Vaspin inhibited proinflammatory cytokine induced activation of nuclear factor-kappa B and its downstream molecules in human endothelial EA.hy926 cells

Aims: In this study, we investigated the effects of visceral adipose tissue-derived serpin (vaspin), a newly discovered adipocytokine, on nuclear factor-kappa B (NF-kappa B) and its downstream molecules in proinflammatory cytokines, tumor necrosis factor-alpha (TNF-alpha) and interleukine-1 (IL-1), stimulated human endothelial EA.hy926 cells to elucidate the role of vaspin in the inflammatory states of endothelium. Methods: A NF-kappa B luciferase reporter system was constructed and stably transfected into human endothelial cell line EA.hy926. Following transfection, EA.hy926 cells were pre-treated with various concentrations of vaspin (0-320 ng/ml) before TNF-alpha and IL-1 stimulation. The transcription activity of NF-kappa B was determined using luciferase reporter assay. Expression levels of NF-kappa B downstream inflammatory cytokines, TNF-alpha, IL-1 and IL-6 were measured by enzyme-linked immunosorbent assay (ELISA). Expressions of adhesion molecules and chemokines, intercellular cell adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and monocyte chemotactic protein-1 (MCP-1) were determined by quantitative real-time PCR (RT-PCR) and western blot in mRNA and protein levels, respectively. Results: Results showed that vaspin inhibited TNF-alpha and IL-1 mediated activation of NF-kappa B and its downstream molecules in a concentration-dependent manner (P < 0.05). Conclusions: We conclude that vaspin protected endothelial cells from proinflammatory cytokines induced inflammation by inhibition of NF-kappa B and its downstream molecules. (C) 2013 Elsevier Ireland Ltd. All rights reserved.