EHRLICHIACIDAL MECHANISM BY MACROPHAGE CA++ MOBILIZATION

巨噬细胞 CA 动员的杀埃里希酸机制

• 批准号: 3145061
• 负责人: YASUKO RIKIHISA
• 金额: $ 16.47万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-08-01 至 1994-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3145061
• 关键词: 3'5' cyclic nucleotide phosphodiesterase; Rickettsia; Rickettsiales disease; calcium channel; calcium channel blockers; calcium flux; calmodulin; cyclic AMP; cyclic GMP; enzyme inhibitors; granulocyte; lysosomes; macrophage; membrane fusion; microfilaments; microtubules; phosphodiesterase inhibitors; phospholipase A2; phospholipase inhibitor; protein kinase C; vesicle /vacuole

Ehrlichiae cause diseases such as human ehrlichiosis and sennetsu ehrlichiosis. Ehrlichiae are a unique group of rickettsiae and are obligate intracellular parasites of macrophages or granulocytes. They multiply in the membrane-bound vacuoles which do not fuse with lysosomes. The goal of this project is to find whether macrophage activating agents kill ehrlichiae and if so how ehrlichiae can be completely eliminated from the host cells. We will test the following specific hypotheses. Hypothesis 1: Ehrlichial infection blocks Ca2+ influx into the macrophage, thus the agents which can mobilize Ca 2+, in infected macrophages kill ehrlichiae. AIM 1: Tests hypothesis 1: a) by examining dose- and time- responses of macrophages in eliminating ehrlichiae in response to these agents, b) by measuring intracellular Ca2+ ion, and c) by examining the effects of Ca2+ channel blockers or Ca2+ antagonists. Hypothesis 2: Ehrlichial killing induced by Ca2+ mobilization is mediated by protein kinase C, calmodulin, phospholipase A2 and/or cAMP, or cGMP. AIM 2: Tests hypothesis 2 by a) evaluating the effects of protein kinase C inhibitors, calmodulin antagonists, Ca2+-dependent phospholipase A2 inhibitors, and agents which increase cAMP and cGMP on the ehrlichiacidal effects of Ca 2+ mobilizing agents, and b) measuring phosphodiesterase, protein kinase C, phospholipase A2 activity and/or cAMP and cGMP. Hypothesis 3: Ca2+ mobilization in the macrophage destroys ehrlichiae by initiation of lysosomal fusion with ehrlichia-containing vacuoles which is accompanied by increased polymerization and reorganization of tubulin and actin in the infected macrophage cytoplasm. AIM 3: Tests hypothesis 3 by a) examining the fusion of lysosomes labeled with electron densetracers or acid phosphatase cytochemistry with ehrlichiae-containing vacuoles, by b) fluorescent labeling of microfilaments and microtubules, and by c) examining the effect of microtubule and microfilament disrupting agents.

埃立克体引起的疾病，如人类埃立克体病和sennetsu 埃立克体病埃立克体是立克次体的一个独特类群， 巨噬细胞或粒细胞的细胞内寄生物。它们繁殖在 不与溶酶体融合的膜泡。的目标 这个项目是为了发现巨噬细胞激活剂是否能杀死 埃里希体，如果是这样，埃里希体是如何从 宿主细胞我们将检验以下具体假设。 假设1：埃立克体感染阻断了Ca2+流入巨噬细胞， 因此，在感染的巨噬细胞中， 埃里希体目的1：检验假设1：a）通过检查剂量和时间- 巨噬细胞在消除埃立克体中的反应 B）通过测量细胞内Ca2+离子，和c）通过检查细胞内Ca2+离子， Ca2+通道阻断剂或Ca2+拮抗剂的作用。 假设2：由Ca2+动员诱导的埃里希体杀伤是介导的 通过蛋白激酶C、钙调蛋白、磷脂酶A2和/或cAMP或cGMP。目的 2：通过a）评估蛋白激酶C的作用来检验假设2 抑制剂、钙调素拮抗剂、Ca 2+依赖性磷脂酶A2 抑制剂和增加杀埃里希体作用的cAMP和cGMP的药剂。 Ca 2+动员剂的作用，和B）测量磷酸二酯酶， 蛋白激酶C、磷脂酶A2活性和/或cAMP和cGMP。 假设3：巨噬细胞中的Ca 2+动员通过以下方式破坏埃立克体： 启动溶酶体与含有埃立克体的空泡融合， 伴随着微管蛋白聚合和重组的增加， 感染的巨噬细胞胞质中的肌动蛋白。目的3：通过a）检验假设3 检查用电子密度示踪剂标记的溶酶体的融合，或 酸性磷酸酶细胞化学，含埃立克体空泡，通过B） 微丝和微管的荧光标记，以及c） 检查微管和微丝破坏剂的作用。