Mechanistic study of the host defense functions of IL-32 in tuberculosis

IL-32在结核病中的宿主防御功能机制研究

• 批准号: 8244621
• 负责人: EDWARD D CHAN
• 金额: --
• 依托单位: VA EASTERN COLORADO HEALTH CARE SYSTEM
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-10-01 至 2015-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8244621
• 关键词: Actins; Address; Aerosols; Affect; Alveolar Macrophages; Animal Model; Antitubercular Agents; Apoptosis; Bacillus (bacterium); C57BL/6 Mouse; Caspase; Caspase-1; Cathepsins; Cause of Death; Cell Death; Cells; Clinical; Communicable Diseases; Complex; Country; Cytoplasm; Development; Disease; Drug Resistant Tuberculosis; Drug resistance; Effectiveness; Evolution; Exhibits; Future; Genes; Genus Mycobacterium; Goals; Growth; Gulf War; Health; Health Resources; Homologous Gene; Host Defense; Host Defense Mechanism; Host resistance; Human; Immune; Immune response; Immunity; Incubated; Individual; Infection; Infectious Agent; Inflammatory; Interferon Type II; Interferons; Interleukins; Investigation; Knock-in Mouse; Knowledge; Korean War; Lead; Leukocytes; Lung; Mediating; Methods; Mission; Modeling; Mouse Strains; Mus; Mycobacterium Infections; Mycobacterium tuberculosis; Mycobacterium tuberculosis H37Rv; Outcome; Pathway interactions; Patients; Phenotype; Play; Process; Proteins; Pulmonary Surfactant-Associated Protein C; Recombinants; Resistance; Risk; Role; Signal Transduction; Spleen; TNF gene; Testing; Time; Tissues; Transgenic Mice; Transgenic Model; Transgenic Organisms; Travel; Tuberculosis; Tumor Necrosis Factor-alpha; United States; Vaccine Therapy; Veterans; Vietnam; Virulent; War; Wild Type Mouse; World War II; actin 2; apoptosis inducing factor; base; caspase-3; clinically relevant; cytokine; design; efficacy testing; improved; in vivo; innovation; interest; killings; macrophage; microbial; mouse model; mycobacterial; pathogen; promoter; protective effect; research study

DESCRIPTION (provided by applicant): The goal of this project is to elucidate the mechanisms by which interleukin-32 (IL-32) enhances immunity against Mycobacterium tuberculosis (MTB). A comprehensive understanding of IL-32 action will not only advance our knowledge of the host-protective immune response to tuberculosis (TB), but also drive future investigations designed to enrich vaccine therapy and cultivate innovative, immune-modifying treatments. IL-32 is a recently described cytokine. We have shown that IL-32 is induced by MTB infection and that it protects host cells against MTB. We discovered that IL-32 inhibits intracellular growth of MTB in THP-1 macrophages through caspase-3-dependent apoptosis and yet-to-be identified caspase-3-independent cell death pathways. While IL-32 plays a potentially critical role in controlling TB, important questions remain. We hypothesize that IL-32 reduces viability of intracellular MTB by inducing macrophages to undergo various types of programmed cell death (PCD), including caspase-3-dependent and caspase-independent apoptosis as well as pyroptosis, a unique form of inflammatory PCD mediated by caspase-1 and inflammasome complexes. Both apoptosis and pyroptosis are known killing mechanisms of intracellular pathogens including mycobacteria. While a mouse homolog of IL-32 has not been found, murine macrophages are activated by human IL-32. RAW 264.7 mouse macrophages incubated with IL-32 have a lower burden of intracellular MTB. To study the anti-TB effect of IL-32 in vivo, we generated transgenic mice that express IL-323 in the lungs under the control of the SPC promoter (SPC-IL-323Tg). Aerosol infection with the hypervirulent MTB W-Beijing HN878 showed that the SPC-IL-323Tg mice have reduced bacterial burden in the lungs and spleen, a more protective immune phenotype, and increased survival compared to wild type (WT) C57BL/6 mice. Recently, our collaborators developed a Tg mouse strain in which IL-323 is under the control of the 2-actin gene promoter. IL-32 is widely expressed in the tissues of these mice, including the lung, spleen, and white blood cells. To begin to address the clinical relevance of IL-32 in humans, we will test the efficacy of IL-32 against a virulent clinical strain of MTB in primary human alveolar macrophages (AM). We propose to more fully describe the mechanisms by which IL-32 kills intracellular MTB as well as the complex immunological pathways affected by IL-32 through the following three aims. Aim 1. Elucidate how programmed cell death pathways contribute to IL-32 induced inhibition of intracellular MTB. We hypothesize that caspase-3-independent apoptosis and caspase-1-mediated pyroptosis are additional mechanisms by which IL-32 exerts its anti-TB effects. Aim 2. Determine how IL-32 transgenic mice are protected against MTB infection. We hypothesize that SPC-IL-323Tg mice will have sustained resistance to MTB at longer times of infection, 2-actin-IL-323Tg mice will also resist MTB infection, and both knock-in models will survive longer and exhibit a more protective immune response with greater induction of PCD in the AM than infected WT mice. Aim 3. Define the anti-mycobacterial activity of IL-32 in primary human cells and determine whether the anti-TB effects of IL-32 can be enhanced. We hypothesize that IL-32 will antagonize intracellular MTB in primary human AM due to its ability to induce caspase-3-dependent apoptosis and caspase-1-mediated pyroptosis. Since W-Beijing strains are less likely to induce host cell apoptosis, we hypothesize that IL-32 will be more efficient in killing MTB H37Rv than W-Beijing HN878. Additionally, enhancing the bioactivity of IL-32 should increase induction of PCD and MTB killing.

