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1.
Neurochem Int ; 52(8): 1416-21, 2008 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-18442871

RESUMEN

In the present study, the purpose is to determine activities of monoamine oxidases (MAO) in the brain of 263K scrapie-infected hamsters during the development of this experimental prion disease. Indeed, MAO activity modifications which have already been related in aging and neurodegenerations is suspected to be involved in the neuron loss process by elevated hydrogen peroxide formation. Monoamine oxidase type A (MAO-A) and B (MAO-B) activities were followed in the brain at different stages of the disease. MAO-A activity did not change significantly during the evolution of the disease. However, concerning the MAO-B activity, a significant increase was observed from 50 days post-infection and through the course of the disease and reached 42.9+/-5.3% at its ultimate stage. Regarding these results, MAO-B could be a potential therapeutic target then we have performed a pre-clinical treatment with irreversible (Selegiline or L-deprenyl) or and reversible (MS-9510) MAO-B inhibitors used alone or in association with an anti-scrapie drug such as MS-8209, an amphotericin B derivative. Our results show that none of the MAO-B inhibitors used was able to delay the onset of the disease. Neither these MAO-B inhibitors nor R-NMDA inhibitors (MK-801) can enhance the effects of MS-8209. The present findings clearly indicate a significant increase of cerebral MAO-B activity in scrapie-infected hamsters. Furthermore, inhibitors of MAO-B do not have any curative or palliative effect on this experimental model indicating that the raise of this activity is probably more a consequence rather than a causal event of the neurodegenerative process.


Asunto(s)
Monoaminas Biogénicas/metabolismo , Encéfalo/enzimología , Monoaminooxidasa/metabolismo , Proteínas PrPSc/metabolismo , Scrapie/enzimología , Anfotericina B/análogos & derivados , Anfotericina B/farmacología , Animales , Encéfalo/fisiopatología , Cricetinae , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Femenino , Mesocricetus , Inhibidores de la Monoaminooxidasa/farmacología , Degeneración Nerviosa/enzimología , Degeneración Nerviosa/fisiopatología , Scrapie/fisiopatología
2.
Fundam Clin Pharmacol ; 21(1): 29-34, 2007 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-17227442

RESUMEN

Type I interferons (IFNs) are widely used to treat viral diseases. Depressive symptoms and suicide attempts are common neuropsychiatric side-effects during treatment with type I IFNs. Activation of indoleamine-2,3-dioxygenase (IDO), the first and rate-limiting enzyme of the kynurenine pathway by IFNs, leads to an increase in tryptophan (Trp) catabolism. Low levels of Trp lead to decrease of serotonin synthesis, which is likely to be related to the depressive symptoms. Ovine type I interferon-tau (IFN-tau) has a more potent antiretroviral effect and is less toxic than human type I IFN-alpha. Effects of IFN-tau and IFN-alpha on IDO expression and activity in primary cultures of human macrophages were compared in parallel to those of IFN-gamma, considered as one of the most potent IDO inducer. We found that both IFN-alpha and IFN-tau were poor inducers of IDO compared with IFN-gamma. However, IDO activation was slightly and significantly lower with ovine IFN-tau than human IFN-alpha, suggesting that ovine IFN-tau might have a lower impact on serotoninergic pathway compared with human IFN-alpha.


Asunto(s)
Antivirales/farmacología , Indolamina-Pirrol 2,3,-Dioxigenasa/genética , Interferón Tipo I/farmacología , Interferón-alfa/farmacología , Macrófagos/efectos de los fármacos , Proteínas Gestacionales/farmacología , Animales , Células Cultivadas , Humanos , Indolamina-Pirrol 2,3,-Dioxigenasa/metabolismo , Macrófagos/enzimología , ARN Mensajero/metabolismo , Ovinos
3.
Nat Biotechnol ; 21(9): 1075-81, 2003 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-12910243

RESUMEN

We have developed a rapid, yeast-based, two-step assay to screen for antiprion drugs. The method allowed us to identify several compounds effective against budding yeast prions responsible for the [PSI+] and [URE3] phenotypes. These inhibitors include the kastellpaolitines, a new class of compounds, and two previously known molecules, phenanthridine and 6-aminophenanthridine. Two potent promoters of mammalian prion clearance in vitro, quinacrine and chlorpromazine, which share structural similarities with the kastellpaolitines, were also active in the assay. The compounds isolated here were also active in promoting mammalian prion clearance. These results validate the present method as an efficient high-throughput screening approach to identify new prion inhibitors and furthermore suggest that biochemical pathways controlling prion formation and/or maintenance are conserved from yeast to humans.


