RESUMO
Central dopaminergic (DA) systems are affected during human immunodeficiency virus (HIV) infection. So far, it is believed that they degenerate with progression of HIV disease because deterioration of DA systems is evident in advanced stages of infection. In this manuscript we found that (a) DA levels are increased and DA turnover is decreased in CSF of therapy-naïve HIV patients in asymptomatic infection, (b) DA increase does not modulate the availability of DA transporters and D2-receptors, (c) DA correlates inversely with CD4+ numbers in blood. These findings show activation of central DA systems without development of adaptive responses at DA synapses in asymptomatic HIV infection. It is probable that DA deterioration in advanced stages of HIV infection may derive from increased DA availability in early infection, resulting in DA neurotoxicity. Our findings provide a clue to the synergism between DA medication or drugs of abuse and HIV infection to exacerbate and accelerate HIV neuropsychiatric disease, a central issue in the neurobiology of HIV.
Assuntos
Dopamina/metabolismo , Infecções por HIV/metabolismo , Infecções por HIV/patologia , Transmissão Sináptica/fisiologia , Ácido 3,4-Di-Hidroxifenilacético/líquido cefalorraquidiano , Adulto , Benzamidas , Antígenos CD4/metabolismo , Estudos de Casos e Controles , Quimiocina CCL2/metabolismo , Galactosefosfatos/metabolismo , HIV/genética , Infecções por HIV/líquido cefalorraquidiano , Infecções por HIV/diagnóstico por imagem , Infecções por HIV/imunologia , Ácido Homovanílico/líquido cefalorraquidiano , Humanos , Masculino , Pessoa de Meia-Idade , Pirrolidinas , Tomografia Computadorizada de Emissão de Fóton Único/métodos , Tropanos , Carga Viral/métodosRESUMO
Stimulation of death receptors such as CD95 or TNF-R1 results in rapid onset of apoptosis. Here we show that inhibition of death receptor-induced apoptosis by the broad range caspase inhibitor ZVAD causes a switch from apoptotic to proinflammatory signaling. In previous studies we have reported that caspase inhibitors induce expression of various proinflammatory cytokines in CD95-stimulated primary T cells, such as TNF-alpha, IFN-gamma and GM-CSF. In this study we provide further evidence for the proinflammatory activity of CD95. Stimulation of CD95 by agonistic antibodies (7C11) resulted in expression of IL-2 in primary T cells, which was further enhanced when caspase activity was blocked by ZVAD. Moreover, CD95 triggered expression of IL-4 and IL-8 when caspase activity was inhibited, but not in the absence of ZVAD. Our findings are of significant importance for the CNS as changes in the cytokine pattern in the periphery affects the entry of various immune cells into the brain. Moreover, invading activated T cells can also directly influence the cytokine profile within the brain, triggering signaling cascades that eventually lead to neuronal cell death. The use of caspase inhibitors to prevent apoptotic cell death should be carefully evaluated in the management of systemic and CNS diseases.
Assuntos
Apoptose , Citocinas/metabolismo , Regulação da Expressão Gênica/fisiologia , Linfócitos T/metabolismo , Clorometilcetonas de Aminoácidos/farmacologia , Anticorpos/farmacologia , Apoptose/efeitos dos fármacos , Interações Medicamentosas , Ensaio de Imunoadsorção Enzimática/métodos , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Modelos Biológicos , Fármacos Neuroprotetores/farmacologia , Linfócitos T/efeitos dos fármacos , Receptor fas/imunologiaRESUMO
The basal ganglia, structures rich in the neurotransmitter dopamine, are primarily affected during human immunodeficiency virus (HIV) infection. The authors measured levels of dopamine and its metabolites, homovanillic acid and 3,4-dihydroxyphenylacetic acid, in brains of uninfected and simian immunodeficiency virus (SIV)-infected rhesus monkeys during the asymptomatic stage of the infection. Moreover, the authors investigated changes in cyclic adenosine monophosphate (cAMP) and cAMP response element-binding protein (CREB), two factors involved in the signaling pathway of dopamine. The brain regions examined were the nucleus accumbens and the corpus amygdaloideum, which are limbic structures of the basal ganglia that are involved in the pathophysiology of psychiatric disorders and substance abuse. Dopamine content was reduced in both regions of SIV-infected monkeys compared to uninfected animals. Moreover, dopamine deficits were associated with a decrease in expression of total CREB. Intracellular concentrations of cAMP were decreased in nucleus accumbens and remained unchanged in corpus amygdaloideum of SIV-infected macaques. Changes in dopamine signaling were not related to pathology or viral load of the investigated animals. The results suggest that dopamine defects precede neurologic deficits and implicate dysfunction of the dopaminergic system in the etiopathogenesis of HIV dementia. Therefore, affective complications in HIV subjects should not be interpreted only as reactive psychological changes. The alterations in the mesolimbic dopaminergic system during asymptomatic stage of SIV infection implicate a biological background for psychiatric disorders in HIV infection.
