 proteins to be significantly altered on HDL after LA, as determined by shotgun proteomics and multidimensional protein identification technology analysis. For example, vitamin D-binding protein (1.74-fold) was increased, while lipopolysaccharide-binding protein (-1.92-fold), platelet-activating factor acetylhydrolase (-2.06), and apolipoprotein A-V (-2.4-fold) were found to be decreased. Gene ontology and KEGG enrichment analysis demonstrated roles of these proteins in response to stress, coagulation, hemostasis, and vesicle-mediated transport. Fast-protein liquid chromatography was used to further purify HDL for miRNA profiling using TaqMan OpenArrays, and we found 8 HDL-miRNAs to be significantly altered -- 3 down (miR-302b -13.9-fold, miR-224 -1.8, miR-572 -2.4-fold) and 5 up (miR-7 9.8-fold, miR-208b 16.2-fold, miR-34a-3p 3-fold, miR-627 2.4-fold, miR-1183 4.8-fold). Both miR-302b and miR-224 decreased; these are miRNAs previously reported to suppress proliferation through targeting AKT2 and apoptosis, respectively. As such, these changes likely alter HDL’s function in FH subjects.

Abstract 447: Acrolein-Modified HDL Impairs Cholesterol Transport and HDL Function

The interaction between scavenger receptor BI (SR-BI) and HDL reduces the risk for cardiovascular disease (CVD) by playing key roles in the reverse cholesterol transport pathway (RCT). In the beginning of RCT, SR-BI stimulates release of free cholesterol (FC) from peripheral macrophages to HDL, whereas in the final step of RCT, HDL binds to hepatic SR-BI for delivery of excess cholesteryl esters (CE) into the liver for excretion via bile. While “functioning” HDL is crucial in protection against CVD, environmental factors such as acrolein, a highly reactive aldehyde found in tobacco smoke, can induce oxidative stress that promotes HDL modifications and renders the particle “dysfunctional”. Although there is a strong correlation between cigarette smoking and CVD risk, much less is known about the effects of smoking on RCT. We hypothesized that acrolein modifications to HDL (acro-HDL) generate a dysfunctional particle by: (i) impairing SR-BI-mediated selective uptake of HDL-CE and efflux of FC and (ii) upregulating inflammatory pathways in macrophages that promote atherogenesis. To test this hypothesis, human HDL was modified with 250μM acrolein and acrolein adducts on HDL were confirmed by immunoblot analysis and mass spectrometry. To evaluate acro-HDL’s functionality at the beginning of RCT, SR-BI-mediated FC release to native- or acro-HDL was measured in SR-BI-transfected COS-7 cells. Our data revealed that there was 30% less efflux of FC in the presence of acro-HDL as compared to native HDL. To assess acro-HDL functionality at the end of RCT, HDL binding and HDL-CE uptake assays were performed and acro-HDL (0-50μg/mL) was 10-25% less efficient at delivering CE to cells via SR-BI. Finally, to determine whether acro-HDL was rendered atherogenic, we used qRT-PCR to demonstrate that acro-HDL, but not native HDL, paralleled the effects of oxidized LDL by upregulating mRNA scavenger receptor expression (CD36 and LOX-1) and inflammatory marker expression (MCP-1, IL-6, ICAM-1) in THP-1 macrophages. Altogether, our data reveal that acrolein modification produces a dysfunctional HDL particle that upregulates inflammatory pathways. More detailed investigations into the impact of acrolein adducts on HDL function may help prevent smoking-related CVD deaths.

HDL from Patients with Inflammatory Disorders has High levels of Oxidized Lipids and is Dysfunctional (LB536)

