Apolipoprotein A-I Mimetic Peptide Research Articles

Apolipoprotein AI mimetic peptide CS-6253 increases the atherosclerotic lesion stability index independent of leukocyte abca1 in bone marrow-derived cells

Apolipoprotein AI mimetic peptide CS-6253 increases the atherosclerotic lesion stability index independent of leukocyte abca1 in bone marrow-derived cells

High-Density Lipoprotein Mimetic Peptide 4F Efficiently Crosses the Blood-Brain Barrier and Modulates Amyloid-β Distribution between Brain and Plasma.

Treatments to elevate high-density lipoprotein (HDL) levels in plasma have decreased cerebrovascular amyloid -β (Aβ) deposition and mitigated cognitive decline in Alzheimer disease (AD) transgenic mice. Since the major protein component of HDL particles, apolipoprotein A-I (ApoA-I), has very low permeability at the blood-brain barrier (BBB), we investigated 4F, an 18-amino-acid ApoA-I/HDL mimetic peptide, as a therapeutic alternative. Specifically, we examined the BBB permeability of 4F and its effects on [125I]Aβ trafficking from brain to blood and from blood to brain. After systemic injection in mice, the BBB permeability of [125I]4F, estimated as the permeability-surface area (PS) product, ranged between 2 and 5 × 10-6 ml/g per second in various brain regions. The PS products of [125I]4F were ∼1000-fold higher compared with those determined for [125I]ApoA-I. Moreover, systemic infusion with 4F increased the brain efflux of intracerebrally injected [125I]Aβ42. Conversely, 4F infusion decreased the brain influx of systemically injected [125I]Aβ42. Interestingly, 4F did not significantly alter the brain influx of [125I]Aβ40. To corroborate the in vivo findings, we evaluated the effects of 4F on [125I]Aβ42 transcytosis across polarized human BBB endothelial cell (hCMEC/D3) monolayers. Treatment with 4F increased the abluminal-to-luminal flux and decreased the luminal-to-abluminal flux of [125I]Aβ42 across the hCMEC/D3 monolayers. Additionally, 4F decreased the endothelial accumulation of fluorescein-labeled Aβ42 in the hCMEC/D3 monolayers. These findings provide a mechanistic interpretation for the reductions in brain Aβ burden reported in AD mice after oral 4F administration, which represents a novel strategy for treating AD and cerebral amyloid angiopathy. SIGNIFICANCE STATEMENT: The brain permeability of the ApoA-I mimetic peptide 4F was estimated to be ∼1000-fold greater than ApoA-I after systemic injection of radiolabeled peptide/protein in mice. Further, 4F treatment increased the brain efflux of amyloid -β and also decreased its brain influx, as evaluated in mice and in blood-brain barrier cell monolayers. Thus, 4F represents a potential therapeutic strategy to mitigate brain amyloid accumulation in cerebral amyloid angiopathy and Alzheimer disease.

ABCA1 (ATP-Binding Cassette Transporter A1) Mediates ApoA-I (Apolipoprotein A-I) and ApoA-I Mimetic Peptide Mobilization of Extracellular Cholesterol Microdomains Deposited by Macrophages.

We examined the function of ABCA1 (ATP-binding cassette transporter A1) in ApoA-I (apolipoprotein A-I) mobilization of cholesterol microdomains deposited into the extracellular matrix by cholesterol-enriched macrophages. We have also determined whether an ApoA-I mimetic peptide without and with complexing to sphingomyelin can mobilize macrophage-deposited cholesterol microdomains. Extracellular cholesterol microdomains deposited by cholesterol-enriched macrophages were detected with a monoclonal antibody, 58B1. ApoA-I and an ApoA-I mimetic peptide 5A mobilized cholesterol microdomains deposited by ABCA1+/+ macrophages but not by ABCA1-/- macrophages. In contrast, ApoA-I mimetic peptide 5A complexed with sphingomyelin could mobilize cholesterol microdomains deposited by ABCA1-/- macrophages. Our findings show that a unique pool of extracellular cholesterol microdomains deposited by macrophages can be mobilized by both ApoA-I and an ApoA-I mimetic peptide but that mobilization depends on macrophage ABCA1. It is known that ABCA1 complexes ApoA-I and ApoA-I mimetic peptide with phospholipid, a cholesterol-solubilizing agent, explaining the requirement for ABCA1 in extracellular cholesterol microdomain mobilization. Importantly, ApoA-I mimetic peptide already complexed with phospholipid can mobilize macrophage-deposited extracellular cholesterol microdomains even in the absence of ABCA1.

