NADPH oxidases are responsible for the failure of nitric oxide to inhibit migration of smooth muscle cells exposed to high glucose.

Our previous studies showed that nitric oxide (NO) fails to inhibit migration of smooth muscle cells (SMC) exposed to high glucose (HG) because of oxidation of the most reactive cysteine, cysteine-674, on the sarco/endoplasmic reticulum ATPase, preventing its S-glutathiolation, thus blocking NO action. This study further addresses the sources of the oxidants responsible for the failure of NO to inhibit SMC migration in HG. NADPH oxidases are the major source of reactive oxygen species (ROS) in SMC. We used small interfering RNA or dominant-negative adenoviral vectors to target components of NADPH oxidase to study their individual roles by measuring serum-induced migration in the presence or absence of NO. In HG, the mRNA levels of Nox1 and Nox4 and the protein level of Nox4 were increased; knocking down Nox1 or Nox4 attenuated the ROS production and restored the inhibition of SMC migration by NO. Blockade of the activation of Rac1 or p47(phox) inhibited serum-induced migration and restored the inhibition of migration by NO. These data indicate that NADPH oxidases are responsible for the failure of NO to inhibit SMC migration caused by HG.