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Derek J Royer, Daniel J Carr; Epithelial cells exhibit myeloid antigens during acute HSV-1 keratitis. Invest. Ophthalmol. Vis. Sci. 2017;58(8):3615.
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© ARVO (1962-2015); The Authors (2016-present)
Herpes simplex virus type 1 (HSV-1) keratitis is associated with significant visual morbidity arising from pathologic corneal fibrosis and neovascularization. We investigated the hypothesis that aberrant epithelial-mesenchymal/myeloid transition (EMT) contributes to corneal pathology during acute HSV-1 keratitis in mice.
Scarified corneas of C57BL/6 mice were infected with 104 plaque forming units of HSV-1, harvested on day six post-infection, and enzymatically digested to generate single-cell suspensions. Cells from processed corneas were phenotypically characterized by flow cytometry (FACS) or immunomagnetically enriched for cells expressing the epithelial cell adhesion molecule EpCAM for transcriptional profiling. Transgenic mice were utilized to explore mechanisms.
Ectopic Ly6G expression was observed in 70% of EpCAM+ cells from healthy and infected corneas. Ectopic Ly6C expression was also observed by FACS in 20% of EpCAM+ cells from HSV-1-infected corneas but was absent in mock-infected and healthy corneal digests. In addition, 20% of the Ly6C+ EpCAM+ cells expressed MHC-II. One-third of the MHC-II+ Ly6C+ cells expressed the inhibitory receptor PD-L1 though none expressed the costimulatory molecule CD86. The epithelioid morphology of EpCAM+ Ly6C+ cells was confirmed by microscopy. Ly6C expression in EpCAM+ cells from HSV-infected corneas was confirmed by qPCR, but classic EMT-associated genes were not detected. Moreover, FACS data show that Ly6C+ EpCAM+ cells lack the mesenchymal marker α smooth muscle actin. The myeloid-like phenotype of EpCAM+ cells was not a direct result of viral infection on an individual cell basis, as determined using a HSV-inducible Cre/Lox reporter model by FACS. Ongoing studies are being conducted to investigate the role of growth factors in driving the HSV-1-associated epithelial cell metaplasia in the cornea.
This study provides the first direct evidence of an epithelial-myeloid transition during viral infection, which may have implications for novel therapeutic development to avert visual morbidity. Importantly, ectopic expression of myeloid antigens on epithelial cells warrants heightened scrutiny of FACS analysis involving cornea digests. Finally, whether MHC-II expression on EpCAM+ corneal cells exacerbates immunopathology or exerts a tolerogenic function remains open to investigation.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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