Functional expression of NADPH oxidase components (alpha- and beta-subunits of cytochrome b558 and 45-kDa flavoprotein) by intrinsic human glomerular mesangial cells.
Functional expression of NADPH oxidase components (alpha- and beta-subunits of cytochrome b558 and 45-kDa flavoprotein) by intrinsic human glomerular mesangial cells.
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NADPH 氧化酶成分（细胞色素 b558 的 α 和 β 亚基和 45-kDa 黄素蛋白）通过内在人肾小球系膜细胞的功能表达。
DOI：
10.1016/s0021-9258(18)54815-8
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发表时间：
1991
期刊：
影响因子：
--
通讯作者：
Klaus Resch
中科院分区：
文献类型：
--
作者：
H. Radeke;A. Cross;J. Hancock;O. Jones;Masaaki Nakamura;V. Kaever;Klaus Resch
最近，我们已经表明，人肾小球系膜细胞（HMCs）释放氧自由基从质膜响应细胞因子。现在，我们已经使用二苯碘鎓，一种共价结合抑制剂的活化45-kDa黄素蛋白，在中性粒细胞放射性标记125 I，并可以确定一个45-kDa的蛋白带中分离的HMC质膜部分。低温差光谱显示在428和558 nm处的峰吸收。HMC膜的直接电位测定法（-340至-160 mV）显示存在低电位细胞色素（76 pmol/mg HMC膜蛋白），鉴定为细胞色素b558。在狭缝印迹中，对α亚基胞外域具有特异性的小鼠单克隆抗体（mAb）7 D 5与HMC呈阳性反应。在蛋白质印迹中，针对α-亚基的细胞质表位的mAb 449鉴定了23-kDa蛋白质;和针对人嗜中性粒细胞的细胞色素b558的大（β）亚基的mAb 48（Verhoeven，A. J.，Bolscher，B. G. J.M.，米尔霍夫湖J.，货车茨维滕河，Keijer，J.，韦宁河美国，和鲁什，D.（1989）Blood 73，1686-1694），在变性的HMC膜蛋白中检测到75 - 100 kDa的涂片。这些数据确定与HMCs，首次提出的NADPH：O2-氧化还原酶在间充质细胞中的三个基本组成部分的表达。
Recently, we have shown that human glomerular mesangial cells (HMCs) release oxygen radicals from the plasma membrane in response to cytokines. Now we have used diphenylene iodonium, a covalent binding inhibitor of activated 45-kDa flavoprotein, in neutrophils radiolabeled with 125I and could identify a 45-kDa protein band in a separated HMC plasma membrane fraction. Low temperature difference spectroscopy showed a peak absorbance at 428 and 558 nm. Direct potentiometry of HMC membranes (-340 to -160 mV) showed the presence of a low potential cytochrome (76 pmol/mg to HMC membrane protein) identified as cytochrome b558. In slot blots, mouse monoclonal antibody (mAb) 7D5, specific for the extracellular domain of the alpha-subunit, showed a positive reaction with HMCs. In Western blots, mAb 449, directed against the cytoplasmic epitope of the alpha-subunit, identified a 23-kDa protein; and mAb 48, raised against the large (beta) subunit of cytochrome b558 of human neutrophils (Verhoeven, A. J., Bolscher, B. G. J. M., Meerhof, L. J., van Zwieten, R., Keijer, J., Weening, R. S., and Roos, D. (1989) Blood 73, 1686-1694), detected a smear between 75 and 100 kDa in denatured HMC membrane protein. These data determined with HMCs, suggest for the first time the expression of three essential components of NADPH:O2- oxidoreductase in mesenchymal cells.