RESUMEN
To test the hypothesis that sidedness of interfacial arginine (Arg) in apoA-I mimetic peptides, similar to that observed in apoA-I (Bashtovyy, D. et al. 2011. Sequence conservation of apolipoprotein A-I affords novel insights into HDL structure-function. J. Lipid Res. 52: 435-450.), may be important for biological activity, we compared properties of 4F and analogs, [K4,¹5>R]4F and [K9,¹³>R]4F, with Lys>Arg substitutions on the right and left side, respectively, of the 4F amphipathic helix. Intraperitoneal administration of these peptides into female apoE null mice (n = 13 in each group) reduced en face lesions significantly compared with controls; 4F and [K4,¹5>R]4F were equally effective whereas [K9,¹³>R]4F was less effective. Turnover experiments indicated that [K4,¹5>R]4F reached the highest, whereas [K9,¹³>R]4F had the lowest, plasma peak levels with a similar half life as the [K4,¹5>R]4F analog. The half life of 4F was two times longer than the other two peptides. The order in their abilities to associate with HDL in human plasma, generation of apoA-I particles with pre-ß mobility from isolated HDL, lipid associating ability, and sensitivity of lipid complexes to trypsin digestion was: 4F>[K4,¹5,>R]4F>[K9,¹³>R]4F. These studies support our hypothesis that the sidedness of interfacial Arg residues in the polar face of apoA-I mimetics results in differential biological properties.
Asunto(s)
Apolipoproteína A-I/química , Arginina/química , Aterosclerosis/tratamiento farmacológico , Peptidomiméticos/química , Peptidomiméticos/farmacología , Animales , Apolipoproteína A-I/deficiencia , Apolipoproteína A-I/genética , Arildialquilfosfatasa/metabolismo , Aterosclerosis/sangre , Aterosclerosis/metabolismo , Fenómenos Químicos , Colesterol/sangre , Femenino , Eliminación de Gen , Guanidina/farmacología , Humanos , Lipoproteínas HDL/metabolismo , Ratones , Oxidación-Reducción , Peptidomiméticos/metabolismo , Peptidomiméticos/uso terapéutico , Fosfatidilcolinas/metabolismo , Estructura Secundaria de Proteína , Desplegamiento Proteico/efectos de los fármacos , Especies Reactivas de Oxígeno/sangre , Liposomas Unilamelares/metabolismoRESUMEN
We have shown that Ac-hE18A-NH2, a dual-domain cationic apolipoprotein-mimetic peptide, reduces plasma cholesterol levels in dyslipidemic mice. Two single-domain cationic peptides based on the lytic class L peptide 18L were developed to test the hypothesis that a single-domain cationic amphipathic peptide can reduce atherosclerosis in apolipoprotein (apo)E null mice when orally administered. To incorporate anti-inflammatory properties, aromatic residues were clustered in the nonpolar face similar to peptide 4F, resulting in modified 18L (m18L). To reduce lytic properties, the Lys residues of 18L were replaced with Arg with the resulting peptide called modified R18L (mR18L). Biophysical studies showed that mR18L had stronger interactions with lipids than did m18L. Peptide mR18L was also more effective than m18L in promoting LDL uptake by HepG2 cells. ApoE null mice received normal chow or chow containing m18L or mR18L for six weeks. A significant reduction in plasma cholesterol and aortic sinus lesion area was seen only in the mR18L group. Plasma from mice administered mR18L, unlike those from the control and m18L groups, did not enhance monocyte adhesion to endothelial cells. Thus oral administration of mR18L reduces plasma cholesterol and lesion formation and inhibits monocyte adhesion.
Asunto(s)
Antiinflamatorios/uso terapéutico , Aneurisma de la Aorta/tratamiento farmacológico , Aterosclerosis/tratamiento farmacológico , Células Endoteliales/efectos de los fármacos , Monocitos/efectos de los fármacos , Proteína Básica de Mielina/uso terapéutico , Fragmentos de Péptidos/uso terapéutico , Péptidos/uso terapéutico , Administración Oral , Animales , Antiinflamatorios/administración & dosificación , Antiinflamatorios/farmacología , Aneurisma de la Aorta/metabolismo , Aneurisma de la Aorta/patología , Arginina/química , Arginina/metabolismo , Aterosclerosis/metabolismo , Aterosclerosis/patología , Cationes , Adhesión Celular/efectos de los fármacos , Colesterol/sangre , Colesterol/metabolismo , Células Endoteliales/citología , Células Endoteliales/metabolismo , Células Hep G2 , Humanos , Metabolismo de los Lípidos/efectos de los fármacos , Lisina/química , Lisina/metabolismo , Ratones , Ratones Noqueados , Monocitos/metabolismo , Monocitos/patología , Proteína Básica de Mielina/administración & dosificación , Proteína Básica de Mielina/farmacología , Fragmentos de Péptidos/administración & dosificación , Fragmentos de Péptidos/farmacología , Péptidos/administración & dosificación , Péptidos/farmacología , Estructura Secundaria de Proteína , Estructura Terciaria de ProteínaRESUMEN
ApoE mimetic peptide possesses the putative receptor binding domain 141-150 (LRKLRKRLLR) of apoE covalently linked to the class A amphipathic helical peptide 18A. It dramatically reduces plasma cholesterol in dyslipidemic mouse and rabbit models. Recycling of apoE mimetic peptide increases the duration of preß-HDL formation leading to extended anti-inflammatory and atheroprotective properties.