Asunto(s)
Fenantridinas/química , Fenantridinas/aislamiento & purificación , Priones/antagonistas & inhibidores , Priones/química , Mapeo de Interacción de Proteínas/métodos , Saccharomyces cerevisiae/aislamiento & purificación , Saccharomyces cerevisiae/metabolismo , Animales , Línea Celular Tumoral , Recuento de Colonia Microbiana/métodos , Humanos , Mamíferos , Ratones , Neuroblastoma , Fenantridinas/metabolismo , Saccharomyces cerevisiae/clasificación , Especificidad de la Especie , Levaduras/clasificación , Levaduras/aislamiento & purificación , Levaduras/metabolismo
4.
Neuroreport ; 17(1): 89-93, 2006 Jan 23.
Artículo en Inglés | MEDLINE | ID: mdl-16361957

RESUMEN

Magnetic resonance spectroscopy studies in animal models of prion disease are very few and concern terminal stages of infection. In order to study earlier stages of the disease, we used in-vivo magnetic resonance spectroscopy in a mouse model of scrapie and, for the first time, in mice infected with a bovine spongiform encephalopathy strain. In bovine spongiform encephalopathy-infected mice, we observed an increase in myo-inositol preceding clinical signs by 20 days, followed by a decrease in N-acetylaspartate at advanced stages. In scrapie-infected mice, changes in N-acetylaspartate and myo-inositol were detected at the beginning of the symptomatic phase. These results show that magnetic resonance spectroscopy is a valuable tool for detecting subtle metabolic changes associated to gliosis and neuronal dysfunction in prion diseases.


Asunto(s)
Espectroscopía de Resonancia Magnética , Enfermedades por Prión/metabolismo , Scrapie/metabolismo , Animales , Ácido Aspártico/análogos & derivados , Ácido Aspártico/metabolismo , Encéfalo/metabolismo , Encéfalo/patología , Bovinos , Modelos Animales de Enfermedad , Femenino , Infecciones/complicaciones , Imagen por Resonancia Magnética/métodos , Ratones , Ratones Endogámicos C57BL , Enfermedades por Prión/patología , Factores de Tiempo
5.
Cancer Res ; 64(2): 719-27, 2004 Jan 15.
Artículo en Inglés | MEDLINE | ID: mdl-14744790

RESUMEN

To define genetic determinants of tumor cell resistance to the cytotoxic action of tumor necrosis factor alpha (TNF), we have applied cDNA microarrays to a human breast carcinoma TNF-sensitive MCF7 cell line and its established TNF-resistant clone. Of a total of 5760 samples of cDNA examined, 3.6% were found to be differentially expressed in TNF-resistant 1001 cells as compared with TNF-sensitive MCF7 cells. On the basis of available literature data, the striking finding is the association of some differentially expressed genes involved in the phosphatidylinositol-3-kinase/Akt signaling pathway. More notably, we found that the PRNP gene coding for the cellular prion protein (PrP(c)), was 17-fold overexpressed in the 1001 cell line as compared with the MCF7 cell line. This differential expression was confirmed at the cell surface by immunostaining that indicated that PrP(c) is overexpressed at both mRNA and protein levels in the TNF-resistant derivative. Using recombinant adenoviruses expressing the human PrP(c,) our data demonstrate that PrP(c) overexpression converted TNF-sensitive MCF7 cells into TNF-resistant cells, at least in part, by a mechanism involving alteration of cytochrome c release from mitochondria and nuclear condensation.


Asunto(s)
Neoplasias de la Mama/patología , Muerte Celular/efectos de los fármacos , Resistencia a Antineoplásicos , Proteínas PrPC/farmacología , Factor de Necrosis Tumoral alfa/toxicidad , Línea Celular Tumoral , ADN Complementario/genética , Enzimas/genética , Femenino , Humanos , Análisis de Secuencia por Matrices de Oligonucleótidos , Transfección
6.
J Interferon Cytokine Res ; 25(8): 444-52, 2005 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-16108727

RESUMEN

Interferon-tau (IFN-tau) is a type I IFN responsible for maternal recognition of the fetus in ruminants. In addition to its physiologic role, IFN-tau also inhibits HIV replication in human lymphocytes and macrophages and displays immunomodulatory effects but lacks the toxicity associated with other type I IFNs. Human IFN-alpha promotes a Th1 response, whereas IFN-tau has anti-inflammatory properties, inducing the production of Th2 cytokines in murine models of experimental autoimmune encephalitis (EAE) or fetal loss. We compared the effects of ovine IFN-tau (OvIFN-tau) and human IFN-alpha (HuIFN-alpha) on cytokine mRNA and protein production in human peripheral blood mononuclear cells (PBMCs) activated with a recall antigen, such as purified protein derivative (PPD) of tuberculin or with a proinflammatory stimulus, such as lipopolysaccharide (LPS). In both cases, IFN-alpha increased IFN-gamma production, whereas IFN-tau did not and thereby promoted Th2 cytokine production. This original property renders IFN-tau a potential candidate for therapeutic applications in immune disorders, such as multiple sclerosis (MS), but its therapeutic use in the treatment of HIV infection should be considered with caution.


