BACKGROUND: Herpes simplex virus (HSV)-infected endothelium is a model for vascular injury and possibly the development of atherosclerosis. In vitro infection of human umbilical vein endothelial cells (HUVEC) by HSV-1 results in a number of changes including the expression of a procoagulant activity (PCA) compatible with that due to tissue factor (TF) synthesis. In this study, we have further characterized this PCA using more stringent assays for TF, and examined whether virus rendered incapable of replication retains the ability to stimulate TF synthesis in HUVEC. EXPERIMENTAL DESIGN: Confluent monolayers of HUVEC were exposed to intact or ultraviolet/heat-inactivated HSV-1. At appropriate time intervals, TF PCA was assessed by clotting assays, and TF antigen by an enzyme-linked immunosorbent assay specific for TF. The appearance of mRNA specific for TF was performed by Northern blotting. RESULTS: TF activity was demonstrated by both 1-stage and 2-stage clotting assays; the dependence of the latter on factor VIIa, and the inhibition by specific blocking antibodies to human TF support the notion that the PCA is indeed due to TF. Furthermore, cellular TF antigen levels were found to parallel TF activity, and there was a transient de novo expression of TF mRNA. Tissue factor PCA in HSV-infected HUVEC remained "encrypted"; that is, full clotting activity was not expressed in the absence of cellular disruption in a situation analogous to that seen in all normal cells thus far examined that express TF PCA. However, this response did not appear to be dependent upon replicative infection of HSV-1 within the endothelial cell since a similar (although lesser) induction of TF PCA was present in cells that had been exposed to virus previously rendered incapable of replication. CONCLUSIONS: HSV-1 induces PCA in HUVEC which is clearly TF-dependent; this response does not require viral replication. These data indicate increased complexity in HSV interactions with vascular endothelium and imply induction of some procoagulant functions by nonproductive infection.
BACKGROUND: Herpes simplex virus (HSV)-infected endothelium is a model for vascular injury and possibly the development of atherosclerosis. In vitro infection of human umbilical vein endothelial cells (HUVEC) by HSV-1 results in a number of changes including the expression of a procoagulant activity (PCA) compatible with that due to tissue factor (TF) synthesis. In this study, we have further characterized this PCA using more stringent assays for TF, and examined whether virus rendered incapable of replication retains the ability to stimulate TF synthesis in HUVEC. EXPERIMENTAL DESIGN: Confluent monolayers of HUVEC were exposed to intact or ultraviolet/heat-inactivated HSV-1. At appropriate time intervals, TF PCA was assessed by clotting assays, and TF antigen by an enzyme-linked immunosorbent assay specific for TF. The appearance of mRNA specific for TF was performed by Northern blotting. RESULTS:TF activity was demonstrated by both 1-stage and 2-stage clotting assays; the dependence of the latter on factor VIIa, and the inhibition by specific blocking antibodies to humanTF support the notion that the PCA is indeed due to TF. Furthermore, cellular TF antigen levels were found to parallel TF activity, and there was a transient de novo expression of TF mRNA. Tissue factor PCA in HSV-infected HUVEC remained "encrypted"; that is, full clotting activity was not expressed in the absence of cellular disruption in a situation analogous to that seen in all normal cells thus far examined that express TF PCA. However, this response did not appear to be dependent upon replicative infection of HSV-1 within the endothelial cell since a similar (although lesser) induction of TF PCA was present in cells that had been exposed to virus previously rendered incapable of replication. CONCLUSIONS:HSV-1 induces PCA in HUVEC which is clearly TF-dependent; this response does not require viral replication. These data indicate increased complexity in HSV interactions with vascular endothelium and imply induction of some procoagulant functions by nonproductive infection.