Asunto(s)
Apolipoproteínas E/química , Arildialquilfosfatasa/química , Peróxido de Hidrógeno/química , Lípidos/química , Lipoproteínas HDL/química , Animales , Antiinflamatorios/farmacología , Aterosclerosis/prevención & control , Línea Celular , Humanos , Ratones , Péptidos/química , Conejos , Factores de TiempoRESUMEN
Recent evidence indicates that inflammation may significantly contribute to the pathogenesis of Alzheimer's disease (AD). Since the apo A-I mimetic peptide D-4F has been shown to inhibit atherosclerotic lesion formation and regress already existing lesions (in the presence of pravastatin) and the peptide also decreases brain arteriole inflammation, we undertook a study to evaluate the efficacy of oral D-4F co-administered with pravastatin on cognitive function and amyloid beta (A beta) burden in the hippocampus of APPSwe-PS1 Delta E9 mice. Three groups of male mice were administered D-4F and pravastatin, Scrambled D-4F (ScD-4F, a control peptide) and pravastatin in drinking water, while drinking water alone served as control. The escape latency in the Morris Water Maze test was significantly shorter for the D-4F+statin administered animals compared to the other two groups. While the hippocampal region of the brain was covered with 4.2+/-0.5 and 3.8+/-0.6% of A beta load in the control and ScD-4F+statin administered groups, in the D-4F+statin administered group A beta load was only 1.6+/-0.1%. Furthermore, there was a significant decrease in the number of activated microglia (p<0.05 vs the other two groups) and activated astrocytes (p<0.05 vs control) upon oral D-4F+statin treatment. Inflammatory markers TNFalpha and IL-1 beta levels were decreased significantly in the D-4F+statin group compared to the other two groups (for IL-1 beta p<0.01 vs the other two groups and for TNF-alpha p<0.001 vs control) and the expression of MCP-1 were also less in D-4F+statin administered group compared to the other two groups. These results suggest that the apo A-I mimetic peptide inhibits amyloid beta deposition and improves cognitive function via exerting anti-inflammatory properties in the brain.
Asunto(s)
Enfermedad de Alzheimer/tratamiento farmacológico , Péptidos beta-Amiloides/metabolismo , Apolipoproteína A-I/administración & dosificación , Cognición/efectos de los fármacos , Hipocampo/metabolismo , Administración Oral , Enfermedad de Alzheimer/complicaciones , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/psicología , Precursor de Proteína beta-Amiloide/genética , Animales , Anticolesterolemiantes/administración & dosificación , Astrocitos/efectos de los fármacos , Astrocitos/fisiología , Quimiocina CCL2/metabolismo , Colesterol/sangre , Modelos Animales de Enfermedad , Quimioterapia Combinada , Hipocampo/efectos de los fármacos , Hipocampo/fisiopatología , Interleucina-1beta/metabolismo , Masculino , Aprendizaje por Laberinto/efectos de los fármacos , Aprendizaje por Laberinto/fisiología , Ratones , Ratones Transgénicos , Microglía/efectos de los fármacos , Microglía/fisiología , Placa Amiloide/efectos de los fármacos , Placa Amiloide/fisiología , Pravastatina/administración & dosificación , Nexinas de Proteasas , Receptores de Superficie Celular/genética , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Anti-atherogenic effects of high density lipoprotein (HDL) and its major protein component apolipoprotein A-I (apoA-I) are principally thought to be due to their ability to mediate reverse cholesterol transport. These agents also possess anti-oxidant properties that prevent the oxidative modification of low density lipoprotein (LDL) and anti-inflammatory properties that include inhibition of endothelial cell adhesion molecule expression. Results of the Framingham study revealed that a reduction in HDL levels is an independent risk factor for coronary artery disease (CAD). Accordingly, there has been considerable interest in developing new therapies that specifically elevate HDL cholesterol. However, recent evidence suggests that increasing circulating HDL cholesterol levels alone is not sufficient as a mode of HDL therapy. Rather, therapeutic approaches that increase the functional properties of HDL may be superior to simply raising the levels of HDL per se. Our laboratory has pioneered the development of synthetic, apolipoprotein mimetic peptides which are structurally and functionally similar to apoA-I but possess unique structural homology to the lipid-associating domains of apoA-I. The apoA-I mimetic peptide 4F inhibits atherogenic lesion formation in murine models of atherosclerosis. This effect is related to the ability of 4F to induce the formation of pre-ß HDL particles that are enriched in apoA-I and paraoxonase. 4F also possesses anti-inflammatory and anti-oxidant properties that are independent of its effect on HDL quality per se. Recent studies suggest that 4F stimulates the expression of the antioxidant enzymes heme oxygenase and superoxide dismutase and inhibits superoxide anion formation in blood vessels of diabetic, hypercholesterolemic and sickle cell disease mice. The goal of this review is to discuss HDL-dependent and -independent mechanisms by which apoA-I mimetic peptides reduce vascular injury in experimental animal models.
