Iron Acquisition by Mycobacterium tuberculosis Within Phagocytes

吞噬细胞内结核分枝杆菌对铁的获取

• 批准号: 8195964
• 负责人: BRADLEY E BRITIGAN
• 金额: --
• 依托单位: OMAHA VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-01 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8195964
• 关键词: Acute; Anemia due to Chronic Disorder; Antibiotics; Antitubercular Agents; Bacteria; Bone Marrow; Cell membrane; Cells; Cellular biology; Chronic; Citrates; Clinical; Confocal Microscopy; Culture Media; Cytoplasm; Data; Defensins; Dendritic Cells; Development; Disease; Drug resistance in tuberculosis; Enzyme-Linked Immunosorbent Assay; FDA approved; Family; Francisella tularensis; Functional disorder; Gallium; Gene Mutation; Genetic; Growth; Health; Heme; Hepatocyte; Hereditary hemochromatosis; Homeostasis; Host Defense; Human; Hypercalcemia of Malignancy; Immunoblot Analysis; In Vitro; Individual; Infection; Interferons; Interleukin-6; Iron; Kinetics; Lactoferrin; Lead; Life; Link; Liver; Locales; Lung; Measures; Mediating; Metabolic Pathway; Metabolism; Methods; Microbe; Microbial Biofilms; Minority; Mission; Modeling; Movement; Multi-Drug Resistance; Mus; Mycobacterium tuberculosis; Myelogenous; Nature; Organism; Pathogenesis; Patients; Peptides; Phagocytes; Phagosomes; Pharmaceutical Preparations; Physiological; Platelet Factor 4; Play; Predisposition; Prevalence; Prevention strategy; Process; Production; Protein Export Pathway; Proteins; Pseudomonas aeruginosa; Pulmonary Tuberculosis; Research Support; Resistance; Reverse Transcriptase Polymerase Chain Reaction; Role; Route; Serum; Severity of illness; Siderophores; Site; Small Interfering RNA; Source; Spleen; Testing; Therapeutic Agents; Transferrin; Transition Elements; Tuberculosis; Veterans; Virulent; Work; base; cell type; cytokine; extracellular; heme oxygenase-1; hepcidin; high risk; in vivo; insight; macrophage; metal transporting protein 1; microbial; microbicide; novel; novel strategies; pathogen; public health relevance; response; trafficking; treatment strategy; tuberculosis drugs; tuberculosis treatment; uptake

DESCRIPTION (provided by applicant): The pathophysiology of tuberculosis (TB) is linked to the ability of the causative organism, Mycobacterium tuberculosis (M.tb), to grow within human macrophages. Lung myeloid dendritic cells are a newly recognized reservoir of M.tb during lung infection. M.tb metabolism in macrophages and dendritic cells may differ. Iron (Fe) acquisition is critical for M.tb growth. In vivo, most extracellular Fe is chelated to transferrin (TF) and lactoferrin (LF), decreasing microbial access to Fe. We find that M.tb in human macrophages can acquire Fe from TF and LF, as well as the macrophage cytoplasm. Fe uptake from extracellular TF and LF may involve Fe moving into the cytoplasm. Some (e.g. hereditary hemochromatosis) conditions that lower intracellular Fe in macrophages, decrease M.tb Fe uptake. Whether other factors that regulate macrophage Fe metabolism (e.g. ferroportin, heme oxygenase 1 (HO-1), hepcidin) impact M.tb Fe acquisition is unknown. Similarly unknown is whether Fe acquisition by M.tb growing in dendritic cells is different from that occurring in macrophages. We hypothesize that: 1) M.tb acquires Fe from an intracellular Fe pool(s) in macrophages and dendritic cells; 2) acquisition of extracellular Fe by M.tb involves initial Fe movement into this pool; 3) the kinetics/route of M.tb Fe uptake varies with the extracellular Fe chelate and host cell type (macrophage vs. dendritic cell) involved; 4) genetic or physiological factors that modulate macrophage and/or dendritic cell cytoplasmic Fe alter Fe available for use by M.tb; and 4) administration of Ga, which disrupts M.tb Fe-dependent metabolism could prove to be a novel therapy for TB. In order to test these hypotheses we will pursue the following specific aims: 1. Delineate and compare the ability of M.tb residing within human macrophages and myeloid dendritic cells to: A) acquire Fe from physiologic extracellular chelates (e.g. TF and LF) and from cytoplasmic sites in the macrophage and dendritic cell; and B) to what extent Fe acquisition from extracellular chelates involves initial Fe movement into these same cytoplasmic sites. 2. Determine: A) if M.tb modulates macrophage Fe metabolism, which in turn alters Fe acquisition and growth by the bacteria; and B) the role of key components of macrophage Fe metabolism (e.g. ferroportin, HFE, HO-1, hepcidin) on M.tb Fe acquisition and growth. 3. Determine the efficacy of Ga as a therapeutic agent against M.tb using a murine model of M.tb infection. If positive results are seen, further ascertain the mechanism(s) of Ga's anti-tuberculous activity. The proposed studies use human macrophages and dendritic cells along with fully virulent M.tb strains to accomplish these aims. Experimental methods employed include: measuring uptake of 59Fe, siRNA, confocal microscopy, immunoblot analysis, RT-PCR, and ELISA and use of a murine model of pulmonary TB. Insight into M.tb Fe metabolism could lead to novel treatments that would benefit veterans.

