Herpesviruses are highly prevalent human pathogens that have the ability to form life-long latent infection in host cells. The persistent reinjury of vessels upon virus reactivation has been suggested to link infection to vascular disease. The goal of this study was to determine the mechanism and role of early initiating events in virus-mediated vasculopathy. We now report that these events are mediated through the expression of tissue factor (TF) and a previously unknown mechanism, that both function in the acceleration of factor VIIa (FVIIa)-dependent activation of factor X (FX) to FXa. The current study identifies herpes simplex virus type 1 (HSV1)-encoded glycoprotein C (gC) as a novel second independent FX activating pathway on the virus surface. Using specific chromogenic assays, an HSV1 gC-deficient virus invariably generated 5 fold less FXa per particle than either wild type or gC-rescued strains. The direct involvement of gC was confirmed using purified recombinant gC, which enhanced FXa production, and like TF, was dependent on FVIIa, Ca 2+ and anionic phospholipid. Differential inhibition of gC-competent and -deficient strains by an anti-TF antibody confirmed simultaneous and independent TF- and gC-dependent FX activating mechanisms on the virus. Hypothesizing that cell signaling by thrombin, the final coagulation protease, may be advantageous to the virus, the effect on Herpesvirus infection was assessed. Using plaque formation assays, a thrombin specific inhibitor, hirudin, was shown to attenuate the serum-dependent increase in infection, demonstrating the importance of virus initiated thrombin production. In agreement, the addition of purified thrombin resulted in an approximate 60--80% increase in infectious events. The same enhancement was facilitated by incubation with a protease activated receptor 1 (PAR1) agonist, TRAP, indicating the effect is mediated through at least PAR1 on the cell surface. Using western blot analysis and chromogenic assays, individual variations in thrombogenic potential associated with each Herpesvirus was also determined and correlated with well-documented clinical observations. Cumulatively, these observations illustrate that Herpesviruses have evolved strategies to mimic and exploit host proteins to generate haemostatic cell signaling enzymes that may ultimately lead to the increased susceptibility of cells to infection and perturbation of the vasculature.
| Identifer | oai:union.ndltd.org:uottawa.ca/oai:ruor.uottawa.ca:10393/29067 |
| Date | January 2003 |
| Creators | Sutherland, Michael R |
| Contributors | Pryzdial, Ed, |
| Publisher | University of Ottawa (Canada) |
| Source Sets | Université d’Ottawa |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | 162 p. |
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