RESUMEN
3-Hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) are currently the drug of choice for the clinical management of elevated low-density lipoprotein (LDL) cholesterol. Although statin treatment provides an overall improvement in outcomes, clinical trial data reveal a significant number of cardiac events despite reaching targeted LDL levels. A low serum high-density lipoprotein (HDL) cholesterol level is an independent predictor of cardiovascular risk. Accordingly, there has been interest in determining whether HDL elevation, in addition to LDL lowering, further reduces risk in patients with coronary artery disease. Several commonly prescribed lipid-lowering therapies modestly raise HDL, but their use may be limited by the development of adverse reactions. Emerging data suggest that HDL quality and function may also be significantly reduced by atherosclerosis and other inflammatory diseases. The goal of this review is to discuss the current status of HDL therapeutics, with emphasis on a novel class of agent, the apolipoprotein A-I mimetic peptides, which improve the functional properties of HDL cholesterol.
Asunto(s)
Enfermedades Cardiovasculares/sangre , Enfermedades Cardiovasculares/prevención & control , HDL-Colesterol/sangre , HDL-Colesterol/efectos de los fármacos , Inhibidores de Hidroximetilglutaril-CoA Reductasas/farmacología , Inhibidores de Hidroximetilglutaril-CoA Reductasas/uso terapéutico , Animales , Apolipoproteína A-I/sangre , Apolipoproteína A-I/farmacología , Apolipoproteína A-I/uso terapéutico , Aterosclerosis/sangre , Aterosclerosis/tratamiento farmacológico , Proteínas de Transferencia de Ésteres de Colesterol/antagonistas & inhibidores , Ácido Clofíbrico/farmacología , Humanos , Lipoproteínas/farmacología , Hígado/efectos de los fármacos , Hígado/metabolismo , Fragmentos de Péptidos/farmacologíaRESUMEN
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.
Asunto(s)
Inflamación/prevención & control , Lipopolisacáridos/antagonistas & inhibidores , Péptidos/farmacología , Proteínas de Fase Aguda/metabolismo , Secuencia de Aminoácidos , Proteínas Portadoras/metabolismo , Adhesión Celular , Células Cultivadas , Células Endoteliales/citología , Humanos , Mediadores de Inflamación/fisiología , Lípido A/farmacología , Receptores de Lipopolisacáridos/fisiología , Lipopolisacáridos/metabolismo , Lipopolisacáridos/toxicidad , Lipoproteínas HDL/farmacología , Glicoproteínas de Membrana/metabolismo , Datos de Secuencia Molecular , Monocitos/fisiología , Péptidos/metabolismo , Fosfatidilcolinas/farmacología , Molécula 1 de Adhesión Celular Vascular/biosíntesisRESUMEN
BACKGROUND: These studies were designed to determine whether the dual-domain peptide with a class A amphipathic helix linked to the receptor-binding domain of apolipoprotein (apo) E (Ac-hE-18A-NH2) possesses both antidyslipidemic and antiinflammatory properties. METHODS AND RESULTS: A single bolus (15 mg/kg IV) of Ac-hE-18A-NH2 that contains LRKLRKRLLR (141- to 150-residue region of apo E) covalently linked to apo A-I mimetic peptide 18A not only reduced plasma cholesterol levels (baseline, 562+/-29.0 mg/dL versus 287.7+/-22.0 mg/dL at 18 hours, P<0.001) in the Watanabe heritable hyperlipidemic rabbit model but also significantly improved arterial endothelial function. This improvement was associated with a reduction in 2 markers of oxidative stress. First, the plasma lipid hydroperoxide content was reduced significantly, an effect associated with a 5-fold increase in HDL paraoxonase activity. Second, the formation of superoxide anion, a scavenger of nitric oxide, was also significantly reduced in arteries of these animals. CONCLUSIONS: Because dyslipidemia and endothelial dysfunction are common features of the atherosclerotic disease process, this unique dual-domain peptide has ideal composite properties that ameliorate key contributory factors to atherosclerosis.
Asunto(s)
Apolipoproteínas E/química , Colesterol/sangre , Hiperlipidemias/tratamiento farmacológico , Péptidos/farmacología , Animales , Apolipoproteínas E/uso terapéutico , Aterosclerosis/tratamiento farmacológico , Aterosclerosis/prevención & control , Endotelio Vascular/efectos de los fármacos , Endotelio Vascular/patología , Endotelio Vascular/fisiología , Hiperlipidemias/patología , Inflamación/tratamiento farmacológico , Errores Innatos del Metabolismo Lipídico/tratamiento farmacológico , Lípidos/sangre , Masculino , Imitación Molecular , Estrés Oxidativo/efectos de los fármacos , Fragmentos de Péptidos/síntesis química , Fragmentos de Péptidos/farmacología , Fragmentos de Péptidos/uso terapéutico , Péptidos/síntesis química , Péptidos/uso terapéutico , Conejos , Superóxidos/análisisRESUMEN
Levels of high density lipoprotein (HDL) and its major protein component, apolipoprotein (apo) A-I, are strongly inversely correlated to risk of atherosclerosis and other vascular diseases. A number of properties of apo A-I may contribute to this protection, including removal of cholesterol from peripheral tissues to the liver (reverse cholesterol transport), anti-inflammatory and anti-oxidative activities, and modulation of vascular function. Apo A-I has lipid-associating domains that form class A amphipathic helices. Peptide analogs that have no sequence homology to the domains in apo A-I but possess the class A motif have been shown to not only associate with phospholipid but also mimic several of the functional properties of apo A-I. Peptide 4F, with four phenylalanines on the non-polar face, was found to be maximally effective in mimicking the positive qualities of apo A-I; this peptide inhibited atherosclerosis, reduced inflammation and oxidation, and improved vascular function in a number of animal models, and when synthesized with D-amino acids is orally bioavailable. Several other classes of peptide mimetics are now being studied, and may contribute to our understanding of the functions of apo E and apo J. The use of peptide mimetics to study apolipoprotein function has proved to be a powerful tool, and may lead to novel therapeutic agents in the prevention of atherosclerosis and other vascular diseases.