HDL from Patients with Inflammatory Disorders has High levels of Oxidized Lipids and is Dysfunctional. Persiah Bakhtiari, Omeed El Boudvarej MD, Susan Hama, Nasim Pourtabatabaei MD, Sheila Safar MD, Samra Vazirian MD, Seyedehsara Seyedali MD, Sepideh Madahian MD, Greg Hough. David Geffen School of Medicine at UCLA, Los Angeles California, 90095 Population studies have shown that plasma levels of HDL inversely correlate with the prevalence of coronary heart disease. HDL levels in an individual does not necessarily correlate with the risk for coronary heart disease. Many patients with high HDL, present with coronary events and many with low HDL do not. We demonstrated that HDL composition and function is important than HDL levels. Under excessive inflammatory pressure unsaturated fatty acids can undergo oxidative modification. Protective enzymes including paraoxonase, LCAT and PLTP are inactivated by oxidative modification reducing HDL anti inflammatory properties. This results in HDL not being able to protect LDL from oxidative modification. Methods: Fasting plasma from patients with coronary disease were analyzed for their ability to prevent LDL modification. Additionally the content of oxidized lipids in HDL was determined using LC‐ESI‐MS/MS. Results: Among coronary patients over 69 percent had non protective HDL while among healthy controls 4.5 percent had dysfunctional HDL. The content of a variety of oxidized lipids in the HDL from patients was orders of magnitude higher than that found in HDL from normal Controls. Conclusion: Until there are effective medications for raising HDL, it should be clear to the public that the best way of dealing with low HDL cholesterol is lowering LDL cholesterol.

Dysfunctional HDL May Increase Heart Disease Risk

Dysfunctional HDL May Increase Heart Disease Risk

Dysfunctional HDL: the journey from savior to slayer

HDL has many functions such as reverse cholesterol transport, anti-inflammation and antioxidation. Different clinical conditions associated with inflammation, oxidation, advanced glycation and protein carbamylation can alter the functionality of HDL, converting normal HDL into dysfunctional HDL, which is no longer cardioprotective. Dysfunctional HDL leads to the impairment of reverse cholesterol transport and might exhibit proinflammatory, pro‑oxidant, prothrombotic and proapoptotic properties, all responsible for the subsequent endothelial dysfunction. This review addresses the current knowledge regarding the loss‑of‑function and gain‑of‑dysfunction responsible for the proatherosclerotic properties of HDL in several pathological conditions associated with systemic inflammation.

A8.28 Reconstituted high density lipoprotein (RHDL) modulates TH1 and TH17 immune responses and pro-inflammatory cytokine production in a murine model of rheumatoid arthritis

Background and ObjectivesHigh density lipoprotein (HDL) has a variety of functions which confer protection from cardiovascular and other human diseases. Increased cardiovascular risk is observed in humans with autoimmune disorders...

Carotid intima media thickness and low high-density lipoprotein (HDL) in South Asian immigrants: could dysfunctional HDL be the missing link?

IntroductionSouth Asian immigrants (SAIs) in the US exhibit higher prevalence of coronary artery disease (CAD) and its risk factors compared with other ethnic populations. Conventional CAD risk factors do not explain the excess CAD risk; therefore there is a need to identify other markers that can predict future risk of CAD in high-risk SAIs. The objective of the current study is to assess the presence of sub-clinical CAD using common carotid artery intima-media thickness (CCA-IMT), and its association with metabolic syndrome (MS) and pro-inflammatory/dysfunctional HDL (Dys-HDL).Material and methodsA community-based study was conducted on 130 first generation SAIs aged 35–65 years. Dys-HDL was determined using the HDL inflammatory index. Analysis was completed using logistic regression and Fisher's exact test.ResultsSub-clinical CAD using CCA-IMT ≥ 0.8 mm (as a surrogate marker) was seen in 31.46%. Age and gender adjusted CCA-IMT was significantly associated with type 2 diabetes (p = 0.008), hypertension (p = 0.012), high-sensitivity C-reactive protein (p < 0.001) and homocysteine (p = 0.051). Both the presence of MS and Dys-HDL was significantly correlated with CCA-IMT, even after age and gender adjustment. The odds of having Dys-HDL with CCA-IMT were 5 times (95% CI: 1.68, 10.78).ConclusionsThere is a need to explore and understand non-traditional CAD risk factors with a special focus on Dys-HDL, knowing that SAIs have low HDL levels. This information will not only help to stratify high-risk asymptomatic SAI groups, but will also be useful from a disease management point of view.

Markers of Systemic Inflammation and Apo-AI Containing HDL Subpopulations in Women with and without Diabetes.

