INVASION OF BRAIN MICROVASCULAR ENDOTHELIAL-CELLS BY GROUP-B STREPTOCOCCI

Group B streptococci (GBS) are the leading cause of meningitis in newb orns, Although meningitis develops following bacteremia, the precise m echanism or mechanisms whereby GBS leave the bloodstream and gain acce ss to the central nervous system (CNS) are not known, We hypothesized that GBS produce meningitis because of a unique capacity to invade hum an brain microvascular endothelial cells (BMEC), the single-cell layer which constitutes the blood-brain barrier. In order to test this hypo thesis, we developed an in vitro model with BMEC isolated from a human , immortalized by simian virus 40 transformation, and propagated in ti ssue culture monolayers. GBS invasion of BMEC monolayers was demonstra ted by electron microscopy. Intracellular GBS were found within membra ne-bound vacuoles, suggesting the organism induced its own endocytic u ptake. GBS invasion of BMEC was quantified with a gentamicin protectio n assay, Serotype III strains, which account for the majority of CNS i solates, invaded BMEC more efficiently than strains from other common GBS serotypes. GBS survived within BMEC for up to 20 h without signifi cant intracellular replication, GBS invasion of BMEC required active b acterial DNA, RNA, and protein synthesis, as,yell as microfilament and microtubule elements of the eukaryotic cytoskeleton, The polysacchari de capsule of GBS attenuated the invasive ability of the organism. At high bacterial densities, GBS invasion of BMEC was accompanied by evid ence of cellular injury; this cytotoxicity was correlated to beta-hemo lysin production by the bacterium, Finally, GBS demonstrated transcyto sis across intact, polar BMEC monolayers grown on Transwell membranes. GBS invasion of BMEC may be a primary step in the pathogenesis of men ingitis, allowing bacteria access to the CNS by transcytosis or by inj ury and disruption of the endothelial blood-brain barrier.