RESUMO
The function of the proteasome has been linked to various pathologies, including cancer and neurodegeneration. Proteasomal inhibition can lead to death in a variety of cell types, however the manner in which this occurs is unclear, and may depend on the particular cell type. In this work we have extended previous findings pertaining to the effects of pharmacological proteasomal inhibitors on PC12 cells, by examining in more detail the induced death pathway. We find that cell death is apoptotic by ultrastructural criteria. Caspase 9 and 3 are processed, cytochrome c is released from the mitochondria and a dominant negative form of caspase 9 prevents death. Furthermore, Bax undergoes a conformational change and is translocated to the mitochondria in a caspase-independent fashion. Total cell levels of Bax however do not change, whereas levels of the BH3-only protein Bim increase with proteasomal inhibition. Transient overexpression of bcl-xL or, to a lesser extent, of bcl-2, significantly decreased apoptotic death and prevented Bax conformational change. We conclude that death elicited by proteasomal inhibition of PC12 cells follows a classical "intrinsic" pathway. Significantly, antiapoptotic bcl-2 family members prevent apoptosis by inhibiting Bax conformational change. Increased levels of Bim may contribute to cell death in this model.
Assuntos
Apoptose , Inibidores de Proteassoma , Proteína X Associada a bcl-2/química , Proteína X Associada a bcl-2/metabolismo , Proteína bcl-X/metabolismo , Acetilcisteína/análogos & derivados , Acetilcisteína/farmacologia , Animais , Apoptose/efeitos dos fármacos , Proteínas Reguladoras de Apoptose/metabolismo , Proteína 11 Semelhante a Bcl-2 , Caspase 9/metabolismo , Citocromos c/metabolismo , Expressão Gênica/efeitos dos fármacos , Genes Dominantes , Proteínas de Membrana/metabolismo , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/enzimologia , Mitocôndrias/metabolismo , Células PC12 , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Estrutura Quaternária de Proteína/efeitos dos fármacos , Transporte Proteico/efeitos dos fármacos , Proteínas Proto-Oncogênicas/metabolismo , Ratos , Proteína bcl-X/genéticaRESUMO
Ubiquitinated inclusions and selective neuronal cell death are considered the pathological hallmarks of Parkinson's disease and other neurodegenerative diseases. Recent genetic, pathological and biochemical evidence suggests that dysfunction of ubiquitin-dependent protein degradation by the proteasome might be a contributing, if not initiating factor in the pathogenesis of these diseases. In neuronal cell culture models inhibition of the proteasome leads to cell death and formation of fibrillar ubiquitin and alpha-synuclein-positive inclusions, thus modeling some aspects of Lewy body diseases. The processes of inclusion formation and neuronal cell death share some common mechanisms, but can also be dissociated at a certain level.
Assuntos
Corpos de Inclusão/fisiologia , Neurônios/fisiologia , Ubiquinona/metabolismo , Doença de Alzheimer/patologia , Animais , Morte Celular/genética , Morte Celular/fisiologia , Humanos , Corpos de Inclusão/genética , Corpos de Inclusão/metabolismo , Proteínas do Tecido Nervoso/biossíntese , Proteínas do Tecido Nervoso/genética , Doenças Neurodegenerativas/patologia , Neurônios/patologia , Doença de Parkinson/patologia , Sinucleínas , alfa-SinucleínaRESUMO
Adrenomedullin is a biologically active peptide released from the vascular wall, which increases blood flow through its vasorelaxant effects and prevents platelet activation by stimulation of nitric oxide synthesis. The present study demonstrates that activated platelets suppress adrenomedullin secretion from vascular endothelial cells by releasing a factor that was identified as transforming growth factor (TGF)-beta1. Adrenomedullin levels were reduced by up to 40% and this effect was completely abrogated by the addition of latency-associated protein (LAP) or TGF-beta1-neutralizing antibody. Inhibition of adrenomedullin secretion in response to platelet aggregation may be an important mechanism in the induction of hemostasis.