Background. Besides their role in reverse cholesterol transport, HDL particles may affect the atherosclerotic process through the modulation of subclinical inflammation. HDL particles differ in size, composition, and, probably, anti-inflammatory properties. This hypothesis has never been explored in diabetic women, frequently having dysfunctional HDL. The potential relationship between lipid profile, Apo-AI containing HDL subclasses distribution, and common inflammatory markers (hsCRP, IL-6) was examined in 160 coronary heart disease- (CHD-) free women with and without type 2 diabetes. Results. Compared to controls, diabetic women showed lower levels of the atheroprotective large α-1, α-2, and pre-α-1 and higher concentration of the small, lipid-poor α-3 HDL particles (P < 0.05 all); diabetic women also had higher hsCRP and IL-6 serum levels (age- and BMI-adjusted P < 0.001). Overall, HDL subclasses significantly correlated with inflammatory markers: hsCRP inversely correlated with α-1 (P = 0.01) and pre-α-1 (P = 0.003); IL-6 inversely correlated with α-1 (P = 0.003), α-2 (P = 0.004), and pre-α-1 (P = 0.002) and positively with α-3 HDL (P = 0.03). Similar correlations were confirmed at univariate regression analysis. Conclusions. More atheroprotective HDL subclasses are associated with lower levels of inflammatory markers, especially in diabetic women. These data suggest that different HDL subclasses may influence CHD risk also through the modulation of inflammation.

HDL-Associated Paraoxonase 1 as a Bridge between Postmenopausal Osteoporosis and Cardiovascular Disease.

The association of postmenopausal osteoporosis (PMOP) with both atherosclerosis and vascular/valvular calcification is well known. Recently, ample evidence has suggested a common etiologic factor, namely, reduced HDL-associated paraoxonase 1 (PON1) activity, as a causative factor in the development of PMOP and cardiovascular disease (CVD). This common etiologic factor not only contributes to atherosclerotic diseases but also to PMOP following an almost identical mechanism including dysfunctional HDL and lipid oxidation. According to recent studies, lipid oxidation might improve osteoblastic transformation of vascular cells and obstruct such transformation in bone cells. The primary objective of this current review was to summarize the evidence revealing the role of HDL-associated PON1 enzyme in PMOP. Additionally, the review aimed to address some of the subjects that need further investigation in order to define whether hyperhomocysteinemia and sensitivity to lipid oxidation may be risk factors for PMOP.

Urokinase-type plasminogen activator (uPA) decreases hepatic SR-BI expression and impairs HDL-mediated reverse cholesterol transport

ObjectivesThe aim of the present study was to investigate the effect of urokinase-type plasminogen activator (uPA) on the expression of the scavenger receptor class B type I (SR-BI) in hepatocytes, and its impact on the removal of HDL-cholesteryl ester (CE) in the liver. Methods and resultsHuh7 hepatoma cell lines were incubated with increasing concentrations of uPA. uPA dose-dependently decreased SR-BI protein expression, as determined by flow cytometry (FACS) and by Western blot assays, and down-regulated SR-BI gene expression. Functionally, uPA decreased both the cellular binding of HDL to Huh7 hepatocytes, and the selective uptake of CE from HDL, as determined by several methods including BODIPY staining, cellular cholesterol determination and chasing radio-labeled CE transfer from HDL to the cells. These results were further confirmed using primary rat hepatocyes. The effect of uPA on hepatic SR-BI expression was mediated via binding to the uPA receptor (uPAR). In vivo, SR-BI protein and gene expressions were found to be increased in hepatocytes derived from the uPAR-KO mice compared to C57Bl/6 mice, and in parallel HDL-cholesterol levels in plasma derived from uPAR-KO mice were decreased. Moreover, deficiency of uPAR significantly accelerated the plasma decay of injected HDL-[3H]CE. ConclusionsThe results of this study suggest that uPA decreases the removal of HDL-CE in the liver via suppression of the hepatic SR-BI expression. Impaired reverse cholesterol transport (RCT) may result in atherogenic dysfunctional HDL metabolism and may contribute to atherosclerosis development.