Apolipoprotein A-I inhibits experimental colitis and colitis-propelled carcinogenesis.

In both humans with long-standing ulcerative colitis and mouse models of colitis-associated carcinogenesis (CAC), tumors develop predominantly in the distal part of the large intestine but the biological basis of this intriguing pathology remains unknown. Herein we report intrinsic differences in gene expression between proximal and distal colon in the mouse, which are augmented during dextran sodium sulfate (DSS)/azoxymethane (AOM)-induced CAC. Functional enrichment of differentially expressed genes identified discrete biological pathways operating in proximal vs distal intestine and revealed a cluster of genes involved in lipid metabolism to be associated with the disease-resistant proximal colon. Guided by this finding, we have further interrogated the expression and function of one of these genes, apolipoprotein A-I (ApoA-I), a major component of high-density lipoprotein. We show that ApoA-I is expressed at higher levels in the proximal compared with the distal part of the colon and its ablation in mice results in exaggerated DSS-induced colitis and disruption of epithelial architecture in larger areas of the large intestine. Conversely, treatment with an ApoA-I mimetic peptide ameliorated the phenotypic, histopathological and inflammatory manifestations of the disease. Genetic interference with ApoA-I levels in vivo impacted on the number, size and distribution of AOM/DSS-induced colon tumors. Mechanistically, ApoA-I was found to modulate signal transducer and activator of transcription 3 (STAT3) and nuclear factor-κB activation in response to the bacterial product lipopolysaccharide with concomitant impairment in the production of the pathogenic cytokine interleukin-6. Collectively, these data demonstrate a novel protective role for ApoA-I in colitis and CAC and unravel an unprecedented link between lipid metabolic processes and intestinal pathologies.

The apolipoprotein-AI mimetic peptide L4F at a modest dose does not attenuate weight gain, inflammation, or atherosclerosis in LDLR-null mice.

ObjectiveHigh density lipoprotein (HDL) cholesterol levels are inversely related to cardiovascular disease risk and associated with a reduced risk of type 2 diabetes. Apolipoprotein A-I (apoA-I; major HDL protein) mimetics have been reported to reduce atherosclerosis and decrease adiposity. This study investigated the effect of L4F mimetic peptide and apoA-I overexpression on weight gain, insulin resistance, and atherosclerosis in an LDL receptor deficient (Ldlr-/-) model fed a high fat high sucrose with cholesterol (HFHSC) diet.MethodsStudies in differentiated 3T3-L1 adipocytes tested whether L4F could inhibit palmitate-induced adipocyte inflammation. In vivo studies used male Ldlr-/- mice fed a HFHSC diet for 12 weeks and were injected daily with L4F (100 µg/mouse) subcutaneously during the last 8 weeks. Wild-type and apoA-I overexpressing Ldlr-/- mice were fed HFHSC diet for 16 weeks.ResultsNeither L4F administration nor apoA-I overexpression affected weight gain, total plasma cholesterol or triglycerides in our studies. While pre-treatment of 3T3-L1 adipocytes with either L4F or HDL abolished palmitate-induced cytokine expression in vitro, L4F treatment did not affect circulating or adipose tissue inflammatory markers in vivo. Neither L4F administration nor apoA-I overexpression affected glucose tolerance. ApoA-I overexpression significantly reduced atherosclerotic lesion size, yet L4F treatment did not affect atherosclerosis.ConclusionOur results suggest that neither L4F (100 µg/day/mouse) nor apoA-I overexpression affects adiposity or insulin resistance in this model. We also were unable to confirm a reduction in atherosclerosis with L4F in our particular model. Further studies on the effect of apoA-I mimetics on atherosclerosis and insulin resistance in a variety of dietary contexts are warranted.

