Abstract
Purpose :
Neuropeptides play an important role in the regulation of ocular surface homeostasis. Tear levels of calcitonin gene-related peptide (CGRP) have been shown to decrease in dry eye patients. In this study, we aimed to investigate the effect of exogenous administration of CGRP on disease severity, corneal epitheliopathy, and inflammatory response in experimental dry eye disease (DED).
Methods :
DED was induced by placing female C57BL/6 mice in the controlled-environment chamber for 14 days. 50 μM of CGRP or albumin (n=10 each) was administered topically three times per day (D) from D0 to D14 after DED induction. Corneal fluorescein staining (CFS) and tear break-up time (TBUT) were performed to evaluate disease severity. In vivo corneal confocal microscopy (IVCM) was performed to evaluate the inflammatory cells in the stroma. Tissues were harvested on D14 after treatment. Cultured human corneal epithelial cells (CECs) were stimulated with hyperosmotic stress (500 mOsm) and treated with CGRP (10-7M). Cell viability, proliferation, migration, and apoptosis were evaluated by MTT, immunostaining, scratch wound assay, and flow cytometry, respectively. mRNA expression of TNF-α, IL-1β, and IL-6 was measured using RT-PCR.
Results :
Compared with albumin control, topical application of CGRP significantly decreased CFS scores (P<0.001) and increased TBUT (P<0.01) of DED mice on days 3, 7, 10, and 14. In the central cornea, the density of hyper-reflective infiltrated cells was 46.2% higher in the albumin group than in the CGRP group (P<0.05) on D14. The number of goblet cells per conjunctival fornix was 21.3% higher in the CGRP group compared to the albumin group (P<0.05). After CGRP treatment, the percentage of Ki67+ cells was 155% higher in the corneal epithelium of the DED mouse (P<0.01) and 45.2% higher in CECs cultured in hyperosmosis (P<0.001), compared to the controls. The CGRP treatment led to a 164% increase in cell viability (P<0.001), a 74.3% higher wound healing rate (P<0.05), an 80.1% reduction of apoptotic cell frequency (P<0.001), and reduced mRNA levels of TNF-α, IL-1β and IL-6 (P<0.05) in CECs cultured in hyperosmosis.
Conclusions :
These results demonstrate that topical CGRP ameliorates DED by enhancing corneal epithelial proliferation and migration, and inhibiting apoptosis and inflammatory responses.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.