Peroxynitrite mediates TNF-α-induced endothelial barrier dysfunction and nitration of actin

American Journal of Physiology - Lung Cellular and Molecular Physiology - Tập 290 Số 4 - Trang L674-L684 - 2006
Paul Neumann1, Nancy Gertzberg, Erin E Vaughan, Joshua Weisbrot, Renee Woodburn, W. Marcus Lambert, Arnold Johnson
1151, 113 Holland Ave., Dept. of Veterans Affairs Medical Center, Albany, NY 12208, USA.

Tóm tắt

We tested the hypothesis that tumor necrosis factor (TNF)-α induces a peroxynitrite (ONOO−)-dependent increase in permeability of pulmonary microvessel endothelial monolayers (PMEM) that is associated with generation of nitrated β-actin (NO2-β-actin). The permeability of PMEM was assessed by the clearance rate of Evans blue-labeled albumin. β-Actin was extracted from PMEM lysate with a DNase-Sepharose column. The extracted β-actin was quantified in terms of its nitrotyrosine/β-actin ratio with antinitrotyrosine and anti-β-actin antibodies, sequentially, by dot-blot assays. The cellular compartmentalization of NO2-β-actin was displayed by showing confocal localization of nitrotyrosine-immunofluorescence with β-actin-immunofluorescence but not with F-actin fluorescence. Incubation of PMEM with TNF (100 ng/ml) for 0.5 and 4.0 h resulted in increases in permeability to albumin. There was an increase in the nitrotyrosine/β-actin ratio at 0.5 h with minimal association of the NO2-β-actin with F-actin polymers. The TNF-induced increase in the nitrotyrosine/β-actin ratio and permeability were prevented by the anti-ONOO− agent Urate. The data indicate that TNF induces an ONOO−-dependent barrier dysfunction, which is associated with the generation of NO2-β-actin.

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American Journal of Physiology - Lung Cellular and Molecular Physiology

Tập 290 Số 4

L674-L684

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