Abstract W MP38: Apolipoprotein A-I (ApoA-I) Mimetic Peptide, D-4F, Regulates miRNA Expression and Improves Functional Outcome after Stroke in Diabetic Rats

Introduction: Diabetes mellitus (DM) in patients is associated with low high-density lipoprotein cholesterol (HDL-C) and impairment of the anti-oxidative capacity of HDL-C. Several miRNAs have been identified as having a physiological role in tissues in which diabetes complications occur. D-4F is an economical ApoA-I mimetic peptide which increases HDL function. We therefore investigated the therapeutic effect and underlying mechanisms of D-4F treatment of stroke induced functional benefit effects in type one DM (T1DM) rats. Methods: T1DM was induced in Wistar rats by streptozotocin (60mg/kg, ip) .Subsequently they were subjected to embolic middle cerebral artery occlusion. D-4F (10 mg/kg i.p.) was administered at 2h, 24h and 48h after stroke. Functional outcomes, brain blood barrier (BBB) leakage and lesion volume were evaluated. Results: D-4F treatment of stroke in T1DM rats significantly decreased Evans Blue dye leakage (15.85±1.52 ng/mg vs contro: 30.57± 5.88 ng/mg) and significantly improved functional outcome compared to non-treatment T1DM-control. D-4F treatment significantly increased miR-124a and miR-181c, but decreased miR-200b expression in the ischemic brain. Using laser capture, D-4F also significantly increased ischemic brain endothelial cell miR-126 expression compared to non-treatment control. In addition, D-4F treatment significantly decreased miR-181c target gene tumor necrosis factor-a (TNF-a), and miR-124a target gene monocyte chemoattractant protein-1 (MCP1) expression. D-4F also decreased proinflammatory nuclear NFkB (26.07±3.17 vs 37.43±3.47), Toll-like receptor 4 (TLR4, 7.26±0.69 vs 10.5±1.2), matrix metalloproteinase 9 (MMP9, 0.16±0.04 vs 0.48±0.09) expression, and increased CD163 (M2 macrophage marker, 120.98±8.13 vs 85.02± 9.47) number compared to T1DM-MCAo control rats. Conclusion: D-4F treatment of stroke in T1DM rats regulates miRNA expression and their target gene and protein expression and induces anti-inflammatory effects and promotes M2 macrophage polarization which may contribute to D-4F decreased BBB leakage and induced functional benefit effects after stroke in T1DM rats.

Improvement of aortic valve stenosis by ApoA‐I mimetic therapy is associated with decreased aortic root and valve remodelling in mice

We have shown that infusions of apolipoprotein A-I (ApoA-I) mimetic peptide induced regression of aortic valve stenosis (AVS) in rabbits. This study aimed at determining the effects of ApoA-I mimetic therapy in mice with calcific or fibrotic AVS. Apolipoprotein E-deficient (ApoE(-/-) ) mice and mice with Werner progeria gene deletion (Wrn(Δhel/Δhel) ) received high-fat diets for 20 weeks. After developing AVS, mice were randomized to receive saline (placebo group) or ApoA-I mimetic peptide infusions (ApoA-I treated groups, 100 mg·kg(-1) for ApoE(-/-) mice; 50 mg·kg(-1) for Wrn mice), three times per week for 4 weeks. We evaluated effects on AVS using serial echocardiograms and valve histology. Aortic valve area (AVA) increased in both ApoE(-/-) and Wrn mice treated with the ApoA-I mimetic compared with placebo. Maximal sinus wall thickness was lower in ApoA-I treated ApoE(-/-) mice. The type I/III collagen ratio was lower in the sinus wall of ApoA-I treated ApoE(-/-) mice compared with placebo. Total collagen content was reduced in aortic valves of ApoA-I treated Wrn mice. Our 3D computer model and numerical simulations confirmed that the reduction in aortic root wall thickness resulted in improved AVA. ApoA-I mimetic treatment reduced AVS by decreasing remodelling and fibrosis of the aortic root and valve in mice.

