RESUMEN
La dermatitis atópica es la forma más frecuente de eccema durante el primer año de vida; sin embargo, cuando la presentación es atípica o se asocia a infecciones, constituye un desafío diagnóstico para el pediatra. Es importante mantener un índice alto de sospecha para detectar inmunodeficiencias primarias asociadas a eccemas graves desde el período neonatal. Un ejemplo de estas es el síndrome de hiperinmunoglobulinemia E (hiper-IgE) autosómico dominante. Este cuadro se caracteriza por la presencia de infecciones cutáneas y respiratorias recurrentes, dermatitis atópica, eosinofilia y aumento de IgE. Se reporta el caso clínico de una niña de 1 mes y 29 días con diagnóstico de hiper-IgE con afección cutánea desde el nacimiento.
Atopic dermatitis is the most common form of eczema often developed before the first year of life. Nevertheless, when the presentation is atypical or related to infections the diagnostic represents a challenge for the pediatricians. It is important to maintain a high index of suspicion for the detection of primary immunodeficiency associated to severe eczema. One of them is the autosomal dominant hyper-IgE syndrome characterized by recurrent skin and respiratory infections, atopic dermatitis, eosinophilia, and high serum IgE concentrations. In this paper, we report a 1 months and 29 days old baby girl diagnosed with hyper-IgE and a skin involvement since birth.
Asunto(s)
Humanos , Femenino , Lactante , Dermatitis Atópica/diagnóstico , Síndrome de Job/diagnóstico , Dermatitis Atópica/inmunología , Eccema/diagnóstico , Eccema/inmunología , Síndrome de Job/complicacionesRESUMEN
Atopic dermatitis is the most common form of eczema often developed before the first year of life. Nevertheless, when the presentation is atypical or related to infections the diagnostic represents a challenge for the pediatricians. It is important to maintain a high index of suspicion for the detection of primary immunodeficiency associated to severe eczema. One of them is the autosomal dominant hyper-IgE syndrome characterized by recurrent skin and respiratory infections, atopic dermatitis, eosinophilia, and high serum IgE concentrations. In this paper, we report a 1 months and 29 days old baby girl diagnosed with hyper-IgE and a skin involvement since birth.
La dermatitis atópica es la forma más frecuente de eccema durante el primer año de vida; sin embargo, cuando la presentación es atípica o se asocia a infecciones, constituye un desafío diagnóstico para el pediatra. Es importante mantener un índice alto de sospecha para detectar inmunodeficiencias primarias asociadas a eccemas graves desde el período neonatal. Un ejemplo de estas es el síndrome de hiperinmunoglobulinemia E (hiper-IgE) autosómico dominante. Este cuadro se caracteriza por la presencia de infecciones cutáneas y respiratorias recurrentes, dermatitis atópica, eosinofilia y aumento de IgE. Se reporta el caso clínico de una niña de 1 mes y 29 días con diagnóstico de hiper-IgE con afección cutánea desde el nacimiento.
Asunto(s)
Dermatitis Atópica , Eccema , Síndrome de Job , Infecciones del Sistema Respiratorio , Indización y Redacción de Resúmenes , Dermatitis Atópica/diagnóstico , Femenino , Humanos , Síndrome de Job/diagnósticoAsunto(s)
Mucosa Nasal/fisiopatología , Rinitis/diagnóstico , Rinitis/fisiopatología , Adulto , Resistencia de las Vías Respiratorias , Estudios de Casos y Controles , Adaptabilidad , Femenino , Humanos , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Sensibilidad y Especificidad , Adulto JovenRESUMEN
OBJECTIVE: To define the natural history, clinical signs, treatment and the modalities of medium- and long-term follow-up of patients operated for sinus mucocele. PATIENTS AND METHOD: Retrospective study of all patients operated for sinus mucocele between January 1993 and December 2009 (n=68). Demographic data, symptoms, medical imaging findings, surgical treatment and results were recorded. RESULTS: The mean age of patients in this series was 53 years (range: 27-82 years, sex ratio: 3/2). The most common site was fronto-ethmoidal. Fifty-one patients (75%) had a history of sinus surgery, essentially for nasal polyposis. Only 15% of mucoceles occurred spontaneously. Presenting symptoms, in decreasing order of frequency, were facial pain or headache (38%), ocular or orbital complications (28%), while 20% of patients were asymptomatic. Surgery was performed by endonasal endoscopic sinus surgery (n=57, 84%) or via a combined, transfacial and endonasal approach, associated with navigation after January 2003. The mean follow-up was 7 years (range: 4 months-16 years). During this follow-up period, 23.5% of patients developed recurrence or a second mucocele after a mean interval of 4 years. CONCLUSION: This study demonstrates the high recurrence rate of mucocele, particularly in multi-operated patients with chronic sinusitis. Long-term, regular, clinical and radiological follow-up is necessary to detect asymptomatic lesions prior to the onset of complications.
