Studies on the production active oxygens by neutrophils and its regulation.

（1）无细胞系统的研究（i）我们溶解了NADPH氧化酶的膜成分，并通过琼脂糖连接的麦胚凝集素（WGA）层析进行分级。尽管每个级分在无细胞活化系统中没有显示出产生O_2^-的活性，从通过柱的级分中重构膜组分的组分，并用N-前一级分可被磷脂如唑菌素替代。(ii)我们发现Mg^2+似乎在G蛋白水平上调节NADPH氧化酶的激活。(iii)由于细胞色素B_的大亚基的氨基酸序列<558>与先前表征的黄素蛋白的氨基酸序列的比对显示，细胞色素的中间和C-末端部分可能分别是FAD和NADPH结合结构域，细胞色素B_<558>似乎是具有NADPH氧化酶的NADPH结合位点的黄素蛋白。(iv)在无细胞体系中研究了胆红素对NADPH依赖的O_（2”）产生的影响，结果表明胆红素通过降低胞浆组分的活性来抑制NADPH氧化酶的活化过程。(2)（1）在体外研究了碘化甲状腺原氨酸对趋化肽FMLP刺激人中性粒细胞产生O_2 ~-的作用，发现甲状腺素和三碘甲状腺原氨酸干扰FMLP与受体的结合，导致中性粒细胞产生O_2 ~-等功能的抑制。(ii)我们表明，通过使用电透化的人中性粒细胞，cAMP抑制NADPH氧化酶的激活不仅在网站之前的蛋白激酶C，但也在另一个网站后的激酶。

(1)Studies in a cell-free system(i) We solubilized the membrane component of the NADPH oxidase and fractionated by chromatography on agarose-linked wheat germ agglutinin(WGA).Although each fraction showed no potency for the O_2^--generating activity in a cell-free activation system,that of the membrane component was reconstituted from the fraction passing through the column and that eluted with N-acetylglucosamine.The former fraction could be replaced by a phospholipid,such as azolectin.(ii) We found that Mg^<2+> seems to modulate the activation of the NADPH oxidase at the level of the G-protein.(iii) Because the alignments of the amino acid sequence of the large subunit of the cytochrome b_<558> with those of previously characterized flavoproteins revealed that the middle and C-terminal portions of the cytochrome are likely to be FAD-and NADPH-binding domains,respectively, cytochrome b_<558> appears to be a flavoprotein with an NADPH-binding site of the NADPH oxidase.(iv) The effect of bilirubin on the NADPH-dependent O_<2"> production was studied in a cell-free system,and the results indicate that bilirubin inhibits the activation process of the NADPH oxidase by decreasing the potency of the cytosolic fraction.(2) Studies with intact or electropermeabilized neutrophils(i) We studied the action of iodinated thyronines of O_2^- production of human neutrophils stimulated by a chemotactic peptide,FMLP,in vitro and found that thyroxine and tri iodothyronine interfere with the binding of FMLP to the receptor,leading to the inhibition of neutrophil functions such as O_2^- production.(ii) We showed by using electropermeabilized human neutrophils that c AMP inhibited the activation of the NADPH oxidase not only the at the site before the protein kinase C but also at another site after the kinase.