Miyamoto Y, Akaike T, Yoshida M, Goto S, Horie H, Maeda H
Department of Microbiology, Kumamoto University School of Medicine, Japan.
Proc Soc Exp Biol Med. 1996 Apr;211(4):366-73. doi: 10.3181/00379727-211-43982.
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-hydroxpyrazolo[3,4-d]pyrimidine (AHPP) and allopurinol competitively inhibited the conversion of xanthine to uric acid catalyzed by XO, with apparent Ki values of 0.17 +/- 0.02 and 0.50 +/- 0.03 micro M respectively; alloxanthine inhibited this conversion in a noncompetitive manner with an apparent Ki value of 3.54 +/- 1.12 microM. 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 O(2-). 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 micromol/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 micromol/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 mumol/300 g rat). On the basis of these results, it seems that XO regulates EDRF/NO via production of O2-.
一氧化氮（NO）如今几乎与内皮源性舒张因子（EDRF）同义，它与超氧阴离子自由基（O2-）反应，形成一种潜在有毒的分子——过氧亚硝酸盐（ONOO-）。由于黄嘌呤氧化酶（XO）似乎是血管系统中产生O2-的主要酶，因此阐明XO对NO/EDRF的调节机制很重要。我们首先在体外研究了三种吡唑并嘧啶衍生物对XO的抑制作用。动力学研究表明，4-氨基-6-羟基吡唑并[3,4-d]嘧啶（AHPP）和别嘌呤醇竞争性抑制XO催化的黄嘌呤向尿酸的转化，表观Ki值分别为0.17±0.02和0.50±0.03μM；别黄嘌呤以非竞争性方式抑制这种转化，表观Ki值为3.54±1.12μM。通过光泽精依赖性化学发光法测定的黄嘌呤/XO系统中的O2-生成受到AHPP最强烈的剂量依赖性抑制；别嘌呤醇本身似乎通过将电子转移到O2从而产生O(2-)来使该酶失活。AHPP以剂量依赖性方式显著增强了兔和自发性高血压大鼠（SHR）主动脉环中EDRF介导的舒张作用，而别嘌呤醇不影响舒张作用，用别黄嘌呤醇仅观察到血管舒张作用的轻微增强。最后，静脉注射AHPP（50.4mg/kg；100μmol/300g大鼠）使SHR大鼠的血压降至初始血压的70%；这个血压几乎是正常大鼠的血压。别嘌呤醇（100μmol/300g大鼠；静脉注射）显示血压短暂下降，静脉注射别黄嘌呤醇（100μmol/300g大鼠）观察到SHR高血压有中度降低（10%）。基于这些结果，似乎XO通过产生O2-来调节EDRF/NO。
