Regulation of tumor necrosis factor-α–induced microvascular endothelial cell hyperpermeability by recombinant B-cell lymphoma-extra large
| Autor: | Rickesha L. Wilson, Ed W. Childs, Felicia A. Hunter, Binu Tharakan, Devendra A. Sawant, Hayden W. Stagg |
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| Rok vydání: | 2013 |
| Předmět: |
Stress fiber
Beta-catenin medicine.medical_treatment bcl-X Protein Vascular permeability Cysteine Proteinase Inhibitors Article Capillary Permeability Adherens junction medicine Animals Edema RNA Small Interfering Lung Cells Cultured beta Catenin Caspase 8 biology Caspase 3 Tumor Necrosis Factor-alpha Endothelial Cells Adherens Junctions Actin cytoskeleton Molecular biology Recombinant Proteins Rats Cell biology Endothelial stem cell Cytokine biology.protein Surgery Tumor necrosis factor alpha Oligopeptides BH3 Interacting Domain Death Agonist Protein |
| Zdroj: | Journal of Surgical Research. 184:628-637 |
| ISSN: | 0022-4804 |
| Popis: | Background Tumor necrosis factor-α (TNF-α), a cytotoxic cytokine, induces endothelial cell barrier dysfunction and microvascular hyperpermeability, leading to tissue edema, a hallmark of traumatic injuries. The objective of the present study was to determine whether B-cell lymphoma-extra large (Bcl-xL), an antiapoptotic protein, would regulate and protect against TNF-α–mediated endothelial cell barrier dysfunction and microvascular hyperpermeability. Methods Rat lung microvascular endothelial cells were grown as monolayers on Transwell membranes, and fluorescein isothiocyanate-bovine albumin flux (5 mg/mL) across the monolayer was measured fluorometrically to indicate changes in monolayer permeability. The rat lung microvascular endothelial cell adherens junctional integrity and actin cytoskeleton was studied using β-catenin immunofluorescence and rhodamine phalloidin dye, respectively. Pretreatment of caspase-8 inhibitor (Z-IETD-FMK, 100 μM) for 1 hour and transfection of Bcl-2-homology domain 3-interacting domain death agonist small interfering RNA (10 μM) for 48 hours were performed to study their respective effects on TNF-α–induced (10 ng/mL; 1-hour treatment) monolayer permeability. Recombinant Bcl-xL protein (2.5 μg/ml) was transfected in rat lung microvascular endothelial cells for 1 hour, and its effect on permeability was demonstrated using a permeability assay. Caspase-3 activity was assayed fluorometrically. Results Z-IETD-FMK pretreatment protected the adherens junctions and decreased TNF-α–induced monolayer hyperpermeability. Bcl-2-homology domain 3-interacting domain death agonist small interfering RNA transfection attenuated the TNF-α–induced increase in monolayer permeability. Recombinant Bcl-xL protein showed protection against TNF-α–induced actin stress fiber formation, an increase in caspase-3 activity, and monolayer hyperpermeability. Conclusions Our results have demonstrated the protective effects of recombinant Bcl-xL protein against TNF-α–induced endothelial cell adherens junction damage and microvascular endothelial cell hyperpermeability. These findings support the potential for Bcl-xL–based drug development against microvascular hyperpermeability and tissue edema. |
| Databáze: | OpenAIRE |
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