Intracellular Signal-transducing elements involved in transendothelial migration of lymphoma cells

To investigate the molecular mechanisms underlying transendothelial migration of tumor cells, an essential process for their hematogenous dissemination, we developed an in vitro model system that allows the separate monitoring of cell adhesion and transmigration processes. This system uses a human pre-B lymphoma cell line, Nalm-6, and a cultured mouse endothelial cell line, KOP2.16. Nalm-6 cells rapidly adhered to KOP2.16 and subsequently transmigrated underneath them. Using this model, we examined the effects on transendothelial migration, of various reagents which specifically interfere with the function of intracellular signal transduction molecules. Treatment of Nalm-6 cells with wortmannin (WMN), herbimycin A, pertussis toxin, or C3 exoenzyme of Clostridium botulinum, which specifically inhibit P13 kinase and/or myosin light chain kinase, herbimycin-sensitive tyrosine kinases, heterotrimeric G proteins, and the small G proteins, and the small G proteins rho/rac, respectively, reduced transmigration in a dose-dependent manner, Pretreatment of KOP2.16 endothelial cells with WMN also reduced transmigration in a dose-dependent manner. Binding of Nalm-6 binding to KOp2.16 was not affected, even when Nalm-6 or KOP2.16 cells were pretreated with these inhibitors, indicating that the reduction of transmigration was not due to a reduction of Nalm-6 to KOP2.16. These results also indicate that the signal transduction pathway(s) involved in transmigration can be dissociated from that of adhesion. Our results support the notion that endothelial cells are not a passive barrier in lymphoma extravasation, but that they assist lymphoma cell extravasation.