Abstract
Purpose :
The secretion of lipids by meibomian glands (MG) plays a crucial role in maintaining the ocular surface health, with their acini intricately innervated by a dense meshwork of nerve fibers. MG dysfunction often manifests clinically following ocular surgeries that damage corneal nerves. This study aimed to assess the impact of corneal nerve cut on MG.
Methods :
Six- to eight-week-old female BALB/c mice underwent circular cutting of the anterior stroma of the right cornea using a 2-mm punch. After two weeks, corneal nerve fibers were evaluated through whole-mount staining with class III β-tubulin antibody. TUNEL assay was employed to assess apoptotic cells in MG, while LipidTox staining examined MG lipogenesis. Immunostaining for IL-6, TNF-α, IL-17, CD45, 4-HNE, and 8-OHdG was performed on bilateral MG to evaluate cytokine expression, inflammatory cell infiltration, and oxidative stress. Flow cytometry analyzed CD4+IL17+, CD4+IFN-γ+, and Foxp3+ regulatory T cells in eye-draining lymph nodes (DLN). Western blot assessed IL-17 receptor expression in murine MG and human meibomian gland epithelial cells (HMGEC). HMGEC cocultured with IL-17 was subjected to western blot analysis for pAKT/AKT and PPAR-γ. In vivo neutralization of IL-17 was performed to evaluate its effects on lipogenesis.
Results :
At two weeks post trephination, central corneal nerve density decreased in both eyes. Lipogenesis reduction was observed in both eyes at 2 and 4 weeks (P<0.05). TUNEL assay revealed increased apoptosis in nerve-cut and contralateral eyes. While oxidative stress and cytoplasmic IL-6, TNF-α showed no increase, MG strongly expressed cytoplasmic IL-17 at 2 weeks in nerve-cut and contralateral eyes compared to normal mice (P=0.04). CD45+ cells infiltrated in the vicinity of MG post-nerve cut. Flow cytometry showed no changes in CD4+IL17+, CD4+IFN-γ+, and Foxp3+ T cell frequencies in the DLN. Western blot confirmed IL-17R expression in murine MG and HMGEC. In vitro, IL-17 treatment reduced pAKT/AKT and PPAR-γ expression in differentiated acinar cells in a dose-dependent manner. In vivo IL-17 neutralization restored MG lipogenesis post-nerve cut.
Conclusions :
Unilateral corneal nerve cut resulted in reduced lipogenesis and increased apoptosis and MG inflammation, particularly with IL-17 expression. This study suggests that corneal nerve injury may disrupt neuroregulation of MG, leading to inflammation and dysfunction.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.