描述（由申请人提供）： 结核病 (TB) 的病理生理学与致病微生物结核分枝杆菌 (M.tb) 在人类巨噬细胞内生长的能力有关。肺骨髓树突状细胞是肺部感染期间新发现的结核分枝杆菌储存库。巨噬细胞和树突状细胞中结核分枝杆菌的代谢可能不同。 铁 (Fe) 的获取对于 M.tb 的增长至关重要。在体内，大多数细胞外铁与转铁蛋白 (TF) 和乳铁蛋白 (LF) 螯合，减少微生物对铁的获取。我们发现人巨噬细胞中的M.tb可以从TF和LF以及巨噬细胞细胞质中获取Fe。从细胞外 TF 和 LF 摄取 Fe 可能涉及 Fe 进入细胞质。某些情况（例如遗传性血色素沉着病）会降低巨噬细胞中细胞内铁的含量，从而减少 M.tb 铁的摄取。调节巨噬细胞 Fe 代谢的其他因素（例如铁转运蛋白、血红素加氧酶 1 (HO-1)、铁调素）是否影响 M.tb Fe 的获取尚不清楚。同样未知的是，树突状细胞中生长的 M.tb 对 Fe 的获取是否与巨噬细胞中发生的不同。 我们假设：1) M.tb 从巨噬细胞和树突状细胞的细胞内 Fe 池中获取 Fe； 2) M.tb 获取细胞外 Fe 涉及初始 Fe 移动到该池中； 3) M.tb Fe 吸收的动力学/途径随细胞外 Fe 螯合物和所涉及的宿主细胞类型（巨噬细胞与树突细胞）的不同而变化； 4) 调节巨噬细胞和/或树突状细胞胞质 Fe 改变 M.tb 可用的 Fe 的遗传或生理因素； 4) 给予 Ga 可以扰乱 M.tb Fe 依赖性代谢，可能被证明是一种新的结核病治疗方法。 为了检验这些假设，我们将追求以下具体目标： 1. 描述并比较人类巨噬细胞和骨髓树突状细胞内 M.tb 的能力： A) 从生理细胞外螯合物（例如 TF 和 LF）以及巨噬细胞和树突状细胞的细胞质位点获取 Fe； B) 从细胞外螯合物获取 Fe 在多大程度上涉及初始 Fe 移动到这些相同的细胞质位点。 2. 确定：A) M.tb 是否调节巨噬细胞铁代谢，进而改变细菌对铁的获取和生长； B) 巨噬细胞铁代谢的关键成分（例如铁转运蛋白、HFE、HO-1、铁调素）对 M.tb Fe 获取和生长的作用。 3. 使用 M.tb 感染的小鼠模型确定 Ga 作为抗 M.tb 治疗剂的功效。如果看到积极的结果，进一步确定Ga的抗结核活性的机制。 拟议的研究使用人类巨噬细胞和树突状细胞以及完全有毒的结核分枝杆菌菌株来实现这些目标。采用的实验方法包括：测量 59Fe 的摄取、siRNA、共聚焦显微镜、免疫印迹分析、RT-PCR 和 ELISA 以及使用肺结核小鼠模型。 对 M.tb Fe 代谢的深入了解可能会带来新的治疗方法，使退伍军人受益。