Asunto(s)
Mucocele/cirugía , Enfermedades de los Senos Paranasales/cirugía , Complicaciones Posoperatorias/cirugía , Adulto , Anciano , Anciano de 80 o más Años , Endoscopía , Senos Etmoidales/patología , Senos Etmoidales/cirugía , Femenino , Estudios de Seguimiento , Seno Frontal/patología , Seno Frontal/cirugía , Humanos , Interpretación de Imagen Asistida por Computador , Hallazgos Incidentales , Imagen por Resonancia Magnética , Masculino , Persona de Mediana Edad , Mucocele/diagnóstico , Neuronavegación , Enfermedades de los Senos Paranasales/diagnóstico , Complicaciones Posoperatorias/diagnóstico , Recurrencia , Reoperación , Tomografía Computarizada por Rayos XRESUMEN
Glatiramer acetate (GA) is an immunomodulator approved for therapy of relapsing-remitting multiple sclerosis (RRMS), but recent findings indicate that it may also have additional, neurotrophic effects. Here, we found that supernatants from human GA-reactive T lymphocytes potentiated oligodendrocyte numbers in rodent and human oligodendrocyte progenitor (OPC) cultures. Effects of Th2-polarized lines were stronger than Th1-polarized cells. Microarray and ELISA analyses revealed that neurotrophic factors induced in Th2- and Th1-polarized GA-reactive lines included IGF-2 and BMP-7 respectively, and functional studies confirmed IGF-2 as trophic for OPCs. Our results support the concept that GA therapy may result in supportive effects on oligodendrocytes in RRMS patients.
Asunto(s)
Adyuvantes Inmunológicos/fisiología , Factor II del Crecimiento Similar a la Insulina/fisiología , Oligodendroglía/inmunología , Péptidos/fisiología , Células Madre/inmunología , Células Madre/fisiología , Células Th2/inmunología , Adyuvantes Inmunológicos/uso terapéutico , Animales , Línea Celular Tumoral , Linaje de la Célula/inmunología , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Acetato de Glatiramer , Humanos , Recuento de Linfocitos/métodos , Ratones , Esclerosis Múltiple Recurrente-Remitente/tratamiento farmacológico , Esclerosis Múltiple Recurrente-Remitente/inmunología , Esclerosis Múltiple Recurrente-Remitente/patología , Oligodendroglía/citología , Oligodendroglía/efectos de los fármacos , Péptidos/uso terapéutico , Células Madre/citología , Células Madre/efectos de los fármacos , Células Th2/citología , Células Th2/efectos de los fármacosRESUMEN
Several studies have suggested that a potential mechanism for estrogen-mediated neuroprotection following experimental stroke is a result of modulating glutamate-mediated excitotoxicity. Our laboratory has shown that in male rats, estrogen injection (systemic or direct intracortical injection) resulted in an immediate depolarization of cortical neurons. Therefore, the present study was designed to investigate whether the estrogen-induced depolarization of cortical neurons was required in mediating the early events associated with this neuroprotection. We tested this hypothesis by co-injecting selective antagonists of the NMDA (MK-801) or AMPA (DNQX) glutamatergic receptors with estrogen. Systemic injection of estrogen significantly attenuated the MK-801-induced decrease in infarct volume following middle cerebral artery occlusion (MCAO). Similarly, when estrogen and MK-801 were co-injected directly into the cortex, no neuroprotection was observed. However, when estrogen or MK-801 was injected centrally 10 min prior to the injection of the other drug, significant neuroprotection was observed. This led us to hypothesize that estrogen-mediated neuroprotection required an initial activation of NMDA receptors. Furthermore, our results suggest that this estrogen-mediated neuroprotection was also associated with a significant increase in m-calpain and activation of an endoplasmic reticulum (ER) specific caspase-12. Finally, the results of current clamp experiments showed that estrogen significantly depolarized cortical neurons as well as enhanced NMDA-induced depolarization. Taken together, these results suggest that estrogen pretreatment may activate NMDA receptors resulting in modification of ER-associated molecular mechanisms involved in neuroprotection following MCAO.