Asunto(s)
Apolipoproteínas/fisiología , Aterosclerosis/tratamiento farmacológico , Péptidos/farmacología , Péptidos/uso terapéutico , Enfermedades Vasculares/tratamiento farmacológico , Animales , Apolipoproteína A-I/uso terapéutico , Apolipoproteínas/química , Aterosclerosis/fisiopatología , Colesterol/metabolismo , HDL-Colesterol/metabolismo , Enfermedad Coronaria/tratamiento farmacológico , Humanos , Lipopolisacáridos/química , Lipopolisacáridos/farmacología , Ratones , Ratones Endogámicos C57BL , Imitación Molecular , Conformación Proteica , Enfermedades Vasculares/fisiopatologíaRESUMEN
OBJECTIVE: We tested for synergy between pravastatin and D-4F by administering oral doses of each in combination that were predetermined to be ineffective when given as single agents. METHODS AND RESULTS: The combination significantly increased high-density lipoprotein (HDL)-cholesterol levels, apolipoprotein (apo)A-I levels, paraoxonase activity, rendered HDL antiinflammatory, prevented lesion formation in young (79% reduction in en face lesion area; P<0.0001) and caused regression of established lesions in old apoE null mice (ie, mice receiving the combination for 6 months had lesion areas that were smaller than those before the start of treatment (P=0.019 for en face lesion area; P=0.004 for aortic root sinus lesion area). After 6 months of treatment with the combination, en face lesion area was 38% of that in mice maintained on chow alone; P<0.00004) with a 22% reduction in macrophage content in the remaining lesions (P=0.001), indicating an overall reduction in macrophages of 79%. The combination increased intestinal apoA-I synthesis by 60% (P=0.011). In monkeys, the combination also rendered HDL antiinflammatory. CONCLUSIONS: These results suggest that the combination of a statin and an HDL-based therapy may be a particularly potent treatment strategy.
Asunto(s)
Apolipoproteína A-I/farmacología , Apolipoproteínas E/genética , Aterosclerosis/tratamiento farmacológico , HDL-Colesterol/inmunología , Inhibidores de Hidroximetilglutaril-CoA Reductasas/farmacología , Pravastatina/farmacología , Factores de Edad , Alimentación Animal , Animales , Antiinflamatorios/farmacología , Apolipoproteína A-I/metabolismo , Aterosclerosis/inmunología , Aterosclerosis/prevención & control , Células Cultivadas , Quimiotaxis de Leucocito/efectos de los fármacos , HDL-Colesterol/metabolismo , Sinergismo Farmacológico , Femenino , Mucosa Intestinal/metabolismo , Macaca fascicularis , Masculino , Ratones , Ratones Mutantes , Monocitos/citologíaRESUMEN
We have shown that the dual domain peptide Ac-hE18A-NH(2), in which LRKLRKRLLR, (141-150 region of human apo E) covalently linked to a class A lipid-associating domain, is able to associate with apo B-containing lipoproteins and enhance their clearance both in vitro and in vivo. We present here the differential effects of this peptide on the plasma cholesterol levels in different mouse models. The peptide intravenously administered (100 microg) into C57BL/6J mice on atherogenic diet, apo E null, and apo E null/LDL-receptor (LDL-R) null double knock out mouse models, was able to rapidly reduce plasma cholesterol levels within 2 min, and the effect persisted for more than 6 h. The reduction was limited to the VLDL and IDL/LDL fractions; HDL was not reduced in any mouse model studied. However, the peptide had no effect on the plasma cholesterol levels in C57BL/6J mice on normal diet, LDL-R null mice on normal chow, and LDL-R null mice on Western diet. Administration to LDL-R null mice of 125I-labelled human lipoproteins incubated with peptide resulted in accelerated human VLDL and LDL clearance with associated increase of radioactivity in the liver. These results, coupled with our earlier in vitro observations, indicate that the Arg-rich peptide-assisted rapid clearance of plasma cholesterol in dyslipidemic mice is due to the peptide targeting apo B-48-containing atherogenic lipoproteins to the liver for increased uptake and degradation.
