Protective Immune Responses to Blastomyces Dermatitidis

芽生菌病是一种主要的全身性真菌病，通常会导致进行性肺部疾病，但在皮炎芽生菌感染期间导致免疫功能衰竭和抵抗的因素尚不明确。我们在原发性肺部感染的小鼠模型中创建了一种WI-1敲除的皮炎杆菌，这种细菌被减弱和控制。给老鼠注射这种药物可以使它们免受再次感染，并唤起绝育免疫。在此，我们建议使用在上一个资助期开发的等基因减毒和野生型菌株和小鼠模型来确定实验性肺芽孢菌病免疫失败和耐药性的细胞和分子基础。我们假设：皮炎杆菌在非免疫宿主中破坏免疫t细胞的产生，导致进行性感染。在接种疫苗的宿主中，t细胞介导抵抗，并在t细胞亚群产生和维持疫苗免疫和记忆的需求方面表现出可塑性。我们的具体目标是：(1)破译免疫抑制细胞因子转化生长因子- β (TGF- β)在进行性原发性感染中t细胞免疫失败的作用，以及CTLA-4 （t细胞活化的负调节因子）在TGF- β诱导的细胞免疫抑制中的作用。(2)描述CD4+和CD8+ t细胞介导疫苗对皮炎免疫的不同机制-细胞因子（ifn - γ, tnf - α），细胞溶解和直接抗菌活性，以及CD28共同刺激和IL-12信号传导在产生和维持这些回忆免疫反应中的需求。(3)从皮炎B. cDNA文库的表达产物和原始的保护性细胞壁抗原的t细胞免疫印迹中，鉴定和克隆保护性t细胞在体外识别的抗原，并证明克隆的重组抗原在体内对皮炎B.具有保护性免疫。了解免疫衰竭和对真菌的抗性机制将有助于在健康人群和艾滋病等宿主防御受损患者中规划疫苗和治疗策略。

Blastomycosis, one of the principal systemic mycoses, often produces a progressive pulmonary disease, but the factors that account for immune failure and resistance during infection with Blastomyces dermatitidis infection are ill defined. We created a WI-1 knockout of B. dermatitidis that is attenuated and controlled in a murine model of primary pulmonary infection. Its administration to mice vaccinates them against re-infection and evokes sterilizing immunity. We propose here to use the isogenic attenuated and wild-type strains and mouse models developed in the last funding period to define the cellular and molecular bases of immune failure and resistance in experimental pulmonary blastomycosis. We hypothesize: that B. dermatitidis subverts generation of immune T-cells in a nonimmune host, leading to progressive infection. In a vaccinated host, T-cells mediate resistance, and show plasticity in the requirements of T-cell subsets for generation and maintenance of vaccine immunity and memory. Our specific aims are to: (1) Decipher the role of the immunosuppressive cytokine transforming growth factor-beta (TGF- beta) in failure of T-cell immunity during progressive primary infection, and the participation of CTLA-4, a negative regulator of T-cell activation, during TGF-beta-induced cellular immune suppression. (2) Delineate differential mechanisms - cytokines (IFN-gamma, TNF-alpha), cytolysis, and direct antimicrobial activity - by which CD4+ and CD8+ T-cells mediate vaccine immunity to B. dermatitidis, and requirements for CD28 co- stimulation and IL-12 signaling in generating and maintaining these recall immune responses. (3) Identify and clone antigens that protective T-cells recognize in vitro from expressed products of a B. dermatitidis cDNA library, and in T-cell immunoblots of a crude, protective cell-wall membrane antigen, and demonstrate that the cloned recombinant antigens confer protective immunity in vivo to B. dermatitidis. Understanding mechanisms of immune failure and resistance to fungi will assist in planning vaccine and treatment strategies in healthy people, and in patients with impairments of host defense such as AIDS.