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A2326 - Targeting Soluble Guanylate Cyclase for Hyperoxia-Induced Neonatal Airway Dysfunction
Author Block: L. Wheeler1, R. D. Britt2, A. Haak2, A. M. Roesler3, S. A. Wicher3, J. Ravix3, T. Green3, L. J. Manlove3, M. A. Thompson3, C. M. Pabelick3, D. J. Tschumperlin2, Y. Prakash3; 1Mayo Clinic Graduate School of Biomedical Sciences, Mayo Clinic, Rochester, MN, United States, 2Department of Physiology and Biomedical Engineering, Mayo Clinic, Rochester, MN, United States, 3Department of Anesthesiology and Perioperative Medicine, Mayo Clinic, Rochester, MN, United States.
Objective: Hyperoxia with or without respiratory support is a necessary intervention for many of the >500,000 premature babies born each year in the US. However, hyperoxia exacerbates airway hyperresponsiveness, airflow obstruction, and wheezing early in life, and enhances the risk of chronic bronchial airway disease. Emerging evidence suggests that detrimental effects of hyperoxia on developing airways are attributable in part to disruption of the NO-sGC-cGMP axis, such that sGC dysfunction may be responsible for the lack of NO-induced bronchodilation. Further downstream long-term consequences of sGC dysfunction, particularly in the context of hyperoxia-induced fibrosis are not known. Here, mechanisms such as the YAP/TAZ pathway have emerged as key integrators of developmental signals including the NO-sGC-cGMP axis ultimately influencing cell proliferation, apoptosis, and cell fate in lung organogenesis. From a therapeutic perspective, there is increasing interest in novel sGC modulators that influence oxidized, dysfunctional sGC. With this background, we tested the effects of novel sGC modulators on the progressive fibrotic remodeling after hyperoxia-induced oxidative stress in developing airway cells. Methods: Human fetal fibroblast cells were isolated from canalicular stage (18-22 week gestation) lung and grown under standard conditions (deidentified samples; Mayo IRB exempt). Cells were serum-starved for 48 h prior to 48 h treatment with sGC modulators BAY 41 (stimulator), BAY 58 (activator), aticiguat, and highly novel sGC modulators (GPHR 312768, 312846, 312849) at 100 nM, 1 µM, and 10 µM with/without exposure to 21% room air, 40% hyperoxia, and additionally 0.2 ng/mL or 2.0 ng/mL TGF-β. Media and cells were harvested for mRNA, protein expression, and enzyme activity using standard techniques. Results: Hyperoxia increased fetal fibroblast proliferation, promoted collagen I, III, and fibronectin deposition, and enhanced MMP-9 activity, while BAY 41, BAY 58, and other sGC modulators attenuated hyperoxia effects. While hyperoxia decreased cGMP, sGC modulators reversed this effect and furthermore increased phospho-VASP at Ser239 (indicator of PKG activity). sGC modulators reduced α-SMA expression five-fold and reduced fiber disorganization within cells. These effects were dose-dependent. In addition, cells pre-treated with sGC compounds showed reduced YAP/TAZ nuclear localization. Effects of sGC modulators were blunted by PKG pre-inhibition. Conclusion: These data show that novel sGC modulators can blunt hyperoxia and TGF-β induced proliferation and pro-fibrotic pathways in developing fibroblasts. Targeting sGC with such novel modulators may help alleviate pathologic remodeling effects of neonatal exposure to hyperoxia. Supported by NIH grants R01 HL056470 (Prakash), K99 HL131682 (Britt), R01 HL138402 (Pabelick) and R25 GM075148 (Ravix).