B17-06 Preventing Airway Disease in Bronchopulmonary Dysplasia: Potential Role of Denitrosylase Enzymes

Rationale Bronchopulmonary dysplasia (BPD) is diagnosed annually in over ten thousand US infants, with a paucity of effective therapies for associated airway hyperreactivity. S-nitrosoglutathione (GSNO) is an endogenous bronchodilator catabolized by denitrosylase enzymes, including GSNO reductase (GSNOR) and recently described aldo-keto reductase family 1 member A1 (AKR1A1) (PMID: 31649033). GSNOR is increased in human BPD airways (PMID:33780653). In a 3-week sustained hyperoxia mouse model of BPD, GSNOR expression was increased, and acute chemical GSNOR inhibition reversed airway hyperreactivity (PMID:27484068). Genetic knockout of GSNOR in mice prevented later BPD changes in this model, including airway hyperreactivity (PMID:33780653). Here we investigate human tracheal aspirate GSNOR expression in at-risk neonates and the effects of GSNOR knockout versus early treatment with a pharmaceutical GSNOR inhibitor in a newborn 5-day hyperoxia mouse model. We hypothesize that (1) in human tracheal aspirate samples, neonatal GSNOR expression is increased in patients diagnosed with Grade 3 BPD, and (2) in the mouse model, both GSNOR knockout and GSNOR pharmacologic inhibition during hyperoxic exposures will prevent airway hyperreactivity after room air recovery. Methods Human Samples: Tracheal aspirates were collected weekly for up to 6 weeks in intubated infants from two academic pediatric hospitals. Aspirates were analyzed by automated capillary simple Western (ProteinSimple, Bio-Techne) to quantify GSNOR normalized to total protein. Mouse Model: Newborn C57BL/6 mice (wild-type or GSNOR knockouts) were exposed to room air or 60% hyperoxia for 5 days. Wild-type pups received daily s.q. saline or GSNOR inhibitor (1 mg/kg N6022) during oxygen exposures. In vivo respiratory system resistance (Rrs) was measured in response to inhaled methacholine (flexiVent, SCIREQ) after recovery in room air to 3 weeks postnatal age. In 6-day wild-type littermates, whole lungs were collected immediately after exposures to quantify expression of AKR1A1 (ProteinSimple). Results In the human tracheal aspirates, GSNOR expression was increased in samples from patients diagnosed with Grade 3 BPD (Fig1a). In the mouse model, both genetic GSNOR knockout and early daily GSNOR inhibitor treatments attenuated oxygen-induced airway hyperreactivity compared to the wild-type hyperoxia group (Fig1b). AKR1A1 expression was increased in the lungs of the wild-type hyperoxia group compared to room air controls (Fig1c). Conclusions These studies translate our observations of elevated airway GSNOR from infants developing BPD with experimental prevention of airway hyperreactivity in oxygen-exposed GSNOR knockouts and GSNOR inhibitor treated wild-types. Additionally, we identify elevated AKR1A1 as a novel denitrosylase mediator in newborn hyperoxia, which warrants further investigation. This abstract is funded by: NIH R01HL172894, R03HL155246, P01L158507, & 5T32HL091816