Macrophages are distributed throughout the body and are crucial for the restoration of damaged tissues. However, their characteristics in the cornea and roles in the repair of corneal injures are unclear. Here we show that corneal macrophages can be classified as CCR2⁻ macrophages, which already exist in the cornea at embryonic day 12.5 (E12.5) and are similar to yolk sac-derived macrophages, microglia, in phenotype and gene expression, and CCR2⁺ macrophages, which do not appear in the cornea until E17.5. At a steady state, CCR2⁻ corneal macrophages have local proliferation capacity and are rarely affected by monocytes; however, following corneal epithelial abrasion, most CCR2⁻ corneal macrophages are replaced by monocytes. In contrast, CCR2⁺ macrophages are repopulated by monocytes under both a steady-state condition and following corneal wounding. Depletion of CCR2⁺ macrophages decreases corneal inflammation after epithelial abrasion, whereas depletion of CCR2⁻ macrophages increases inflammation of the injured cornea. Loss of either cell type results in a delay in corneal healing. These data indicate that there are two unique macrophage populations present in the cornea, both of which participate in corneal wound healing by balancing the inflammatory response.

The cornea is a special interface between the internal ocular tissue and the external environment that provides a powerful chemical, physical, and biological barrier against the invasion of harmful substances and pathogenic microbes. This protective effect is determined by the unique anatomical structure and cellular composition of the cornea, especially its locally resident innate immune cells, such as Langerhans cells (LCs), mast cells (MCs), macrophages, γδ T lymphocytes, and innate lymphoid cells. Recent studies have demonstrated the importance of these immune cells in terms of producing different cytokines and other growth factors in corneal homeostasis and its pathologic conditions. This review paper briefly describes the latest information on these resident immune cells by specifically analyzing research from our laboratory.

Inflammation and reepithelialization after corneal abrasion are critical for the rapid restoration of vision and the prevention of microbial infections. However, the endogenous regulatory mechanisms are not completely understood. Here we report that the manipulation of autonomic nervous system (ANS) regulates the inflammation and healing processes. The activation of sympathetic nerves inhibited reepithelialization after corneal abrasion but increased the influx of neutrophils and the release of inflammatory cytokines. Conversely, the activation of parasympathetic nerves promoted reepithelialization and inhibited the influx of neutrophils and the release of inflammatory cytokines. Furthermore, we observed that CD64⁺CCR2⁺ macrophages in the cornea preferentially expressed the β-2 adrenergic receptor (AR), whereas CD64⁺CCR2⁻ macrophages preferentially expressed the α-7 nicotinic acetylcholine receptor (α7nAChR). After abrasion, the topical administration of a β2AR agonist further enhanced the expression of the proinflammatory genes in the CD64⁺CCR2⁺ cell subset sorted from injured corneas. In contrast, the topical administration of an α7nAChR agonist further enhanced the expression of the anti-inflammatory genes in the CD64⁺CCR2⁻ subset. Thus crosstalk between the ANS and local macrophage populations is necessary for the progress of corneal wound repair. Manipulation of ANS inputs to the wounded cornea may represent an alternative approach to the treatment of impaired wound healing.

Purpose: In mammalian corneal epithelium, mitosis shows a distinct circadian pattern. However, how this circadian pattern is maintained, and how it or its disruption influence renewal and regeneration remain unclear. Methods: C57BL/6 mice were maintained under 12-hour light/12-hour dark (LD), 12-hour light/12-hour light (LL), 12-hour dark/12-hour dark (DD), or reversed LD (DL, 12-hour dark/12-hour light; jet-lag defined as a shift of 12 hours) conditions. Mitotic cells in corneal epithelium were enumerated and analyzed via immunofluorescence at different zeitgeber times (ZTs). Expression of core clock genes (Clock, Bmal1, Period2, Cry1, and Rev-erbα) was qualified via quantitative RT-PCR. The rate and quality of healing at different ZT times and after administration of two small-molecule modifiers of the circadian clock, KL001 and SR8278, was evaluated. Results: In this study, photic cues were found to influence the 24-hour rhythm of corneal clock gene expression and epithelial cell mitosis in mice. Disruption of the circadian clock by exposure to constant light, constant dark, or jet-lag conditions modified the normal 24-hour patterns of corneal epithelial mitosis and corneal clock gene expression. The time of day of wound occurrence affected the rate and quality of corneal healing, with both of these parameters peaking during the more mitotically active hours of the morning. The two small-molecule modifiers of the circadian clock, KL001 and SR8278, had negative and positive effects on corneal wound healing, respectively. Conclusions: Circadian rhythms significantly influence corneal epithelium renewal and repair in mice. Our findings reveal possible opportunities for biological rhythm-based interventional strategies to control corneal healing and restore corneal homeostasis.