High density lipoproteins and endothelial functions: mechanistic insights and alterations in cardiovascular disease

Prospective population studies in the primary prevention setting have shown that reduced plasma levels of HDL cholesterol are associated with an increased risk of coronary disease and myocardial infarction. Experimental and translational studies have further revealed several potential anti-atherogenic effects of HDL, including protective effects on endothelial cell functions. HDL has been suggested to protect endothelial cell functions by prevention of oxidation of LDL and its adverse endothelial effects. Moreover, HDL from healthy subjects can directly stimulate endothelial cell production of nitric oxide and anti-inflammatory, anti-apoptotic, and anti-thrombotic effects as well as endothelial repair processes. However, several recent clinical trials using HDL cholesterol-raising agents, such as torcetrapib, dalcetrapib, and niacin, did not demonstrate a significant reduction of cardiovascular events in patients with coronary disease. Of note, growing evidence suggests that the vascular effects of HDL can be highly heterogeneous and vasoprotective properties of HDL are altered in patients with coronary disease. Characterization of underlying mechanisms and understanding of the clinical relevance of this "HDL dysfunction" is currently an active field of cardiovascular research. Notably, in some recent studies no clear association of higher HDL cholesterol levels with a reduced risk of cardiovascular events was observed in patients with already established coronary disease. A greater understanding of mechanisms of action of HDL and its altered vascular effects is therefore critical within the context of HDL-targeted therapies. In this review, we will address different effects of HDL on endothelial cell functions potentially relevant to atherosclerotic vascular disease and explore molecular mechanisms leading to "dysfunctional HDL".

Distinct Genetic Backgrounds in Quantitative Traits Preceding Type 2 Diabetes: One Reason for Missing Heritability

Genome-wide association studies (GWAS) have identified variants associated with type 2 diabetes (T2D), but less than five percent of the genetic variance associated with T2D is accounted for in GWAS studies performed using T2D as endpoint. These findings suggest relevance of assessing preceding quantitative traits (QTs) or sub-phenotypes and their respective markers for contributing to the appearance of glycemic thresholds defining T2D. The QTs first appear in gestation and childhood when birth weight and childhood weight gain have an influence on T2D. Changes in fetal growth are associated with metabolic programming for T2D in adulthood. Subsequently gains in body fat leading to obesity, ectopic lipid deposition in liver and muscle, dyslipidemia and hypertension express in the β-cells, hypothalamus, adipocytes, myocytes, liver and kidney, and are associated with worsening insulin resistance and β-cell failure preceding overt T2D. Obesity, an increasingly common trait is associated with gene variants that regulate energy balance but have no association with T2D with few exceptions. For instance, FTO gene variants are associated with early onset of obesity and have also been associated with T2D and hypertension suggesting pleiotropism. Non-alcoholic fatty liver disease (NAFLD) has an independent genetic background and is considered as a new addition to the metabolic syndrome. Association of NAFLD with dyslipidemia, Review Article British Journal of Medicine & Medical Research, 4(2): 590-620, 2014 591 cardiovascular disease and hepatic insulin resistance contributes to progression to T2D. The classic dyslipidemia encountered in insulin resistant states consisting of increased triglyceride, low high-density lipoprotein cholesterol (HDL-C) and molding of triglyceridecontaining lipoproteins to form atherogenic LDL particles and dysfunctional HDL particles, has strong environmental and genetic association. In addition evidence is accumulating to show that the abnormal lipoproteins have a direct effect on β-cells leading to failure and possible acceleration of diabetes onset. Consequently known variants affecting either the dyslipidemia or β-cell function have a compound effect on T2D. Like other preceding traits the genetic background for commonly encountered hypertension is independent of diabetes although it remains a predictive trait. However, increasing evidence supports a role for the renin-angiotensin system in causing oxidative stress and insulin resistance. Finally when glucose intolerance has progressed to overt T2D, reversibility is usually no longer possible; therefore recognition of preceding prediabetic glycemic thresholds and both their genetic and environmental associations may facilitate and serve as signals for prevention. Thus a better understanding of the sequential expression of the respective QTs and genetic variation affecting their metabolic effects leading to T2D could initiate more effective prevention strategies.

