Toll-like receptors and bacterial endophthalmitis

摘要 细菌性眼内炎是眼球穿通伤和眼内伤的一种威胁视力的并发症。 手术，特别是白内障手术，是#年老年人口中最常见的眼科手术 美国。大约每1000名患者中就有3名在白内障后发展为细菌性眼内炎。 做手术。随着美国老龄人口预计将大幅增长，白内障手术的数量 执行的也将显著增加，导致发病率成比例增加 眼内炎。视网膜的视觉特性对炎症引起的损伤高度敏感。 因此，快速检测和清除入侵病原体对于最大限度地减少视网膜损伤至关重要。这个 在初步数据中提出的最新发现显示，视网膜对TLR2激动剂有反应 Pam3Cys通过产生天然免疫的介体，并在玻璃体内注射Pam3Cys之前 细菌感染，完全防止金黄色葡萄球菌(SA)眼内炎的发展 C57BL/6(B6)小鼠。这导致了TLR2在视网膜先天免疫中起关键作用的假设 对金黄色葡萄球菌的反应以及通过TLR的激活和信号传递决定了疾病的结局。这个 本建议的目的是阐明TLR2激活阻止发育的机制。 金黄色葡萄球菌眼内炎。提出了三个具体的目标：1)确定TLR2在视网膜先天发育中的作用 对SA的反应，以及Pam3Cys预处理如何调制TLR2信号。天生的反应 将在正常的TLR2配体上进行测试，SA挑战B6小鼠视网膜和培养的视网膜(小胶质细胞， 星形胶质细胞，穆勒和RPE)细胞，通过评估TLR介导的细胞信号和促炎因子的产生 细胞因子/趋化因子，2)确定TLR2配体诱导保护性刺激的机制 视网膜先天免疫力。TLR2在感染前的激活主要通过下调TLR2来诱导保护机制 调节视网膜中的致炎细胞因子和上调抗菌肽 在随后的细菌挑战中。TLR2-/-和MyD88-/-小鼠将被用来确定这些 机制依赖于TLR2/MyD88，对其他细菌有活性，并在小鼠模型中有效。 玻璃体内注射相关的SA眼内炎。3)确定放线菌素相关的作用 抗菌肽(JAMP)在SA眼内炎期间视网膜固有反应中的作用TLR信令 参与抽筋诱导的途径和抽筋调节细菌清除和 将使用培养的视网膜细胞和抽筋基因敲除(Cnlp-/-)小鼠来测试视网膜完整性的保存。 这些目标的完成应该有助于对视网膜固有反应的理解 微生物病原体，并可能导致识别新的治疗靶点，以防止手术- 伴发眼内炎。

ABSTRACT Bacterial endophthalmitis is a vision-threatening complication of penetrating eye injury and intraocular surgery, notably cataract surgery, the most common ophthalmic procedure performed in older populations in the United States. Approximately, 3 out of 1000 patients develop bacterial endophthalmitis after cataract surgery. As the aged population in the US is expected to grow dramatically, the number of cataract surgeries performed will also increase significantly, resulting in a proportional increase in the incidence of endophthalmitis. The visual properties of the retina are highly sensitive to inflammation-caused damage therefore, a rapid detection and clearance of invading pathogens is critical in minimizing retinal damage. The recent discovery presented in the preliminary data revealed that the retina responds to the TLR2 agonist Pam3Cys by producing the mediators of innate immunity, and that intravitreal injection of Pam3Cys, prior to bacterial infection, completely prevented the development of Staphylococcus aureus (SA) endophthalmitis in C57BL/6 (B6) mice. This leads to the hypothesis that TLR2 plays a critical role in retinal innate immune response to S. aureus and that activation and signaling through TLR determines the disease outcome. The objective of this proposal is to elucidate the mechanisms by which TLR2 activation prevents the development of SA endophthalmitis. Three specific aims are proposed: 1) To determine the role of TLR2 in retinal innate response against SA, and how TLR2 signaling is modulated by Pam3Cys pretreatment. The innate response will be tested in normal, TLR2 ligand, and the SA challenged B6 mouse retinas and cultured retinal (microglia, astrocytes, Muller and RPE) cells by assessing TLR-mediated cell signaling and production of proinflammatory cytokines/chemokines, 2) To determine the mechanisms of TLR2 ligand-induced stimulation of protective retinal innate immunity. Activation of TLR2 prior to infection induces protective mechanisms mainly by down- regulating proinflammatory (Th1) cytokines and up-regulating antimicrobial peptides (AMPs) in the retina upon subsequent bacterial challenge. TLR2-/- and MyD88-/- mice will be used to determine whether these mechanisms are TLR2/MyD88 dependent, active against other bacteria, and are effective in a mouse model of intravitreal injection-associated SA endophthalmitis. 3) To determine the role of cathelicidin related antimicrobial peptide (CRAMP) in retinal innate responses during SA endophthalmitis. The TLR signaling pathways involved in CRAMP induction and mechanisms by which CRAMP modulates bacterial clearance and preservation of retinal integrity, will be tested using cultured retinal cells and CRAMP knockout (Cnlp-/-) mice. Completion of these aims should provide insight into the understanding of the retinal innate response to microbial pathogens, and may lead to the identification of new therapeutic targets for preventing surgery- associated endophthalmitis.