Publications

Intermittent hypoxia (IH) has been implicated in the cardiovascular consequences of obstructive sleep apnea (OSA). However, the lack of suitable experimental systems has precluded assessment as to whether IH is detrimental, protective, or both for the endothelium. The aim of the work was to determine the effects of frequency and amplitude of IH oxygenation swings on aortic endothelial wound healing. Monolayers of human primary endothelial cells were wounded and subjected to constant oxygenation (1%, 4%, 13%, or 20% O2) or IH at different frequencies (0.6, 6, or 60 cycles/h) and magnitude ranges (13–4% O2 or 20–1% O2), using a novel well-controlled system, with wound healing being measured after 24 h. Cell monolayer repair was similar at 20% O2 and 13% O2, but was considerably increased (approximately twofold) in constant hypoxia at 4% O2. The magnitude and frequency of IH considerably modulated wound healing. Cycles ranging 13–4% O2 at the lowest frequency (0.6 cycles/h) accelerated endothelial wound healing by 102%. However, for IH exposures consisting of 20% to 1% O2 oscillations, wound closure was reduced compared with oscillation in the 13–4% range (by 74% and 44% at 6 cycles/h and 0.6 cycles/h, respectively). High-frequency IH patterns simulating severe OSA (60 cycles/h) did not significantly modify endothelial wound closure, regardless of the oxygenation cycle amplitude. In conclusion, the frequency and magnitude of hypoxia cycling in IH markedly alter wound healing responses and emerge as key factors determining how cells will respond in OSA. NEW & NOTEWORTHY Intermittent hypoxia (IH) induces cardiovascular consequences in obstructive sleep apnea (OSA) patients. However, the vast array of frequencies and severities of IH previously employed in OSA-related experimental studies has led to controversial results on the effects of IH. By employing an optimized IH experimental system here, we provide evidence that the frequency and magnitude of IH markedly alter human aortic endothelial wound healing, emerging as key factors determining how cells respond in OSA.

JTD Keywords: Sleep apnea, Repair, Endothelium, Hypoxia, Reoxygenation

High frequency intermittent hypoxia is one of the most relevant injurious stimuli experienced by patients with obstructive sleep apnea (OSA). Given that the conventional setting for culturing cells under intermittent hypoxia conditions is limited by long equilibration times, we designed a simple bioreactor capable of effectively subjecting cultured cells to controlled high-frequency hypoxic/normoxic stimuli. The bioreactor's operation is based on exposing cells to a medium that is bubbled with the appropriate mixture of gases into two separate containers, and from there it is directed to the cell culture dish with the aid of two bidirectional peristaltic pumps. The device was tested on human alveolar epithelial cells (A549) and mouse melanoma cells (B16-F10), subjecting them to patterns of intermittent hypoxia (20s at 5% O 2 and 50s at 20% O 2), which realistically mimic OSA of up to severe intensity as defined by the apnea hypopnea index. The proposed bioreactor can be easily and inexpensively assembled and is of practical use for investigating the effects of high-rate changes in oxygen concentration in the cell culture medium.

JTD Keywords: Hypoxia-reoxygenation, Obstructive sleep apnea, Oxygen partial pressure