MECHANISMS OF CELL REPLICATION IN ASBESTOS CANCERS

本项目将重点研究青石棉的分子机制， 和温石棉诱发小鼠胸膜增生 间皮细胞（RPM）和气管支气管上皮细胞（HTE）， 间皮瘤和支气管癌的祖细胞。 以来 增加的细胞增殖与以下因素有因果关系和/或促成以下因素： 许多癌症，我们假设石棉纤维提供了一个持久的 有丝分裂刺激是肿瘤发展所必需的， 石棉对c-fos和c-jun原癌基因的持续诱导。 到 确定青石棉和温石棉是否通过类似的 或独特的过程中的各种细胞类型，我们将研究的机制， 使用抑制剂激活c-fos和c-jun基因表达 蛋白激酶C（PKC）和ADP-核糖转移酶。 活性物质清除剂 氧物种（AOS）和铁螯合纤维将被用来测试 假设石棉相关c-fos和c-jun诱导可能 由AOS介导。 此外，使用非石棉的比较实验 纤维[耐火陶瓷（RCF）和玻璃（MMVF-10）]， 石棉尺寸，长尺寸（>10微米）与短尺寸（<2微米） 温石棉纤维和类似化学品的颗粒（钠闪石、玻璃） 在一定浓度范围内的石棉组合物将决定 诱导c-fos和c-jun所固有的纤维特性。无论 这些原癌基因表达的增加与细胞凋亡有因果关系。 石棉的扩散将使用瞬态和瞬态两种方法进行评估。 评估青石棉和温石棉影响的转染试验 纤维对HTE细胞中AP-1依赖性报告基因表达的影响 c-fos和c-jun蛋白在细胞培养物中的免疫定位 接触石棉。 在后面的实验中，我们将使用 免疫细胞化学以确定c-fos和c-jun蛋白是否 仅在增生的间皮和气管上皮中表达 通过增殖细胞的表达鉴定的上皮细胞 核抗原（PCNA）或掺入5 ′-溴脱氧尿苷 （BrdU）转化为DNA。 使用暴露于以下物质后的细胞的比较实验： 体外纤维的增殖和细胞毒性浓度， 大鼠吸入苯妥英钠后c-fos和c-jun蛋白的原位定位 我们可以将这些基因产物的表达 细胞增殖和/或细胞毒性。 机制知识 石棉对石棉靶细胞c-fos和c-jun的诱导作用 诱发的癌症将为预防和治疗开辟途径 接近疾病。 该项目涉及博士之间的合作。 莫斯曼，一位在石棉和AOS方面有专长的细胞生物学家，海因茨博士， 一位在细胞复制方面有专长的分子生物学家，莱斯利博士， 一位擅长免疫细胞化学的肺部病理学家

This project will focus on the molecular mechanisms by which crocidolite and chrysotile asbestos induce proliferation of rodent pleural mesothelial (RPM) and tracheobronchial epithelial (HTE) cells, the progenitor cells of mesotheliomas and bronchogenic carcinomas. Since increased cell proliferation is causally related and/or contributory to many cancers, we hypothesize that asbestos fibers provide a persistent mitogenic stimulus necessary for tumor development as is suggested by the persistent induction of c-fos and c-jun protooncogenes by asbestos. To determine if crocidolite and chrysotile induce proliferation by similar or unique processes in various cell types, we will examine the mechanisms of activation of c-fos and c-jun gene expression using inhibitors of protein kinase C (PKC) and ADP-ribose transferase. Scavengers of active oxygen species (AOS) and iron-chelated fibers will be used to test the hypothesis that asbestos-associated induction of c-fos and c-jun may be mediated by AOS. Moreover, comparative experiments using nonasbestos fibers [refractory ceramic (RCF) and glass (MMVF-10)] of similar dimension to asbestos, sized long (>10 microm) vs. short (<2 micro) chrysotile fibers, and particles (riebeckite, glass) of similar chemical composition to asbestos at a range of concentrations will determine the properties of fibers intrinsic to induction of c-fos and c-jun. Whether increased expression of these protooncogenes is causally related to cell proliferation by asbestos will be assessed using both transient transfection assays to assess the effects of crocidolite and chrysotile fibers on the expression of an AP-1 dependent reporter gene in HTE cells and immunolocalization of c-fos and c-jun proteins in cell cultures exposed to asbestos. In the latter experiments, we will use immunocytochemistry to determine if c-fos and c-jun proteins are expressed exclusively in proliferating mesothelial and tracheal epithelial cells as identified by the expression of proliferating cell nuclear antigen (PCNA) or the incorporation of 5'-bromo-deoxyuridine (BrdU) into DNA. Comparative experiments using cells after exposure to both proliferative and cytotoxic concentrations of fibers in vitro and in situ localization of c-fos and c-jun proteins after inhalation of fibers by rats will allow us to link expression of these gene products with cell proliferation and/or cytotoxicity. Knowledge of the mechanisms of induction of c-fos and c-jun by asbestos in target cells of asbesto- induced cancers will open up avenues for preventive and therapeutic approaches to disease. This project involves collaborations between Dr. Mossman, a cell biologist with expertise in asbestos and AOS, Dr. Heintz, a molecular biologist with expertise in cell replication, and Dr. Leslie, a lung pathologist with expertise in immunocytochemistry.