Asunto(s)
Interferón Tipo I/farmacología , Interferón gamma/biosíntesis , Linfocitos/efectos de los fármacos , Linfocitos/metabolismo , Proteínas Gestacionales/farmacología , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Citocinas/genética , Humanos , Interferón Tipo I/inmunología , Interferón-alfa/farmacología , Interferón gamma/inmunología , Lipopolisacáridos/farmacología , Linfocitos/citología , Linfocitos/inmunología , Proteínas Gestacionales/inmunología , ARN Mensajero/genética , ARN Mensajero/metabolismo
7.
J Neuropathol Exp Neurol ; 62(5): 475-85, 2003 May.
Artículo en Inglés | MEDLINE | ID: mdl-12769187

RESUMEN

Recent experimental studies showed that activated macrophages/microglia (AMM) express excitatory amino acid transporters (EAATs), suggesting that, in addition to their neurotoxic properties, they also have a neuroprotective role by clearing extracellular glutamate and producing antioxidant glutathione. To test this hypothesis in human, the brain of 12 HIV-positive patients and 3 controls were immunostained for EAAT-1. EAAT-1 was expressed by AMM in all HIV-infected cases but not in HIV-negative controls. Expression varied according to the disease stage. In 5 cases with active HIV-encephalitis (HIVE), AMM strongly expressed EAAT-1 in the white matter and basal ganglia, analogous to HLA-DR and CD68 expression. There was weaker expression in the cortex and perineuronal microglial cells were not involved. In a case with "burnt out" HIVE following highly active antiretroviral therapy (HAART), EAAT-1 expression was mild, identical to that of HLA-DR and CD68 in the white matter and cortex and involved perineuronal microglial cells. In 3 AIDS patients without HIVE and in 3 pre-AIDS cases, EAAT-1 expression in the white matter was weaker than HLA-DR and CD68 expression; there was stronger correlation in the gray matter where perineuronal microglial cells were stained predominantly. Our findings in humans tend to confirm that AMM, particularly perineuronal microglial cells, play a neuroprotective role in the early stages of HIV infection and, possibly, following treatment. This is in keeping with the early microglial activation seen in pre-AIDS cases, and the late occurrence of neuronal loss. It may also explain the reversible cognitive disorders following treatment in some cases.


Asunto(s)
Encéfalo/metabolismo , Transportador 1 de Aminoácidos Excitadores/metabolismo , Infecciones por VIH/metabolismo , Macrófagos/metabolismo , Microglía/metabolismo , Fármacos Neuroprotectores/metabolismo , Complejo SIDA Demencia/metabolismo , Complejo SIDA Demencia/patología , Adulto , Anciano , Biomarcadores , Encéfalo/citología , Encéfalo/patología , Proteína Ácida Fibrilar de la Glía/metabolismo , Infecciones por VIH/patología , Antígenos HLA-DR/metabolismo , Humanos , Masculino , Persona de Mediana Edad
8.
J Neuropathol Exp Neurol ; 63(10): 1058-71, 2004 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-15535133

RESUMEN

The mechanisms of neuronal apoptosis in prion diseases are unclear. Experimental studies suggest that it may result from 2 associated mechanisms: glutamate-mediated excitotoxicity and oxidative stress. Recent studies showed that activated macrophages/microglia (AMM) express excitatory amino acid transporters (EAATs) in HIV infection, suggesting that they may play a neuroprotective role by clearing extra-cellular glutamate and producing anti-oxidant glutathione. In order to test this hypothesis in prion diseases, samples from cerebral cortex, striatum, thalamus, and cerebellum from 14 patients with Creutzfeldt-Jakob disease (8 sporadic, 2 familial, 2 iatrogenic, and 2 variant), and 4 with fatal familial insomnia (3 homozygous Met/Met at codon 129 of the PRNP gene, 1 heterozygous Met/Val), and 3 controls were immunostained for EAAT-1, GFAP, HLA-DR, CD68, IL-1, caspase 3, and PrP. In prion diseases, EAAT-1 immunopositivity was found in affected areas. Only AMM, interstitial, perivascular, perineuronal (sometimes around apoptotic neurons), or close to reactive astrocytes, expressed EAAT-1. Astrocyte EAAT-1 expression was scarcely detectable in controls and was not detected in prion disease cases. The proportion of AMM expressing EAAT-1 did not correlate with the severity of neuronal apoptosis, spongiosis, astrocytosis, microgliosis, or PrP deposition, but only with disease duration. Occasional EAAT-1 expressing AMM were found in patients with short survival, whereas diffuse EAAT-1 expression by AMM was observed in cases with long survival (24 to 33 months) that most often were heterozygous for Met/Val at codon 129 of the PRNP gene. Our findings suggest that AMM may develop a partial neuroprotective function in long-lasting prion diseases, although it does not seem to efficiently prevent neurological and neuropathological deterioration. Whether this neuroprotective function of microglia is the cause or the effect of longer survival needs to be clarified.


