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: 7685175
• 负责人: BRADLEY E BRITIGAN
• 金额: --
• 依托单位: CINCINNATI VA MEDICAL CENTER RESEARCH
• 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 2013-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7685175
• 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. PUBLIC HEALTH RELEVANCE: Project Narrative - Relevance to VA Mission: Veterans are at high risk for pulmonary TB. There is a growing problem in treating tuberculosis in that the causative organism, Myocobacterium tuberculosis, is becoming increasingly resistant to available antibiotics. The ability to acquire iron (Fe) from the infected host is required for M. tuberculosis to be able to grow and therefore cause infection. Insight into M. tuberculosis Fe metabolism could lead to development of novel approaches to interfere with the organism's ability to acquire Fe treatments that could lead to the development of new kinds of antibiotics. This application will explore one potential mechanism of doing this using gallium. Thus, increased understanding of how M. tuberculosis acquires and uses Fe during infection could lead to the development of new antibiotics to be used in treating TB, something that would be of great benefit to 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。肺髓样树突状细胞是肺感染过程中结核分枝杆菌的新认识的储库。巨噬细胞和树突状细胞中的结核分枝杆菌代谢可能不同。 铁（Fe）的获得对于结核分枝杆菌生长至关重要。在体内，大多数细胞外铁螯合转铁蛋白（TF）和乳铁蛋白（LF），减少微生物获得铁。我们发现结核分枝杆菌在人巨噬细胞中可以从TF和LF以及巨噬细胞胞质中获得铁。从细胞外TF和LF摄取Fe可能涉及Fe移动到细胞质中。一些（如遗传性血色病）条件，降低细胞内铁在巨噬细胞，减少结核分枝杆菌铁摄取。调节巨噬细胞铁代谢的其他因素（例如，膜铁转运蛋白、血红素加氧酶1（HO-1）、铁调素）是否影响结核分枝杆菌铁的获取尚不清楚。同样未知的是，在树突状细胞中生长的结核分枝杆菌获得的铁是否与巨噬细胞中发生的不同。 我们假设：1）结核分枝杆菌从巨噬细胞和树突状细胞中的细胞内铁池获得铁; 2）结核分枝杆菌获得细胞外铁涉及初始铁移动到该池中; 3）结核分枝杆菌铁摄取的动力学/途径随细胞外铁螯合物和宿主细胞类型而变化（巨噬细胞与树突状细胞）参与; 4）调节巨噬细胞和/或树突状细胞胞质Fe的遗传或生理因素改变了结核分枝杆菌可利用的Fe;和4）施用Ga，其破坏结核分枝杆菌Fe依赖性代谢，可以证明是用于TB的新疗法。 为了检验这些假设，我们将追求以下具体目标：1。描述并比较驻留在人巨噬细胞和髓样树突细胞内的M. t B：A）从生理性细胞外螯合物（例如TF和LF）以及从巨噬细胞和树突细胞中的细胞质位点获得Fe的能力;和B）从细胞外螯合物获得Fe涉及初始Fe移动到这些相同细胞质位点的程度。2.确定：A）M. tB是否调节巨噬细胞Fe代谢，这反过来改变细菌的Fe获取和生长;和B）巨噬细胞Fe代谢的关键组分（例如膜铁转运蛋白、HFE、HO-1、铁调素）对M. tB b Fe获取和生长的作用。3.使用结核分枝杆菌感染的鼠模型确定Ga作为针对结核分枝杆菌的治疗剂的功效。如果观察到阳性结果，则进一步确定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沿着完全毒性的结核分枝杆菌菌株来实现这些目标。所采用的实验方法包括：测量59 Fe、siRNA、共聚焦显微镜、免疫印迹分析、RT-PCR和ELISA的摄取以及肺TB的小鼠模型的使用。 深入了解结核分枝杆菌的铁代谢可能会导致新的治疗方法，这将有利于退伍军人。 公共卫生关系： 项目叙述-与VA使命的相关性：退伍军人患肺结核的风险较高。在治疗结核病方面存在日益严重的问题，因为病原体结核分枝杆菌对可用的抗生素的耐药性越来越强。M需要从感染的宿主获得铁（Fe）的能力。结核病能够生长并因此引起感染。对M.结核病铁代谢可能导致开发新的方法来干扰生物体获得铁治疗的能力，这可能导致开发新型抗生素。本申请将探索使用镓实现这一点的一种潜在机制。因此，增加了对M。结核病在感染过程中获得和使用铁可能会导致新抗生素的开发，用于治疗结核病，这将对退伍军人大有好处。