Immunopathology of the Pneumocystis Life Cycle Stages

肺孢子虫生命周期阶段的免疫病理学

• 批准号: 8397516
• 负责人: Melanie T Cushion
• 金额: --
• 依托单位: CINCINNATI VA MEDICAL CENTER RESEARCH
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-01 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8397516
• 关键词: 3-Dimensional; Aging; Alveolar Macrophages; Anidulafungin; Animals; Binding; Biological Assay; CD8B1 gene; Cell Wall; Chronic Disease; Chronic Obstructive Airway Disease; Clinical; Comorbidity; Complication; Cyst; Development; Ensure; Equilibrium; Gene Expression; Glucans; Goals; Growth; HIV; Human; Immune; Immune response; Immune system; In Vitro; Infection; Infectious Agent; Inflammatory Response; Leukocytes; Life; Life Cycle Stages; Lung; Malignant neoplasm of lung; Mammals; Molecular; Molecular Profiling; Mus; Mycoses; Natural History; Opportunistic Infections; Organism; Partner in relationship; Pathogenesis; Patients; Persons; Phagocytosis; Phase; Pneumocystis; Pneumocystis Infections; Pneumocystis carinii Pneumonia; Pneumonia; Population; Rattus; Registries; Reproduction spores; Research Personnel; Risk; Role; Route; Series; Severity of illness; Silver Methenamine; Staging; Staining method; Stains; System; T-Lymphocyte; Thick; Travel; Veterans; asexual; beta-1,3-D-glucan; cytokine; effective therapy; fungus; immunopathology; immunosuppressed; in vivo; infected vector rodent; inhibitor/antagonist; insight; mortality; receptor; reconstitution; response; tool; transmission process

DESCRIPTION (provided by applicant): Fungal organisms in the genus Pneumocystis, were first clinically noticed because they caused an oftentimes lethal pneumonia (PCP) in humans and other mammals with compromised immune status. Today, the niche of these fungi have expanded to include patients with underlying chronic diseases like COPD, where their presence has been suggested to be a co-morbidity factor. Limited progress has been made in understanding the life cycle, transmission, and natural history of Pneumocystis, due in large part to the absence of a continuous in vitro culture system. The strategies used by these organisms to grow and survive in the context of an intact or immune-debilitated host are largely unknown. The actual agent of infection, the transmissive form, has not been identified, nor has its mode of travel, generally thought to be via an airborne spore. The life cycle of Pneumocystis involves trophic forms (trophs), thought to be the vegetative, asexually dividing life cycle stage; as well as several other developmental stages including the cyst form, which contains 8 ascospores. The cyst is distinct from the trophs in that it contains a thick cell wall comprised mostly of B-1,3 glucan; is larger (5-8um vs 1-4um); and stains with the fungal wall stain, methenamine silver. Cyst formation is thought to result from mating of trophs, but the interplay between the asexual and sexual phases is not clear. During the tenure of the previous Merit Review, we found that the treatment of Pneumocystis infections with echinocandins suppressed the formation of B-1,3-glucan thereby dramatically shifting the mixed cyst and troph populations in immunosuppressed mice and rats to one made almost exclusively of trophic forms. Strikingly, the mortality was markedly reduced in these mice, yet large populations of trophs were present in their lungs. Moreover, echinocandin treated mice were unable to transmit the infection. In the present proposal, we will use the echinocandins as molecular tools to dissect the life cycle in a manner that has never been available to investigators before. It is our central hypothesis that the asexual and sexual cycles act in concert to maintain a balance within the mammalian lung that permits a long and sustainable infection that ensures sustained transmission without demise of the host. We posit that cysts are the agents of transmission and the process of cyst formation requires B-1,3-glucan. However, the presence of cysts via B-1,3 glucan induces a detrimental inflammatory response in the host that increases severity of disease. In the present proposal, we will identify the immune responses evoked by each different population and dissect the life cycle of Pneumocystis using the echinocandins. Towards these goals, we propose the following specific aims: (1) Dissect the Pneumocystis life cycle using the echinocandin, anidulafungin, as a molecular tool by transcriptional analyses and in vivo studies to assess transmission and identify the infectious agent of Pneumocystis infection; (2) Characterize the P. murina cyst and trophic form interactions with alveolar macrophages through a series of binding and phagocytosis assays; and (3) Investigate the role of P. murina cysts in the deleterious pulmonary hyperinflammatory response following immune reconstitution by evaluating cytokine and cellular profiles in the lungs of cyst replete and cyst depleted populations of P. murina.

描述（由申请人提供）： 肺孢子菌属的真菌生物首先在临床上引起注意，因为它们经常在人类和其他免疫状态受损的哺乳动物中引起致命性肺炎（PCP）。如今，这些真菌的生存范围已经扩大到包括患有慢性阻塞性肺病等潜在慢性疾病的患者，它们的存在被认为是一种共病因素。在了解肺孢子虫的生命周期、传播和自然史方面取得的进展有限，这在很大程度上是由于缺乏连续的体外培养系统。这些生物体在完整或免疫衰弱的宿主环境中生长和生存所使用的策略在很大程度上是未知的。真正的感染因子，即传播形式，尚未确定，其传播方式也尚未确定，通常认为是通过空气传播的孢子。肺孢子菌的生命周期涉及营养形式（营养体），被认为是营养、无性分裂的生命周期阶段；以及其他几个发育阶段，包括囊肿形式，其中包含 8 个子囊孢子。包囊与营养体的不同之处在于它含有厚的细胞壁，主要由 B-1,3 葡聚糖组成；更大（5-8um vs 1-4um）；并用真菌壁染色剂乌洛托品银进行染色。包囊的形成被认为是营养体交配的结果，但无性阶段和有性阶段之间的相互作用尚不清楚。 在上一次优异评审期间，我们发现用棘白菌素治疗肺孢子虫感染可抑制 B-1,3-葡聚糖的形成，从而显着地将免疫抑制小鼠和大鼠中的混合囊肿和营养物群体转变为几乎完全由营养形式。引人注目的是，这些小鼠的死亡率显着降低，但它们的肺部却存在大量的营养物质。此外，棘白菌素治疗的小鼠无法传播感染。在本提案中，我们将使用棘白菌素作为分子工具，以研究人员以前从未使用过的方式剖析生命周期。 我们的中心假设是，无性周期和有性周期协同作用以维持哺乳动物肺部内的平衡，从而允许长期且可持续的感染，从而确保持续传播而不会导致宿主死亡。我们假设包囊是传播媒介，包囊形成过程需要 B-1,3-葡聚糖。然而，B-1,3 葡聚糖引起的囊肿会在宿主体内诱发有害的炎症反应，从而增加疾病的严重程度。在本提案中，我们将识别每个不同群体引起的免疫反应，并使用棘白菌素剖析肺孢子虫的生命周期。为了实现这些目标，我们提出以下具体目标：（1）利用棘白菌素、阿尼芬净作为分子工具，通过转录分析和体内研究来剖析肺孢子虫的生命周期，以评估传播情况并识别肺孢子虫感染的传染源； (2) 通过一系列结合和吞噬测定来表征鼠鼠包囊和营养形式与肺泡巨噬细胞的相互作用； (3)通过评估包囊充满和囊肿耗尽的鼠类种群肺部的细胞因子和细胞谱，研究鼠类包囊在免疫重建后有害的肺部高炎症反应中的作用。