Abstract 2817: Potential Role and Molecular Mechanisms of Oxidative Stress in Atrial Tachycardia Remodeling
Atrial tachycardia remodeling (ATR) plays an important role in the pathophysiology of atrial fibrillation (AF). Oxidative stress (OxS) occurs in the atria of AF patients and antioxidants may prevent AF. This study determined whether OxS contributes to AF by mediating effects of ATR, with the use of in vivo and in vitro models.
Methods: Studies were performed in 1) atrial tissues and cardiomyocytes (ACMs) from15 dogs subjected to atrial tachypacing (400 bpm × 1 wk, ATP) and 15 control (CTL) dogs; 2) ACMs subjected for 24 hrs to tachypacing in vitro at 3 Hz (P3 cells), with 1 Hz paced ACMs (P1 cells) as CTLs. ICaL was recorded with whole cell patch clamp, Ca2+ transients with Indo-1 AM, superoxide production with dihydroethidium fluorescence (DHEF), mRNA and protein expression by qPCR and immunoblot respectively.
Results: In vivo ATP reduced ICaL at +10 mV from 5.6±0.6 to 2.1±0.6 pA/pF*** (***P<0.001) and increased DHEF from 3.5±0.4 to 22.5±1.1 AU***. Incubation of ATP ACMs with the antioxidant 2-MPG restored ICaL to 4.1±0.4 pA/pF*** and decreased DHEF to 7.1±0.8 AU***, without affecting CTL ACMs. In vitro 3 Hz pacing decreased ICaL by 61%*** and increased DHEF 2-fold (P<0.01) vs P1 cells. Antioxidants 2-MPG and N-acetylcysteine (NAC) reversed 3-Hz pacing effects on DHEF and ICaL. OxS with H2O2 (100 μM × 24 hr) mimicked P3 effects (52%↑*** ICaL, 167%↓*** DHEF) in P1 cells. H2O2 caused Ca2+i loading, increasing the Ca2+ transient by 37%***. CaMKII autophosphorylation and CaMKII phospholamban-phosphorylation were enhanced in H2O2-exposed P1 cells, indicating that OxS causes Ca2+-dependent CaMKII-activation. The CaMKII blocker KN-93 prevented ICaL downregulation and DHEF enhancement by 3-Hz pacing, suggesting that CaMKII contributes to ATR-induced OxS and ICaL downregulation. H0O2-induced ICaL downregulation was prevented by: decreasing Ca2+i loading by ICaL blockade (nimodipine) or Ca2+ chelation (BAPTA-AM); inhibition of calmodulin (W7) or CaMKII (KN93); suppression of free-radical generation via NADPH-oxidase (apocynin) and OxS suppression with antioxidants (2-MPG or NAC).
Conclusions: ATR causes OxS, which enhances cellular Ca2+ loading and activates CaMKII, creating a positive feedback system that contributes to ATR-induced ICaL downregulation.