ABSTRACT: Excessive free radical production by immune cells has been linked to cell death and tissue injury during sepsis. Peroxynitrite is a short-lived oxidant and a potent inducer of cell death that has been identified in several pathological conditions. Caffeic acid phenethyl ester (CAPE) is an active component of honeybee products and exhibits antioxidant, anti-inflammatory, and immunomodulatory activities. The present study examined the ability of CAPE to scavenge peroxynitrite in RAW 264.7 murine macrophages stimulated with lipopolysaccharide/interferon-γ that was used as an in vitro model. Conversion of 123-dihydrorhodamine to its oxidation product 123-rhodamine was used to measure peroxynitrite production. Two mouse models of sepsis (endotoxemia and cecal ligation and puncture) were used as in vivo models. The level of serum 3-nitrotyrosine was used as an in vivo marker of peroxynitrite. The results demonstrated that CAPE significantly improved the viability of lipopolysaccharide/interferon-γ–treated RAW 264.7 cells and significantly inhibited nitric oxide production, with effects similar to those observed with an inhibitor of inducible nitric oxide synthase (1400W). In addition, CAPE exclusively inhibited the synthesis of peroxynitrite from the artificial substrate SIN-1 and directly prevented the peroxynitrite-mediated conversion of dihydrorhodamine-123 to its fluorescent oxidation product rhodamine-123. In both sepsis models, CAPE inhibited cellular peroxynitrite synthesis, as evidenced by the absence of serum 3-nitrotyrosine, an in vivo marker of peroxynitrite. Thus, CAPE attenuates the inflammatory responses that lead to cell damage and, potentially, cell death through suppression of the production of cytotoxic molecules such as nitric oxide and peroxynitrite. These observations provide evidence of the therapeutic potential of CAPE treatment for a wide range of inflammatory disorders.
*Departments of Anesthesiology, †Ophthalmology, ‡Medical Microbiology, §Rehabilitation Medicine, and ∥Parasitology, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia; and ¶Department of Molecular Biology and Genetics, Faculty of Arts and Science, Canik Basari University, Samsun, Turkey
Received 17 Dec 2013; first review completed 8 Jan 2014; accepted in final form 18 Mar 2014
Address reprint requests to Mustafa Kassim Abdulazez Kassim, Department of Anaesthesiology, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia. E-mail: Mustafakassim@um.edu.my.
This study was supported in part by grants (PV009/2011B, RG225/10HTM, HIR E000056, and RG515/13HTM) from the University of Malaya.
There are no competing interests to declare.