E. coli K1 is the most common gram negative pathogen causing neonatal sepsis and meningitis. During E. coli K1 infection accompanied with bacterial invasion and hematogenous spreading, brain microvascular endothelial cells (BMEC) are directly targeted by E. coli K1 and their virulence factors. Among a number of known virulence factors contributing to E. coli K1 invasion of BMEC, IbeA is the major invasin which is unique to pathogenic E. coli K1 but not present in nonpathogenic E. coli K12. Therefore, we tested the hypothesis that IbeA, an outer membrane protein, induced human BMEC apoptosis. IbeA was shown to be an outer membrane protein, which shares significant sequence homology to another E. coli K1 virulence factor CglE. We investigated the potential of recombinant His-tagged IbeA to induce human BMEC (HBMEC) apoptosis. The TUNEL and caspase assays were used to measure apoptosis. The results showed that IbeA was able to induce apoptosis in HBMEC. Furthermore, caspase-8 was found to be activated by IbeA and IbeA+ E. coli K1. However, the ibeA mutant 10A-23 and another E. coli K1 invasion protein IbeB were unable to stimulate caspase 8. Hence, caspase-8-dependence of IbeA-induced endothelial apoptosis may bear relevance to novel insight into the pathogenesis of E. coli K1-induced sepsis and meningitis.