The Role of IL-17 in fungal keratitis

描述（由申请人提供）：IL-17和Th17细胞在宿主对真菌感染的应答中具有重要作用。在目前的研究中，我们研究了IL-17在曲霉菌和镰刀菌感染角膜的小鼠模型中的作用，这是全球失明和视力障碍的主要原因。C57BL/6小鼠经皮下或腹腔途径免疫。通过任一途径免疫的小鼠表现出快速的真菌清除，并显着减少角膜疾病相比，未免疫的，感染的小鼠，严重的角膜混浊和持续的真菌存在。细胞内流式细胞术显示免疫后脾脏中真菌特异性Th1和Th17细胞的发展，其在感染后选择性地募集到角膜（Th17细胞在48小时，Th1细胞在72小时）。 中和IL-17免疫后，或感染的IL-17-/-小鼠抑制的保护性表型，而IFN-？中和没有效果。 在感染后24小时，在免疫小鼠的角膜中检测到产生IL-17的中性粒细胞的离散群体，但未免疫小鼠的角膜中未检测到，并且在脾脏和骨髓中检测到。此外，此时中性粒细胞耗竭损害IL-17的产生并阻断保护性免疫。产生IL-17的中性粒细胞产生的活性氧水平升高，体外杀死菌丝的能力增强，表明这类细胞在调节真菌感染中具有重要作用。来自角膜、脾和骨髓的FACS分选的IL-17嗜中性粒细胞的定量PCR显示IL-17转录物，表明从头合成。此外，当与来自免疫小鼠的脾细胞上清液孵育时，幼稚骨髓嗜中性粒细胞表达IL-17 RNA，并且在抗IL-6和抗IL-23两者存在下消除表达。相反，用IL-16和IL-23两者刺激，而不是用单个Ab刺激IL-17表达。总之，这些数据表明，产生IL-17的中性粒细胞是由免疫小鼠中的IL-6和IL-23产生的，并被募集到真菌感染的角膜，导致快速的真菌清除，这可能是由于活性氧的产生增加或与Th17细胞的相互作用。这些发现显示了IL-17、Th17和产生IL-17的中性粒细胞在调节真菌感染中的作用。在所提出的实验中，我将确定Th17细胞和IL-17产生细胞在真菌性角膜炎的保护性免疫中的相对贡献。) 公共卫生相关性： 项目叙述：真菌性角膜炎是角膜溃疡、视力损害和失明的主要原因之一。在许多情况下，目前针对这种感染的抗真菌治疗不足以解决疾病。然而，拟议研究的结果可能会导致这种使人衰弱的感染的适当监管和新疗法。

DESCRIPTION (provided by applicant): IL-17 and Th17 cells have an important role in the host response to fungal infections. In the current study, we examined the role of IL-17 in a murine model of Aspergillus and Fusarium infections of the cornea, which are a major cause of blindness and visual impairment worldwide. C57BL/6 mice immunized by intratracheal or subcutaneous routes. Mice immunized by either route demonstrated rapid fungal clearance and significantly reduced corneal disease compared with unimmunized, infected mice which severe corneal opacity and a persistent fungal presence. Intracellular flow cytometry showed development of fungal-specific Th1 and Th17 cells in the spleen following immunization, which were selectively recruited to the cornea after infection (Th17 cells at 48h, and Th1 cells at 72h). Neutralization of IL-17 following immunization, or infection of IL-17-/- mice inhibited the protective phenotype, whereas IFN-? neutralization had no effect. A discrete population of IL-17 producing neutrophils was detected in the corneas 24h after infection corneas of immunized, but not unimmunized mice, and were detected in the spleen and bone marrow. Further, neutrophil depletion impaired IL-17 production at this time and blocked protective immunity. IL-17 producing neutrophils produced elevated ROS and had increased capacity to kill hyphae in vitro, indicating that this population of cells has an important role in regulating fungal infectio. Quantitative PCR of the FACS sorted IL-17 neutrophils from the cornea, spleen and bone marrow showed IL-17 transcripts, indicating de novo synthesis. Further, naive bone marrow neutrophils expressed IL-17 RNA when incubated with splenocyte supernatants from immunized mice, and expression was ablated in the presence of both anti-IL-6 and anti-IL-23. Conversely, stimulation with both IL-16 and IL-23, but not with single Abs stimulated IL-17 expression. Together, these data indicate that IL-17 producing neutrophils are generated by IL-6 and IL-23 in immunized mice, and are recruited to fungal infected corneas, resulting in rapid fungal clearance possibly due to elevated production of reactive oxygen species or interactions with Th17 cells. These findings show a role for IL-17, Th17, and IL-17 producing neutrophils in regulating fungal infections. In the proposed experiments I will determine the relative contribution of Th17 cells and IL-17 producing cells in protective immunity in fungal keratitis. ) PUBLIC HEALTH RELEVANCE: Project narrative: Fungal keratitis is one of the leading causes of corneal ulcers, visual impairment, and blindness. In many cases the current anti-fungal treatments for this infection are not adequate in resolving the disease. However, the results from the proposed studies could lead to proper regulation and novel therapies for this debilitating infection.