Regulation of endothelial heme oxygenase activity during hypoxia is dependent on chelatable iron

Abstract

The regulation of heme oxygenase (HO) activity and its dependence on iron was studied in bovine aortic endothelial cells (BAEC) subjected to hypoxia-reoxygenation (H/R). HO activity was induced by hypoxia (10 h) and continued to increase during the reoxygenation phase. HO-1 protein levels were strongly induced by hypoxia from undetectable levels and remained elevated at least 8 h postreoxygenation. Addition of the Fe(3+) chelator desferrioxamine mesylate (DFO) or the Fe(2+) chelator o-phenanthroline during hypoxia alone or during the entire H/R period inhibited the induction of HO activity and HO-1 protein levels. However, DFO had no effect and o-phenanthroline had a partial inhibitory effect on HO activity and protein levels when added only during reoxygenation. Loading of BAEC with Fe(3+) enhanced the activation of the HO-1 gene by H/R, whereas loading with L-aminolevulinic acid, which stimulates heme synthesis, had little effect. These results suggest that chelatable iron participates in regulating HO expression during hypoxia.

Heme oxygenase-1 protects human hepatocytes in vitro against warm and cold hypoxia

“…An increase in HO-1 mRNA or protein expression after hypoxia was observed in rat lung, liver, heart, or in rat aortic vascular smooth muscle cells [11]. In bovine aortic endothelial cells, increase in HO-1 activity, mRNA and protein expression was also observed after hypoxic treatment of cells [33,34]. These results show, similar to our study, that hypoxia results in a time-dependent increase of HO-1 mRNA and protein expression, although the timings of induction and downregulation may vary among tissue, cell cultures or varies species.…”

Iron Released by Sodium Nitroprusside Contributes to Heme Oxygenase-1 Induction via the cAMP-Protein Kinase A-Mitogen-Activated Protein Kinase Pathway in RAW 264.7 Cells

“…Therefore, different effects of DFO on SNP-and SNAPexposed cells suggest that induction of HO-1 by these chemicals is under different regulatory mechanisms. Indeed, free iron is known to have effects on the gene expression of several proteins involved in iron metabolism, transferrin receptor, and ferritin (Suematsu et al, 1994;Raju and Maines, 1996;Brenneisen et al, 1998), and on enzymes potentially involved in oxidant or inflammatory conditions-inducible nitric-oxide synthase (Tenhunen et al, 1969), the aconitases (Tenhunen et al, 1970), and HO-1 (Ryter et al, 2000). Furthermore, evidence from the results of carboxy-PTIO or HCB, a known scavenger of NO, strongly indicates that iron is more critical than NO in SNP induction of HO-1 in RAW 264.7 cells.…”

Kaposi sarcoma-associated herpesvirus (KSHV) induces heme oxygenase-1 expression and activity in KSHV-infected endothelial cells

“…49 Briefly, following overnight incubation in complete media alone or with 10 M CuMP or CrMP, monolayers were rinsed with PBS and scraped directly into 300 L sonication buffer (0.25 M sucrose, 20 mM Tris-HCl, 50 g/mL Pefabloc SC, 4 g/mL leupeptin; pH 7.4) sonicated on ice 2 times for 30 seconds and centrifuged for 20 minutes at 18 000g. The protein concentration of the resultant supernatant was determined using Bradford reagent as described above.…”

“…HO activity was measured by the spectrophotometric determination of bilirubin production as described elsewhere. 49 Final reaction concentrations were 1 mg endothelial cell protein extract, 50 M heme, 2 mg/mL pooled rat liver cytosol, 1 mM MgCl 2 , 3 units glucose-6-dehydrogenase, 1 mM glucose-6-phosphate, and 2 mM NADP ϩ in 0.5 mL 0.1 M potassium phosphate buffer (pH 7.4) for 30 minutes at 37°C. The reaction was stopped by the addition of 2 volumes of chloroform.…”