Increased catalytic activity of CBS (cystathionine β-synthase) was recently shown to mediate vasodilation of the cerebral microcirculation, which is initiated within minutes of the onset of acute hypoxia. To test whether chronic hypoxia was a stimulus for increased CBS expression, U87-MG human glioblastoma and PC12 rat phaeochromocytoma cells were exposed to 1% or 20% O2 for 24–72 h. CBS mRNA and protein expression were increased in hypoxic cells. Hypoxic induction of CBS expression was abrogated in cells transfected with vector encoding shRNA targeting HIF (hypoxia-inducible factor) 1α or 2α. Exposure of rats to hypobaric hypoxia (0.35 atm; 1 atm=101.325 kPa) for 3 days induced increased CBS mRNA, protein and catalytic activity in the cerebral cortex and cerebellum, which was blocked by administration of the HIF inhibitor digoxin. HIF-binding sites, located 0.8 and 1.2 kb 5′ to the transcription start site of the human CBS and rat Cbs genes respectively, were identified by ChIP assays. A 49-bp human sequence, which encompassed an inverted repeat of the core HIF-binding site, functioned as a hypoxia-response element in luciferase reporter transcription assays. Thus HIFs mediate tissue-specific CBS expression, which may augment cerebral vasodilation as an adaptive response to chronic hypoxia.
- cerebral circulation
- cystathionine β-synthase (CBS)
- hydrogen sulfide
- hypoxia-inducible factor (HIF)
Abbreviations: CBS, cystathionine β-synthase; CTH, cystathionine γ-lyase; DMEM, Dulbecco’s modified Eagle’s medium; EV, empty vector; HBSS, Hepes-buffered saline solution; HIF, hypoxia-inducible factor; HO, haem oxygenase; HRE, hypoxia-response element; IP, intraperitoneal; LDHA, lactate dehydrogenase A; MPST, 3-mercaptopyruvate sulfurtransferase; qRT-PCR, quantitative reverse transcription–PCR; RPL13A, ribosomal protein L13a; SLC2A1, solute carrier 2A1; VEGF, vascular endothelial growth factor; WT, wild-type
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