RESUMO
The CD53 antigen is a member of the tetraspanin membrane protein family that is expressed in the lymphoid-myeloid lineage. Its biological role remains unknown. Using microarrays, we identified CD53 as one of the principal genes up-regulated by exposure of macrophages to LPS. Northern blot analysis confirmed the induction of CD53 in RAW264.7 macrophages treated with LPS or SNAP (a nitric oxide donor). Cells stably transfected with sense CD53 cDNA had increased levels of intracellular GSH and lower levels of peroxide, and were more resistant to H2O2 and to UVB irradiation. Cells harboring antisense CD53 had the opposite properties. We propose that the induction of CD53 is a major mechanism by which macrophages protect themselves against LPS-induced oxidative stress and UVB irradiation.
Assuntos
Antígenos CD/metabolismo , Antígenos de Diferenciação de Linfócitos T/metabolismo , Lipopolissacarídeos/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/efeitos da radiação , Estresse Oxidativo , Penicilamina/análogos & derivados , Animais , Transporte Biológico , Northern Blotting , Western Blotting , Linhagem Celular , Sobrevivência Celular , DNA Complementar/metabolismo , Citometria de Fluxo , Glutationa/metabolismo , Peróxido de Hidrogênio/farmacologia , Macrófagos/metabolismo , Camundongos , Doadores de Óxido Nítrico/farmacologia , Análise de Sequência com Séries de Oligonucleotídeos , Penicilamina/farmacologia , Reação em Cadeia da Polimerase , RNA/metabolismo , Espécies Reativas de Oxigênio , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Tetraspanina 25 , Fatores de Tempo , Transfecção , Raios Ultravioleta , Regulação para Cima , gama-Glutamiltransferase/metabolismoRESUMO
Nitric oxide (NO) is involved in many physiological processes and also causes pathological effects by inducing apoptosis. It can enhance or suppress apoptosis depending on its concentration and the cell type involved. In this report, we used cDNA microarray analysis to show that SNAP, an NO donor, strongly induces Bcl-2/adenovirus E1B 19kDa-interacting protein 3 (BNIP3) in macrophages. BNIP3 is a mitochondrial pro-apoptotic protein that contains a Bcl-2 homology 3 domain and a COOH-terminal transmembrane (TM) domain. Macrophages activated by LPS/IFN-gamma produce nitric oxide synthase 2 (NOS2) and release endogenous NO. Expression of BNIP3 was also induced in macrophages by LPS/IFN-gamma, and the induction was blocked by a NOS2 inhibitor, S-methyl-isothiourea. Peritoneal macrophages from NOS2-null mice failed to produce BNIP3 in response to LPS/IFN-gamma. We conclude that BNIP3 expression in macrophages is controlled by the intracellular level of nitric oxide. Overexpression of BNIP3 but not of BNIP3 deltaTM, a BNIP3 mutant without the TM domain and C-terminal tail, led to apoptosis of the cells. Promoter analysis showed that the region between -281 and -1 of the 5'-upstream enhancer region of murine BNIP3 was sufficient for NO-dependent expression of BNIP3.