A Role for Antithrombin Binding Heparan Sulfate in Blood Coagulation and Corneal Angiogenesis Induced by Herpes Simplex Virus Type–1 Infection

Abstract

Abstract: : Purpose: Novel biological functions of modified forms of heparan sulfate (HS) ranging from regulation of blood coagulation and angiogenesis to generation of HSV–1 gD receptors has wide range of clinical implications. Recently, cloning of an enzyme 3OST–5 isoform showed a unique binding for both antithrombin (AT) and HSV–1 gD. Deletion of the AT binding site on cell surface HS could result in the development of degenerative cardiac valve disease. The goal of this study was to investigate the role for anticoagulant HS in HSV–1 entry into ocular cell and characterize the peptides isolated via phage display technique against HSV–1 gD and 3OST–5 on HSV–1 entry in ocular cells. Methods: RT–PCR was used to check the expression of different isofroms of 3OSTs family members in ocular cell types including corneal epithelium, corneal fibroblasts (CF) and trabecular meshwork (TM) cells. Results obtained from RT–PCR showed the expression of gD receptors including 3OSTs. Ocular cells were then exposed to reporter virus to study viral entry. Phages isolated against HSV–1 gD and 3OST–5 from phage library were tested for blocking viral entry into ocular cells. Results: Using RT–PCR assay TM and CF cells were found to express gD receptors. Interestingly, in TM cells 3OSTs members (3OST–3, and 3OST–5 but not 3OST–1) were found to mediate HSV–1 entry. Multiple assays further showed the susceptibility of TM cells to HSV–1 entry. The peptides isolated against gD and 3OST–5 isoform from phage library showed the blocking of HSV–1 entry in ocular cells. Conclusions: RT–PCR results showed the expression of gD receptors in TM and CF cells including anticoagulant HS. Entry assays and florescent microscopy showed the susceptible nature of TM to HSV–1 entry. The peptides isolated against gD and 3OST–5 isoform will help in understanding the structural and functional analysis of HS and future drug design.

Keywords: herpes simplex virus • trabecular meshwork • receptors