Asunto(s)
Corteza Cerebral/patología , Estrógenos/administración & dosificación , Infarto de la Arteria Cerebral Media/tratamiento farmacológico , Neuronas/patología , Fármacos Neuroprotectores/administración & dosificación , Receptores de N-Metil-D-Aspartato/fisiología , Análisis de Varianza , Animales , Presión Sanguínea/efectos de los fármacos , Calpaína/metabolismo , Caspasa 12/metabolismo , Maleato de Dizocilpina/farmacología , Interacciones Farmacológicas , Activación Enzimática/efectos de los fármacos , Agonistas de Aminoácidos Excitadores/farmacología , Frecuencia Cardíaca/efectos de los fármacos , Técnicas In Vitro , Infarto de la Arteria Cerebral Media/patología , Infarto de la Arteria Cerebral Media/fisiopatología , Masculino , Potenciales de la Membrana/efectos de los fármacos , Neuronas/efectos de los fármacos , Técnicas de Placa-Clamp/métodos , Quinoxalinas/farmacología , Ratas , Ratas Sprague-DawleyRESUMEN
Recently, adenosine has been proposed to be a "metabolic" switch that may sense and direct immune and inflammatory responses. Inflammation and pro-inflammatory cytokine production are important in development of HIV-1 associated dementia, a devastating consequence of HIV-1 infection of the CNS. The HIV-1 protein Tat induces cell death in the CNS and activates local inflammatory responses partially by inducing calcium release from the endoplasmic reticulum. Because activation of adenosine receptors decreases production of the pro-inflammatory cytokine TNF-alpha in several experimental paradigms both in vitro and in vivo, we hypothesized that adenosine receptor activation would control both increased intracellular calcium and TNF-alpha production induced by Tat. Treatment of primary monocytes with Tat significantly increased the levels of intracellular calcium released from IP3 stores. Activation of adenosine receptors with CGS 21680 inhibited Tat-induced increases of intracellular calcium by 90 +/- 8% and was dependent on protein phosphatase activity because okadaic acid blocked the actions of CGS 21680. Tat-induced TNF-alpha production was inhibited 90 +/- 6% by CGS 21680 and concurrent treatment with okadaic acid blocked the inhibitory actions of CGS 21680. Using a model monocytic cell line, CGS 21680 treatment increased cytosolic serine/threonine phosphatase. Together, these data indicate that A2A receptor activation increases protein phosphatase activity, which blocks IP3 receptor-regulated calcium release and reduction of intracellular calcium inhibits TNF-alpha production in monocytes.
Asunto(s)
Adenosina/análogos & derivados , Productos del Gen tat/metabolismo , VIH-1/patogenicidad , Inflamación/inmunología , Fosfoproteínas Fosfatasas/metabolismo , Receptor de Adenosina A2A/metabolismo , Transducción de Señal , Adenosina/farmacología , Calcio/metabolismo , Células Cultivadas , VIH-1/metabolismo , Humanos , Monocitos/inmunología , Fenetilaminas/farmacología , Factor de Necrosis Tumoral alfa/metabolismo , Células U937/inmunología , Productos del Gen tat del Virus de la Inmunodeficiencia HumanaRESUMEN
Herpesvirus infections can frequently lead to acute inflammation, yet the mechanisms regulating this event remain poorly understood. In order to determine some of the immunological mechanisms regulated by human herpesvirus infections, we studied the gene expression profile of lymphocytes infected with human herpesvirus 6 (HHV-6) by using a novel immunomicroarray. Our nylon-based immunomicroarray contained more than 1,150 immune response-related genes and was highly consistent between experiments. Experimentally, we found that independently of the HHV-6 strain used to infect T cells, multiple proinflammatory genes were increased and anti-inflammatory genes were decreased at the mRNA and protein levels. HHV-6 strains A and B increased expression of the genes for interleukin-18 (IL-18), the IL-2 receptor, members of the tumor necrosis factor alpha superfamily receptors, mitogen-activated protein kinase, and Janus kinase signaling proteins. As reported previously, CD4 protein levels were also increased significantly. Specific type 2 cytokines, including IL-10, its receptor, and IL-14, were downregulated by HHV-6 infection and, interestingly, amyloid precursor proteins and type 1 and 2 presenilins. Thus, T cells respond to HHV-6 infection by inducing a type 1 immune response that may play a significant role in the development and progression of diseases associated with HHV-6, including pediatric, hematologic, transplant, and neurologic disorders.
