Abstract
Tissue intermittent hypoxia (IH) occurs in obstructive sleep apnea, sickle cell anemia, physical exercise and other conditions. Poor gas solubility and slow diffusion through culture media hampers mimicking IH-induced transitions of O-2 in vitro.
We aimed to develop a system enabling exposure of cultured cells to IH and to validate such exposure by real-time O-2 measurements and cellular responses. Standard 24-well culture plates and plates with bottoms made from a gas permeable film were placed in a heated cabinet.
Desired cycling of O-2 levels was induced using programmable solenoids to purge mixtures of 95% N-2 + 5% CO2 or 95% O-2 + 5% CO2. Dissolved oxygen, gas pressure, temperature, and water evaporation were measured during cycling.
IH-induced cellular effects were evaluated by hypcoda inducible factor (HIF) and NF-kappa B luciferase reporters in HEK296 cells and by insulin secretion in rat insulinoma cells. Oxygen cycling in the cabinet was translated into identical changes of O-2 at the well bottom in gas permeable, but not in standard cultureware.
Twenty-four hours of IH exposure increased HIF (112%), NF-kappa B (111%) and insulin secretion (44%). Described system enables reproducible and prolonged IH exposure in cultured cells while controlling for important environmental factors.