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反应中起着重要作用。在目前的研究中，我们研究了IL-17在角膜曲霉菌和镰刀菌感染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。经气管或皮下途径免疫C57BL/6小鼠。与未免疫的、感染了严重角膜混浊和持续真菌存在的小鼠相比，任何一种途径免疫的小鼠都表现出快速的真菌清除和显著的减少角膜疾病。细胞内流式细胞术显示，免疫后小鼠的脾中出现了真菌特异性的Th1和Th17细胞，并在感染后选择性地被招募到角膜(Th17细胞在48h，Th1细胞在72h)。 免疫后中和IL-17或感染IL-17-/-小鼠可抑制保护性表型，而干扰素？中和不起作用。免疫小鼠感染后24小时，角膜中可检测到大量产生IL-17的中性粒细胞，而未免疫小鼠角膜中未检测到IL-17产生中性粒细胞。此外，中性粒细胞耗尽会在此时损害IL-17的产生，并阻止保护性免疫。产生IL-17的中性粒细胞产生较高的ROS，并在体外增强了对菌丝的杀灭能力，表明该细胞群在调节真菌感染方面具有重要作用。流式细胞仪对角膜、脾和骨髓中性粒细胞进行定量聚合酶链式反应，结果显示IL-17转录本，提示IL-17的从头合成。此外，幼稚的骨髓中性粒细胞在与免疫小鼠的脾细胞上清液孵育时表达IL-17RNA，并且在抗IL-6和抗IL-23的存在下表达被消融。相反，同时使用IL-16和IL-23刺激，但不单独使用单抗刺激可刺激IL-17的表达。综上所述，这些数据表明，在免疫的小鼠中，产生IL-17的中性粒细胞是由IL-6和IL-23产生的，并被招募到真菌感染的角膜，导致真菌的快速清除，可能是由于活性氧类的产生增加或与Th17细胞的相互作用。这些发现表明，IL-17、Th17和IL-17产生的中性粒细胞在调节真菌感染方面发挥了作用。在拟议的实验中，我将确定Th17细胞和IL-17产生细胞在真菌性角膜炎保护性免疫中的相对贡献。) 公共卫生相关性： 项目简介：真菌性角膜炎是导致角膜溃疡、视力障碍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这种疾病。然而，拟议研究的结果可能会导致对这种虚弱感染的适当调节和新的治疗方法。

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.