描述（由申请人提供）： 该项目的目标是阐明白细胞介素 32 (IL-32) 增强对结核分枝杆菌 (MTB) 免疫力的机制。对 IL-32 作用的全面了解不仅将增进我们对结核病 (TB) 的宿主保护性免疫反应的了解，而且还将推动旨在丰富疫苗疗法和培育创新的免疫修饰疗法的未来研究。 IL-32 是最近描述的一种细胞因子。我们已经证明 IL-32 是由 MTB 感染诱导的，并且它可以保护宿主细胞免受 MTB 的侵害。我们发现 IL-32 通过 caspase-3 依赖性细胞凋亡和尚未确定的 caspase-3 独立细胞死亡途径抑制 THP-1 巨噬细胞中 MTB 的细胞内生长。虽然 IL-32 在控制结核病方面发挥着潜在的关键作用，但重要的问题仍然存在。我们假设 IL-32 通过诱导巨噬细胞经历各种类型的程序性细胞死亡 (PCD) 来降低细胞内 MTB 的活力，包括 caspase-3 依赖性和 caspase 非依赖性细胞凋亡以及细胞焦亡（一种由 caspase-1 和炎性体复合物介导的独特形式的炎症性 PCD）。细胞凋亡和焦亡都是已知的细胞内病原体（包括分枝杆菌）的杀死机制。 虽然尚未发现 IL-32 的小鼠同源物，但小鼠巨噬细胞可被人 IL-32 激活。与 IL-32 一起孵育的 RAW 264.7 小鼠巨噬细胞具有较低的细胞内 MTB 负担。为了研究 IL-32 体内的抗结核作用，我们培育了在 SPC 启动子 (SPC-IL-323Tg) 控制下在肺部表达 IL-323 的转基因小鼠。高毒力 MTB W-Beijing HN878 的气溶胶感染表明，与野生型 (WT) C57BL/6 小鼠相比，SPC-IL-323Tg 小鼠肺和脾脏中的细菌负担减少，免疫表型更具保护性，并且存活率提高。最近，我们的合作者开发了一种 Tg 小鼠品系，其中 IL-323 受到 2-肌动蛋白基因启动子的控制。 IL-32 在这些小鼠的组织中广泛表达，包括肺、脾和白细胞。 为了开始解决 IL-32 在人类中的临床相关性，我们将在原代人肺泡巨噬细胞 (AM) 中测试 IL-32 对 MTB 临床毒株的功效。我们建议通过以下三个目标更全面地描述 IL-32 杀死细胞内 MTB 的机制以及受 IL-32 影响的复杂免疫途径。目标 1. 阐明程序性细胞死亡途径如何促进 IL-32 诱导的细胞内 MTB 抑制。我们假设 caspase-3 独立的细胞凋亡和 caspase-1 介导的细胞焦亡是 IL-32 发挥其抗结核作用的额外机制。目标 2. 确定 IL-32 转基因小鼠如何免受 MTB 感染。我们假设 SPC-IL-323Tg 小鼠在较长的感染时间内对 MTB 具有持续的抵抗力，2-actin-IL-323Tg 小鼠也能抵抗 MTB 感染，并且两种敲入模型都将比感染的 WT 小鼠存活更长时间并表现出更强的保护性免疫反应，AM 中 PCD 的诱导更强。目标 3. 确定 IL-32 在原代人细胞中的抗分枝杆菌活性，并确定 IL-32 的抗结核作用是否可以增强。我们假设 IL-32 会拮抗原代人 AM 中的细胞内 MTB，因为它能够诱导 caspase-3 依赖性细胞凋亡和 caspase-1 介导的细胞焦亡。由于W-Beijing菌株不太可能诱导宿主细胞凋亡，因此我们假设IL-32比W-Beijing HN878更有效地杀死MTB H37Rv。此外，增强 IL-32 的生物活性应该会增加 PCD 和 MTB 杀灭的诱导。