Assuntos
Encéfalo/metabolismo , AMP Cíclico/metabolismo , Dopamina/metabolismo , Transdução de Sinais/fisiologia , Síndrome de Imunodeficiência Adquirida dos Símios/fisiopatologia , Ácido 3,4-Di-Hidroxifenilacético/análise , Ácido 3,4-Di-Hidroxifenilacético/metabolismo , Animais , Encéfalo/patologia , Encéfalo/virologia , Cromatografia Líquida de Alta Pressão , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Dopamina/análise , Eletroforese em Gel de Poliacrilamida , Ácido Homovanílico/análise , Ácido Homovanílico/metabolismo , Immunoblotting , Imuno-Histoquímica , Macaca mulatta , Síndrome de Imunodeficiência Adquirida dos Símios/patologia , Vírus da Imunodeficiência Símia/fisiologiaRESUMO
Drug abuse and human immunodeficiency virus (HIV) infection seem to cause cumulative damage in the central nervous system (CNS). Elevated extracellular dopamine is thought to be a prime mediator of the reinforcing effects of addictive substances. To investigate the possible role of increased dopamine availability in the pathogenesis of HIV dementia, simian immunodeficiency virus (SIV)-infected monkeys were treated with dopaminergic drugs (selegiline or L-DOPA). Both substances increased intracerebral SIV expression, combined with aggravation of infection-related neuropathology and ultrastructural alterations of dendrites in dopaminergic areas (spongiform polioencephalopathy) in asymptomatic animals. Moreover, this treatment resulted in enhanced TNF-alpha expression in the brains of SIV-infected animals. These findings indicate a synergistic interaction between dopamine and SIV infection on microglia activation, leading to increased viral replication and production of neurotoxic substances. Our results suggest that increased dopamine availability through dopaminergic medication or addictive substances may potentiate HIV dementia.
Assuntos
Dopaminérgicos/efeitos adversos , Síndrome de Imunodeficiência Adquirida dos Símios/virologia , Vírus da Imunodeficiência Símia/efeitos dos fármacos , Animais , Contagem de Células/métodos , Viroses do Sistema Nervoso Central/patologia , Viroses do Sistema Nervoso Central/virologia , Relação Dose-Resposta a Droga , Expressão Gênica/efeitos dos fármacos , Imuno-Histoquímica/métodos , Hibridização In Situ/métodos , Inflamação/etiologia , Inflamação/patologia , Inflamação/virologia , Interleucina-1/metabolismo , Interleucina-6/metabolismo , Levodopa/efeitos adversos , Macaca mulatta , Monoaminoxidase/metabolismo , Selegilina/efeitos adversos , Síndrome de Imunodeficiência Adquirida dos Símios/patologia , Vírus da Imunodeficiência Símia/fisiologia , Coloração e Rotulagem/métodos , Estatísticas não Paramétricas , Fatores de Tempo , Fator de Necrose Tumoral alfa/metabolismo , Replicação Viral/efeitos dos fármacosRESUMO
The role of the viral burden in the brain for the pathogenesis of human immunodeficiency virus-associated neurological disorders is still unclear. To address this issue, we have quantified the viral load in plasma, cerebrospinal fluid (CSF) and brain tissue of macaques infected with simian immunodeficiency virus (SIV). We discovered that the viral strain used for infection determines the replicative capacity in microglial cells as well as the extent of neuropathological lesions and the occurrence of neurological symptoms. Moreover, the viral load in the brain parenchyma correlated with the development of overt neurological disease whereas the one in plasma did not. By comparing the viral load in three different compartments, we demonstrated that the viral burden in the CSF is influenced both by the viral replication in the periphery as well as in the brain parenchyma. According to these results, it is not the absolute amount of viral load in the CSF but rather the viral antigen contributed by the viral production within the brain which correlates with the development of neurological disease. In longitudinal studies, we observed that this autochthonous virus production, as evidenced by a ratio of the viral load in CSF to the one in plasma, takes place for a prolonged period of time before overt neurological signs are manifested. This finding suggests that this ratio could be used as a prognostic marker for immunodeficiency virus-induced neurological disease.