Asunto(s)
Encéfalo/metabolismo , Síndrome de Creutzfeldt-Jakob/metabolismo , Transportador 1 de Aminoácidos Excitadores/metabolismo , Insomnio Familiar Fatal/metabolismo , Macrófagos/metabolismo , Microglía/metabolismo , Adulto , Anciano , Amiloide/genética , Encéfalo/patología , Estudios de Casos y Controles , Niño , Codón , Síndrome de Creutzfeldt-Jakob/genética , Síndrome de Creutzfeldt-Jakob/patología , Femenino , Heterocigoto , Humanos , Insomnio Familiar Fatal/genética , Insomnio Familiar Fatal/patología , Masculino , Metionina , Persona de Mediana Edad , Proteínas Priónicas , Priones , Precursores de Proteínas/genética , Índice de Severidad de la Enfermedad , Factores de Tiempo , Valina
9.
Brain Pathol ; 13(2): 211-22, 2003 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-12744474

RESUMEN

It is now widely accepted that neuronal damage in HIV infection results mainly from microglial activation and involves apoptosis, oxidative stress and glutamate-mediated neurotoxicity. Glutamate toxicity acts via 2 distinct pathways: an excitotoxic one in which glutamate receptors are hyperactivated, and an oxidative one in which cystine uptake is inhibited, resulting in glutathione depletion and oxidative stress. A number of studies show that astrocytes normally take up glutamate, keeping extracellular glutamate concentration low in the brain and preventing excitotoxicity. This action is inhibited in HIV infection, probably due to the effects of inflammatory mediators and viral proteins. Other in vitro studies as well as in vivo experiments in rodents following mechanical stimulation, show that activated microglia and brain macrophages express high affinity glutamate transporters. These data have been confirmed in chronic inflammation of the brain, particularly in SIV infection, where activated microglia and brain macrophages also express glutamine synthetase. Recent studies in humans with HIV infection show that activated microglia and brain macrophages express the glutamate transporter EAAT-1 and that expression varies according to the disease stage. This suggests that, besides their recognized neurotoxic properties in HIV infection, these cells also have a neuroprotective function, and may partly make up for the inhibited astrocytic function, at least temporarily. This hypothesis might explain the discrepancy between microglial activation which occurs early in the disease, and neuronal apoptosis and neuronal loss which is a late event. In this review article, we discuss the possible neuroprotective and neurotrophic roles of activated microglia and macrophages that may be generated by the expression of high affinity glutamate transporters and glutamine synthetase, 2 major effectors of glial glutamate metabolism, and the implications for HIV-induced neuronal dysfunction, the underlying cause of HIV dementia.


Asunto(s)
Sistema de Transporte de Aminoácidos X-AG/genética , Glutamato-Amoníaco Ligasa/genética , Infecciones por VIH/metabolismo , Macrófagos/metabolismo , Microglía/metabolismo , Simportadores/genética , Complejo SIDA Demencia/fisiopatología , Animales , Encéfalo/metabolismo , Encéfalo/patología , Regulación de la Expresión Génica , Proteínas de Transporte de Glutamato en la Membrana Plasmática , Infecciones por VIH/inmunología , Infecciones por VIH/patología , Humanos , Macrófagos/inmunología , Macrófagos/patología , Ratones , Microglía/inmunología , Microglía/patología , Fármacos Neuroprotectores/metabolismo , Ratas
10.
Retrovirology ; 1: 29, 2004 Sep 25.
Artículo en Inglés | MEDLINE | ID: mdl-15447786

RESUMEN

BACKGROUND: The aim of this study was to evaluate gene therapy for AIDS based on the transduction of circulating lymphocytes with a retroviral vector giving low levels of constitutive macaque interferon beta production in macaques chronically infected with a pathogenic isolate of SIVmac251. RESULTS: Two groups of three animals infected for more than one year with a pathogenic primary isolate of SIVmac251 were included in this study. The macaques received three infusions of their own lymphocytes transduced ex vivo with the construct encoding macaque IFN-beta (MaIFN-beta or with a vector carrying a version of the MaIFN-beta gene with a deletion preventing translation of the mRNA. Cellular or plasma viremia increased transiently following injection in most cases, regardless of the retroviral construct used. Transduced cells were detected only transiently after each infusion, among the peripheral blood mononuclear cells of all the animals, with copy numbers of 10 to 1000 per 106 peripheral mononuclear cells. CONCLUSION: Long-term follow-up indicated that the transitory presence of such a small number of cells producing such small amounts of MaIFN-beta did not prevent animals from the progressive decrease in CD4+ cell count typical of infection with simian immunodeficiency virus. These results reveal potential pitfalls for future developments of gene therapy strategies of HIV infection.


