title: Effect of chemical stabilizers of hypoxia-inducible factors on early lung development.
abstract: Low oxygen stimulates pulmonary vascular development and airway branching and involves hypoxia-inducible factor (HIF). HIF is stable and initiates expression of angiogenic factors under hypoxia, whereas normoxia triggers hydroxylation of the HIF-1alpha subunit by prolyl hydroxylases (PHDs) and subsequent degradation. Herein, we investigated whether chemical stabilization of HIF-1alpha under normoxic (20% O(2)) conditions would stimulate vascular growth and branching morphogenesis in early lung explants. Tie2-LacZ (endothelial LacZ marker) mice were used for visualization of the vasculature. Embryonic day 11.5 (E11.5) lung buds were dissected and cultured in 20% O(2) in the absence or presence of cobalt chloride (CoCl(2), a hypoxia mimetic), dimethyloxalylglycine (DMOG; a nonspecific inhibitor of PHDs), or desferrioxamine (DFO; an iron chelator). Vascularization was assessed by X-gal staining, and terminal buds were counted. The fine vascular network surrounding the developing lung buds seen in control explants disappeared in CoCl(2)- and DFO-treated explants. Also, epithelial branching was reduced in the explants treated with CoCl(2) and DFO. In contrast, DMOG inhibited branching but stimulated vascularization. Both DFO and DMOG increased nuclear HIF-1alpha protein levels, whereas CoCl(2) had no effect. Since HIF-1alpha induces VEGF expression, the effect of SU-5416, a potent VEGF receptor (VEGFR) blocker, on early lung development was also investigated. Inhibition of VEGFR2 signaling in explants maintained under hypoxic (2% O(2)) conditions completely abolished vascularization and slightly decreased epithelial branching. Taken together, the data suggest that DMOG stabilization of HIF-1alpha during early development leads to a hypervascular lung and that airway branching proceeds without the vasculature, albeit at a slower rate.
authors.affiliation: CIHR Group in Lung Development, Hospital for Sick Children Research Institute, Department of Pediatrics and Institute of Medical Sciences, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Freek A
authors.initials: FA
authors.lastname: Groenman
authors.forename: Martin
authors.initials: M
authors.lastname: Rutter
authors.forename: Jinxia
authors.initials: J
authors.lastname: Wang
authors.forename: Isabella
authors.initials: I
authors.lastname: Caniggia
authors.forename: Dick
authors.initials: D
authors.lastname: Tibboel
authors.forename: Martin
authors.initials: M
authors.lastname: Post
mesh_terms: D000600:Amino Acids, Dicarboxylic / Q000494:pharmacology*; D000818:Animals; D002712:Chlorides; D003035:Cobalt / Q000494:pharmacology*; D003676:Deferoxamine / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004622:Embryo, Mammalian / Q000166:cytology / Q000187:drug effects; D004847:Epithelial Cells / Q000166:cytology; D005290:Ferric Compounds / Q000494:pharmacology; D051793:Hypoxia-Inducible Factor 1 / Q000378:metabolism*; D066298:In Vitro Techniques; D007211:Indoles / Q000494:pharmacology; D008168:Lung / Q000187:drug effects* / Q000196:embryology*; D051379:Mice; D009024:Morphogenesis / Q000187:drug effects; D018919:Neovascularization, Physiologic / Q000187:drug effects; D011758:Pyrroles / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000037:antagonists & inhibitors
chemical_list: D000600:Amino Acids, Dicarboxylic; D002712:Chlorides; D005290:Ferric Compounds; D051793:Hypoxia-Inducible Factor 1; D007211:Indoles; D011758:Pyrroles; D003035:Cobalt; C116890:Semaxinib; D040301:Vascular Endothelial Growth Factor Receptor-2; C018021:cobaltous chloride; D003676:Deferoxamine; C024555:ferric chloride; C040947:oxalylglycine