Differential level of calcitonin gene-related peptide (CGRP) in the corneas versus trigeminal ganglia (TG) of herpes simplex virus-1 (HSV-1) infected mice regulates the neutrophil influx in these tissues.

Abstract

Purpose : This study aimed to measure the amount of CGRP neuropeptide in the corneas and TGs of HSV-1 infected mice and correlates their level of expression with the influx of neutrophils in the respective tissues.

Methods : Eight-twelve weeks C57BL/6J mice received topical application of HSV-1 McKrae onto the corneal surface after mild scratching in 3 ul of PBS. Mice were euthanized at 3- and 10 days post-infection (DPI). The acid extraction of dissected corneas and TGs was done using 2% acetic acid. CGRP amount was quantitated using an EIA assay kit. The flow cytometry assay was carried out on the single-cell suspension prepared from the corneas and TGs, and the frequency of neutrophils and monocytes among the myeloid cells was determined in these tissues. RT-qPCR assay was performed to measure the expression of CGRP receptor, RAMP1 in uninfected and HSV-1 infected corneas.

Results : Our results showed a significant decrease (P= 0.0079) in CGRP in HSV-1 infected corneas at 10- than 3-DPI. However, there was no significant difference in the amount of CGRP in the TGs of HSV-1 infected mice at these two time points. The scratch control corneas exhibited significantly higher levels of CGRP than HSV-1 infected corneas at 10-DPI. When compared between TGs and corneas from HSV-1 infected mice, at 3-DPI, the CGRP level was 6-fold lower in the corneas than in the TGs of HSV-1 infected mice. By 10-DPI, a 14-fold decrease in CGRP level was determined in the corneas compared to the TGs of HSV-1 infected mice. CGRP has been reported to modulate the influx of neutrophils in inflamed tissue. Our results showed that at 10 days post-infection, 78.9% (median) of CD11b+ myeloid cells in the HSV-1 infected corneas were neutrophils (Ly6G^hiLy6c^lo). On the contrary, 7.87% (median) of CD11b+ myeloid cells in the TGs of HSV-1 infected mice were neutrophils. The ratio of neutrophils to monocytes in the corneas was more than 1, whereas it was less than 1 in the TGs of HSV-1-infected mice at 10-DPI. Our RT-qPCR results showed the presence of CGRP receptor (RAMP1) in uninfected and HSV-1 infected corneas.

Conclusions : Together, our results showed an inverse correlation between the amounts of CGRP and the influx of neutrophils in the corneas and TGs of HSV-1-infected mice.

This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.