Asunto(s)
Perfilación de la Expresión Génica , Genes Virales , Herpesvirus Humano 6/genética , Linfocitos T/metabolismo , Secuencia de Bases , Antígenos CD4/genética , Línea Celular , Cartilla de ADN , Regulación hacia Abajo , Ensayo de Inmunoadsorción Enzimática , Humanos , Interleucina-18/genética , Análisis de Secuencia por Matrices de Oligonucleótidos , Reacción en Cadena de la Polimerasa , Linfocitos T/virología , Regulación hacia ArribaRESUMEN
The ubiquitous neuromodulator adenosine inhibits the production of several proinflammatory cytokines through activation of specific cell-surface adenosine receptors. We demonstrated recently that antisense oligonucleotides to tumor necrosis factor-alpha (TNF-alpha) are neuroprotective in a rat model of intracerebral hemorrhage. Therefore, we hypothesized that activation of adenosine receptors would provide protection against intracerebral hemorrhage-induced TNF-alpha production and inflammatory events. In vitro experiments showed that adenosine A1, A2A, and A3 receptor subtypes were present on U937 cells, and activation of these subtypes inhibited TNF-alpha production with a rank order of A2A > > A1 > A3. Prolonged treatment of U937 cells with the A2A receptor agonist 2-p-(carboxyethyl)phenethylamino-5'-N-ethylcarboxamidoadenosine hydrochloride (CGS 21680) desensitized adenosine A2A, A1, and A3 receptors. CGS 21680 administration directly into the striatum immediately prior to the induction of intracerebral hemorrhage inhibited TNF-alpha mRNA and, 24 hours following induction, reduced parenchymal neutrophil infiltration (p < 0.001) and TUNEL-positive cells (p < 0.002) within and bordering the hematoma. These results suggest that pharmacological strategies targeting A2A receptors may provide effective inhibition of acute neurotoxic proinflammatory events that occur following intracerebral hemorrhage.
Asunto(s)
Adenosina/análogos & derivados , Apoptosis , Hemorragia Cerebral/patología , Agonistas del Receptor Purinérgico P1 , Receptores Purinérgicos P1/metabolismo , Adenosina/administración & dosificación , Adenosina/farmacología , Adenosina/uso terapéutico , Adenilil Ciclasas/metabolismo , Animales , Apoptosis/efectos de los fármacos , Hemorragia Cerebral/tratamiento farmacológico , Hemorragia Cerebral/enzimología , Hemorragia Cerebral/metabolismo , Quimiotaxis de Leucocito/efectos de los fármacos , Humanos , Etiquetado Corte-Fin in Situ , Inflamación/tratamiento farmacológico , Inflamación/enzimología , Inflamación/metabolismo , Inflamación/patología , Inyecciones Intraventriculares , Masculino , Neutrófilos/citología , Neutrófilos/efectos de los fármacos , Neutrófilos/inmunología , Fenetilaminas/administración & dosificación , Fenetilaminas/farmacología , Fenetilaminas/uso terapéutico , Fitohemaglutininas/farmacología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas , Ratas Sprague-Dawley , Receptor de Adenosina A2A , Receptor de Adenosina A3 , Acetato de Tetradecanoilforbol/farmacología , Factor de Necrosis Tumoral alfa/biosíntesis , Factor de Necrosis Tumoral alfa/genética , Células U937RESUMEN
BACKGROUND AND PURPOSE: Tumor necrosis factor-alpha (TNF-alpha) expression is increased in brain after cerebral ischemia, although little is known about its abundance and role in intracerebral hemorrhage (ICH). A TNF-alpha-specific antisense oligodeoxynucleotide (ORF4-PE) was used to study the extent to which TNF-alpha expression influenced neurobehavioral outcomes and brain damage in a collagenase-induced ICH model in rat. METHODS: Male Sprague-Dawley rats were anesthetized, and ICH was induced by intrastriatal administration of heparin and collagenase. Immediately before or 3 hours after ICH induction, ORF4-PE was administered directly into the site of ICH. TNF-alpha mRNA and protein levels were measured by reverse transcriptase-polymerase chain reaction and immunoblot analyses. Cell death was measured by terminal deoxynucleotidyl transferase-mediated uridine 5'triphosphate-biotin nick end labeling (TUNEL). Neurobehavioral deficits were measured for 4 weeks after ICH. RESULTS: ICH induction (n=6) elevated TNF-alpha mRNA and protein levels (P:<0.01) at 24 hours after the onset of injury compared with sham controls (n=6). Immunohistochemical labeling indicated that ICH was accompanied by elevated expression of TNF-alpha in neutrophils, macrophages, and microglia. Administration of ORF4-PE (2.0 nmol) directly into striatal parenchyma, 15 minutes before (n=4) or 3 hours after (n=6) ICH, decreased levels of TNF-alpha mRNA (P:<0.001) and protein (P:<0. 01) in the brain tissue surrounding the hematoma compared with animals treated with saline alone (n=6). Mean+/-SEM striatal cell death (cells per high-powered field) was also reduced in animals receiving ORF4-PE (34.1+/-5.0) compared with the saline-treated ICH group (80.3+/-7.50) (P:<0.001). ORF4-PE treatment improved neurobehavioral deficits observed at 24 hours (P:<0.001) after induction of ICH (n=6) compared with the untreated ICH group (n=6). This improvement was maintained at 28 days after hemorrhage induction (P:<0.001). CONCLUSIONS: These results indicate a pathogenic role for TNF-alpha during ICH and demonstrate that reducing TNF-alpha expression using antisense oligodeoxynucleotides is neuroprotective.
