IMMUNOTOXICITY OF POLYCYCLIC AROMATIC HYDROCARBONS

多环芳烃的免疫毒性

• 批准号: 2154124
• 负责人: Scott W Burchiel
• 金额: $ 22.91万
• 依托单位: UNIVERSITY OF NEW MEXICO
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-07-01 至 1997-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2154124
• 关键词: B lymphocyte; CD3 molecule; CD4 molecule; SDS polyacrylamide gel electrophoresis; T lymphocyte; air pollution; antigen receptors; apoptosis; benzanthracenes; biological signal transduction; carbopolycyclic compound; cell adhesion molecules; environmental toxicology; enzyme activity; enzyme inhibitors; flow cytometry; human tissue; hydrocarbons; immunoprecipitation; immunotoxicity; laboratory mouse; leukocyte activation /transformation; phosphorylation; protein kinase; protooncogene; radiotracer; western blottings

Polycyclic aromatic hydrocarbons (PAHs) are important environmental pollutants that inhibit humoral and cell mediated immunity in mice and perhaps in humans. The precise mechanisms responsible for immunosuppression are not known. Previous studies by this laboratory have shown that a methylated PAH (7,12-dimethylbenz(a)anthracene, DMBA) interferes with early events associated with lymphocyte activation in murine B and T cells exposed in vivo and in vitro. Exposure of resting B and T cells (human and mouse) to low concentrations (3muM) of DMBA causes an increase in free intracellular levels of Ca+2 and a significant increase in the phosphorylation of several cell proteins associated with cell activation. This application examines the significance of these events in the immunotoxicity of PAHs. Higher concentrations of DMBA (10-30 muM) continue to increase baseline Ca+2 in lymphocytes, but inhibit phosphorylation in T cells induced by antigen receptor stimulation. One of the primary goals of this application is to study the mechanisms of alteration of Ca+2 and phosphorylation in B and T cells following in vivo or in vitro exposure to methylated (DMBA and 3-methylcholanthrene) and nonmethylated PAHs (benzo(a)pyrene and benz(a)anthracene) compared to nonimmunosuppressive PAHs (benzo(e)pyrene and anthracene). Studies are proposed to examine the mechanism of PAH-induced cell death in lymphocytes. Data is presented that high doses of DMBA produce lymphocytotoxicity via an apoptosis-like mechanism, and that cell to cell interactions (cell density) may play an important role in this process. In vivo studies will be conducted using B6C3F1 mice treated orally by gavage with PAHs daily for 5 days at doses of 5-150 mg/kg. Studies will also be conducted with lymphoid cells obtained from mice, human/murine lymphoid cell lines, and normal human peripheral blood T cells exposed to PAHs in vitro at concentrations of 3-30 muM. The effects of PAHs on phosphorylation of antigen receptor- associated molecules (pp 32,34,35,37 and phospholipase C-gamma) important in B cell activation produced by anti-IgM/IgD will be studied by SDS-PAGE analysis of P-32 labeled digitoninized cells and anti-phosphotyrosine Western blots. the type of phosphorylation of proteins (ser/thr/tyr) will be characterized in whole cell lysates and in specific immunoprecipitated proteins. Parallel studies of anti-CD3/CD4 induced Ca+2 mobilization examined by flow cytometry and phosphorylation of molecules important in T cell activation (CD3, and phospholipase C-gamma) will also be performed. the effects of PAHs on several kinases (PKA, PKC, and PTK, as well as src- related proto-oncogenes) and phosphatases (including CD45) important in cell activation will be studied. Several possible mechanisms will be explored for PAH-induced increases in resting cell Ca+2 levels, including inhibition of Ca+2-ATPase, direct release of stored Ca+2, and indirect stimulation of release by increased IP3 production. The results of these studies are likely to give broad insights into mechanisms of immunotoxicity produced by environmental and other agents that alter Ca+2 homeostasis and signal transduction in lymphoid cells.

多环芳烃（PAHS）是重要的环境 抑制小鼠的体液和细胞介导的免疫力的污染物， 也许在人类中。 负责的确切机制 免疫抑制尚不清楚。 该实验室以前的研究 表明甲基化的PAH（7,12-二甲基苯子（A）蒽，DMBA） 干扰与淋巴细胞激活相关的早期事件 鼠B和T细胞在体内和体外暴露。 休息b的暴露 DMBA的低浓度（3MUM）和T细胞（人和小鼠）原因 自由细胞内Ca+2的提高和显着增加 在与细胞相关的几种细胞蛋白的磷酸化中 激活。 本申请研究了这些事件的重要性 PAH的免疫毒性。 较高浓度的DMBA（10-30 MUM） 继续增加淋巴细胞中的基线Ca+2，但抑制 由抗原受体刺激诱导的T细胞中的磷酸化。 之一 该应用的主要目标是研究 体内B和T细胞中B和T细胞中Ca+2的改变和磷酸化的改变 或体外暴露于甲基化（DMBA和3-甲基胆碱）和 与非甲基化的PAH（苯并（A）pyrene和苯并（A）蒽）相比 非免疫抑制性PAHS（苯并（E）pyrene和anthracene）。 研究是 提议检查淋巴细胞中PAH诱导的细胞死亡的机制。 数据表明，高剂量的DMBA通过A的淋巴细胞毒性 细胞凋亡样机制以及细胞相互作用（细胞密度） 可能在此过程中起重要作用。 体内研究将是 每天使用用PAHS烤口口服的B6C3F1小鼠进行5 剂量为5-150 mg/kg的天数。 研究还将用淋巴样进行 从小鼠，人/鼠淋巴样细胞系获得的细胞，正常 在浓度下暴露于PAHS的人外周血T细胞 3-30妈妈。 PAHS对抗原受体磷酸化的影响 - 相关分子（PP 32,34,35,37和磷脂酶C-Gamma） 在由抗IGM/IgD产生的B细胞激活中，将通过SDS-PAGE研究 P-32标记的数字量化细胞和抗磷酸酪氨酸的分析 蛋白质印迹。 蛋白质（Ser/Thr/Tyr）的磷酸化类型将 在全细胞裂解物和特定免疫沉淀中的特征 蛋白质。 抗CD3/CD4诱导CA+2动员的平行研究 通过流式细胞仪和T中很重要 还将进行细胞活化（CD3和磷脂酶C-Gamma）。 PAH对几种激酶（PKA，PKC和PTK以及SRC-）的影响 相关的原始基因）和磷酸酶（包括CD45） 将研究细胞激活。 将有几种可能的机制 探索了PAH引起的静息细胞Ca+2级别的增加，包括 抑制Ca+2-ATPase，直接释放储存的Ca+2和间接 通过增加的IP3产生来刺激释放。 这些结果 研究可能会对免疫毒性机制提供广泛的见解。 由环境和其他改变CA+2稳态的代理商生产的 淋巴样细胞中的信号转导。