Functionally Defective High-Density Lipoprotein and Paraoxonase: A Couple for Endothelial Dysfunction in Atherosclerosis

The endothelium is the primary target for biochemical or mechanical injuries caused by the putative risk factors of atherosclerosis. Endothelial dysfunction represents the ultimate link between atherosclerotic risk factors that promote atherosclerosis. HDL-C is thought to exert at least some parts of its antiatherogenic facilities via stimulating endothelial NO production, nearby inhibiting oxidative stress and inflammation. HDL-C is capable of opposing LDL's inductive effects and avoiding the ox-LDL's inhibition of eNOS. Paraoxonase 1 (PON1) is an HDL-associated enzyme esterase which appears to contribute to the antioxidant and antiatherosclerotic capabilities of HDL-C. “Healthy HDL,” namely the particle that contains the active Paraoxonase 1, has the power to suppress the formation of oxidized lipids. “Dysfunctional HDL,” on the contrary, has reduced Paraoxonase 1 enzyme activity and not only fails in its mission but also potentially leads to greater formation of oxidized lipids/lipoproteins to cause endothelial dysfunction. The association of HDL-C PON1 and endothelial dysfunction depends largely on the molecules with exact damaging effect on NO synthase coupling. Loss of nitric oxide bioavailability has a pivotal role in endothelial dysfunction preceding the appearance of atherosclerosis. Analyses of HDL-C and Paraoxonase1 would be more important in the diagnosis and treatment of atherosclerosis in the very near future.

Apolipoprotein A-II is a key regulatory factor of HDL metabolism as appears from studies with transgenic animals and clinical outcomes

The structure and metabolism of HDL are linked to their major apolipoproteins (apo) A-I and A-II. HDL metabolism is very dynamic and depends on the constant remodeling by lipases, lipid transfer proteins and receptors. HDL exert several cardioprotective effects, through their antioxidant and antiinflammatory capacities and through the stimulation of reverse cholesterol transport from extrahepatic tissues to the liver for excretion into bile. HDL also serve as plasma reservoir for C and E apolipoproteins, as transport vehicles for a great variety of proteins, and may have more physiological functions than previously recognized. In this review we will develop several aspects of HDL metabolism with emphasis on the structure/function of apo A-I and apo A-II. An important contribution to our understanding of the respective roles of apo A-I and apo A-II comes from studies using transgenic animal models that highlighted the stabilizatory role of apo A-II on HDL through inhibition of their remodeling by lipases. Clinical studies coupled with proteomic analyses revealed the presence of dysfunctional HDL in patients with cardiovascular disease. Beyond HDL cholesterol, a new notion is the functionality of HDL particles. In spite of abundant literature on HDL metabolic properties, a major question remains unanswered: which HDL particle(s) confer(s) protection against cardiovascular risk?

Impaired HDL function in obese adolescents: Impact of lifestyle intervention and bariatric surgery

HDL regulates endothelial function via stimulation of nitric oxide production. It is documented that endothelial function is impaired in obese adolescents, and improved by lifestyle interventions (LI). HDL function in obese adolescents and the impact of LI or Roux-en-Y gastric bypass surgery (RYGB) was assessed. HDL was isolated from 14 adolescents with normal body mass index (HDLcontrol ), 10 obese (HDLobese ) before and after 6 month LI, and five severe obese adolescents before and one year after RYGB. HDL-mediated phosphorylation of endothelial nitric oxide synthase (eNOS)-Ser(1177) , eNOS-Thr(495) , and PKC-ßII was evaluated. In addition the HDL proteome was analyzed. HDLobese -mediated eNOS-Ser(1177) phosphorylation was reduced, whereas eNOS-Thr(495) phosphorylation increased significantly when compared to HDLcontrol . No impact of obesity was observed on PKC-ßII phosphorylation. LI and RYGB had no impact on HDL-mediated phosphorylation of eNOS and PKC-ßII. A principle component plot analysis of the HDL particle separated controls and severe obese, whereas the interventions did not trigger sufficient differences to the HDL proteome to permit distinction. These results demonstrated that HDL-function is impaired in obese adolescents, and that LI or RYGB did not correct this dysfunction. This might be an argument for developing earlier prevention strategies in obese adolescents to avoid HDL dysfunction.

SP0145 HDL as a Pro-Inflammatory Agent

Although HDL levels are traditionally inversely related to atherosclerosis, the relationship is complex and involves both HDL quantity and function. HDL has been described as “chameleon-like,” anti-inflammatory in the basal...

