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nNOS and Nox4 go nuclear: nNOS-derived and NADPH oxidase-derived reactive oxygen/nitrogen species promote oxidative nuclear damage in alveolar epithelial cells

Emerging evidence implicates a role for angiotensin II (Ang II)-stimulated reactive oxygen and nitrogen species (ROS/RNS) formation in acute lung injury (ALI). However, details of the mechanism are lacking. We hypothesized that compartmentalized generation of superoxide (O2-) and nitric oxide (•NO) may be key events in the Ang II-stimulated progression of ALI. In the present study, we found that Ang II markedly enhanced ROS/RNS production 7.4-fold, an effect blocked by the specific nNOS inhibitor N(G)-propyl-L-arginine, the NADPH oxidase inhibitor apocynin, or small interfering RNA (siRNA)-specific gene silencing targeted against nNOS or Nox4. nNOS/Nox4 transiently co-immunoprecipitates, and co-localizes at the peri-nuclear region 15 minutes post Ang II stimulation. Subsequently, confocal and western blot analyses show that nNOS/Nox4 translocates to the nucleus, suggesting that nNOS/Nox4 may directly regulate nuclear signaling. Furthermore, PAR polymers, which are undetectable in resting conditions, were generated following Ang II stimulation, an effect blocked with apocynin or N(G)-propyl-L-arginine. In conclusion, these data suggest Ang II causes nNOS/Nox4 to co-localize at the peri-nuclear region of A549 cells, where superoxide produced by Nox4, and •NO produced by nNOS immediately react to form peroxynitrite, which leads to subsequent nuclear oxidative damage as evidenced by increased PAR polymer formation. Furthermore, these experiments demonstrate inflammatory-stimulated nuclear translocalization of nNOS/Nox4, which has important implications for direct ROS/RNS-mediated nuclear activities. Therefore, inhibition of nNOS/Nox4 may be an effective therapeutic target in patients with ALI.

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Connelly, R., Schmalstieg, F. & Traber, D. nNOS and Nox4 go nuclear: nNOS-derived and NADPH oxidase-derived reactive oxygen/nitrogen species promote oxidative nuclear damage in alveolar epithelial cells. Crit Care 11, P17 (2007). https://doi.org/10.1186/cc5177

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Keywords

  • Nitric Oxide
  • Acute Lung Injury
  • Peroxynitrite
  • Alveolar Epithelial Cell
  • Apocynin