Asunto(s)
Colesterol/sangre , Hiperlipidemias/sangre , Lipoproteínas/farmacología , Fragmentos de Péptidos/farmacología , Animales , Apolipoproteínas E/deficiencia , Dieta Aterogénica , Electroforesis en Gel Bidimensional , Femenino , Humanos , Inyecciones Intravenosas , Lípidos/sangre , Lipoproteínas/administración & dosificación , Lipoproteínas/sangre , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Fragmentos de Péptidos/administración & dosificación , Receptores de LDL/deficienciaRESUMEN
BACKGROUND: Apolipoprotein E (ApoE) is the major apolipoprotein present in the high-density lipoprotein-like particles in the central nervous system (CNS). ApoE is involved in various protective functions in CNS including cholesterol transport, anti-inflammatory, and antioxidant effects. An ApoE peptide would be expected to exert protective effects on neuroinflammation. OBJECTIVE: To determine the effects of an ApoE mimetic peptide Ac-hE18A-NH2 on amyloid-ß pathology. METHOD: Using human APP/PS1ΔE9 transgenic mice and in vitro studies, we have evaluated the effect of an ApoE mimetic peptide, Ac-hE18A-NH2, on amyloid plaque deposition and inflammation. RESULTS: Administration of Ac-hE18A-NH2 to APP/PS1ΔE9 mice for 6 weeks (50 µg/mouse, 3 times a week) significantly improved cognition with a concomitant decrease in amyloid plaque deposition and reduced activated microglia and astrocytes, and increased brain ApoE levels. Oligomeric Aß42 (oAß42) and oxidized PAPC (ox-PAPC) inhibited secretion of ApoE in U251 cells, a human astrocyte cell line, and this effect was ameliorated in the presence of peptide Ac-hE18A-NH2. The peptide also increased Aß42 uptake in a cell line of human macrophages. CONCLUSIONS: Peptide Ac-hE18A-NH2 attenuates the effects of oxidative stress on ApoE secretion, inhibits amyloid plaque deposition, and thus could be beneficial in the treatment of Alzheimer's disease.
Asunto(s)
Neuropatías Amiloides/tratamiento farmacológico , Antipsicóticos/uso terapéutico , Encéfalo/metabolismo , Lipoproteínas/uso terapéutico , Fragmentos de Péptidos/uso terapéutico , Neuropatías Amiloides/complicaciones , Neuropatías Amiloides/metabolismo , Neuropatías Amiloides/patología , Péptidos beta-Amiloides/metabolismo , Precursor de Proteína beta-Amiloide/genética , Animales , Apolipoproteínas E/metabolismo , Encéfalo/efectos de los fármacos , Línea Celular Transformada , Colesterol/sangre , Trastornos del Conocimiento/tratamiento farmacológico , Trastornos del Conocimiento/etiología , Trastornos del Conocimiento/genética , Modelos Animales de Enfermedad , Ensayo de Inmunoadsorción Enzimática , Hipocampo/efectos de los fármacos , Hipocampo/metabolismo , Hipocampo/patología , Humanos , Masculino , Aprendizaje por Laberinto/efectos de los fármacos , Ratones , Ratones Transgénicos , Mutación/genética , Fragmentos de Péptidos/metabolismo , Placa Amiloide/tratamiento farmacológico , Placa Amiloide/etiología , Placa Amiloide/genética , Presenilina-1/genética , TransfecciónRESUMEN
OBJECTIVE: We investigated two apoE mimetic peptides with similar long-term plasma cholesterol reducing abilities for their effects on atherosclerotic lesions in Western diet-fed female LDL-receptor (LDL-R) null mice. METHODS AND RESULTS: Single doses of peptides Ac-hE18A-NH(2) and mR18L were administered retro-orbitally to LDL-R null mice on Western diet and plasma cholesterol was measured at 10 min, 4 h, and 24 h post administration. Peptide mR18L and not Ac-hE18A-NH(2) reduced plasma cholesterol levels significantly at 4 h post administration. However, multiple administrations (100 µg/mouse twice weekly for 8 weeks) resulted in a similar reduction in plasma cholesterol. Only the plasma from the Ac-hE18A-NH(2) group had significantly reduced reactive oxygen species levels at the end of the treatment protocol. Both mR18L and Ac-hE18A-NH(2) showed reduced atherosclerotic lesion areas. However, peptide Ac-hE18A-NH(2) was significantly more effective in inhibiting atherosclerosis. Both peptides reduced total plaque macrophage load compared to the saline treated animals, with peptide Ac-hE18A-NH(2) having a greater reduction. Incubation of HepG2 cells and THP-1 monocyte-derived macrophages with both peptides in the presence of oxidized phospholipid showed that Ac-hE18A-NH(2) promotes the secretion of apoE from the cells whereas mR18L does not. CONCLUSIONS: Despite similar reductions in plasma cholesterol levels, Ac-hE18A-NH(2) was more effective in inhibiting lesions than mR18L, possibly due to its ability to promote the secretion of apoE from hepatocytes and macrophages.
