Increased interleukin-8 expression by cigarette smoke extract in endothelial cells
Introduction
Cigarette smoking is a major risk factor in the development of lung inflammatory diseases and cancer. Our previous study had shown that passive cigarette smoking and cigarette smoke extract worsened alcohol-induced gastric mucosal damage and the action was found to be due to the increase of neutrophil infiltration (Chow et al., 1997). It is well established that cigarette smoking can increase the incidence of cancer and invasion of tumor cells. In addition, neutrophil adhesion, infiltration and angiogenic processes are the important steps in the inflammatory process and cancer development.
IL-8 is a member of the C-X-C chemokine family which exerts chemotactic effect primarily on neutrophils (Hebert and Baker, 1993, Graves and Jiang, 1995). It has been reported that IL-8 and tumor biology have been related (Opdenakker and Van Damme, 1992). Koch et al. (1992) also suggest that IL-8 is a potent angiogenic factor. It is shown that cigarette smoke extract can induce interleukin-8 release from human bronchial epithelial cells (Mio et al., 1997). It is well known that endothelial cells play an important role in inflammatory processes and tumor development, and have the potential to secrete IL-8 in physiological conditions. However, it is unknown whether cigarette smoking can increase the secretion of IL-8 from endothelial cells. In this experiment, we aimed to examine whether cigarette smoke extract could stimulate the release of IL-8 from endothelial cell line and also investigate the possible signal transduction pathway involved in this experimental model.
Section snippets
Reagents
All chemicals and reagents were purchased from Sigma™ (Sigma Chemical Company, St Louis, USA) unless specified. Protein kinase C (PKC) inhibitor GF109203X (0.05 and 5.0 μM), protein kinase A (PKA) inhibitor H8 (0.05 and 5.0 μM), and protein tyrosine kinase (PTK) inhibitor genistein (10 and 100 μM) were bought from Calbiochem™ (San Diego, USA).
Cigarette smoke extract preparation
The commercial brand ‘Camel™’ (R.J. Reynolds, Winston-Salem, NC, USA) was used in the present study. We prepared different fractions of cigarette smoke
Effects of CSE on IL-8 expression
The ability of CSE to increase IL-8 secretion in ECV304 was evaluated by analyzing ECV304 supernatants for IL-8 using an IL-8 ELISA kit. In Fig. 1, both chloroform soluble extract (Chlo E) and ethanol soluble extract (EtOH E) showed a dose-dependent increase of IL-8 expression in ECV304 cell line. To determine the IL-8 time response to CSE, kinetic expression of IL-8 from CSE-treated ECV304 was studied at different time intervals. It was found that IL-8 levels in the supernatants of
Discussion
It is well established that IL-8, a potent leukocyte chemotactic factor, is produced in many types of cells, including monocytes, keratinocytes, lymphocytes, eosinophils, fibroblasts mesothelial cells and endothelial cells. A unique feature of IL-8 when compared to other chemotactic factors, is its specificity for neutrophils both in vitro and in vivo, and is comparatively weaker on other cell types. It is shown that endogenous IL-8 secreted by activated endothelial cells can induce the
Acknowledgements
This research was supported by a RGC grant (HKU 7277/97M) awarded by the Hong Kong Research Grant council. The authors also thanked Dr Song-Ze Ding for his technical advice on the preparation of different kinase inhibitors.
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