Assuntos
Antígenos Virais/análise , Encéfalo/virologia , Encefalite Viral/virologia , Produtos do Gene gag/análise , Microglia/virologia , Síndrome de Imunodeficiência Adquirida dos Símios/virologia , Vírus da Imunodeficiência Símia/patogenicidade , Complexo Relacionado com a AIDS/líquido cefalorraquidiano , Complexo Relacionado com a AIDS/diagnóstico , Animais , Antígenos Virais/sangue , Antígenos Virais/líquido cefalorraquidiano , Encéfalo/patologia , Linfócitos T CD4-Positivos/virologia , Modelos Animais de Doenças , Encefalite Viral/sangue , Encefalite Viral/líquido cefalorraquidiano , Ensaio de Imunoadsorção Enzimática , Produtos do Gene gag/sangue , Produtos do Gene gag/líquido cefalorraquidiano , Hibridização In Situ , Macaca mulatta , Prognóstico , RNA Viral/análise , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Síndrome de Imunodeficiência Adquirida dos Símios/sangue , Síndrome de Imunodeficiência Adquirida dos Símios/líquido cefalorraquidiano , Vírus da Imunodeficiência Símia/genética , Vírus da Imunodeficiência Símia/isolamento & purificação , Carga Viral , Replicação ViralRESUMO
HIV infection is associated with a marked vulnerability of the dopaminergic system. We found recently that dopaminergic substances increase brain pathology in the simian model of HIV infection. In the current study we used the chronically HIV-infected T-lymphoblasts ACH-2 to elucidate the effects of dopamine (DA) on HIV infection. Cells were exposed to various concentrations of DA for 24 hours. Flow cytometry measurements demonstrated that DA induced a concentration-dependent HIV activation. To study the mechanism of action of DA, cells were treated besides DA with glutathione, one of the main components of cellular defense mechanisms against oxidative stress as well as its indirect precursor N-acetylcysteine. Treatment with these antioxidants attenuated DA-induced-HIV activation indicating that changes in cellular redox states might have been the causative factor for the observed effect. Our data suggest that HIV activation is tightly linked to intracellular oxidant/antioxidant levels and that excessive DA exposure may modulate cellular vulnerability to HIV.