Asunto(s)
Apolipoproteínas E/metabolismo , Aterosclerosis/prevención & control , Lipoproteínas/farmacología , Fragmentos de Péptidos/farmacología , Péptidos/farmacología , Receptores de LDL/genética , Animales , Apolipoproteínas E/química , Colesterol/sangre , Femenino , Células Hep G2 , Hepatocitos/efectos de los fármacos , Hepatocitos/metabolismo , Humanos , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Ratones , Especies Reactivas de Oxígeno/sangreRESUMEN
OBJECTIVE: The apolipoprotein E mimetic peptide Ac-hE18A-NH(2), capable of reducing plasma cholesterol and possessing anti-inflammatory properties, was compared with the well-studied anti-atherogenic apoA-I mimetic peptide 4F for reducing lesion formation in female apoE null mice with already existing lesions. METHODS AND RESULTS: In initial experiments, Ac-hE18A-NH(2) was administered retro-orbitally two or three times weekly for 6-8 weeks, while peptide 4F was administered intraperitoneally every day for the same period. Age matched controls were injected with saline every day. At the end of the treatment period, plasma cholesterol levels of Ac-hE18A-NH(2) administered mice were significantly lower than in 4F and control mice. However, both 4F and Ac-hE18A-NH(2) showed reduced lesion areas in en face lesion analysis to a similar extent compared to the control group, while paraoxonase-1 (PON-1) activity was increased only in the Ac-hE18A-NH(2) group. In the third experiment, both peptides were administered at the same dose, frequency, and route of administration. The reduction in en face lesions with Ac-hE18A-NH(2) was significantly greater than the 4F and control groups, although lesions in 4F-treated mice were also significantly reduced compared with controls. Both peptide groups had significantly reduced plasma lipid hydroperoxides, but only the Ac-hE18A-NH(2) group had significantly reduced serum amyloid A levels. HDL and plasma inflammatory indices were significantly reduced in both peptide groups compared with controls. CONCLUSIONS: Although both peptides had similar anti-inflammatory properties, Ac-hE18A-NH(2) was more effective in inhibiting lesions than 4F at the same dose, frequency, and route of administration, perhaps due to its cholesterol reducing properties.
Asunto(s)
Anticolesterolemiantes/farmacología , Enfermedades de la Aorta/prevención & control , Apolipoproteína A-I/farmacología , Apolipoproteínas E/deficiencia , Aterosclerosis/prevención & control , Lipoproteínas/farmacología , Fragmentos de Péptidos/farmacología , Factores de Edad , Envejecimiento , Animales , Antiinflamatorios/farmacología , Anticolesterolemiantes/administración & dosificación , Antioxidantes/farmacología , Enfermedades de la Aorta/sangre , Enfermedades de la Aorta/genética , Enfermedades de la Aorta/patología , Apolipoproteína A-I/administración & dosificación , Apolipoproteínas E/genética , Arildialquilfosfatasa/sangre , Aterosclerosis/sangre , Aterosclerosis/genética , Aterosclerosis/patología , Colesterol/sangre , Modelos Animales de Enfermedad , Esquema de Medicación , Femenino , Inyecciones Intraperitoneales , Inyecciones Intravenosas , Peróxidos Lipídicos/sangre , Lipoproteínas/administración & dosificación , Ratones , Ratones Noqueados , Fragmentos de Péptidos/administración & dosificación , Proteína Amiloide A Sérica/metabolismo , Factores Sexuales , Factores de TiempoRESUMEN
Apolipoprotein E (apoE) is a ligand for clearance of lipoprotein remnants such as chylomicrons and very low-density lipoproteins. It has anti-atherogenic and anti-inflammatory properties. Therefore, there is extensive ongoing research to create peptides that can mimic properties of apoE. A number of synthetic peptides that encompass different regions of apoE have been studied for inhibiting inflammatory states, including Alzheimer disease. However, peptides that clear atherogenic lipoproteins, analogous to apoE, via enhanced hepatic uptake have not been previously reviewed. Toward this end, we describe the design and studies of a dual-domain apoE mimetic peptide, Ac-hE18A-NH(2). This peptide consists of residues 141-150, the putative receptor-binding region of human apoE, covalently linked to a well characterized class A amphipathic helix, 18A, which has no sequence homology to any other exchangeable apolipoprotein sequences. It demonstrates dramatic effects in reducing plasma cholesterol levels in dyslipidemic mouse and rabbit models. We discuss the scientific rationale and review the literature for the design and efficacy of the peptide. Analogous to apoE, this peptide bypasses the low-density lipoprotein receptor for the hepatic uptake of atherogenic lipoproteins via heparan sulfate proteoglycan (HSPG). ApoE mimetics such as Ac-hE18A-NH(2) may therefore restore or replace ligands in genetically induced hyperlipidemias to enable reduction in atherogenic lipoproteins via HSPG even in the absence of functional low-density lipoprotein receptors. Therefore, this and similar peptides may be useful in the treatment of dyslipidemic disorders such as familial hyperlipidemia and atherosclerosis.
