Nitric oxide overproduction derived from inducible nitric oxide synthase increases cardiomyocyte apoptosis in human atrial fibrillation

Abstract

Background Inflammation and oxidant stress have been suggested to be involved in the structural remodeling in atrial fibrillation (AF), and inducible nitric oxide synthase (iNOS) is associated with inflammation and oxidant stress. To study whether iNOS could contribute to atrial remodeling in AF, we investigated the relationship between inflammation, oxidant stress, nitric oxide (NO) and its synthase, and cardiomyocytic apoptosis in the right atrium in human AF. Methods Fifteen patients with permanent AF (PmAF) and 17 patients with sinus rhythm (SR), who underwent mitral valve replacement surgery were investigated. Western blotting and immunohistochemical staining were used to detect the expression of endothelial nitric oxide synthase (eNOS), iNOS and 3-nitrotyrosine (3NT; a biological marker of peroxynitrite) in the right atrium. The occurrence of cardiomyocytic apoptosis was determined by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL). Caspase 3 staining for the activated, cleaved protease was also performed. In addition, concentrations of NO 2 −/NO 3 − (NO X ) both in plasma and in the right atrium were measured by a NO X Detection Kit. Finally, plasma levels of high-sensitivity C-reactive protein (hs-CRP) were routinely measured. Results and conclusions Levels of hs-CRP were far more enhanced in patients with PmAF compared to the controls. Plasma levels of NO X were significantly lower in PmAF patients than SR patients, but the production of NO X in the local right atrium increased obviously. Furthermore, iNOS and 3NT expressions increased dramatically in the right atrium in PmAF patients, whereas the expression of eNOS did not change apparently. In addition, when patients were further divided into a higher hs-CRP group (≥ 5 mg/L) and a lower hs-CRP group (< 5 mg/L) according to the hs-CRP level, significant upregulation of iNOS was found in the higher hs-CRP group. Apoptosis index and caspase 3 staining were also prominently enhanced in PmAF patients compared with SR patients. More importantly, we demonstrated in this study that a higher expression of 3NT was associated with an increased expression of iNOS/eNOS ( r = 0.74, P < 0.05) and an enhanced apoptosis index ( r = 0.69, P < 0.05). In conclusion, the results presented novel evidence that imbalanced expression of iNOS/eNOS could contribute to protein nitration and cardiomyocyte apoptosis in human AF, in which condition inflammation may be an important participant.