Cytochemical localization of hydrogen peroxide production in the rat uterus

A reduced nicotinamide adenine dinucleotide phosphate (NAD(P)H)-dependent H2O2-generating activity of the rat uterus was investigated both electron cytochemically and biochemically. We tried to cytochemically demonstrate H2O2 generation from the oxidation of reduced NADH or NADPH using the cerium method. NADPH oxidation resulted in electron-dense deposits on the apical plasma membrane covering the microvilli of the surface epithelium of the lightly fixed endometrium. In control specimens incubated in a medium from which substrate was omitted, no such deposits were observed. The reduction of ferricytochrome c due to NADH oxidation was spectrophotometrically detected in the lightly fixed uterus. Absorption at 550 nm increased with the addition of NADH, but not with that of NAD. The reaction was weakened by preheating and adversely affected by the addition of superoxide dismutase, but it was not inhibited by adding 50 mM sodium azide. These results suggest that a kind of NAD(P)H oxidase, generating H2O2 via superoxide formation, may possibly be present on the apical plasma membrane of the rat endometrial epithelium.