Outer membrane protein A-promoted actin condensation of brain microvascular endothelial cells is required for Escherichia coli invasion

Escherichia coli is the most common gram-negative bacterium that causes men ingitis during the neonatal period. We have previously shown that the entry of circulating E. coli organisms into the central nervous system is due to their ability to invade the blood-brain barrier, which is composed of a la yer of brain microvascular endothelial cells (BMEC). In this report, we sho w by transmission electron microscopy that E. coli transmigrates through BM EC in an enclosed vacuole without intracellular multiplication. The microfi lament-disrupting agents cytochalasin D and latrunculin A completely blocke d E. coli invasion of BMEC. Cells treated with the microtubule inhibitors n ocodazole, colchicine, vincristin, and vinblastine and the microtubule-stab ilizing agent taxol also exhibited 50 to 60% inhibition of E. coli invasion . Confocal laser scanning fluorescence microscopy showed F-actin condensati on associated with the invasive E. coli but no alterations in microtubule d istribution. These results suggest that E. coli uses a microfilament-depend ent phagocytosis-like endocytic mechanism for invasion of BMEC. Previously we showed that OmpA expression significantly enhances the E. coli invasion of BMEC. We therefore examined whether OmpA expression is related to the re cruitment of F-actin, OmpA(+) E. coli induced the accumulation of actin in BMEC to a level similar to that induced by the parental strain, whereas Omp A(-) E. coli did not. Despite the presence of OmpA, a noninvasive E. coli i solate, however, did not show F-actin condensation. OmpA(+)-E. coli-assocci ated condensation of F-actin was blocked by synthetic peptides correspondin g to the N-terminal extracellular domains of OmpA as well as BMEC receptor analogues for OmpA, chitooligomers (GlcNAc beta 1-4GlcNAc oligomers). These findings suggest that OmpA interaction is critical for the expression or m odulation of other bacterial proteins that will subsequently cause actin ac cumulation for the uptake of bacteria.