The cornea of the eye differs from other mucosal surfaces in that it lacks a viable bacterial microbiome and by its unusually high density of sensory nerve endings. Here, we explored the role of corneal nerves in preventing bacterial adhesion. Pharmacological and genetic methods were used to inhibit the function of corneal sensory nerves or their associated transient receptor potential cation channels TRPA1 and TRPV1. Impacts on bacterial adhesion, resident immune cells, and epithelial integrity were examined using fluorescent labeling and quantitative confocal imaging. TRPA1/TRPV1 double gene-knockout mice were more susceptible to adhesion of environmental bacteria and to that of deliberately-inoculated Pseudomonas aeruginosa. Supporting the involvement of TRPA1/TRPV1-expressing corneal nerves, P. aeruginosa adhesion was also promoted by treatment with bupivacaine, or ablation of TRPA1/TRPV1-expressing nerves using RTX. Moreover, TRPA1/TRPV1-dependent defense was abolished by enucleation which severs corneal nerves. High-resolution imaging showed normal corneal ultrastructure and surface-labeling by wheat-germ agglutinin for TRPA1/TRPV1 knockout murine corneas, and intact barrier function by absence of fluorescein staining. P. aeruginosa adhering to corneas after perturbation of nerve or TRPA1/TRPV1 function failed to penetrate the surface. Single gene-knockout mice showed roles for both TRPA1 and TRPV1, with TRPA1^−/− more susceptible to P. aeruginosa adhesion while TRPV1^−/− corneas instead accumulated environmental bacteria. Corneal CD45+/CD11c+ cell responses to P. aeruginosa challenge, previously shown to counter bacterial adhesion, also depended on TRPA1/TRPV1 and sensory nerves. Together, these results demonstrate roles for corneal nerves and TRPA1/TRPV1 in corneal resistance to bacterial adhesion in vivo and suggest that the mechanisms involve resident immune cell populations.

中文翻译：

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神经相关的瞬时受体电位离子通道有助于体内细菌粘附的内在抵抗力

眼睛的角膜与其他粘膜表面的不同之处在于它缺乏可行的细菌微生物组，并且具有异常高密度的感觉神经末梢。在这里，我们探讨了角膜神经在防止细菌粘附中的作用。采用药理学和遗传学方法抑制角膜感觉神经或其相关瞬时受体电位阳离子通道TRPA1和TRPV1的功能。使用荧光标记和定量共聚焦成像检查对细菌粘附、常驻免疫细胞和上皮完整性的影响。TRPA1/TRPV1双基因敲除小鼠更容易粘附环境细菌和故意接种的铜绿假单胞菌。支持表达 TRPA1/TRPV1 的角膜神经的参与，用布比卡因治疗或使用 RTX 消融表达 TRPA1/TRPV1 的神经也促进了铜绿假单胞菌的粘附。此外，TRPA1/TRPV1 依赖的防御被切断角膜神经的去核所废除。高分辨率成像显示正常的角膜超微结构和小麦胚芽凝集素对 TRPA1/TRPV1 敲除小鼠角膜的表面标记，以及没有荧光素染色的完整屏障功能。神经或 TRPA1/TRPV1 功能扰动后粘附在角膜上的铜绿假单胞菌未能穿透表面。单基因敲除小鼠表现出 TRPA1 和 TRPV1 的作用，TRPA1 ^-/-更容易受到铜绿假单胞菌的粘附，而 TRPV1 ^-/-角膜反而积累了环境细菌。角膜 CD45+/CD11c+ 细胞对铜绿假单胞菌攻击的反应（先前显示可对抗细菌粘附）也取决于 TRPA1/TRPV1 和感觉神经。总之，这些结果证明了角膜神经和 TRPA1/TRPV1 在角膜对体内细菌粘附的抵抗中的作用，并表明这些机制涉及常驻免疫细胞群。