Role of NADPH oxidase in chronic intermittent hypoxia‐induced respiratory dysfunction: Insights from pharmacological and transgenic approaches

Abstract

Obstructive sleep apnoea syndrome (OSAS) is characterized by exposure to chronic intermittent hypoxia (CIH), as a consequence of repetitive occlusions of the upper airway in patients during sleep. CIH evokes redox changes culminating in impaired upper airway and diaphragm muscle function. Excessive ROS are also associated with impaired respiratory control, which manifests as destabilised breathing during sleep. There is a paucity of information regarding the molecular mechanisms underlying these effects. We sought to investigate the putative role of the superoxide‐generating NADPH oxidase 2 (NOX2) enzyme in CIH‐induced respiratory muscle dysfunction and respiratory maladaptation.A mouse model of CIH was generated by the cycling of gas from normoxia (21% O2) for 210 seconds to hypoxia (5% O2 at the nadir) over 90 seconds for 8hr/day for 2 weeks. Adult male (C57BL/6J) mice were assigned to one of 5 groups: normoxic controls (sham), CIH‐exposed, CIH+apocynin (NOX2 inhibitor, 2mM) given in the drinking water throughout the CIH and NOX2 null (B6.129S‐Cybbtm1Din/J) sham or CIH exposed. On day 15, whole body plethysmography was used to measure breathing parameters in unrestrained, unaesthetized mice in room air. Flow signals recorded were analysed offline and an apnoea was defined as ≥ 2 missed breaths. Sternohyoid and diaphragm muscle contractile function was examined ex vivo. Gene expression was examined by qRT‐PCR. Western blot was used to measure protein expression. NOX enzyme activity was determined using a spectrophotometric assay.CIH decreased sternohyoid and diaphragm muscle peak specific force by ~45% compared with sham exposure. CIH increased NOX enzyme activity in the sternohyoid, with no alterations in gene or protein expression of NOX subunits compared with sham. CIH decreased the expression of p22phox and rac genes and increased protein expression of NOX4 in the diaphragm, while NOX activity remained unchanged compared with sham. Administration of apocynin and NOX2 gene knockout completely prevented CIH‐induced sternohyoid and diaphragm muscle weakness. Basal minute ventilation remained unchanged following CIH exposure however the number of apnoeas per hour was increased compared with sham. Apocynin intervention reduced the frequency of apnoeas compared with the CIH group. Apnoea index was increased in NOX2 null mice exposed to CIH compared with NOX2 null sham mice reminiscent of that in wild‐type mice.Mice show signs of profound respiratory muscle dysfunction following exposure to 2 weeks of CIH. The putative NOX inhibitor, apocynin, prevents CIH‐induced respiratory muscle weakness. Studies in NOX2 null mice reveal that NOX2 is necessary for this CIH‐induced respiratory muscle weakness. CIH‐induced increase in the propensity for apnoea may be of biological relevance as it may underpin progression in the severity of OSAS pathology (i.e. mild‐to‐moderate OSAS). The reduction in apnoea frequency following treatment with apocynin implicates ROS in the manifestation of CIH‐induced respiratory disturbances in a non‐NOX2 dependant manner. Our results have implications for human OSAS and point to antioxidant intervention as a potential therapeutic strategy.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.