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Satvik Hadigal, Ghadah Karasneh, Deepak Shukla; Role of Syndecan-1 in HSV-1 infection of the corneal epithelium.. Invest. Ophthalmol. Vis. Sci. 2016;57(12):5876.
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© ARVO (1962-2015); The Authors (2016-present)
During a productive herpetic infection in the cornea, cell surface heparan-sulfate proteoglycans, exemplified by syndecan-1, trap newly exiting viral progenies and inhibit their release (1). Therefore, we tested the hypothesis that during a productive infection of the human corneal epithelial (HCE) cells, matrix metalloproteases such as MMP-7 accelerate Syndecan-1 shedding to enhance viral egress and viral spread.
Herpes simplex virus-1 (strains: Mckrae, KOS, K26GFP) were used for the infection of HCE cells. Cell surface syndecan-1 level was determined by flow cytometry using an anti syndecan-1 primary antibody (ebiosciences Cat # 12-1389-41). Shed syndecan-1 ectodomain was measured using slot blot and applying supernatant to cationic membrane and incubating in primary syndecan-1 antibody. Images of infected cells were taken with Carl Zeiss LSM 710 confocal microscope using a 63X oil magnification. Viral plaque assays were used to determine changes in viral egress patterns.
We found that syndecan-1 plays an important role in HSV-1 release from HCE cells. At 24h post infection or beyond we found significant loss of syndecan-1 cell surface expression and an increase in the shedding of syndecan-1 ectodomain. The rate of syndecan-1 ectodomain shedding was proportional to the higher release of HSV-1 virions in the culture supernatant. Overall, there was a net loss of syndecan-1 levels in infected cells as judged by immunofluorescence microscopy. As a possible mechanism of this loss we found that MMP-7 levels were significantly enhanced in infected cells and the greatest increase at MMP-7 levels was observed at the cell surface by flow cytometry (4), which was further confirmed by immunofluorescence imaging of infected HCE cells. Involvement of MMP-7 was somewhat specific as many other MMPs examined did not show significant increase in expression. Chemically induced enhancement of syndecan-1 shedding also enhanced HSV-1 release and inhibitors of MMP-7 such as TIMP-1 showed the opposite effects.
Our study identifies a new host protein, MMP-7, used by herpes simplex virus infection to cleave syndecan-1 proteoglycan for efficient viral egress. Among other MMPs tested in HCE cells, syndecan-1 was specifically cleaved by MMP-7. Inhibitors of MMP-7 inhibited viral egress significantly and hence MMP-7 could potentially be used as a target against HSV infection in the cornea.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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