Abstract
Purpose:
Herpes simplex virus-1 (HSV-1) can cause a lytic infection of the cornea resulting in the damage to the epithelium. The lytic release of the virions from the infected cells exacerbates the damage to the cornea. Our hypothesis is that heparanase, an enzyme, is a novel mediator of HSV-1 release from cells and thus, a new target for anti-viral therapy.
Methods:
Human corneal epithelial cells were cultured and infected with HSV-1 virions. The effect of heparanase was studies using heparanase expression construct, antibodies and exogenous heparinase treatments. The release of the virions was evaluated at various time points by plaque assays. Western blot analysis was performed to evaluate the changes in active and inactive haparanse expression. The connection between haparanase and syndecan shedding was assessed by dot blot analysis.
Results:
We demonstrate that HSV-1 infection results in enhanced expression of heparanase, an enzyme that hydrolyzes heparan sulfate. We found a direct role for enhanced heparanase expression and exogenous treatments with the enzyme in HSV-1 release from infected cells. Enhanced expression of heparanase did not affect HSV-1 entry into host cell, or HSV-1 induced cell-to-cell fusion, suggesting that heparanase activation is tightly regulated, and possibly more sensitive to HS modification utilized for virus egress compared to those that are utilized for virus entry. Furthermore, HSV-1 resulted in an increase in active heparanase expression, which was accompanied by a decrease in the inactive heparanase expression. Active heparanase has also been shown to mediate heparan sulfate proteoglycans (HSPGs) shedding including syndecans; where syndecan ectodomain is released in a soluble form to the extracellular space. Enhancing syndecans shedding by the shedding agonist PMA resulted in increased HSV-1 release from infected cells to the culture supernatant. This suggests that not only heparanase induces virus release from infected cells through HS cleavage, but also by mediating the shedding of HSPGs including syndecans.
Conclusions:
Our study defines a novel mechanism for viral release from the cells of the corneal epithelium and implicates heparanse, an enzyme, in control of the release. Thus, heparanse may be used as a new target to control corneal complications originating from HSV-1 infection.
Keywords: 545 herpes simplex virus •
482 cornea: epithelium •
636 pathobiology