2 DISTINCT PHOSPHOLIPASES-C OF LISTERIA-MONOCYTOGENES INDUCE CERAMIDEGENERATION, NUCLEAR FACTOR-KAPPA-B ACTIVATION, AND E-SELECTIN EXPRESSION IN HUMAN ENDOTHELIAL-CELLS

Infection of endothelial cells by Listeria monocytogenes is an essenti al step in the pathogenesis of listeriosis, We recently reported that L. monocytogenes induces up-regulation of E-selectin and other endothe lial adhesion molecules and subsequent polymorphonuclear leukocyte (PM N) adhesion into cultured human endothelial cells. In the present stud y, we characterized the mechanisms of enhanced E-selectin expression u sing L, monocytogenes wild type (EGD), the isogenic in-frame deletion mutants for phosphatidylcholine (PC)- and phosphatidylinositol (PI)-sp ecific phospholipases EGD Delta plcA and EGD Delta plcB, as well as th e nonvirulent control strain Listeria innocua. Infection of endothelia l cells with EGD Delta plcA or EGD Delta plcB for 6 h induced, as comp ared with EGD wild type, intermediate levels of E-selectin mRNA and pr otein as well as PMN rolling and adhesion at a shear rate of 1 dyne/cm (2), indicating that both bacterial phospholipases are required for a maximal effect. Similarly, ceramide content and NF-kappa B activity we re increased in L, monocytogenes-exposed endothelial cells, but only t o intermediate levels for PC- or PI-phospholipase C (PLC)-deficient li sterial mutants. Phospholipase effects could be mimicked by exogenousl y added ceramides or bacterial sphingomyelinase. The data presented in dicate that PI-PLC and PC-PLC are important virulence factors for L, m onocytogenes infections that induce accumulation of ceramides that in turn may act as second messengers to control host cell signal-transduc tion pathways leading to persistent NF-kappa B activation, increased E -selectin expression, and enhanced PMN rolling/adhesion, The ability o f L. monocytogenes to stimulate PMN adhesion to endothelial cells may be an important mechanism in the pathogenesis of severe listeriosis.