Assuntos
HIV/crescimento & desenvolvimento , Linfócitos T/virologia , Ativação Viral/efeitos dos fármacos , Acetilcisteína/farmacologia , Linhagem Celular , Citometria de Fluxo , Produtos do Gene gag/análise , Glutationa/farmacologia , HIV/efeitos dos fármacos , Proteína do Núcleo p24 do HIV/análise , Humanos , CinéticaRESUMO
Upon inoculation into neonatal rats, murine leukemia virus (MuLV) NT40 causes a non-inflammatory degeneration of the central nervous system. While microglia cells appear to be the major target cells within the brain parenchyma for neurovirulent MuLV, degenerating neurons do not express retroviral gene products. In order to protect rats from neuronal damage we treated retrovirally infected rats once with monoamine oxidase (MAO) B inhibitor Selegiline which--under different conditions--exerts neuroprotective effects. Unexpectedly, when administered at 17 days post-infection (d.p.i.) a single intraperitoneal dose of Selegilin (1 mg/kg bodyweight) significantly shortened the incubation period for neurological disease. In contrast, Selegiline given in a lower dosage (0.05 mg/kg bodyweight) and/or at a different time point (13 d.p.i.) at the low (0.05 mg/kg bodyweight) and the high dose (1.0 mg/kg bodyweight) had no effect on the outcome of neurological disease. Animals treated with Selegiline (1.0 mg/kg bodyweight at 17 d.p.i.) contained higher amounts of viral loads in the CNS, higher numbers of brain cells expressing major histocompatibility complex class II molecules, and exhibited inhibition of MAO-B in comparison to untreated yet infected (control) animals. Supposedly, Selegiline activated the major target cell population of the CNS for MuLV-NT40, microglia, with the consequence of enhanced susceptibility for retroviral infection and triggered endogenous mechanism(s) involved in the pathogenesis of retroviral neurodegeneration.
Assuntos
Vírus da Leucemia Murina/metabolismo , Inibidores da Monoaminoxidase/toxicidade , Doenças Neurodegenerativas/virologia , Infecções por Retroviridae/virologia , Selegilina/toxicidade , Infecções Tumorais por Vírus/virologia , Animais , Animais Recém-Nascidos , Encéfalo/metabolismo , Encéfalo/virologia , Antígenos de Histocompatibilidade Classe II/metabolismo , Humanos , Doenças Neurodegenerativas/metabolismo , Doenças Neurodegenerativas/prevenção & controle , RNA Viral/metabolismo , Ratos , Ratos Endogâmicos F344 , Infecções por Retroviridae/metabolismo , Infecções por Retroviridae/prevenção & controle , Infecções Tumorais por Vírus/metabolismo , Infecções Tumorais por Vírus/prevenção & controleRESUMO
The aim of the present study was to assess the toxic potential of drugs of abuse and other neuropharmacological agents in the pathogenesis of AIDS dementia complex (ADC), the neurological complication of AIDS. Neuroblastoma and glioblastoma cell lines expressing the dopamine transporter, as well as primary macrophages exposed to human immunodeficiency virus-1 (HIV-1), were used to investigate the possibility of any synergistic effect between the mode of toxicity of such substances and virus exposure. The drugs of abuse used in our experiments were cocaine and morphine, which exert their action, among others, on the dopaminergic system. Effects were compared to treatment with dopamine itself and a typical dopaminergic drug used pharmaceutically, selegiline. In macrophage cultures, glutathione (GSH) was upregulated strongly after treatment with dopamine, morphine or selegiline, and this effect was enhanced when cells were pre-exposed to virus. This upregulation is discussed as a compensatory reaction to an oxidative signal. When hydrogen peroxide plus iron sulfate was used as a strong oxidant in macrophages, GSH concentrations decreased as a result of cell injury. Cell numbers remained constant in all treatment groups. In contrast, in both neuroblastoma and glioblastoma cell lines, the modulation of GSH concentrations by neurotropic substances was accompanied by significant cell loss, which was exacerbated by HIV-1 pretreatment. Selegiline did not change cell numbers when incubated alone. However, when incubated following treatment with HIV-1 cell death was highly significant. Ascorbic acid (AA), included as antioxidant, totally restored cell loss in cultures treated with dopamine. However, no effect was observed in combined treatment of AA and morphine or selegiline. The results demonstrate a synergistic role in cellular toxicity due to neurotropic substances and HIV-1, and suggest that neuropharmacological agents may contribute to the pathogenesis of ADC.