Asunto(s)
Síndrome de Inmunodeficiencia Adquirida/tratamiento farmacológico , Terapia Genética/métodos , Interferón beta/genética , Interferón beta/uso terapéutico , Transfusión de Linfocitos , Linfocitos/inmunología , Síndrome de Inmunodeficiencia Adquirida del Simio/tratamiento farmacológico , Virus de la Inmunodeficiencia de los Simios/inmunología , Animales , Secuencia de Bases , Recuento de Linfocito CD4 , Linfocitos T CD8-positivos/inmunología , Cartilla de ADN , ADN Viral/genética , ADN Viral/aislamiento & purificación , Vectores Genéticos , Interferón beta/administración & dosificación , Macaca fascicularis , Masculino , Regiones Promotoras Genéticas , ARN Viral/sangre , ARN Viral/genética , ARN Viral/aislamiento & purificación , Retroviridae , Eliminación de Secuencia , Síndrome de Inmunodeficiencia Adquirida del Simio/inmunología , Virus de la Inmunodeficiencia de los Simios/genética , Carga Viral
11.
FEBS Lett ; 529(1): 17-21, 2002 Oct 02.
Artículo en Inglés | MEDLINE | ID: mdl-12354606

RESUMEN

Transmissible spongiform encephalopathy (TSE) agents or prions induce neurodegenerative fatal diseases in humans and in some mammalian species. Human TSEs include Creutzfeldt-Jakob disease (CJD), Gerstmann-Sträussler-Scheinker syndrome, kuru and fatal familial insomnia. In animals, scrapie in sheep and goats, feline spongiform encephalopathy, transmissible mink encephalopathy, chronic wasting disease in wild ruminants, and bovine spongiform encephalopathy (BSE), which appeared in the UK in the mid-1980s [Wells, G.A.H. et al. (1987) Vet. Rec. 121, 419-420], belong to the TSE group. Prions have biological and physicochemical characteristics that differ significantly from those of other microorganisms; for example, they are resistant to inactivation processes that are effective against conventional viruses, including those that alter nucleic acid structure or function. Alternatively, infectivity is highly susceptible to procedures that modify protein conformation. Today, the exact nature of prions remains unknown even though it is likely that they consist of protein only. At the biochemical level, TSEs are characterised by the accumulation, within the central nervous system of the infected individual, of an abnormal isoform of a particular protein from the host, the prion protein [Prusiner, S.B. (1982) Science 216, 136-144]. TSEs are transmissible among their species of origin, but they can also cross the species barrier and induce chronic infection and/or disease in other species. Transmissibility has been proven in natural situations such as the outbreak of CJD among patients treated with pituitary-derived hormones and the appearance of BSE that affected UK cattle in the mid-1980s.


Asunto(s)
Enfermedades por Prión/etiología , Enfermedades por Prión/fisiopatología , Priones/fisiología , Animales , Humanos , Enfermedades por Prión/transmisión , Priones/química , Salud Pública
12.
Microbes Infect ; 6(2): 157-63, 2004 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-14998513

RESUMEN

Changes in the fine balance between matrix metalloproteinases and their tissue inhibitors, which drives extracellular matrix turnover, may be critical to central nervous system inflammation in HIV infection as well as in neurotoxicity. Although they do not produce virus when infected by HIV, astrocytes may be directly affected by the virion, because some viral proteins are known to transduce signaling in brain cells and are also sensitive to the major proinflammatory cytokine TNFalpha. We therefore studied the effects of HIV and TNFalpha on MMP-2, MMP-9 and their inhibitors, TIMP-1 and TIMP-2, in astrocytes, by zymography and ELISA, respectively, or by RT-PCR for both of them. HIV slightly increased the production of pro-MMP-2 and pro-MMP-9 by astrocytes, in a dose-dependent manner. TNFalpha strongly induced pro-MMP-9. TIMP-1 and TIMP-2 levels were affected only slightly, if at all, by HIV and TNFalpha. Thus, astrocyte/HIV contact may lead to extracellular matrix activation, which may be strongly amplified by the inflammatory response. Our data strongly suggest that, besides their physiological production of MMP-2, astrocytes would be a major source of MMP-9 in the inflamed brain.


Asunto(s)
Astrocitos/efectos de los fármacos , VIH/fisiología , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , Inhibidor Tisular de Metaloproteinasa-1/metabolismo , Inhibidor Tisular de Metaloproteinasa-2/metabolismo , Factor de Necrosis Tumoral alfa/farmacología , Astrocitos/enzimología , Astrocitos/virología , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Humanos , Metaloproteinasa 2 de la Matriz/genética , Metaloproteinasa 9 de la Matriz/genética , Inhibidores de la Metaloproteinasa de la Matriz , Sistemas Neurosecretores/fisiología , Inhibidor Tisular de Metaloproteinasa-1/genética , Virión/fisiología
13.
Antivir Ther ; 9(4): 519-28, 2004 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-15456083

