Toshio Obata

Thyroid hormone-inducible monoamine oxidase inhibitor in rat liver cytosol

An endogenous inhibitor of monoamine oxidase (MAO) was separated by gel-filtration from 105,000 g supernate of T4-treated rat liver cytosol. The inhibition by this inhibitor was concentration-dependent and more potent for A-form MAO than for B-form MAO. The mode of inhibition was competitive either with 5-hydroxytryptamine or beta-phenylethylamine. The molecular weight of this inhibitor was estimated to be 600-700 by gel filtration. The pI value was determined to be 3.0 by isoelectric focusing. This inhibitor was proved to be heat-stable and resistant to protease treatment. MAO inhibition activity was much lower in the cytosol of thyroidectomized, non-T4-treated rats than T4-treated rats, suggesting that this inhibitor is induced by thyroid hormone T4. MAO activity in rat liver might be regulated by the level of this inhibitor.

In vivo monitoring of norepinephrine and hydroxyl free radical generation by ferrous iron in the myocardium with a microdialysis technique

1. We examined in vivo monitoring of norepinephrine and hydroxyl radical generation in rat myocardium with a microdialysis technique. For this purpose, we designed the microdialysis probe holding system which includes loose fixation of the tube and synchronization of the movement of the heart and the probe. 2. The hydroxyl free radical (.OH) reacts with salicylate and generates 2,3- and 2,5-dihydroxybenzoic acid (DHBA) which can be measured electrochemically in picomole quantity by high performance liquid chromatography (HPLC). 3. After probe implantation, norepinephrine concentration of dialysate decreased over the first 150 min and then reached an almost steady level. A positive linear correlation between the ferrous iron and .OH formation trapped as 2,3-DHBA (R2 = 0.960) and 2,5-DHBA (R2 = 0.982) was observed using the microdialysis technique. 4. The present results indicate that non-enzymatic oxidation in the extracellular fluid may play a key role in hydroxyl radical generation by ferrous iron.

Effect of ferrous iron on the generation of hydroxyl free radicals by liver microdialysis perfusion of salicylate

1. We applied in vivo microdialysis technique to examine the effect of Fe2+, Fe3+, Cu2+ and Zn2+ on free radical-generation of rat liver. The hydroxyl free radical (.OH) reacts with salicylate and generates 2,3- and 2,5-dihydroxybenzoic acid (DHBA) which can be measured electrochemically in picomole quantity by HPLC-EC procedure. 2. The relative rates of recovery of 2,3- and 2,5-DHBA at 1 microliter/min in vitro were an average of 10.1 +/- 0.8% and 10.5 +/- 9%, respectively. 3. When the metal ion infused through the dialysis probe, Fe2+ but not Fe3+, Cu2+ and Zn2+ caused an increase in the formation of DHBA of rat liver. 4. After the Ringer solution containing 10 mumole/kg was injected into the penile vein, the levels of 2,3- and 2,5-DHBA were increased about 60% and 40%, respectively. 5. These results indicate that non-enzymatic oxidation in the extracellular fluid may play a key role in Fe2+ generation of -OH in liver. Free radical formation processes may contribute to in vivo free radical formation induced by Fe2+.

モルモットの各種臓器ミトコンドリア中に含まれるType B MAOの分子量について

Molecular weights of mitochondrial type B monoamine oxidase (MAO) in guinea pig brain, liver and kidney were estimated, and their identities and multiplicity were studied. We ascertained what concentration of 3H-pargyline bound to type B MAO specifically from the inhibition curve toward serotonin (5-HT) and beta-phenylethylamine (beta-PEA) by pargyline. Pargyline irreversibly binds to FAD in MAO at a one to one molecular ratio. 3H-pargyline bound to type B MAO specifically and irreversibly by incubation for 5 hr at 37 degrees C, and SDS-disc electrophoresis was carried out using 3H-pargyline as a tracer. The molecular weight of MAO was estimated after specific binding of pargyline was corrected for non-specific binding. The molecular weight of type B MAO in every organ was found to be 60,000, giving a single peak after solubilization with 6% SDS, but several peaks at higher molecular weight were found in each organ after solubilization with 2% SDS. In the brain, there appeared to be a peak of 100,000, and it was suggested that the MAO existed as a dimer which was composed of a FAD containing subunit and a low molecular weight subunit containing no FAD. In the liver, there appeared to be peaks of 120,000 and 240,000, and it was suggested that the MAO existed as a dimer and tetramer. In the kidney, there appeared to be a peak of 180,000, and MAO was suggested to exist as a trimer.