4F, apolipoprotein AI mimetic peptide, attenuates acute lung injury and improves survival in endotoxemic rats

4F, apolipoprotein AI mimetic peptide, mimics anti-inflammatory properties of high-density lipoprotein (HDL). The aim of this study was to investigate whether 4F attenuates acute lung injury and improves survival in endotoxemic rats and to determine whether the therapeutic benefits of 4F are associated with the stimulation of sphingosine-1-phosphate receptor 1 (S1P1), the activation of Akt, the down-regulation of the nuclear factor-κB (NF-κB) pathway, and the suppression of cell adhesion molecules. To induce endotoxemia in rats, lipopolysaccharide (LPS, 10 mg/kg) was injected into a tail vein and 10 minutes later, vehicle or 4F (10 mg/kg) was administered intraperitoneally, respectively. We observed the survival of subjects for 72 hours. At 6-hour post-LPS, we killed animals and measured S1P1 expression, phosphorylated Akt/Akt ratio, cytoplasmic phosphorylated inhibitor κB-α/inhibitor κB-α ratio, nuclear NF-κB p65 expression and DNA-binding activity, endothelial leukocyte adhesion molecule-1 (E-selectin) and intercellular adhesion molecule-1 expression, myeloperoxidase activity, and histologic damages in lung tissues. We also measured serum HDL cholesterol level. 4F improved survival in endotoxemic rats. 4F restored LPS-induced diminution of serum HDL cholesterol level and increased lung S1P1 expression and phosphorylated Akt/Akt ratio in LPS-treated rats. Furthermore, 4F suppressed inhibitor κB-α degradation, NF-κB activation, E-selectin and intercellular adhesion molecule-1 expression, and myeloperoxidase activity, and attenuated histologic damages in lung tissues. 4F attenuated acute lung injury and improved survival in endotoxemic rats. The therapeutic benefits of 4F were found to be associated with the stimulation of S1P1, the activation of Akt, the down-regulation of the NF-κB pathway, and the suppression of cell adhesion molecules.

ApoA-1 mimetic restores adiponectin expression and insulin sensitivity independent of changes in body weight in female obese mice

Background:We examined the ability of the apolipoprotein AI mimetic peptide L-4F to improve the metabolic state of female and male ob mice and the mechanisms involved.Methods:Female and male lean and obese (ob) mice were administered L-4F or vehicle for 6 weeks. Body weight was measured weekly. Fat distribution, serum cytokines and markers of cardiovascular dysfunction were determined at the end of treatment.Results:L-4F significantly decreased serum interleukin (IL)-6, tumor necrosis factor-α and IL-1β. L-4F improved vascular function, and increased serum adiponectin levels and insulin sensitivity compared with untreated mice. In addition, L-4F treatment increased heme oxygenase (HO)-1, pAKT and pAMPK levels in kidneys of ob animals. pAKT and pAMPK levels were significantly reduced in the presence of an HO inhibitor. Interestingly, L4F did not alter body weight in female mice, but caused a significant reduction in males.Conclusions:L-4F treatments reduced cardiovascular risk factors and improved insulin sensitivity in female ob mice independent of body fat changes. Reduced inflammatory cytokine levels accompanied by increased HO activity, serum adiponectin and improved insulin sensitivity suggest that L-4F may promote the conversion of visceral fat to a healthier phenotype. Therefore, L-4F appears to be a promising therapeutic strategy for treating both cardiovascular risk factors and insulin resistance in obese patients of either gender.

Apolipoprotein A-I (ApoA-I) Mimetic Peptide P2a by Restoring Cholesterol Esterification Unmasks ApoA-I Anti-Inflammatory Endogenous Activity In Vivo

The acute-phase protein haptoglobin (Hpt) binds apolipoprotein A-I (ApoA-I) and impairs its action on lecithin-cholesterol acyltransferase, an enzyme that plays a key role in reverse cholesterol transport. We have previously shown that an ApoA-I mimetic peptide, P2a, displaces Hpt from ApoA-I, restoring the enzyme activity in vitro. The aim of this study was to evaluate whether P2a displaces Hpt from ApoA-I in vivo and whether this event leads to anti-inflammatory activity. Mice received subplantar injections of carrageenan. Paw volume was measured before the injection and 2, 4, 6, 24, 48, 72, and 96 h thereafter. At the same time points, concentrations of HDL cholesterol (C) and cholesterol esters (CEs) were measured by high-performance liquid chromatography, and Hpt and ApoA-I plasma levels were evaluated by enzyme-linked immunosorbent assay. Western blotting analysis for nitric-oxide synthase and cyclooxygenase (COX) isoforms was also performed on paw homogenates. CEs significantly decreased in carrageenan-treated mice during edema development and negatively correlated with the Hpt/ApoA-I ratio. P2a administration significantly restored the CE/C ratio. In addition, P2a displayed an anti-inflammatory effect on the late phase of edema with a significant reduction in COX2 expression coupled to an inhibition of prostaglandin E(2) synthesis, implying that, in the presence of P2a, CE/C ratio rescue and edema inhibition were strictly related. In conclusion, the P2a effect is due to its binding to Hpt with consequent displacement of ApoA-I that exerts anti-inflammatory activity. Therefore, it is feasible to design drugs that, by enhancing the physiological endogenous protective role of ApoA-I, may be useful in inflammation-based diseases.