Abstract 124: Increased Selective Clearance of Cholesteryl-Ester in Patients with Rheumatoid Arthritis is Reversed by Treatment with Tofacitinib

Inflammation is associated with dysfunctional HDL. Rheumatoid arthritis (RA) is a chronic inflammatory disease with significantly elevated cardiovascular risk in spite of low LDL and HDL cholesterol (C) levels. Using stable isotope methods, we examined underlying metabolic mechanisms for the reduced cholesterol levels in active RA and for the changes with tofacitinib (tofa) treatment. Apolipoprotein and cholesterol kinetics were assessed in 36 RA patients at baseline and following 6 weeks of tofa treatment compared to baseline values in 33 non-RA controls matched for age, gender, and menopausal status. All subjects received constant IV infusions of [3,4] 13 C 2 cholesterol and U- 13 C 6 leucine with collection of fasting blood samples hourly for 22 hours. Whole-body cholesterol efflux and cholesteryl ester (CE) and apolipoprotein production and fractional catabolic rates (FCR) were determined by modeling isotope enrichments. At baseline, total-C, HDL-C, LDL-C, CE and apoA-I concentrations were lower and CE FCR was elevated in RA. No differences were seen in CE production rate, whole body cholesterol efflux or HDL-apoA-I and LDL-apoB production and FCRs. RA patients had elevated serum HDL-SAA, myeloperoxidase and reduced in vitro LCAT activity and mass. Following 6 weeks of tofa therapy, cholesterol and apoAI approached levels of controls and apoB concentrations increased. Tofa treatment reduced CE FCR, increased HDL-apoA-I production rate with no changes in LDL-apoB production rates or LDL-apoB and HDL-apoA-I FCRs. Increased CE FCR in the absence of changes in HDL-apoA-I or LDL-apoB particle clearance suggest increased selective CE uptake, presumably by SR-B1, which was normalized by tofa. Change in CE FCR with treatment was negatively correlated with changes in HDL size and concentration of large HDL particles, but not with LDL size or composition. Change in CE production rate with treatment was positively correlated with change in LCAT mass. This is the first study to evaluate the effect of chronic inflammation on in vivo cholesterol and lipoprotein kinetics in humans. Inflammation increases selective CE uptake explaining the reduced cholesterol levels in RA. Reducing inflammation with tofa restored selective CE uptake to normal rates.

Abstract 405: Therapeutic Potential of Oscimum Tenuiflorum in Prevention and Treatment of Atherosclerosis

Background Current experimental and clinical studies show that Oscimum tenuiflorum (commonly known as basil or Tulsi) possess many health benefits. Oscimum is also suggested to be anti-oxidative and anti-inflammatory. Eugenol, an ortho-methoxy phenol, is one of its components. Myeloperoxidase (MPO), an oxidative enzyme, has been implicated in the pathogenesis of atherosclerosis. MPO dependent oxidation of lipoproteins has been implicated in foam cell formation, dysfunctional HDL and abnormalities in reverse cholesterol transport (RCT). In this study we tested whether eugenol and basil extract could inhibit the oxidation of lipoproteins (LDL &amp; HDL) by MPO. Hypothesis Whole leaf extract of Ocimum tenuiflorum and eugenol would inhibit the oxidation of LDL and HDL by myeloperoxidase and by copper. Methods Basil leaf extract was prepared with water, lyophilized and reconstituted in water. Lipoproteins (LDL &amp; HDL) were isolated from human plasma and their oxidation was performed using copper or MPO in the presence or absence of the basil extract and eugenol. MPO activity was measured using guaiacol and TMB as substrates. Results Basil extract and eugenol inhibited MPO enzyme activity against synthetic substrates. In addition, they also inhibited the oxidation of LDL and HDL both by copper and MPO, as measured by conjugated diene formation, TBARS formation, as well as by electrophoretic mobility assays. Conclusion In lieu of the reported anti-inflammatory properties of basil leaves, and the importance of oxidative and inflammatory pathways in atherosclerosis, such simple non-pharmacological agents could prove to be valuable adjunt therapeutic agents in the treatment of cardiovascular diseases.

