Escherichia coli invasion of brain endothelial cells

Huang, Sheng-He

Abstract

Despite the availability of antibiotics over the last sixty years, neonatal sepsis and meningitis (NSM) remains a major cause of death in newborns. E. coli is the most common gram-negative bacteria causing NSM. IbeA has been identified as a major invasin that allows E. coli to penetrate across the blood-brain barrier (BBB) in vitro [microvascular endothelial cells (BMEC)] and in vivo (neonatal murine model). IbeA and its homologue are unique to the pathogen but not present in nonpathogenic E. coli K12. The genetic island GimA encoding ibeA and cglE is significantly more prevalent in E. coli strains causing early human NSM and avian diseases. Bacterial meningitis is accompanied by pathogen penetration and polymorphonuclear neutrophil (PMN) transmigration across the BBB, which is mainly formed by BMEC. The underlying mechanisms for these pathogenicities remain to be elucidated. Our work has shown that IbeA/E mediate the pathogenicities (bacterial invasion and PMN transmigration across the BBB) of E. coli K1 in vitro (BMEC) and in vivo (meningitis in murine neonates). Vimentin is the common receptor on BMEC and PMN surface that contributes to IbeA/E-mediated pathogenicities. Disruption of lipid rafts with cholesterol depleting agents (e.g., filipin and methyl β-cyclodextrin) and vimentin (Vim) targeting with its inhibitor withaferin A are able to block IbeA/E+ E. coli K1-induced bacterial invasion and PMN transmigration across BMEC. Based on these findings, we hypothesize that Vim-enriched lipid rafts (Vim/LR) are common signaling platforms and therapeutic targets for IbeA/E-induced pathogenicities. E. coli K1 entry is mediated by IbeA/vimentin-induced signaling in BMEC. Vimentin in both BMEC and PMN is required for E. coli K1-stimulated PMN transmigration across the BBB. These hypotheses will be tested with the following specific aims.
In Aim 1, we will study whether IbeA/E-induced signaling is essential for bacterial invasion in vitro and in vivo by defining the Vim/LR-dependent signaling platform in BMEC and genetic blocking of the Vim gene.
In Aim 2, we will examine how IbeA/Vim signaling contributes to transcellular migration of PMN across the BBB using the cell culture systems and animal models (Vim + and Vim -), and molecular genetic approaches.
In Aim 3, we will determine the therapeutic efficacy of Vim/LR-targeted inhibitors in vitro (BMEC), ex vivo (organotypic hippocampal culture) and in vivo (neonatal mouse model). In summary, the uniqueness of this proposal is to define the dual virulence functions of IbeA/E: bacterial invasion and PMN transmigration across the BBB, which are the hallmarks of bacterial meningitis. These two pathogenicities are dependent on IbeA/E-induced signaling via Vim/LRs. Vim/LRs may be exploited as the potential therapeutic targets for blocking E. coli K1 invasion and PMN transmigration across HBMEC.

Public Health Relevance

Neonatal sepsis and meningitis (NSM) remains a major cause of death in newborns. IbeA and its homologue CglE have been identified as important virulence factors that allow pathogenic E. coli, the most common gram-negative bacteria causing NSM, to penetrate across the tissue barriers to cause NSM. The uniqueness of this proposal is the studies on the disease-causing mechanisms and novel therapeutic strategies for blocking the dual virulence functions of IbeA and CglE: bacterial invasion and white blood cell transmigration across the blood-brain barrier.