Asunto(s)
Hemorragia Cerebral/tratamiento farmacológico , Fármacos Neuroprotectores/administración & dosificación , Oligonucleótidos Antisentido/administración & dosificación , Factor de Necrosis Tumoral alfa/antagonistas & inhibidores , Animales , Apoptosis/efectos de los fármacos , Conducta Animal/efectos de los fármacos , Hemorragia Cerebral/inducido químicamente , Hemorragia Cerebral/patología , Colagenasas , Cuerpo Estriado/efectos de los fármacos , Cuerpo Estriado/patología , Modelos Animales de Enfermedad , Encefalitis/inmunología , Encefalitis/patología , Heparina , Inmunohistoquímica , Etiquetado Corte-Fin in Situ , Imagen por Resonancia Magnética , Masculino , Microinyecciones , Infiltración Neutrófila/efectos de los fármacos , Infiltración Neutrófila/inmunología , ARN Mensajero/biosíntesis , Ratas , Ratas Sprague-Dawley , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Índice de Severidad de la Enfermedad , Factor de Necrosis Tumoral alfa/biosíntesis , Factor de Necrosis Tumoral alfa/genéticaRESUMEN
Infection of the brain by lentiviruses, including human immunodeficiency virus (HIV) and feline immunodeficiency virus (FIV), causes inflammation and results in neurodegeneration. Molecular diversity within the lentivirus envelope gene has been implicated in the regulation of cell tropism and the host response to infection. Here, we examine the hypothesis that envelope sequence diversity modulates the expression of host molecules implicated in lentivirus-induced brain disease, including matrix metalloproteinases (MMP) and related transcription factors. Infection of primary macrophages by chimeric HIV clones containing brain-derived envelope fragments from patients with HIV-associated dementia (HAD) or nondemented AIDS patients (HIV-ND) showed that MMP-2 and -9 levels in conditioned media were significantly higher for the HAD clones. Similarly, STAT-1 and JAK-1 levels were higher in macrophages infected by HAD clones. Infections of primary feline macrophages by the neurovirulent FIV strain (V(1)CSF), the less neurovirulent strain (Petaluma), and a chimera containing the V(1)CSF envelope in a Petaluma background (FIV-Ch) revealed that MMP-2 and -9 levels were significantly higher in conditioned media from V(1)CSF- and FIV-Ch-infected macrophages, which was associated with increased intracellular STAT-1 and JAK-1 levels. The STAT-1 inhibitor fludarabine significantly reduced MMP-2 expression, but not MMP-9 expression, in FIV-infected macrophages. Analysis of MMP mRNA and protein levels in brain samples from HIV-infected persons or FIV-infected cats showed that MMP-2 and -9 levels were significantly increased in lentivirus-infected brains compared to those of uninfected controls. Elevated MMP expression was accompanied by significant increases in STAT-1 and JAK-1 mRNA and protein levels in the same brain samples. The present findings indicate that two lentiviruses, HIV and FIV, have common mechanisms of MMP-2 and -9 induction, which is modulated in part by envelope sequence diversity and the STAT-1/JAK-1 signaling pathway.
Asunto(s)
Encéfalo/virología , Genes env/genética , Variación Genética , Infecciones por Lentivirus/enzimología , Lentivirus/genética , Metaloproteinasas de la Matriz/metabolismo , Complejo SIDA Demencia/enzimología , Complejo SIDA Demencia/virología , Animales , Encéfalo/enzimología , Gatos , Células Cultivadas , Proteínas de Unión al ADN/metabolismo , Regulación de la Expresión Génica , VIH/genética , VIH/metabolismo , VIH/fisiología , Infecciones por VIH/enzimología , Infecciones por VIH/virología , Humanos , Virus de la Inmunodeficiencia Felina/genética , Virus de la Inmunodeficiencia Felina/metabolismo , Virus de la Inmunodeficiencia Felina/fisiología , Lentivirus/metabolismo , Lentivirus/fisiología , Infecciones por Lentivirus/virología , Macrófagos/enzimología , Macrófagos/virología , Proteínas Tirosina Quinasas/metabolismo , Proteínas Recombinantes de Fusión/metabolismo , Factor de Transcripción STAT1 , Transducción de Señal , Transactivadores/metabolismo , Proteínas del Envoltorio Viral/metabolismo , Replicación ViralRESUMEN
HIV-1 protein Tat is neurotoxic and increases macrophage and microglia production of TNF-alpha, a cytopathic cytokine linked to the neuropathogenesis of HIV dementia. Others have shown that intracellular calcium regulates TNF-alpha production in macrophages, and we have shown that Tat releases calcium from inositol 1,4, 5-trisphosphate (IP3) receptor-regulated stores in neurons and astrocytes. Accordingly, we tested the hypothesis that Tat-induced TNF-alpha production was dependent on the release of intracellular calcium from IP3-regulated calcium stores in primary macrophages. We found that Tat transiently and dose-dependently increased levels of intracellular calcium and that this increase was blocked by xestospongin C, pertussis toxin, and by phospholipase C and type 1 protein kinase C inhibitors but not by protein kinase A or phospholipase A2 inhibitors. Xestospongin C, BAPTA-AM, U73122, and bisindolylmalemide significantly inhibited Tat-induced TNF-alpha production. These results demonstrate that in macrophages, Tat-induced release of calcium from IP3-sensitive intracellular stores and activation of nonconventional PKC isoforms play an important role in Tat-induced TNF-alpha production.
