Granzyme and Granulysin Mediated Death of Trypanosoma cruzi

颗粒酶和颗粒溶素介导的克氏锥虫死亡

• 批准号: 9229525
• 负责人: RICARDO TOSTES GAZZINELLI
• 金额: $ 62.65万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-03-01 至 2021-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9229525
• 关键词: Acute; Affect; Apoptosis; Arrhythmia; Bacteria; Blood Transfusion; CD8-Positive T-Lymphocytes; Cardiac; Cardiomyopathies; Cause of Death; Cell Death; Cell membrane; Cells; Cessation of life; Chagas Disease; Cholesterol; Chronic; Cleaved cell; Complex; Country; Cytoplasmic Granules; Cytotoxic T-Lymphocytes; Data; Electron Transport; Enzymes; GZMA gene; Goals; Granzyme; Host resistance; Human; Immigration; Immune; Immune response; Infection; Infiltration; Inflammatory Response; Interferon Type II; Killer Cells; Latin America; Leishmania major; Life; Listeria monocytogenes; Lymphocyte; Mammalian Cell; Mammals; Mediating; Membrane; Modeling; Morphology; Mus; Natural Immunity; Natural Killer Cells; Organ Transplantation; Outcome; Parasites; Pathology; Pathway interactions; Patients; Peptide Hydrolases; Pharmaceutical Preparations; Plasma; Play; Preventive vaccine; Principal Investigator; Process; Proteins; Public Health; Role; Serine Protease; Severity of illness; Superoxides; Surrogate Markers; T-Lymphocyte; Testing; Time; Tissues; Toxic Megacolon; Toxoplasma gondii; Transgenes; Transgenic Mice; Trypanosoma cruzi; VDAC1 gene; Vaccinated; Vaccines; Virulence Factors; Wild Type Mouse; adaptive immunity; base; cell killing; cytotoxic; disorder risk; drug development; fungus; granulysin; human disease; immunological synapse; in vivo; insight; killings; macrophage; neglect; new therapeutic target; novel; pathogen; perforin; prevent; programs; public health relevance; response; transmission process; vaccine development; vaccine trial; vaccine-induced immunity

 DESCRIPTION (provided by applicant): When killer lymphocytes recognize cells infected with intracellular pathogens, they release their cytotoxic granule contents to induce apoptosis of the infected target cell. However, what happens to intracellular parasites during this process is unclear. Host cell apoptosis is triggered by cytotoxic granule proteases (granzymes, Gzm), delivered into the target cell by the cholesterol-dependent membrane perturbing protein, perforin (PFN). Cytotoxic granules of humans and some other mammals, but not rodents, contain another pore-forming protein, granulysin (GNLY), which preferentially disrupts cholesterol-poor bacterial, fungal and parasite membranes. We recently found that when human killer lymphocytes recognize bacteria-infected cells, they rapidly kill intracellular bacteria. GNLY delivers Gzms into bacteria where they proteolytically attack bacterial electron transport chain complex I to generate toxic superoxide anion and also proteolytically destroy bacterial oxidative defense enzymes. Preliminary data show that GNLY also delivers Gzms into protozoan parasites (Trypanosoma cruzi, Toxoplasma gondii, Leishmania major), which are rapidly killed, also in a superoxide-dependent manner. Destruction of intracellular parasites relies on the concerted action of all three cytotoxic effector molecules - Gzms, PFN and GNLY. Mice that express a GNLY transgene in killer lymphocytes are able to survive a T. cruzi challenge that is lethal to wild-type (WT) mice. This proposal uses T. cruzi as a model to investigate the hypothesis that GNLY, Gzm and PFN-mediated killing of intracellular protozoan parasites provides strong immune protection against intracellular parasites. We hypothesize that T. cruzi amastigotes are killed before the host cell, which limits the spread of infection. We also hypothesize that killer cells rapidly induce programs of cell death in T. cruzi that share some features with death pathways activated by Gzms in mammalian cells or bacteria. We also hypothesize that direct parasite killing helps control acute infection, enhance vaccine protection and reduce the chagasic pathology of chronic infection. This proposal joins the expertise of two principal investigators, who are leaders in studying (1) how killer cells destroy infected host cels and their pathogens and (2) the immune response to T. cruzi. The aims of the proposal are to define the features of killer cell-mediated parasite death, identify T. cruzi Gzm substrates that are important in causing death and explore how direct parasite killing contributes to immune defense by comparing infection in WT mice that do not express GNLY with GNLY-transgenic mice.

 描述（由申请人提供）：当杀伤淋巴细胞识别出被细胞内病原体感染的细胞时，它们释放其细胞毒性颗粒内容物以诱导被感染的靶细胞凋亡。然而，在此过程中细胞内寄生虫发生了什么尚不清楚。宿主细胞凋亡由细胞毒性颗粒蛋白酶（颗粒酶，Gzm）触发，由胆固醇依赖性膜扰动蛋白穿孔素（PFN）递送至靶细胞。人类和其他一些哺乳动物（但啮齿动物除外）的细胞毒性颗粒含有另一种成孔蛋白颗粒溶素（GNLY），它会优先破坏缺乏胆固醇的细菌、真菌和寄生虫的细胞膜。我们最近发现，当人类杀伤淋巴细胞识别出细菌感染的细胞时，它们会迅速杀死细胞内的细菌。 GNLY 将 Gzms 传递到细菌中，在那里它们通过蛋白水解攻击细菌电子传递链复合物 I，产生有毒的超氧阴离子，并通过蛋白水解破坏细菌氧化防御酶。初步数据显示，GNLY 还将 Gzms 传递到原生动物寄生虫（克氏锥虫、弓形虫、大型利什曼原虫）中，这些寄生虫也以超氧化物依赖性方式被迅速杀死。细胞内寄生虫的破坏依赖于所有三种细胞毒性效应分子 - Gzms、PFN 和 GNLY 的协同作用。在杀伤淋巴细胞中表达 GNLY 转基因的小鼠能够在对野生型 (WT) 小鼠致命的克氏锥虫攻击中存活下来。该提案使用 T. cruzi 作为模型来研究以下假设：GNLY、Gzm 和 PFN 介导的细胞内原生动物寄生虫杀伤提供了针对细胞内寄生虫的强大免疫保护。我们假设克氏锥虫无鞭毛体在宿主细胞之前被杀死，这限制了感染的传播。我们还假设，克氏锥虫中的杀伤细胞会快速诱导细胞死亡程序，其与哺乳动物细胞或细菌中 Gzms 激活的死亡途径具有一些共同特征。我们还假设直接杀死寄生虫有助于控制急性感染，增强疫苗保护并减少慢性感染的恰加斯病理。该提案结合了两位主要研究人员的专业知识，他们是研究（1）杀伤细胞如何破坏受感染的宿主细胞及其病原体以及（2）对克氏锥虫的免疫反应的领导者。该提案的目的是定义杀伤细胞介导的寄生虫死亡的特征，识别对导致死亡很重要的克氏锥虫 Gzm 底物，并通过比较不表达 GNLY 的 WT 小鼠与 GNLY 转基因小鼠的感染，探索直接杀死寄生虫如何有助于免疫防御。