Involvement of a local Fenton reaction in the reciprocal modulation by O2 of the glucagon-dependent activation of the phosphoenolpyruvate carboxykinase gene and the insulin-dependent activation of the glucokinase gene in rat hepatocytes

H₂O₂ mimicked the action of periportal pO₂ in the modulation by O₂ of the glucagon-dependent activation of the phosphoenolpyruvate carboxykinase (PCK) gene and the insulin-dependent activation of the glucokinase (GK) gene. H₂O₂ can be converted in the presence of Fe²⁺ in a Fenton reaction into hydroxyl anions and hydroxyl radicals (^•OH). The hydroxyl radicals are highly reactive and might interfere locally with transcription factors. It was the aim of the present study to investigate the role of and to localize such a Fenton reaction. Hepatocytes cultured for 24 h were treated under conditions mimicking periportal or perivenous pO₂ with glucagon or insulin plus the iron chelator desferrioxamine (DSF) or the hydroxyl radical scavenger dimethylthiourea (DMTU) to inhibit the Fenton reaction. PCK mRNA was induced by glucagon maximally under conditions of periportal pO₂ and half-maximally under venous pO₂. GK mRNA was induced by insulin with reciprocal modulation by O₂. DSF and DMTU reduced the induction of PCK mRNA to about half-maximal and increased the induction of GK mRNA to maximal under both O₂ tensions. Hydroxyl radical formation was maximal under arterial pO₂. Perivenous pO₂, DSF and DMTU each decreased the formation of ^•OH to about 70% of control. The Fenton reaction could be localized in a perinuclear space by confocal laser microscopy and three-dimensional reconstruction techniques. In the same compartment, iron could be detected by electron-probe X-ray microanalysis. Thus a local Fenton reaction is involved in the O₂ signalling, which modulated the glucagon- and insulin-dependent PCK gene and GK gene activation.