Asunto(s)
Canales de Calcio/fisiología , Calcio/metabolismo , Productos del Gen tat/fisiología , VIH-1/fisiología , Macrófagos/metabolismo , Receptores Citoplasmáticos y Nucleares/fisiología , Factor de Necrosis Tumoral alfa/biosíntesis , Señalización del Calcio/inmunología , Células Cultivadas , Relación Dosis-Respuesta Inmunológica , Estrenos/farmacología , Humanos , Inositol 1,4,5-Trifosfato/metabolismo , Receptores de Inositol 1,4,5-Trifosfato , Líquido Intracelular/metabolismo , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Toxina del Pertussis , Pirrolidinonas/farmacología , Factores de Tiempo , Factor de Necrosis Tumoral alfa/antagonistas & inhibidores , Fosfolipasas de Tipo C/fisiología , Factores de Virulencia de Bordetella/farmacología , Productos del Gen tat del Virus de la Inmunodeficiencia HumanaRESUMEN
Many cases of early-onset inherited Alzheimer's disease (AD) are caused by mutations in the presenilin-1 (PS1) gene. PS1 mutations may perturb cellular Ca(2+) homeostasis and thereby render neurons vulnerable to excitotoxicity and apoptosis. We now report that PC12 cells expressing PS1 mutations and primary hippocampal neurons from PS1 mutant knockin mice exhibit greatly increased levels of ryanodine receptors (RyR) and enhanced Ca(2+) release following stimulation with caffeine. Double-labeling immunostaining and co-immunoprecipitation analyses indicate that PS1 and RyR are colocalized and interact physically. Caffeine treatment sensitizes neurons expressing mutant PS1 to apoptosis induced by amyloid beta-peptide, a neurotic peptide linked to the pathogenesis of AD. When taken together with recent evidence for alterations in RyR in brains of AD patients, our data suggest that PS1 mutations may promote neuronal degeneration in AD by increasing transcription and translation of RyR and altering functional properties of ryanodine-sensitive Ca(2+) pools.
Asunto(s)
Calcio/metabolismo , Corteza Cerebral/citología , Proteínas de la Membrana/genética , Neuronas/metabolismo , Canal Liberador de Calcio Receptor de Rianodina/metabolismo , Animales , Cafeína/farmacología , Células Cultivadas , Hipocampo/citología , Homeostasis/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Mutación , Células PC12 , Presenilina-1 , Ratas , TransfecciónRESUMEN
Following decapitation, during the brief time period of postmortem brain tissue collection, significant changes in neuro-metabolite levels can occur. To circumvent such changes, we routinely kill rats using 1 to 2 sec pulses of focused high-energy microwave irradiation (10 kW). The effects of high-energy microwave irradiation on total RNA and mRNA integrity in brain however, are unknown. Total RNA recovery, per gram wet weight, in brain regions of microwaved rats was less than 50% of that in rats killed by decapitation. Formaldehyde agarose gel electrophoresis showed that ribosomal RNA components were highly degraded in all brain regions of microwaved rats. In contrast, poly A+mRNA, as measured by poly A+driven cDNA synthesis and Northern analysis, in brain samples of microwaved or decapitated rats was of equal integrity and quantity when expressed per mg tissue weight. Furthermore, positive RT-PCR products for GAPDH and TNF-alpha were observed in brain regions of both microwaved and decapitated rats. These observations indicated that high-energy focused microwave irradiation does not reduce mRNA abundance and integrity. Thus, this method of animal sacrifice can be used to simultaneously study, accurately and precisely, levels of brain metabolites as well as molecular biological events in discrete brain regions of experimental animals without postmortem interference.
