Effects of intermittent hypoxia and hyperoxia on angiogenesis and lung development in newborn mice

Abstract

Background: Premature birth disrupts hypoxia driven microvascular development that directs alveolar and lung growth. Changes in oxygen exposure after birth can perturb the regulation of angiogenesis leading to bronchopulmonary dysplasia (BPD). We studied the effects of intermittent hypoxia or hyperoxia on HIF and angiogenic gene expression and lung development in newborn mice.

Methods: Newborn litters were randomized within 12 h of birth to 12% O2 (4 h), 50% O2 (4 h) or 12% O2 (2 h)/50% O2 (2 h) followed by room air (RA) recovery for 20 h. Mice in RA were the control group. The mice were exposed to 6 such cycles (D1-D6) and sacrifice on D7. Whole lung mRNA was isolated and gene expression performed by qRT-PCR (HIF1α/2α/1β; PHD2, Ang1, Tie2, Vegf, VegfR1 & VegfR2) and analyzed by PCR array data analysis web portal. HIF-1α, prolyl hydroxylase-2 and VEGF protein were analyzed in whole lung by ELISA. Lung morphology was assessed by H&E sections and radial alveolar counts; cell proliferation by Ki67 immunostaining.

Results: HIF-1α mRNA and VEGF protein were significantly downregulated in the 50% O2 group; VEGF mRNA and protein were significantly downregulated in the 12% O2-50% O2 group; Ang-1 and its receptor mRNA expression were downregulated in 12% O2 and 12% O2-50% O2 groups. 50% O2 (hyperoxia) and 12% O2-50% O2 (hypoxia-hyperoxia) groups demonstrated alveolar simplification by RAC and the same groups had decreased cell proliferation by Ki67 staining compared to RA and hypoxia (12% O2) groups.

Conclusions: Downregulation of HIF and angiogenic gene expression with associated changes in lung histology following intermittent hypoxia-hyperoxia is likely an important contributing factor in the development of BPD.

Keywords: Oxygen; VEGF; hyperoxia; hypoxia; hypoxia inducible factor.