RESUMEN

OBJECTIVES: To investigate whether P-glycoprotein (P-gp) and multidrug resistance proteins (MRPs), which limit the bioavailability of HIV protease inhibitors (PIs) and nucleoside reverse transcriptase inhibitors (NRTIs), modulate the anti-HIV activity of NRTIs, non-NRTIs and PIs in vitro. DESIGN: We used primary cultures of major HIV target cells: human monocyte-derived macrophages (MDMs) and lymphocytes. METHODS: P-gp and MRP expression in response to long-term zidovudine (3'-azido-3'-deoxythymidine; AZT) or indinavir treatment was quantified by RT-PCR. MDM and lymphocytes were infected in vitro with HIV-1/Ba-L and HIV-1-LAI, respectively, and treated with antiretroviral drugs. We evaluated the activity of these drugs in combination with PSC833, a P-gp inhibitor, and/or probenecid, an MRP1 inhibitor. Intracellular AZT triphosphate derivative (AZT-TP) was quantified by HPLC-MSMS. P-gp ATPase activity was measured with inside-out native membrane vesicles enriched in P-gp. RESULTS: Levels of MDR1, mrp4 and mrp5 mRNA were high following AZT treatment. In infected MDM, PSC833 and probenecid increased the anti-HIV activity of AZT and indinavir. AZT (5 nM) decreased HIV replication by 34% alone and by 72% in combination with P-gp/MRP inhibitors. Indinavir (10 nM) gave 14% inhibition alone and 81% in combination. The increase in anti-HIV activity of AZT was correlated with an increase in intracellular AZT-TP concentration. However, unlike PIs, neither AZT nor its metabolites interacted with P-gp. CONCLUSION: AZT increases the expression of multidrug transporters, thereby decreasing its pharmacological activity. The cellular efflux of AZT probably involves MRP4 or MRP5. In contrast, increases in indinavir anti-HIV activity require the inhibition of both P-gp and MRP1.


Asunto(s)
Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/metabolismo , Subfamilia B de Transportador de Casetes de Unión a ATP/metabolismo , Inhibidores de la Proteasa del VIH/farmacología , VIH-1/efectos de los fármacos , Indinavir/farmacología , Leucocitos Mononucleares/efectos de los fármacos , Macrófagos/efectos de los fármacos , Inhibidores de la Transcriptasa Inversa/farmacología , Zidovudina/farmacología , Subfamilia B de Transportador de Casetes de Unión a ATP/antagonistas & inhibidores , Subfamilia B de Transportador de Casetes de Unión a ATP/genética , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/antagonistas & inhibidores , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/genética , Células Cultivadas , Ciclosporinas/farmacología , VIH-1/metabolismo , Humanos , Leucocitos Mononucleares/metabolismo , Leucocitos Mononucleares/virología , Macrófagos/metabolismo , Macrófagos/virología , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/genética , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/metabolismo , Probenecid/farmacología , ARN Mensajero/análisis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de Tiempo
14.
J Med Chem ; 47(5): 1183-92, 2004 Feb 26.
Artículo en Inglés | MEDLINE | ID: mdl-14971898

RESUMEN

New formamidine-3TC (3TC = 2',3'-dideoxy-3'-thiacytidine) analogues have been synthesized through various methods, and their antiviral activities (HIV, HBV) have been evaluated in vitro. Anti-HIV-1 in acutely infected MT-4 cells and peripheral blood monocellular cells (PBMCs) showed that compounds substituted by N,N-diarylformamidine side chains at the 4-N nucleic base position (compounds 3 and 8-11) had at least equivalent anti-HIV activity as 3TC (EC50 = 0.5 and 11.6 microM, respectively). Moreover, the newly synthesized compounds demonstrated higher anti-HBV activity (EC50 ranging from 0.01 to 0.05 microM) compared to the parent nucleoside 3TC (EC50 = 0.2 microM). It should be underlined that these new promising derivatives inhibited HIV in cells of a macrophage lineage, which are known to be cellular reservoir for HIV. These results were particularly of interest, since the antiviral activities appeared not to be mediated through the formamidine bond hydrolysis and consequently the release of free 3TC. These new analogue series were found to be highly stable to hydrolysis even after prolonged incubation in different biological media (t(1/2) ranged from 48 to 120 h). This enzymatic stability, coupled to the fact that no delay in the antiviral response was observed compared to the free 3TC antiviral response, suggest that this new N,N-diarylformamidine nucleoside series should not be considered as classical prodrugs.


Asunto(s)
Amidinas/síntesis química , Antivirales/síntesis química , Lamivudine/análogos & derivados , Lamivudine/síntesis química , Amidinas/farmacología , Fármacos Anti-VIH/síntesis química , Fármacos Anti-VIH/farmacología , Antivirales/farmacología , Extractos Celulares , Línea Celular , Medios de Cultivo , Estabilidad de Medicamentos , VIH-1/efectos de los fármacos , Virus de la Hepatitis B/efectos de los fármacos , Humanos , Hidrólisis , Técnicas In Vitro , Lamivudine/farmacología , Monocitos/efectos de los fármacos , Monocitos/virología , Relación Estructura-Actividad
15.
J Med Chem ; 47(7): 1789-95, 2004 Mar 25.
Artículo en Inglés | MEDLINE | ID: mdl-15027871

RESUMEN

We synthesized a series of N-(N-acetyl-L-cysteinyl)-S-acetylcysteamine (10) analogues bearing various acyl groups on thiol cysteine or cysteamine residues, to investigate the structure-activity relationship for pro-GSH and anti-HIV properties in human macrophages. The S-substituents were ranked in the following order of efficacy: H > or = acetyl > isobutyryl > pivaloyl > benzoyl. We found that none of these derivatives had pro-GSH or antiviral activities in vitro higher than that of 10, but several displayed similar levels of anti-HIV activity, making them possible candidates for use as adjuvant therapies in conjunction with HAART, for treating neurological aspects of HIV infection.


