Novel characterization of CXCR4 expressing cells in uninfected and herpes simplex virus-1 infected corneas

To characterize CXCR4-expressing cells in uninfected and herpes simplex virus-1 (HSV-1) infected corneas. The corneas of C57BL/6J mice were infected with HSV-1 McKrae. The RT-qPCR assay detected CXCR4 and CXCL12 transcripts in uninfected and HSV-1-infected corneas. Immunofluorescence staining for CXCR4 and CXCL12 protein was performed in the frozen sections of herpes stromal keratitis (HSK) corneas. Flow cytometry assay characterized the CXCR4-expressing cells in uninfected and HSV-1-infected corneas. Flow cytometry data showed CXCR4 expressing cells in the separated epithelium and stroma of uninfected corneas. In the uninfected stroma, CD11b + F4/80+ macrophages are the predominant CXCR4-expressing cells. In contrast, most CXCR4 expressing cells in the uninfected epithelium were CD207 (langerin)+, CD11c+, and expressed MHC class II molecule, documenting the Langerhans cells (LCs) phenotype. After corneal HSV-1 infection, CXCR4 and CXCL12 mRNA levels increased significantly in HSK corneas than in uninfected corneas. Immunofluorescence staining showed CXCR4 and CXCL12 protein localization in the newly formed blood vessels in the HSK cornea. Furthermore, the infection resulted in LCs proliferation, causing an increase in their numbers in the epithelium at 4 days post-infection (p.i.). However, by 9-day p.i., the LCs numbers declined to the counts observed in naïve corneal epithelium. Our results also showed neutrophils and vascular endothelial cells as the prominent CXCR4-expressing cell types in the stroma of HSK corneas. Together, our data demonstrate the expression of CXCR4 on resident antigen presenting cells in the uninfected cornea and on infiltrating neutrophils and newly formed blood vessels in the HSK cornea.