Toll-like receptors and bacterial endophthalmitis

摘要 细菌性眼内炎是眼球穿通伤和眼内感染的一种威胁视力的并发症， 手术，特别是白内障手术，是老年人中最常见的眼科手术， 美国的大约每1000名患者中有3名发生白内障后细菌性眼内炎 手术随着美国老年人口的急剧增长，白内障手术的数量 也将大大增加，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的病原体对于最小化视网膜损伤至关重要。的 在初步数据中提出的最近发现揭示了视网膜对TLR 2激动剂的反应 Pam 3Cys通过产生先天免疫介质，以及在免疫前玻璃体内注射Pam 3Cys， 细菌感染，完全防止了金黄色葡萄球菌（SA）眼内炎的发展， C57 BL/6（B6）小鼠。这导致了TLR 2在视网膜先天免疫中起关键作用的假设。 响应S。金黄色葡萄球菌和激活和信号通过TLR决定疾病的结果。的 本提案的目的是阐明TLR 2激活阻止发展的机制， SA眼内炎。本研究的目的有三：1）确定TLR 2在视网膜先天性损害中的作用， 对SA的应答，以及Pam 3Cys预处理如何调节TLR 2信号传导。先天免疫反应 将在正常、TLR 2配体和SA攻击的B6小鼠视网膜和培养的视网膜（小胶质细胞， 星形胶质细胞、Muller和RPE）细胞通过评估TLR介导的细胞信号传导和促炎性细胞因子的产生来检测。 2）为了确定TLR 2配体诱导的保护性刺激的机制， 视网膜先天免疫感染前TLR 2的激活主要通过下调- 调节视网膜中的促炎（Th 1）细胞因子和上调抗微生物肽（AMP） 在随后的细菌挑战中。将使用TLR 2-/-和MyD 88-/-小鼠来确定这些小鼠是否是正常的。 这种机制是TLR 2/MyD 88依赖性的，对其他细菌有活性，并且在小鼠模型中有效。 玻璃体内注射相关SA眼内炎。3)确定cathelicidin相关的作用 抗微生物肽（CRAMP）在SA眼内炎期间视网膜先天反应中的作用。TLR信号 参与CRAMP诱导的途径和CRAMP调节细菌清除的机制， 将使用培养的视网膜细胞和CRAMP敲除（Cnlp-/-）小鼠测试视网膜完整性的保存。 这些目标的完成应该提供深入了解视网膜的先天反应， 微生物病原体，并可能导致识别新的治疗靶点，以防止手术- 相关性眼内炎。

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.