Asunto(s)
Química Encefálica/genética , Microondas , ARN Mensajero/aislamiento & purificación , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Adenosina/aislamiento & purificación , Animales , Northern Blotting , Cromatografía Líquida de Alta Presión , Gliceraldehído-3-Fosfato Deshidrogenasas/genética , Masculino , Cambios Post Mortem , ARN Mensajero/análisis , Ratas , Ratas Sprague-Dawley , Factor de Necrosis Tumoral alfa/genéticaRESUMEN
ABSTRACT Exon sequences upstream of splice sites play a critical role in mRNA processing, which is dependent on spliceosome interactions with these sites. Using antisense oligodeoxynucleotides (ODN), we targeted these and other sequences of the proinflammatory tumor necrosis factor-alpha (TNF-alpha) gene because it is multiply spliced and has been difficult to regulate with ODN in the past. ODN targeting exon sequences upstream of the donor splice sites of internal exons 2 (ORF4) and 3 (ORF6) significantly reduced TNF-alpha levels in stimulated U937 cells by 62%+/-7% and 51%+/-9%, respectively, in a dose-dependent manner but did not affect interleukin-6 (IL-6) levels. In contrast, ODN targeting the exon sequences downstream of the acceptor splice sites of exons 1, 2, and 3 failed to reduce TNF-alpha levels significantly under the same conditions. End-phosphorothioated ORF4 (ORF4-PE) significantly reduced TNF-alpha mRNA levels by greater than 80% (p < 0.001) and protein levels by 60% (p < 0.001) in U937 cells. ORF4-PE reduced newly synthesized TNF-alpha protein levels by >80% in lipopolysaccharide (LPS)-stimulated human macrophages, by greater than 60% in phorbol myristate acetate/phyto-hemagglutinin (PMA/PHA)-stimulated human peripheral blood mononuclear cells (PBMC), and by approximately 50% in LPS-stimulated murine monocytes. These results suggest that exon sequences flanking donor splice sites are highly susceptible target domains for antisense inhibition of TNF-alpha gene expression.
Asunto(s)
Oligodesoxirribonucleótidos Antisentido/farmacología , Factor de Necrosis Tumoral alfa/biosíntesis , Animales , Células Cultivadas , Relación Dosis-Respuesta a Droga , Diseño de Fármacos , Exones , Expresión Génica/efectos de los fármacos , Humanos , Leucocitos Mononucleares/citología , Leucocitos Mononucleares/efectos de los fármacos , Macrófagos/citología , Macrófagos/efectos de los fármacos , Ratones , Monocitos/citología , Monocitos/efectos de los fármacos , Empalme del ARN , Transducción de Señal , Factor de Necrosis Tumoral alfa/genética , Células U937/citología , Células U937/efectos de los fármacosRESUMEN
Cytokines, including tumor necrosis factor-alpha (TNF alpha) and interleukin-6 (IL-6), have been implicated in the pathogenesis of multiple sclerosis (MS). The production and release of these cytokines are regulated in part by specific purinergic (adenosine) cell surface receptors. To determine the extent to which the adenosine A1 receptor influenced cytokine expression in peripheral blood mononuclear cells (PBMCs) from MS and control patients, we measured plasma adenosine and TNF alpha levels, A1 receptor messenger RNA (mRNA) and protein amounts, and the effects of activation of A1 receptors on TNF alpha and IL-6 production by PBMCs. Plasma levels of TNF alpha were significantly higher and adenosine levels were significantly lower in MS patients compared with control subjects. Levels of TNF alpha and IL-6 in mitogen-stimulated PBMC culture supernatants from MS patients or control patients were similar. Conversely, treatment of PBMCs with the adenosine A1 receptor agonist R-phenylisopropyladenosine (R-PIA) (1 microM) significantly inhibited mitogen-stimulated production of TNF alpha but not IL-6 in control subjects and significantly inhibited production of IL-6 but not TNF alpha in MS patients. The effects of R-PIA were selectively blocked by the A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX). A1 receptor protein levels were decreased significantly in PBMCs from MS patients. Taken together, these results suggest that decreased levels of adenosine and its A1 receptor modulate TNF alpha and IL-6 levels and may contribute to the pathogenesis of MS.
