The interactions of leukocytes with the CAM’s on the brain endothelial cells play an important role in their CNS transmigration. Since our results demonstrated increase of three specific CAMs, we next analyzed the role of these CAMs in mediating the adhesion of leukocytes to the HBMVE cells. As seen in Fig. 4, under mockinfected conditions only a small number of monocytes adhered to the HBMVE cells, which increased significantly to the WNVinfected HBMVE cells. However, GSI-IX Pretreatment of HBMVE cells with a cocktail of blocking antibodies against ICAM-1, VCAM-1 and E-selectin before the co-culture dramatically reduced the number of monocytes adhered to the infected HBMVE cells. We also observed similar results of increased attachment of lymphocytes to the WNV-infected HBMVE cells, which was significantly reduced in the presence of blocking antibodies to CAMs. After demonstrating the role of CAMs in leukocyte-HBMVE cell adhesion, we subsequently analyzed their role in the disruption of in vitro BBB model during the transmigration. As demonstrated in Fig. 5, incubation of the WNV-infected inserts with monocytes in the presence of a cocktail of neutralizing antibodies against CAMs reversed the decrease in the TEER values. A similar trend was observed during the incubation of lymphocytes, which showed a significant reduction in the percentage change in the TEER values in the presence of the blocking antibodies. Next, we determined whether blocking leukocytes adhesion to the endothelial cells using antibodies against CAMs also interferes with the leukocyte transmigration across the BBB model. Using fluorescent-tagged monocytes, we observed that the transendothelial migration of monocytes across the uninfected BBB model was minimal after 2 hrs of incubation. Pretreatment of the in vitro BBB model with tumor necrosis factor-a increased the number of migrated monocytes by.2 fold as represented by the increased fluorescence of the leukoTracker dye in the lower chamber. Similarly, compared to the controls, WNV infection of the BBB model increased the number of monocytes crossing the BBB model by 2.5-fold. Presence of blocking antibodies had no effect on the minimal monocytes migration across the uninfected models. Collectively, our results indicate that infection of HBMVE cells with WNV increased the adhesion and migratory capacity of leukocytes resulting in the disruption of the BBB model. Infection with WNV in the mouse model of WNV-encephalitis is characterized by BBB disruption and massive leukocyte infiltration into the brain. The actual basis of BBB disruption, however, has not been elucidated. We previously demonstrated that WNV infection in the HBMVE cells can be one of the routes of cell-free virus entry into the brain, however it does not compromise the integrity of the BBB model, suggesting that events other than direct virus infection are responsible for the BBB disruption. Here, we demonstrate that transmigration of leukocytesacross the WNV-infected BBB model compromises its integrity, WNV-induced ICAM-1, VCAM-1 and E-selectin are critical in mediating.