Effect of CRH on NO bioavailability, ROS production and antioxidant defense systems in endothelial EAhy926 cells

Abstract

Local or ‘Immune’ Corticotropin-Releasing Hormone (CRH) is secreted in peripheral tissues and plays a direct immunomodulatory role as an endocrine or paracrine mediator of inflammation. The present study was undertaken to determine whether CRH affects the endothelial redox state. Accordingly, intracellular reactive oxygen species (ROS) content and peroxynitrite levels, endothelial nitric oxide synthase (eNOS) activity and nitric oxide (NO) levels as well as catalase activity, superoxide dismutase (SOD) activity and glutathione (GSH) levels were measured in the presence or absence of selective CRH receptor-1 and CRH receptor-2 inhibitors in endothelial EAhy926 cells exposed in vitro in 10⁻⁷ M CRH for 2 h. CRH acting through both receptors induced a significant increase of ROS content (p < 0.001), catalase activity (p < 0.001) and SOD activity (p < 0.001), accompanied by a simultaneous significant decrease of eNOS activity and NO levels (p < 0.001), as well as a significant increase in nitrotyrosine (peroxynitrite) levels (p < 0.05). The data indicate that CRH may act as a regulator of pro-inflammatory mechanisms inducing adaptation of endothelial cell function to local stress.

Keywords::

• CRH
• reactive oxygen species
• nitric oxide
• eNOS
• catalase
• GSH
• endothelial cells
• inflammation

Acknowledgements

We express our sincere thanks to Professor A. Margioris and Assistant Professor Ch. Tsatsanis for their substantial help and the donation of EAhy926 endothelial cells.

Declaration of interest: This paper is part of the 03ED research project, implemented within the framework of the ‘Reinforcement Programme of Human Research Manpower’ (PENED) and co-financed by National and Community Funds (25% from the Greek Ministry of Development-General Secretariat of Research and Technology and 75% from E.U.-European Social Fund). The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper.

This paper was first published online on Early Online on 20 May 2010.