RESUMEN
apoE is the primary lipid carrier within the CNS and the strongest genetic risk factor for late onset Alzheimer's disease (AD). apoE is primarily lipidated via ABCA1, and both are under transcriptional regulation by the nuclear liver X receptor (LXR). Considerable evidence from genetic (using ABCA1 overexpression) and pharmacological (using synthetic LXR agonists) studies in AD mouse models suggests that increased levels of lipidated apoE can improve cognitive performance and, in some strains, can reduce amyloid burden. However, direct synthetic LXR ligands have hepatotoxic side effects that limit their clinical use. Here, we describe a set of small molecules, previously annotated as antagonists of the purinergic receptor, P2X7, which enhance ABCA1 expression and activity as well as apoE secretion, and are not direct LXR ligands. Furthermore, P2X7 is not required for these molecules to induce ABCA1 upregulation and apoE secretion, demonstrating that the ABCA1 and apoE effects are mechanistically independent of P2X7 inhibition. Hence, we have identified novel dual activity compounds that upregulate ABCA1 across multiple CNS cell types, including human astrocytes, pericytes, and microglia, through an indirect LXR mechanism and that also independently inhibit P2X7 receptor activity.
Asunto(s)
Transportador 1 de Casete de Unión a ATP/agonistas , Apolipoproteínas E/agonistas , Antagonistas del Receptor Purinérgico P2X/farmacología , Receptores Purinérgicos P2X7/metabolismo , Bibliotecas de Moléculas Pequeñas/farmacología , Transportador 1 de Casete de Unión a ATP/metabolismo , Adamantano/análogos & derivados , Adamantano/química , Adamantano/farmacología , Animales , Apolipoproteínas E/metabolismo , Aziridinas/química , Aziridinas/farmacología , Benzamidas/química , Benzamidas/farmacología , Células Cultivadas , Humanos , Ratones , Ratones Noqueados , Estructura Molecular , Naftoquinonas/química , Naftoquinonas/farmacología , Antagonistas del Receptor Purinérgico P2X/química , Receptores Purinérgicos P2X7/deficiencia , Bibliotecas de Moléculas Pequeñas/química , Sulfonamidas/química , Sulfonamidas/farmacología , Regulación hacia Arriba/efectos de los fármacosRESUMEN
BACKGROUND: phospholipid transfer protein (PLTP) plays important roles in lipoprotein metabolism and atherosclerosis and is expressed by macrophages and macrophage foam cells (MFCs). The aim of the present study was to determine whether the major protein from HDL, apoA-I, affects PLTP derived from MFCs. RESULTS: as cell model we used human THP-1 monocytes incubated with acetylated LDL, to generate MFC. The addition of apoA-I to the cell media increased apoE secretion from the cells, in a concentration dependent fashion, without affecting cellular apoE levels. In contrast, apoA-I had no effect on PLTP synthesis and secretion, but strongly induced the PLTP activity in the media. ApoA-I also increased phospholipid transfer activity of PLTP isolated from human plasma. This effect was dependent on apoA-I concentration but independent on apoA-I lipidation status. ApoE, ApoA-II and apoA-IV, but not immunoglobulins or bovine serum albumin, also increased PLTP activity. We also report that apoA-I protects PLTP from heat inactivation. CONCLUSION: apoA-I enhances the phospholipid transfer activity of PLTP secreted from macrophage foam cells without affecting the PLTP mass.
Asunto(s)
Apolipoproteína A-I/farmacología , Células Espumosas/metabolismo , Proteínas de Transferencia de Fosfolípidos/metabolismo , Apolipoproteínas E/agonistas , Apolipoproteínas E/metabolismo , Células Cultivadas , Calor , Humanos , Sustancias Protectoras , Desnaturalización ProteicaRESUMEN
Cholesterol efflux from macrophages is a key process in reverse cholesterol transport and, therefore, might inhibit atherogenesis. 9-cis-ß-carotene (9-cis-ßc) is a precursor for 9-cis-retinoic-acid (9-cis-RA), which regulates macrophage cholesterol efflux. Our objective was to assess whether 9-cis-ßc increases macrophage cholesterol efflux and induces the expression of cholesterol transporters. Enrichment of a mouse diet with ßc from the alga Dunaliella led to ßc accumulation in peritoneal macrophages. 9-cis-ßc increased the mRNA levels of CYP26B1, an enzyme that regulates RA cellular levels, indicating the formation of RA from ßc in RAW264.7 macrophages. Furthermore, 9-cis-ßc, as well as all-trans-ßc, significantly increased cholesterol efflux to high-density lipoprotein (HDL) by 50% in RAW264.7 macrophages. Likewise, food fortification with 9-cis-ßc augmented cholesterol efflux from macrophages ex vivo. 9-cis-ßc increased both the mRNA and protein levels of ABCA1 and apolipoprotein E (APOE) and the mRNA level of ABCG1. Our study shows, for the first time, that 9-cis-ßc from the diet accumulates in peritoneal macrophages and increases cholesterol efflux to HDL. These effects might be ascribed to transcriptional induction of ABCA1, ABCG1, and APOE. These results highlight the beneficial effect of ßc in inhibition of atherosclerosis by improving cholesterol efflux from macrophages.