Asunto(s)
Apolipoproteínas E/metabolismo , Colesterol/sangre , Lipoproteínas/farmacología , Fragmentos de Péptidos/farmacología , Animales , Modelos Animales de Enfermedad , Diseño de Fármacos , Dislipidemias/tratamiento farmacológico , Dislipidemias/fisiopatología , Humanos , Ratones , Péptidos/farmacología , Conejos , Receptores de LDL/metabolismoRESUMEN
Apolipoprotein E (apoE) exerts prominent anti-inflammatory effects and undergoes recycling by target cells. We previously reported that the peptide Ac-hE18A-NH(2), composed of the receptor binding domain (LRKLRKRLLR) of apoE covalently linked to the Class A amphipathic peptide 18A, dramatically lowers plasma cholesterol and lipid hydroperoxides and enhances paraoxonase activity in dyslipidemic animal models. The objective of this study was to determine whether this peptide, analogous to apoE, exerts anti-inflammatory effects and undergoes recycling under in vitro conditions. Pulse chase studies using [(125)I]-Ac-hE18A-NH(2) in THP-1 derived macrophages and HepG2 cells showed greater amounts of intact peptide in the cells at later time points indicating recycling of the peptide. Ac-hE18A-NH(2) induced a 2.5-fold increase in prebeta-HDL in the conditioned media of HepG2 cells. This effect persisted for 3 days after removal of the peptide from culture medium. Ac-hE18A-NH(2) also induced the secretion of cell surface apoE from THP-1 macrophages. In addition, the peptide increased cholesterol efflux from THP-1 cells by an ABCA1 independent mechanism. Moreover, Ac-hE18A-NH(2) inhibited LPS-induced vascular cell adhesion molecule-1 (VCAM-1) expression, and reduced monocyte adhesion in human umbilical vein endothelial cells (HUVECs). It also reduced the secretion of interleukin-6 (IL-6) and monocyte chemoattractant protein-1 (MCP-1) from THP-1 macrophages even when administered post-LPS and abolished the 18-fold increase in LPS-induced mRNA levels for MCP-1 in THP-1 cells. Taken together, these results suggest that addition of the putative apoE receptor-domain to the Class A amphipathic peptide 18A results in a peptide that, similar to apoE, recycles, thus enabling the potentiation and prolongation of its anti-atherogenic and anti-inflammatory effects. Such a peptide has great potential as a therapeutic agent in the management of atherosclerosis and other inflammatory diseases.
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Antiinflamatorios/farmacología , Lipoproteínas/farmacología , Fragmentos de Péptidos/farmacología , Animales , Células Cultivadas , Lipoproteínas de Alta Densidad Pre-beta/biosíntesis , Lipoproteínas de Alta Densidad Pre-beta/efectos de los fármacos , Humanos , Inflamación/prevención & control , Péptidos/metabolismo , Conejos , Factores de TiempoRESUMEN
OBJECTIVE: We recently described anti-atherogenic properties of the dual domain peptide Ac-hE18A-NH(2) derived by covalently linking the heparin binding domain 141-150 of apoE to 18A, a class A amphipathic helical peptide. In this paper we have compared the properties of Ac-hE18A-NH(2) with the non-heparin binding 151-160 region of apoE linked to 18A (Ac-nhE18A-NH(2)). METHODS AND RESULTS: Both peptides were highly helical in solution and in association with lipids. Ac-hE18A-NH(2) and not Ac-nhE18A-NH(2) enhanced uptake of low density lipoprotein (LDL) in HepG2 cells. While Ac-hE18A-NH(2) retarded the electrophoretic mobility of LDL, Ac-nhE18A-NH(2) slightly enhanced mobility. Ac-hE18A-NH(2) reduced monocyte association with endothelial cells, while Ac-nhE18A-NH(2) increased it. Ac-hE18A-NH(2) also reduced lipid hydroperoxide content of LDL while Ac-nhE18A-NH(2) increased it. A single administration of Ac-hE18A-NH(2) (100 µg/mouse) into apoE null mice dramatically reduced cholesterol (from 600 mg/dL to 180 mg/dL at 5 min and to 60 mg/dL at 5h) while Ac-nhE18A-NH(2) had no effect. Administration (100 µg/mouse/day, three days a week) into apoE null mice for six weeks showed Ac-hE18A-NH(2) group having a moderate aortic sinus lesion reduction compared with the control group (-15.1%), while the Ac-nhE18A-NH(2) administered group had increased lesion area (+33.0% vs controls and 36.1% vs Ac-hE18A-NH(2)). Plasma from mice administered Ac-hE18A-NH(2) for six weeks showed a significant reduction in plasma cholesterol and triglyceride levels and increase in paraoxonase-1 (PON-1) activity compared to controls, while Ac-nhE18A-NH(2) caused no change in plasma cholesterol and decreased PON-1 activity. CONCLUSION: It is proposed that Ac-hE18A-NH(2) reduced lesion progression in apoE null mice due to its anti-inflammatory and lipoprotein clearing properties, while Ac-nhE18A-NH(2) exhibited pro-atherogenic effects.