Asunto(s)
Monocitos/metabolismo , Esclerosis Múltiple/sangre , Receptores Purinérgicos P1/metabolismo , Adenosina/sangre , Adulto , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Masculino , Persona de Mediana Edad , Receptores Purinérgicos P1/sangre , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Human immunodeficiency virus type 1 (HIV-1) infection of the brain results in viral replication primarily in macrophages and microglia. Despite frequent detection of viral genome and proteins in the brains of AIDS patients with and without HIV dementia, only 20% of AIDS patients become demented. To investigate the role of viral envelope gene variation in the occurrence of dementia, we examined regions of variability in the viral envelope gene isolated from brains of AIDS patients. Brain-derived HIV-1 V1-V2 envelope sequences from seven demented and six nondemented AIDS patients displayed significant sequence differences between clinical groups, and by phylogenetic analysis, sequences from the demented group showed clustering. Infectious recombinant viruses containing brain-derived V3 sequences from both clinical groups were macrophagetropic, and viruses containing brain-derived V1, V2, and V3 sequences from both clinical groups spread efficiently in macrophages. In an indirect in vitro neurotoxicity assay using supernatant fluid from HIV-1-infected macrophages, recombinant viruses from demented patients induced greater neuronal death than viruses from nondemented patients. Thus, the HIV-1 envelope diversity observed in these patient groups appeared to influence the release of neurotoxic molecules from macrophages and might account in part for the variability in occurrence of dementia in AIDS patients.
Asunto(s)
Complejo SIDA Demencia/patología , Complejo SIDA Demencia/virología , Encéfalo/patología , Encéfalo/virología , Genes env , VIH-1/genética , VIH-1/patogenicidad , Neuronas/patología , Complejo SIDA Demencia/etiología , Secuencia de Aminoácidos , Muerte Celular , Células Cultivadas , Productos del Gen env/genética , Variación Genética , VIH-1/aislamiento & purificación , Células HeLa , Humanos , Macrófagos/metabolismo , Macrófagos/virología , Datos de Secuencia Molecular , Neurotoxinas/metabolismo , Especificidad de Órganos , Filogenia , Homología de Secuencia de AminoácidoRESUMEN
Activation and infection by HIV-1 of glial cells and infiltrating macrophages are cardinal features of AIDS-related neurological disease. Tumor necrosis factor-alpha (TNF-alpha) is released by these cell types, and increased TNF-alpha mRNA and protein levels are associated with the development and severity of HIV-induced neurological disease. HIV-1 proteins have been implicated in HIV neuropathogenesis including Tat which has been shown to be a potent inducer of TNF-alpha. We review our data showing the induction of TNF-alpha by Tat in primary human fetal astrocytes, human peripheral blood mononuclear cells, macrophages, and astrocytic and macrophage cell lines. TNF-alpha induction was NF-kappaB dependent and was eliminated by inhibiting protein kinase A, phospholipase C and protein tyrosine kinase activity. In addition, we examined the molecular diversity of the tat genome in the brains of HIV-infected patients from different HIV-1 clades. Comparison of matched brain- and spleen-derived tat sequences indicated that homology among brain-derived clones was greater than that between the brain- and spleen-derived clones. The brain-derived tat sequences were markedly heterogeneous in regions which influence viral replication and intracellular transport. Future studies using Tat, encoded by different sequences, will be necessary to determine the functional significance of tat molecular diversity. Nonetheless, these studies suggest that Tat is an important inducer of TNF-alpha production and thus may play a key role in the pathogenesis of HIV-related neurological disease.
Asunto(s)
Complejo SIDA Demencia/inmunología , Productos del Gen tat/genética , Variación Genética , VIH-1/genética , Factor de Necrosis Tumoral alfa/genética , Astrocitoma , Relación Dosis-Respuesta Inmunológica , Regulación Viral de la Expresión Génica , Productos del Gen tat/farmacología , VIH-1/inmunología , Humanos , Células Jurkat/virología , Datos de Secuencia Molecular , FN-kappa B/genética , FN-kappa B/inmunología , ARN Mensajero/análisis , Homología de Secuencia de Aminoácido , Factor de Necrosis Tumoral alfa/inmunología , Células U937/virología , Proteínas Virales/genética , Proteínas Virales/inmunología , Productos del Gen tat del Virus de la Inmunodeficiencia HumanaRESUMEN
Several studies have suggested an association between human herpesvirus 6 (HHV-6) infection and multiple sclerosis. As HHV-6 is predominantly a T-cell tropic virus, we examined the frequency of detection of HHV-6 genome in peripheral blood mononuclear cells from relapsing-remitting (n = 32) and chronic progressive (n = 14) patients and from healthy (n = 17) and neurological (n = 7) controls. Two sensitive polymerase chain reaction assays were used to target different regions within the HHV-6 genome. Depending on the polymerase chain reaction assay used, the detection of HHV-6 genome ranged from 11.7 to 23.5% (controls), 3.1 to 23.0% (relapsing-remitting), and 14.2 to 28.5% (chronic progressive). Although these observations do not exclude a pathogenic role for HHV-6 in multiple sclerosis, they indicate a lack of correlation between HHV-6 infection of peripheral blood mononuclear cells and the development of multiple sclerosis.