Assuntos
Complexo AIDS Demência/etiologia , Glutationa/fisiologia , HIV-1/patogenicidade , Glicoproteínas de Membrana , Proteínas de Membrana Transportadoras , Proteínas do Tecido Nervoso , Transtornos Relacionados ao Uso de Substâncias/complicações , Proteínas de Transporte/metabolismo , Sobrevivência Celular , Células Cultivadas , Cocaína/farmacologia , Progressão da Doença , Dopamina/farmacologia , Proteínas da Membrana Plasmática de Transporte de Dopamina , Inibidores da Captação de Dopamina/farmacologia , Humanos , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Macrófagos/virologia , Morfina/farmacologia , Entorpecentes/farmacologia , Estresse Oxidativo , Selegilina/farmacologia , Células Tumorais Cultivadas , Regulação para CimaRESUMO
The U-373 MG glioblastoma and the IMR-32 neuroblastoma cell lines were found to express the dopamine (DA) and vesicular monoamine transporters, using reverse transcriptase-polymerase chain reaction (RT-PCR). To further characterize the DA transporter, [3H]GBR-12935 binding and [3H]DA uptake studies were performed. Specific binding of [3H]GBR-12935 to U-373 MG and IMR-32 cells is saturable as saturation experiments indicated. Scatchard analysis revealed two binding sites on U-373 MG as well as on IMR-32 cells. The high-affinity sites exhibited a KD of 2.95 and 0.42 nM and a Bmax of 6.4 and 0.83 fmol/mg protein for U-373 MG and IMR-32 cells, respectively. The low-affinity sites exhibited a KD of 144 and 251 nM and a Bmax of 37.5 and 119 fmol/mg protein for the same cells, respectively. The high-affinity binding of both types of cells probably represents the "classic" DA uptake site identified in other studies from human and rat striatal membranes or synaptosomes, while the low-affinity binding may represent a mazindol-insensitive binding site (the "piperazine acceptor site"). [3H]DA uptake was 0.55 +/- 0.16 and 1.08 +/- 0.33 pmol/mg protein for U-373 MG and IMR-32 cells, respectively. Since the DA transporter has been implicated as an important site for drugs and toxins, the above-mentioned cell lines may be a useful tool in the study of the mechanism of action of DA transporter modulating substances.
Assuntos
Neoplasias Encefálicas/metabolismo , Proteínas de Transporte/biossíntese , Glioblastoma/metabolismo , Glicoproteínas de Membrana/biossíntese , Proteínas de Membrana Transportadoras , Proteínas do Tecido Nervoso , Neuroblastoma/metabolismo , Neuropeptídeos , Animais , Linhagem Celular , Dopamina/metabolismo , Proteínas da Membrana Plasmática de Transporte de Dopamina , Humanos , Cinética , Piperazinas/metabolismo , Piperazinas/farmacologia , Reação em Cadeia da Polimerase , RNA Mensageiro/biossíntese , Ratos , Proteínas Vesiculares de Transporte de Aminas BiogênicasRESUMO
Microglia are the major target for human immunodeficiency virus (HIV) infection within the central nervous system. Because only a few cells are productively infected, it has been suggested that an aberrant cytokine production by this cell population may be an indirect mechanism leading to the development of neurological disorders in HIV-infected patients. Therefore we decided to study the secretion pattern of several interleukins (IL) by microglial cells and peripheral blood macrophages isolated from uninfected and simian immunodeficiency virus (SIV)-infected Rhesus monkeys. We found that uninfected, unstimulated primate microglia produce more IL-6 and less TNF alpha than peripheral blood macrophages, but generate comparable levels of IL-1 beta and IL-8. After infection with SIV in vitro, synthesis of all cytokines tested is increased compared to uninfected cultures and to peripheral blood macrophages. Microglia isolated from infected animals produce more IL-8 and TNF alpha than the uninfected cultures and display a strongly increased capacity to secrete TNF alpha upon stimulation with lipopolysaccharide. In addition, production of IL-6 by in vivo-infected microglia increases with time in culture to very high levels despite the fact that only a few cells contained replicating virus. These findings clearly show that the cytokine production of microglia is impaired after SIV infection both in vitro and in vivo and that a low level of viral replication is sufficient for these alterations to occur. In conclusion, the results of this study further support a possible role of cytokines in the pathogenesis of neuro-AIDS.