RESUMEN
Endothelin-1 (ET-1), a vasoactive and mitogenic peptide mainly produced by vascular endothelial cells, may be involved in the progression of several human tumors. Here, we present an immunohistochemical analysis of the expression pattern of ET-1 receptor subtypes (ET(A)-R and ET(B)-R) and a functional study of their potential role in human oligodendrogliomas and oligoastrocytomas. By comparison, we assessed the corresponding expression patterns of glioblastomas. Interestingly, a nuclear localization of ET-1 receptor subtypes (associated or not with a cytoplasmic labeling) was constantly observed in tumor cells from all three glioma types. Moreover, we noted a distinct receptor distribution in the different gliomas: a nuclear expression of ET(B)-R by tumor cells was found to be restricted to oligodendrogliomas and oligoastrocytomas, while a nuclear expression of ET(A)-R was only detected in tumor cells from some glioblastomas. Using primary cultures of oligodendroglial tumor cells, we confirmed the selective expression of nuclear ET(B)-R, together with a plasma membrane expression, and further demonstrated that this receptor was functionally coupled to intracellular signaling pathways known to be involved in cell survival and/or proliferation: extracellular signal-regulated kinase and focal adhesion kinase activation, actin cytoskeleton reorganization. In addition, impairment of ET(B)-R activation in these cells by in vitro treatment with an ET(B)-R-specific antagonist induced cell death. These data point to ET-1 as a possible survival factor for oligodendrogliomas via ET(B)-R activation and suggest that ET(B)-R-specific antagonists might constitute a potential therapeutic alternative for oligodendrogliomas.
Asunto(s)
Neoplasias Encefálicas/metabolismo , Endotelina-1/metabolismo , Oligodendroglioma/metabolismo , Receptor de Endotelina A/metabolismo , Receptor de Endotelina B/metabolismo , Citoesqueleto de Actina/metabolismo , Antihipertensivos/farmacología , Antihipertensivos/uso terapéutico , Astrocitoma/tratamiento farmacológico , Astrocitoma/metabolismo , Neoplasias Encefálicas/tratamiento farmacológico , Membrana Celular/metabolismo , Núcleo Celular/metabolismo , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Citoplasma/metabolismo , Antagonistas de los Receptores de la Endotelina B , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Quinasa 1 de Adhesión Focal , Proteína-Tirosina Quinasas de Adhesión Focal , Glioblastoma/tratamiento farmacológico , Glioblastoma/metabolismo , Humanos , Inmunohistoquímica , Oligodendroglioma/tratamiento farmacológico , Oligopéptidos/farmacología , Oligopéptidos/uso terapéutico , Piperidinas/farmacología , Piperidinas/uso terapéutico , Proteínas Tirosina Quinasas/metabolismo , Células Tumorales CultivadasRESUMEN
mRNA, Western analysis and immunohistochemistry were used to study the expression of the small heat shock protein (HSP) 27 in the rat septum and hippocampus following fimbria-fornix lesions, a model of neurodegeneration and regeneration. (HSP) 27 mRNA level was increased 2.5-fold in the medial septum 3 days after lesion and this increase persisted for 10 days. In the hippocampus, after an initial 15-fold increase at 3 days post-injury, HSP27 mRNA returned to basal levels 10 days after the lesion. Three and 10 days after lesion, HSP27 protein levels were increased in the septum (4.5 and 5-fold, respectively) and hippocampus (65 and 10-fold, respectively). The morphology of the HSP27 positive cells was indistinguishable from that of GFAP-immunoreactive cells. In addition, in the septum of injured rats, occasional neurons were heavily labelled with anti-HSP27 antibodies. Thus, up-regulation of HSP27, particularly in glial cells, may be a component of glial input in the processes on degeneration/regeneration which occur in this model.
Asunto(s)
Fórnix/patología , Proteínas de Choque Térmico/metabolismo , Hipocampo/metabolismo , Proteínas de Neoplasias/metabolismo , Tabique del Cerebro/metabolismo , Animales , Western Blotting , Regulación de la Expresión Génica , Proteínas de Choque Térmico HSP27 , Proteínas de Choque Térmico/genética , Hipocampo/química , Inmunohistoquímica , Masculino , Proteínas de Neoplasias/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas , Ratas Sprague-Dawley , Tabique del Cerebro/químicaRESUMEN
Changes in gene expression within the hippocampus induced by denervation after electrolytic fimbria-fornix lesion in rat were compared to morphological and biochemical alterations. Fimbria-fornix lesion results in degeneration of hippocampal cholinergic terminals as evidenced by a sustained (2 days to 1 month) decrease in cholineacetyltransferase (ChAT) activity (50%). These changes were accompanied by a decrease in growth associated protein 43 (GAP-43) immunoreactivity in all hippocampal layers 4 days after lesion followed by a subsequent increase and return to normal levels by 20 days postinjury. This increase in GAP-43 expression in the hippocampus between 7 to 20 days after lesion may reflect heterotypic sprouting. TUNEL-positive cells were revealed by in situ assay within the hippocampus at 10 days, but not at 3 days, after lesion. Two subtracted cDNA libraries from the dorsal hippocampus of control and injured rats (at 3 and 10 days postlesion) were constructed in order to search for new genes potentially implicated in degeneration/regeneration phenomena. We analysed 1,536 clones from each library by differential screening and found a total of 46 up-regulated genes. Among the 15 known genes, 6 coded for proteins involved in signal transduction pathways. The upregulation of growth arrest DNA damage induced gene (GADD153), brain-specific RING finger protein, JNK interacting protein (JIP-1), protein kinase A (PKA), and Na+K+ ATPase was studied by quantitative polymerase chain reaction (PCR). Two of these genes, GADD153 and JIP-1, have been previously shown to participate in cell modifications induced by stress and apoptosis.
Asunto(s)
Apoptosis/genética , Hipocampo/patología , Degeneración Nerviosa/genética , Animales , Unión Competitiva/genética , Colina O-Acetiltransferasa/metabolismo , Fragmentación del ADN , Cartilla de ADN , Desnervación , Modelos Animales de Enfermedad , Fórnix/enzimología , Fórnix/patología , Proteína GAP-43/análisis , Proteína GAP-43/inmunología , Expresión Génica/fisiología , Biblioteca de Genes , Pruebas Genéticas , Hipocampo/enzimología , Técnicas para Inmunoenzimas , Etiquetado Corte-Fin in Situ , Masculino , Degeneración Nerviosa/enzimología , Plasticidad Neuronal/genética , Hibridación de Ácido Nucleico/métodos , Reacción en Cadena de la Polimerasa/métodos , Ratas , Ratas Sprague-DawleyRESUMEN
Opioid peptides, deriving from prodynorphin, proenkephalin and proopiomelanocortin genes, have been shown to modulate brain development. Prodynorphin gene expression was studied here by in situ hybridization in the developing rat hypothalamus using oligodeoxynucleotide probes. Prodynorphin mRNA-synthetizing cells were observed in the ventromedial hypothalamic nucleus, the supraoptic and the paraventricular nuclei from embryonic days 16, 18 and 21, respectively. We detected no transient expression of prodynorphin gene in the rat hypothalamus. Prodynorphin mRNA-containing cells were also observed prenatally in the striatum, the cortex, the hippocampus and the amygdala. When compared with data from the literature, our results suggest that translation may immediately follow transcription of prodynorphin gene in the supraoptic nucleus. The presence of prodynorphin mRNA in the developing rat hypothalamus also raises the possibility of an involvement of prodynorphin-derived peptides in developmental processes and/or in the maturation of adult neural regulations.