Regulation of the expression of soluble guanylyl cyclase by reactive oxygen species

Abstract

Background and purpose: Superoxide anions produced during vascular disease scavenge nitric oxide (NO), thereby reducing its biological activity. The aim of the present study was to investigate whether reactive oxygen species (ROS) have a direct effect on soluble guanylyl cyclase (sGC) subunit levels and function and to ascertain the mechanism(s) involved. Experimental approach: Rat aortic smooth muscle cells (RASM) or freshly isolated vessels were exposed to reactive oxygen species (ROS)-generating agents and sGC subunit expression was determined at the mRNA and/or protein level. cGMP accumulation was also determined in RASM exposed to ROS. Key results: Incubation of smooth muscle cells with H 2 O 2 , xanthine/xanthine oxidase (X/XO) or menadione sodium bisulphite (MSB) significantly decreased protein levels of a1 and b1 subunits of sGC and reduced SNP-induced cGMP formation. Similarly, sGC expression was reduced in freshly isolated vessels exposed to ROS-generating agents. The ROS-triggered inhibition of a1 and b1 levels was not blocked by proteasome inhibitors, suggesting that decreased sGC protein was not due to protein degradation through this pathway. Real time RT-PCR analysis demonstrated a 68% reduction in steady state mRNA levels for the a1 subunit following exposure to H 2 O 2 . In addition, a1 promoter-driven luciferase activity in RASM decreased by 60% after H 2 O 2 treatment. Conclusion and implications:We conclude that oxidative stress triggers a decrease in sGC expression and activity that results from reduced sGC steady state mRNA levels. Altered sGC expression is expected to contribute to the changes in vascular tone and remodeling observed in diseases associated with ROS overproduction.

Aldosterone Increases Oxidant Stress to Impair Guanylyl Cyclase Activity by Cysteinyl Thiol Oxidation in Vascular Smooth Muscle Cells

“…More recently, it was shown that rat aortic smooth muscle cells exposed to elevated levels of H 2 O 2 (500 mol/liter) for 24 h demonstrated decreased GC ␣ 1 -and ␤ 1 -subunit mRNA and protein expression; however, exposure to this concentration of H 2 O 2 was associated with a 25% decrease in cell viability. In addition, when these investigators exposed cells to a lower concentration of H 2 O 2 (150 mol/ liter), they saw no change in ␣ 1 -subunit protein expression and an increase in ␤ 1 -subunit expression (39). In our experiments, the levels of H 2 O 2 generated by aldosterone were similar to the lower concentration of H 2 O 2 studied by these investigators, and aldosterone treatment at these concentrations did not influence cell viability.…”

Soluble Guanylate Cyclase Redox State Under Hypoxia or Hypoxia/Reoxygenation in Isolated Monkey Coronary Arteries

“…However, it is also important to remember that more prolonged exposure to reactive oxygen species leads to downregulation of sGC expression. Gerassimou et al have confirmed that sGC expression is reduced in freshly isolated rat aortic strips by exposure to superoxide-generating agents or hydrogen peroxide for a long period (23). In any case, taken together, the response to agents activating the reduced sGC would be diminished sooner or later if the generation of reactive oxygen and nitrogen species was stimulated.…”

Impairment by Hypoxia or Hypoxia/Reoxygenation of Nitric Oxide–Mediated Relaxation in Isolated Monkey Coronary Artery: the Role of Intracellular Superoxide

“…The report by Wu and de Champlain has noted an inhibitory effect of superoxide on cGMP formation in vascular smooth muscle cells (24). Other investigators have also reported that exposure of smooth muscle cells to superoxide generating agents causes a decrease in sGC expression and attenuates cGMP formation induced by NO donors (25). In addition, NO-donors used in this study require enzymatic catalysis by mitochondrial aldehyde dehydrogenase 2 (ALDH2) or by P450 in the endoplasmic reticulum to release NO (26).…”