Asunto(s)
Acetilcisteína/análogos & derivados , Acetilcisteína/síntesis química , Fármacos Anti-VIH/síntesis química , Antioxidantes/síntesis química , Macrófagos/efectos de los fármacos , Acetilcisteína/farmacología , Fármacos Anti-VIH/química , Fármacos Anti-VIH/farmacología , Antioxidantes/farmacología , Glutatión/metabolismo , Humanos , Técnicas In Vitro , Macrófagos/virología , Relación Estructura-Actividad
16.
J Med Chem ; 47(25): 6410-9, 2004 Dec 02.
Artículo en Inglés | MEDLINE | ID: mdl-15566310

RESUMEN

The HIV-1 central nervous system infection leads to the onset of neurological impairments called AIDS dementia complex (ADC). PAF plays an important role in this pathology, as it is an HIV-1-induced neurotoxin produced by infected or activated macrophages and microglia, in the brain. We previously reported that PAF-antagonists bearing a trisubstituted piperazine presented in vitro anti-HIV-1 activity in human macrophages. To improve the pharmacological activities of our lead compound, 1a, we modified its carbamate function and evaluated both its antiretroviral and anti-PAF activities. One carbamate derivative (10c) demonstrated a similar antiviral activity but a higher anti-PAF potency, whereas 4a, with an ureide function, presents an increased antiviral activity and can be considered as a pure antiretroviral drug, as it does not present PAF-antagonism. Moreover, we measured the ability of 1a to cross the blood-brain barrier, using the in situ mouse brain perfusion method and its plasmatic concentrations after iv and po administration. The transport parameter measured (K(in)) proves that 1a is able to cross this biological barrier, but a pharmacokinetic study reveals its weak bioavailability in rats.


Asunto(s)
Fármacos Anti-VIH/síntesis química , VIH-1/efectos de los fármacos , Piperazinas/síntesis química , Factor de Activación Plaquetaria/antagonistas & inhibidores , Animales , Fármacos Anti-VIH/química , Fármacos Anti-VIH/farmacología , Disponibilidad Biológica , Barrera Hematoencefálica/metabolismo , Carbamatos/síntesis química , Carbamatos/química , Carbamatos/farmacología , Células Cultivadas , Humanos , Técnicas In Vitro , Macrófagos/efectos de los fármacos , Macrófagos/virología , Masculino , Ratones , Permeabilidad , Piperazinas/química , Piperazinas/farmacología , Agregación Plaquetaria/efectos de los fármacos , Conejos , Ratas , Ratas Sprague-Dawley , Relación Estructura-Actividad , Replicación Viral/efectos de los fármacos
17.
Immunol Lett ; 90(2-3): 223-8, 2003 Dec 15.
Artículo en Inglés | MEDLINE | ID: mdl-14687729

RESUMEN

As natural killer T (NKT) cells have been implicated in the regulation of multiple sclerosis (MS), we investigated expression of the Valpha24JalphaQ canonical rearrangement in MS patients during relapses. We observed major changes in the entire blood Valpha24(+) T-cell repertoire. Seven of the eight patients showed a marked decrease in Valpha24(+) transcript number and a decrease in the diversity of the Valpha24(+) T-cell repertoire, with the exception of a few expanded clones. These perturbations, exacerbated in patient MS (A), led to circulating NKT cell defects.


Asunto(s)
Células Asesinas Naturales/inmunología , Esclerosis Múltiple/inmunología , Receptores de Antígenos de Linfocitos T alfa-beta/inmunología , Linfocitos T/inmunología , Linfocitos T/metabolismo , Secuencia de Aminoácidos , Células Cultivadas , Humanos , Esclerosis Múltiple/sangre , Esclerosis Múltiple/patología , Receptores de Antígenos de Linfocitos T alfa-beta/química , Receptores de Antígenos de Linfocitos T alfa-beta/genética , Recurrencia
18.
AIDS Res Hum Retroviruses ; 19(1): 41-7, 2003 Jan 01.
Artículo en Inglés | MEDLINE | ID: mdl-12596719

RESUMEN

Non-B HIV-1 samples collected in France between 1999 and 2001 were sequenced in the env, reverse transcriptase (RT), and protease genes (1) to characterize further the non-B strains circulating in the country, (2) to assess the importance of recombination, and (3) to describe the polymorphism of RT and protease genes and appreciate a possible impact on susceptibility to antiretroviral (ARV) drugs. The results show that, within a background of CRF02_AG predominance, there is a high genetic diversity of non-B isolates, including intersubtype recombinants. There is an extensive polymorphism of protease and RT genes compared with B consensus sequences; we have so far no data indicating that these non-B isolates may have reduced sensitivity to ARV drugs.