Asunto(s)
Aterosclerosis/prevención & control , HDL-Colesterol/metabolismo , Suplementos Dietéticos , Reguladores del Metabolismo de Lípidos/uso terapéutico , Macrófagos Peritoneales/metabolismo , Regulación hacia Arriba , beta Caroteno/análogos & derivados , Transportador 1 de Casete de Unión a ATP/agonistas , Transportador 1 de Casete de Unión a ATP/genética , Transportador 1 de Casete de Unión a ATP/metabolismo , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 1/agonistas , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 1/genética , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 1/metabolismo , Animales , Apolipoproteínas E/agonistas , Apolipoproteínas E/genética , Apolipoproteínas E/metabolismo , Aterosclerosis/inmunología , Aterosclerosis/metabolismo , Aterosclerosis/patología , Células Cultivadas , Chlorophyta/química , HDL-Colesterol/sangre , Inducción Enzimática , Reguladores del Metabolismo de Lípidos/metabolismo , Macrófagos Peritoneales/inmunología , Macrófagos Peritoneales/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Fitoplancton/química , Células RAW 264.7 , Receptores de LDL/genética , Receptores de LDL/metabolismo , Ácido Retinoico 4-Hidroxilasa/química , Ácido Retinoico 4-Hidroxilasa/genética , Ácido Retinoico 4-Hidroxilasa/metabolismo , beta Caroteno/metabolismo , beta Caroteno/uso terapéuticoRESUMEN
The apolipoprotein E (APOE) gene is the most highly associated susceptibility locus for late onset Alzheimer's Disease (AD), and augmenting the beneficial physiological functions of apoE is a proposed therapeutic strategy. In a high throughput phenotypic screen for small molecules that enhance apoE secretion from human CCF-STTG1 astrocytoma cells, we show the chrysanthemic ester 82879 robustly increases expressed apoE up to 9.4-fold and secreted apoE up to 6-fold and is associated with increased total cholesterol in conditioned media. Compound 82879 is unique as structural analogues, including pyrethroid esters, show no effect on apoE expression or secretion. 82879 also stimulates liver x receptor (LXR) target genes including ATP binding cassette A1 (ABCA1), LXRα and inducible degrader of low density lipoprotein receptor (IDOL) at both mRNA and protein levels. In particular, the lipid transporter ABCA1 was increased by up to 10.6-fold upon 82879 treatment. The findings from CCF-STTG1 cells were confirmed in primary human astrocytes from three donors, where increased apoE and ABCA1 was observed along with elevated secretion of high-density lipoprotein (HDL)-like apoE particles. Nuclear receptor transactivation assays revealed modest direct LXR agonism by compound 82879, yet 10 µM of 82879 significantly upregulated apoE mRNA in mouse embryonic fibroblasts (MEFs) depleted of both LXRα and LXRß, demonstrating that 82879 can also induce apoE expression independent of LXR transactivation. By contrast, deletion of LXRs in MEFs completely blocked mRNA changes in ABCA1 even at 10 µM of 82879, indicating the ability of 82879 to stimulate ABCA1 expression is entirely dependent on LXR transactivation. Taken together, compound 82879 is a novel chrysanthemic ester capable of modulating apoE secretion as well as apoE-associated lipid metabolic pathways in astrocytes, which is structurally and mechanistically distinct from known LXR agonists.
Asunto(s)
Transportador 1 de Casete de Unión a ATP/genética , Apolipoproteínas E/genética , Astrocitos/efectos de los fármacos , Receptores X del Hígado/genética , Piretrinas/farmacología , Receptores de LDL/genética , Transportador 1 de Casete de Unión a ATP/agonistas , Transportador 1 de Casete de Unión a ATP/metabolismo , Animales , Apolipoproteínas E/agonistas , Apolipoproteínas E/metabolismo , Astrocitos/citología , Astrocitos/metabolismo , Línea Celular Tumoral , Ésteres , Fibroblastos/citología , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Regulación de la Expresión Génica , Ensayos Analíticos de Alto Rendimiento , Humanos , Metabolismo de los Lípidos/efectos de los fármacos , Receptores X del Hígado/agonistas , Receptores X del Hígado/metabolismo , Ratones , Receptores Nucleares Huérfanos/genética , Receptores Nucleares Huérfanos/metabolismo , Cultivo Primario de Células , ARN Mensajero/genética , ARN Mensajero/metabolismo , Receptores de LDL/agonistas , Receptores de LDL/metabolismo , Transducción de SeñalRESUMEN
Human apolipoprotein E (apoE) consists of a single polypeptide chain with 299 amino acids and is best known for its role in the transport of cholesterol and other lipids between peripheral tissues and the liver. However, more direct effects of apoE on the vascular wall may well contribute to arterial protection from atherosclerosis. This review will focus on: (a) the ability of apoE to direct cholesterol efflux mechanisms with the aid of apoA1 and the ATP binding cassette transporter 1 (ABC1); (b) the ability of apoE to prevent platelet aggregation by facilitating the production of endogenous nitric oxide (NO); (c) the ability of apoE to inhibit the proliferation of T-lymphocytes by internalization of the IL-2 receptor; and (d) the ability of apoE to inhibit proliferation of endothelial cells by out competing growth factors for interaction with cell surface heparan sulfate proteoglycans (HSPG's). The characterization of apoE and its many functions has provided insight into the ultimate potential of this protein as a possible therapeutic agent for the treatment of atherosclerosis. This review will examine key scientific advances, which focus on possible therapeutic strategies that encompass the use of apoE in the amelioration of atherosclerosis.