Apolipoprotein A-I possesses an anti-obesity effect associated with increase of energy expenditure and up-regulation of UCP1 in brown fat

Apolipoprotein A-I (ApoA-I) is the most abundant protein constituent of high-density lipoprotein (HDL). Reduced plasma HDL and ApoA-I levels have been found to be associated with obesity and metabolic syndrome in human beings. However, whether or not ApoA-I has a direct effect on obesity is largely unknown. Here we analysed the anti-obesity effect of ApoA-I using two mouse models, a transgenic mouse with overexpression of ApoA-I and the mice administered with an ApoA-I mimetic peptide D-4F. The mice were induced to develop obesity by feeding with high fat diet. Both ApoA-I overexpression and D-4F treatment could significantly reduce white fat mass and slightly improve insulin sensitivity in the mice. Metabolic analyses revealed that ApoA-I overexpression and D-4F treatment enhanced energy expenditure in the mice. The mRNA level of uncoupling protein (UCP)1 in brown fat tissue was elevated by ApoA-I transgenic mice. ApoA-I and D-4F treatment was able to increase UCP1 mRNA and protein levels as well as to stimulate AMP-activated protein kinase (AMPK) phosphorylation in brown adipocytes in culture. Taken together, our results reveal that ApoA-I has an anti-obesity effect in the mouse and such effect is associated with increases in energy expenditure and UCP1 expression in the brown fat tissue.

307 Pro-Inflammatory HDL in Idiopathic and Connective Tissue Disease-Associated Pulmonary Arterial Hypertension (PAH) Is Improved Ex Vivo by an Apolipoprotein-AI Mimetic Peptide: A Potential “Target” for Therapies

307 Pro-Inflammatory HDL in Idiopathic and Connective Tissue Disease-Associated Pulmonary Arterial Hypertension (PAH) Is Improved Ex Vivo by an Apolipoprotein-AI Mimetic Peptide: A Potential “Target” for Therapies

Regression of aortic valve stenosis by ApoA‐I mimetic peptide infusions in rabbits

Aortic valve stenosis (AVS) is the most common valvular heart disease, and standard curative therapy remains open heart surgical valve replacement. The aim of our experimental study was to determine if apolipoprotein A-I (ApoA-I) mimetic peptide infusions could induce regression of AVS. Fifteen New Zealand White male rabbits received a cholesterol-enriched diet and vitamin D(2) until significant AVS was detected by echocardiography. The enriched diet was then stopped to mimic cholesterol-lowering therapy and animals were allocated randomly to receive saline (control group, n=8) or an ApoA-I mimetic peptide (treated group, n=7), three times per week for 2 weeks. Serial echocardiograms and post mortem valve histology were performed. Aortic valve area increased significantly by 25% in the treated group after 14 days of treatment (P=0.012). Likewise, aortic valve thickness decreased by 21% in the treated group, whereas it was unchanged in controls (P=0.0006). Histological analysis revealed that the extent of lesions at the base of valve leaflets and sinuses of Valsalva was smaller in the treated group compared with controls (P=0.032). The treatment also reduced calcification, as revealed by the loss of the positive relationship observed in the control group (r=0.87, P=0.004) between calcification area and aortic valve thickness. Infusions of ApoA-I mimetic peptide lead to regression of experimental AVS. These positive results justify the further testing of high-density lipoprotein (HDL)-based therapies in patients with valvular aortic stenosis. Regression of aortic stenosis, if achieved safely, could transform the clinical treatment of this disease.

