Abstract
Nitric oxide (NO), now almost synonymous with endothelium-derived relaxing factor (EDRF), reacts with superoxide anion radical (O2
-) and forms a potentially toxic molecular species, peroxynitrite (ONOO-). Because xanthine oxidase (XO) seems to be a major O2
--producing enzyme in the vascular system, it is important to clarify the mechanism of XO regulation of NO/EDRF. We first characterized the inhibition of XO in vitro by three types of pyrazolopyrimidine derivatives. Kinetic studies indicated that 4-amino-6-hydroxypyrazolo[3,4-d]pyrimidine (AHPP) and allopurinol competitively inhibited the conversion of xanthine to uric acid catalyzed by XO, with apparent K
i values of 0.17±0.02 and 0.50±0.03 μM, respectively; alloxanthine inhibited this conversion in a noncompetitive manner with an apparent K
i value of 3.54±1.12 μM. O2
- generation in the xanthine/XO system assayed by lucigenin-dependent chemiluminescence was suppressed most strongly by AHPP in a dose-dependent fashion; allopurinol itself appears to reduce the enzyme by transfer of an electron to O2, thus generating O2
-. AHPP significantly augmented EDRF-mediated relaxation of aortic rings from both rabbits and spontaneously hypertensive rats (SHR) in a dose-dependent manner, whereas allopurinol did not affect the relaxation and only marginal potentiation of the vasorelaxation was observed with alloxanthine. Finally, iv injection of AHPP (50.4 mg/kg; 100 μmol/300 g rat) reduced the blood pressure of SHR rats to 70% of the initial pressure; this pressure is almost the blood pressure of normal rats. Allopurinol (100 μmol/300 g rat; iv) showed transient decrease in blood pressure and moderate reduction of hypertension of SHR (10%) was observed with iv injection of alloxanthine (100 μmol/300 g rat). On the basis of these results, it seems that XO regulates EDRF/NO via production of O2
-. [P.S.E.B.M. 1996, Vol 211]