Abstract 548: Scavenger Receptor Bi-deficiency-induced Hypercholesterolemia Impairs Lymphocyte Homeostasis

Objective- Deficiency in Scavenger receptor BI (SR-BI), an HDL receptor, leads to a specific HDL-c-elevated hypercholesterolemia and impairs lymphocyte homeostasis in mice. However, how SR-BI-deficiency-induced hypercholesterolemia contributes to the adaptive immunity defects remains unknown. Hypothesis- SR-BI-deficiency-induced hypercholesterolemia leads to the impaired lymphocyte homeostasis. Methods and Results- We utilized two animal models combined with cell culture to test our hypothesis. First, in a high fat/high cholesterol (HF) diet mouse model (n&gt;10 per group), we introduced a 2-fold increase in plasma cholesterol concentration. For HF diet SR-BI +/+ mice, the increased cholesterol was distributed in VLDL/LDL fraction, but the lymphocyte homeostasis was not affected. In contrast, for HF diet SR-BI -/- mice, in addition to VLDL/LDL-c, HDL-c was also further elevated. These mice exhibited marked disruption in lymphocyte homeostasis, as characterized by a 1.6-fold increase in T cell number (2.7-fold vs SR-BI +/+ group), a 2.1-fold increase in B cell number (4.2-fold vs SR-BI +/+ group), exacerbated T cell activation and further accelerated B cell proliferation compared with control diet SR-BI -/- mice. Then, in a bone marrow transplantation model (n=5~6 per group), we found that HF diet SR-BI -/- recipients whether receiving SR-BI +/+ or SR-BI -/- BMCs displayed higher lymphocyte numbers, greater lymphocyte activation and more lymphocyte proliferation than respective HF diet SR-BI +/+ controls, indicating that non-hematopoietic SR-BI-deficiency environment was the dominant factor leading to the impaired adaptive immunity. Finally, in a series of in vitro lymphocyte activation experiments (n=4 per group, repeated twice), we found that HDL from control diet or HF diet SR-BI +/+ mice exhibited an inhibitory effect on T cell or B cell proliferation under TCR or BCR stimulation, but HDL from HF diet SR-BI -/- mice completely lost that function, indicating that the dysfunctional HDL in HF diet SR-BI -/- mice was responsible for impaired lymphocyte homeostasis. Conclusion- The SR-BI-deficiency-induced hypercholesterolemia is the leading factor for impaired lymphocyte homeostasis in SR-BI -/- mice.

Abstract 119: LCAT Activation in Combination with the CETP Inhibitor Anacetrapib Results in Enhanced Cholesterol Efflux and a Favorable Lipid Profile in Dyslipidemic Hamsters

Lecithin cholesterol acyltransferase (LCAT) is a key enzyme involved in the metabolism of HDL. LCAT synthesizes cholesterol esters (CE) in plasma by catalyzing the transfer of the sn-2 fatty acid of phosphatidylcholine to cholesterol on HDL. The hydrophobic CE moves to the core of the HDL particle which results in HDL maturation and a progressive enlargement of the particle. Cholesteryl ester transfer protein (CETP) is a plasma protein that facilitates the transport of CE from HDL to ApoB-containing lipoproteins. CETP inhibition has been identified as a potential strategy for lowering LDL-c and raising HDL cholesterol levels for the treatment of CVD. The purpose of the current study was to test the hypothesis that CETP inhibition in the context of LCAT activation might yield an unfavorable lipoprotein profile (e.g. dysfunctional HDL). We measured plasma lipoprotein (via FPLC), particle size and ex vivo cholesterol efflux from dyslipidemic hamsters chronically treated with a small molecule LCAT activator (20 and 60 mpk/day) in the presence or absence of the CETP inhibitor anacetrapib (20mpk/day). As expected, anacetrapib significantly reduced LDL-c (-45%) and increased HDL-c (+63%) after chronic treatment. LCAT activation reduced TG (-35%) and increased HDL-c (+40-68%) but had no effect on LDL-c. Both treatments led to an apparent increase in HDL particle size as measured by FPLC. Additionally, LCAT activation showed a significant reduction in ABCA1-mediated efflux, presumably due to the resulting increase in HDL particle size. Animals that received both anacetrapib and the LCAT activator maintained the favorable lipid profile observed with single treatment. Moreover, HDL from hamsters treated with both LCATa and CETPi demonstrated a significant increase in ABCG1- and SRB1-mediated efflux whereas single treatment did not exhibit a measurable effect. We provide evidence that LCAT activation in the context of a CETP inhibitor, maintains a favorable lipid profile (lower LDL-c) and appear s to have a synergistic effect on cholesterol efflux.