Pharmacologic augmentation of high-density lipoproteins: mechanisms of currently available and emerging therapies

With the limited effects of low-density lipoprotein-based lipid intervention, more attention is being paid to drugs that augment or mimic high-density lipoprotein's beneficial effects. A thorough understanding of the anti-atherogenic effects of high-density lipoprotein, and the mechanisms of existing or emerging high-density lipoprotein-based therapies, is essential for rational strategy for the prevention of cardiovascular disease. High-density lipoprotein mediates its beneficial effects through reverse cholesterol transport and direct anti-inflammatory effects of apolipoprotein AI and other component parts. Currently available drugs increase high-density lipoprotein-C through increasing apoAI synthesis (statins, fibrates) and decreasing apolipoprotein AI catabolism (niacin). Cholesteryl ester transfer protein inhibitors dramatically raise high-density lipoprotein-C, but clinical data are still required to verify their cardioprotective effects. Novel therapies such as apolipoprotein AImilano, apolipoprotein AI mimetic peptide, and exogenous phospholipids show tremendous promise as treatments for atherosclerosis. High-density lipoprotein and its defining functional protein apoAI prevent atherosclerosis through reverse cholesterol transport and other direct effects. Research has led to the development of novel therapies that increase high-density lipoprotein-C or that mimic direct anti-atherogenic effects of apolipoprotein AI. As these emerging therapies find a place in clinical medicine, we can anticipate preventing a much higher degree of cardiovascular events.

Inhibition of Lipopolysaccharide-Induced Inflammatory Responses by an Apolipoprotein AI Mimetic Peptide

Previous studies suggest that high-density lipoprotein and apoAI inhibit lipopolysaccharide (LPS)-induced inflammatory responses. The goal of the current study was to test the hypothesis that the apoAI mimetic peptide L-4F exerts antiinflammatory effects similar to apoAI. Pretreatment of human umbilical vein endothelial cells (HUVECs) with LPS induced the adhesion of THP-1 monocytes. Incubation of cells with LPS and L-4F (1 to 50 microg/mL) reduced THP-1 adhesion in a concentration-dependent manner. This response was associated with a significant reduction in the synthesis of cytokines, chemokines, and adhesion molecules. L-4F reduced vascular cell adhesion molecule-1 expression induced by LPS or lipid A, whereas a control peptide (Sc-4F) showed no effect. In contrast to LPS treatment, L-4F did not inhibit IL-1beta- or tumor necrosis factor-alpha-induced vascular cell adhesion molecule-1 expression. The inhibitory effect of L-4F on LPS induction of inflammatory markers was associated with reduced binding of LPS to its plasma carrier molecule, lipopolysaccharide binding protein, and decreased binding of LPS to HUVEC monolayers. LPS and L-4F in HUVEC culture medium were fractionated by fast protein liquid chromatography and were localized to the same fractions, suggesting a physical interaction between these molecules. Proinflammatory responses to LPS are associated with the binding of lipid A to cell surface receptors. The current studies demonstrate that L-4F reduces the expression of inflammatory markers induced by LPS and lipid A and suggest that apoAI peptide mimetics may be useful in the treatment of inflammation associated with endotoxemia.

Two Homologous Apolipoprotein AI Mimetic Peptides: RELATIONSHIP BETWEEN MEMBRANE INTERACTIONS AND BIOLOGICAL ACTIVITY

Two related 18-amino acid, class A, amphipathic helical peptides termed 3F-2 and 3F14 were chosen for this study. Although they have identical amino acid compositions and many similar biophysical properties, 3F-2 is more potent than 3F14 as an apolipoprotein AI mimetic peptide. The two peptides exhibit similar gross conformational properties, forming structures of high helical content on a membrane surface. However, the thermal denaturation transition of 3F-2 is more cooperative, suggesting a higher degree of oligomerization on the membrane. Both 3F-2 and 3F14 promote the segregation of cholesterol in membranes containing phosphatidylcholine and cholesterol, but 3F-2 exhibits a greater selectivity for partitioning into cholesterol-depleted regions of the membrane. Magic angle spinning/NMR studies indicate that the aromatic residues of 3F-2 are stacked in the presence of lipid. The aromatic side chains of this peptide also penetrate more deeply into membranes of phosphatidylcholine with cholesterol compared with 3F14. Using the fluorescent probe, 1,3-dipyrenylpropane, we monitored the properties of the lipid hydrocarbon environment. 3F-2 had a greater effect in altering the properties of the hydrocarbon region of the membrane. The results are consistent with our proposed model of the effect of peptide shape on the nature of the difference in peptide insertion into the bilayer.