Lund J, Devereux T, Glaumann H, Gustafsson J A
Department of Medical Nutrition, Karolinska Institute, Huddinge University Hospital, Sweden.
Drug Metab Dispos. 1988 Jul-Aug;16(4):590-9.
Enriched cell populations from rat lung were isolated by use of elutriation. An in vitro ligand binding assay as well as a Western immunoblot assay were used to determine the levels of a binding protein for certain polychlorinated biphenyls (PCBs) in cytosolic preparations from these cell populations. The cell population enriched in Clara cells (30% Clara cells) contained by far the largest amount of the PCB-binding protein, 759 +/- 81 pmol/mg of cytosolic protein as judged by an in vitro ligand binding assay. Western immunoblot analysis of cytosolic preparations from the enriched cell preparations, using antibodies to the PCB-binding protein, showed levels of immunoreactive material in these fractions that corresponded to the level of the PCB-binding protein as determined by the in vitro ligand binding assay. By use of the peroxidase-antiperoxidase method of immunoperoxidase staining, antibodies to the PCB-binding protein were found to stain the Clara cells in sections of paraffin-embedded rat lungs. Intense immunoperoxidase staining of the material lining the airway epithelium was also observed. The protein was predominantly localized to the secretory granules in the apical cytoplasm of the Clara cells as determined using antibodies to the protein, protein A-gold, and electron microscopy. Previous studies have shown a selective in vivo accumulation of methylsulfonyl-PCBs in Clara cells of rodent lung and the present investigation, demonstrating the presence in the Clara cells of a secretory Mr 13,000 protein that binds methylsulfonyl-PCBs with high affinity, gives further support to the contention that the protein is an important factor in determining the in vivo disposition of these compounds.
通过淘析法从大鼠肺中分离出富集细胞群体。采用体外配体结合试验以及蛋白质免疫印迹试验，来测定这些细胞群体胞质制剂中某些多氯联苯（PCB）结合蛋白的水平。富含克拉拉细胞（克拉拉细胞占30%）的细胞群体含有迄今为止数量最多的PCB结合蛋白，通过体外配体结合试验判断，其含量为759±81 pmol/mg胞质蛋白。使用针对PCB结合蛋白的抗体，对富集细胞制剂的胞质制剂进行蛋白质免疫印迹分析，结果显示这些组分中免疫反应性物质的水平，与通过体外配体结合试验测定的PCB结合蛋白水平相对应。采用免疫过氧化物酶染色的过氧化物酶-抗过氧化物酶方法，发现针对PCB结合蛋白的抗体可使石蜡包埋的大鼠肺切片中的克拉拉细胞着色。还观察到气道上皮衬里物质有强烈的免疫过氧化物酶染色。使用针对该蛋白的抗体、蛋白A-金以及电子显微镜确定，该蛋白主要定位于克拉拉细胞顶端细胞质中的分泌颗粒。先前的研究表明，甲基磺酰基-PCBs在啮齿动物肺的克拉拉细胞中有选择性的体内蓄积，而本研究表明克拉拉细胞中存在一种分泌型的分子量为13,000的蛋白，它能以高亲和力结合甲基磺酰基-PCBs，这进一步支持了该蛋白是决定这些化合物体内处置的重要因素这一论点。