Asunto(s)
Fármacos Anti-VIH/farmacología , Genes env/genética , Variación Genética , Infecciones por VIH/virología , Proteasa del VIH/genética , Transcriptasa Inversa del VIH/genética , VIH-1/efectos de los fármacos , VIH-1/genética , Adulto , Sustitución de Aminoácidos , Fármacos Anti-VIH/uso terapéutico , Farmacorresistencia Viral , Femenino , Francia , Infecciones por VIH/tratamiento farmacológico , Proteasa del VIH/química , Transcriptasa Inversa del VIH/química , VIH-1/clasificación , Humanos , Masculino , Persona de Mediana Edad , Recombinación Genética , Inhibidores de la Transcriptasa Inversa/farmacología , Inhibidores de la Transcriptasa Inversa/uso terapéutico , Análisis de Secuencia de ADN
19.
AIDS Res Hum Retroviruses ; 19(4): 307-11, 2003 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-12804006

RESUMEN

The beneficial effects of highly active antiretroviral therapy (HAART) in HIV-infected patients may be limited by inadequate compliance and viral resistance, but also by host cell factors, such as P-glycoprotein (P-gp) and intracellular kinases involved in the phosphorylation of nucleoside reverse transcriptase inhibitors. We investigated the effects of infection and HAART (zidovudine [AZT], lamivudine [3TC], and indinavir [IDV] on the expression of P-gp and cell kinases involved in the phosphorylation of AZT and 3TC in SHIV89.6P-infected cynomolgus macaques. Under unstimulated conditions, we observed a decrease in P-gp mRNA levels in the peripheral blood and lymph node mononuclear cells of infected macaques, which was accentuated by HAART. SHIV infection also resulted in the overexpression of thymidine kinase mRNA, which was abolished by HAART. In conclusion, retroviral infection and HAART modulate in vivo at the transcriptional level the expression of host cell factors that may affect the efficacy of HAART.


Asunto(s)
Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/metabolismo , Terapia Antirretroviral Altamente Activa , Regulación de la Expresión Génica , Infecciones por VIH/tratamiento farmacológico , VIH-1 , Virus de la Inmunodeficiencia de los Simios , Timidina Quinasa/metabolismo , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/genética , Animales , Modelos Animales de Enfermedad , Infecciones por VIH/inmunología , Infecciones por VIH/virología , VIH-1/genética , VIH-1/fisiología , Humanos , Leucocitos Mononucleares/metabolismo , Ganglios Linfáticos/citología , Ganglios Linfáticos/inmunología , Recuento de Linfocitos , Macaca fascicularis , ARN Viral/sangre , Virus de la Inmunodeficiencia de los Simios/genética , Virus de la Inmunodeficiencia de los Simios/fisiología , Timidina Quinasa/genética , Transcripción Genética
20.
J Mol Neurosci ; 18(3): 271-81, 2002 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-12059046

RESUMEN

Amphotericine B (AmB), a macrolide polyene antibiotic, is one of a few drugs that has shown therapeutic properties in scrapie-infected hamster. Its beneficial effect on survival time is mostly marked when animals are treated with its derivative MS-8209. To explore the MS-8209 effect at the cellular level, we investigated at the light and electron microscopy levels, the sequential appearance and distribution of PrP concurrently with histopathological changes in hamsters that were infected intracerebrally with the 263 K scrapie strain and treated or not with the drug. The first histopathological modifications and PrP immunostaining were observed in the thalamus and at the inoculation site where the drug caused a delay in the appearance of lesions and PrP accumulation. Using immunoelectron microscopy, at 70 d postinfection, the inoculation site of untreated animals showed an accumulation of PrP in plaque areas constitued by filaments mixed with alterated membrane structures and in developed lysosomal system of reactive astrocytes. Most of the numerous lysosomes containing PrP showed intra-organelle filaments. In contrast, in MS-8209 treated animals, the number of lysosomes was significantly lower (p < 0.0038), with very few organelles harboring PrP. Our results suggest that in this scrapie model, MS-8209 treatment delays the disease by preventing the replication of the scrapie agent at the inoculation site where the astrocytes appear to be the first cells producing abnormal PrP. The lysosomal system of these astrocytes could constitute a privileged target for MS-8209.


Asunto(s)
Anfotericina B/análogos & derivados , Anfotericina B/farmacología , Astrocitos/efectos de los fármacos , Lisosomas/efectos de los fármacos , Scrapie/patología , Anfotericina B/administración & dosificación , Animales , Astrocitos/metabolismo , Astrocitos/ultraestructura , Encéfalo/citología , Encéfalo/patología , Cricetinae , Femenino , Gliosis/patología , Humanos , Lisosomas/metabolismo , Lisosomas/ultraestructura , Mesocricetus , Priones/química , Priones/metabolismo , Scrapie/metabolismo
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