CCR6⁺ γδ T cells in the cornea are protective during early HSV-1 infection.

Abstract

Purpose : Herpes Simplex Virus-1 (HSV-1) is the leading cause of infectious blindness in developed countries. In the U.S., the annual incidence of new ocular infections is 11.8 per 100,000. Both mice and humans develop similar pathology after intracorneal (i.c) HSV-1 infection, making mice a useful model to understand underlying mechanisms of disease. Mice lacking γδ T cells exhibit reduced interleukin 17A (IL-17A), increased virus titers and develop lethal encephalitis after i.c. infection. To understand the diversity of the γδ T cell response required for immunoprotection, we investigated whether a particular subset of γδ T cells was responsible for IL-17A production following i.c. HSV-1 infection.

Methods : C57Bl/6 mice received 1x10⁶ PFUs HSV-1 injected into the stroma of the cornea. One to two days post infection (P.I.) corneas were excised and digested in Liberase^TM. The cells were immunostained for surface markers and for intracellular cytokines, and analyzed using flow cytometry. ELISAs were used to measure the levels of secreted IL-1α and CCL20.

Results : By 24 hours P.I. there is a significant increase in the number of CCR6⁺ IL-17A⁺ γδ T cells compared to uninfected corneas (n=8, P<0.01). A second subset of γδ T cells lacking CCR6 were also increased in corneas following infection but do not produce IL-17A. To determine whether IL-1α and IL-23 are capable of mediating IL-17A production in the cornea, we performed flow cytometric analysis: CCR6⁺ γδ T cells expressed IL-1R1 and IL-23R. In addition, the CCR6⁺ γδ T cell subset possessed an activated cell phenotype (CD44^hi CD62L^hi). Importantly, both CCL20, the ligand for CCR6, and IL-1α were detected by ELISA within 24 hours P.I. To test whether CCR6⁺ γδ T cells confer protection, mice with γδ T cells lacking CCR6 were infected i.c. with HSV-1. CCR6 deficient (CCR6^-/-) mice had significantly higher corneal pathology scores at days 3 (n=12-13, p<0.05) and 7 (n=12-13, P<0.001) P.I. compared to wild-type mice after infection.

Conclusions : Two major subsets of γδ T cells (based on CCR6 expression) participate in the response to i.c. HSV-1 infection. This study supports the concept that CCR6⁺ γδ T cells are responsible for IL-17A production via recruitment by CCL20 and signaling by IL-1α and IL-23.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.