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Apolipoproteínas E/genética , Aterosclerosis/etiología , Células Endoteliales/efectos de los fármacos , Lipoproteínas/farmacología , Monocitos/efectos de los fármacos , Fragmentos de Péptidos/farmacología , Animales , Apolipoproteínas E/química , Apolipoproteínas E/deficiencia , Arildialquilfosfatasa/metabolismo , Colesterol/sangre , Femenino , Células Hep G2 , Proteoglicanos de Heparán Sulfato/metabolismo , Humanos , Lipoproteínas LDL/efectos de los fármacos , Lipoproteínas LDL/metabolismo , Ratones , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Triglicéridos/sangreRESUMEN
Recently, attention has been focused on pharmacological treatments that increase HDL cholesterol to prevent coronary artery disease. Despite three decades of extensive research of human apolipoprotein A-I (apoA-I), the major protein component of HDL, the molecular basis for its antiatherogenic and anti-inflammatory functions remain elusive. Another protein component of HDL, apoA-II, has structural features similar to those of apoA-I but does not possess atheroprotective properties. To understand the molecular basis for the effectiveness of apoA-I, we used model synthetic peptides. We designed analogs of the class A amphipathic helical motif in apoA-I that is responsible for solubilizing phospholipids. None of these analogs has sequence homology to apoA-I, but all are similar in their lipid-associating structural motifs. Although all of these peptide analogs interact with phospholipids to form peptide:lipid complexes, the biological properties of these analogs are different. Physical-chemical and NMR studies of these peptides have enabled the delineation of structural requirements for atheroprotective and anti-inflammatory properties in these peptides. It has been shown that peptides that interact strongly with lipid acyl chains do not have antiatherogenic and anti-inflammatory properties. In contrast, peptides that associate close to the lipid head group (and hence do not interact strongly with the lipid acyl chain) are antiatherogenic and anti-inflammatory. Understanding the structure and function of apoA-I and HDL through studies of the amphipathic helix motif may lead to peptide-based therapies for inhibiting atherosclerosis and other related inflammatory lipid disorders.
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Antiinflamatorios/uso terapéutico , Antioxidantes/uso terapéutico , Apolipoproteína A-I/uso terapéutico , Apolipoproteínas A/uso terapéutico , Animales , Apolipoproteína A-I/química , Aterosclerosis/tratamiento farmacológico , Materiales Biomiméticos/uso terapéutico , HDL-Colesterol/fisiología , Humanos , Modelos Moleculares , Estructura Secundaria de Proteína , Relación Estructura-ActividadRESUMEN
Two homologous apoA-I mimetic peptides, 3F-2 and 3F(14), differ in their in vitro antiatherogenic properties (Epand, R. M., Epand, R. F., Sayer, B. G., Datta, G., Chaddha, M., and Anantharamaiah, G. M. (2004) J. Biol. Chem. 279, 51404-51414). In the present work, we demonstrate that the peptide 3F-2, which has more potent anti-inflammatory activity in vitro when administered intraperitoneally to female apoE null mice (20 microg/mouse/day) for 6 weeks, inhibits atherosclerosis (lesion area 15,800 +/- 1000 microm(2), n = 29), whereas 3F(14) does not (lesion area 20,400 +/- 1000 microm(2), n = 26) compared with control saline administered (19,900 +/- 1400 microm(2), n = 22). Plasma distribution of the peptides differs in that 3F-2 preferentially associates with high density lipoprotein, whereas 3F(14) preferentially associates with apoB-containing particles. After intraperitoneal injection of (14)C-labeled peptides, 3F(14) reaches a higher maximal concentration and has a longer half-time of elimination than 3F-2. A study of the effect of these peptides on the motional and organizational properties of phospholipid bilayers, using several NMR methods, demonstrates that the two peptides insert to different extents into membranes. 3F-2 with aromatic residues at the center of the nonpolar face partitions closer to the phospholipid head group compared with 3F(14). In contrast, only 3F(14) affects the terminal methyl group of the acyl chain, decreasing the (2)H order parameter and at the same time also decreasing the molecular motion of this methyl group. This dual effect of 3F(14) can be explained in terms of the cross-sectional shape of the amphipathic helix. These results support the proposal that the molecular basis for the difference in the biological activities of the two peptides lies with their different interactions with membranes.
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Apolipoproteína A-I/química , Aterosclerosis/metabolismo , Animales , Biomimética , Carbono/química , Femenino , Membrana Dobles de Lípidos/química , Lipoproteínas/química , Espectroscopía de Resonancia Magnética , Ratones , Ratones Endogámicos C57BL , Modelos Moleculares , Péptidos/química , Fosfolípidos/química , Unión ProteicaRESUMEN
PURPOSE OF REVIEW: Recent publications related to the potential use of synthetic peptides for the management of lipid disorders and their vascular complications are reviewed. RECENT FINDINGS: The potential use of synthetic peptides for the management of lipid disorders and their vascular complications has emerged in recent years. These peptides are models of apolipoproteins, but are much smaller in size than the apolipoproteins. Oral peptides that improve the antiinflammatory properties of HDLs have been shown to potently inhibit atherosclerosis in mouse models. Injection of a peptide with a class A amphipathic helix in a rat model of diabetes dramatically reduced endothelial sloughing and improved vasoreactivity. Injected synthetic peptides have also been described that dramatically lower plasma cholesterol and restore endothelial function in a rabbit model of familial hypercholesterolemia. These studies suggest the therapeutic potential for synthetic peptides in the management of lipid disorders and their vascular complications. SUMMARY: Synthetic peptides much smaller than exchangeable human plasma apolipoproteins but with physical and chemical characteristics similar to the plasma apolipoproteins have shown promise in the management of lipid disorders and their vascular complications in animal models. The initial success of these animal studies suggests that synthetic peptides have the potential to emerge as a new therapeutic class of agents in the management of patients with lipid disorders.