Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2018 Dec 1;365(24):fny274.
doi: 10.1093/femsle/fny274.

Virulence Factor-Dependent Basolateral Invasion of Choroid Plexus Epithelial Cells by Pathogenic Escherichia Coli in Vitro

Affiliations
Free PMC article

Virulence Factor-Dependent Basolateral Invasion of Choroid Plexus Epithelial Cells by Pathogenic Escherichia Coli in Vitro

Rebekah Rose et al. FEMS Microbiol Lett. .
Free PMC article

Abstract

Escherichia coli is the most common Gram-negative causative agent of neonatal meningitis and E. coli meningitis is associated with high morbidity and mortality. Previous research has been carried out with regard to the blood-brain barrier and thereby unveiled an assortment of virulence factors involved in E. coli meningitis. Little, however, is known about the role of the blood-cerebrospinal fluid (CSF) barrier (BCSFB), in spite of several studies suggesting that the choroid plexus (CP) is a possible entry point for E. coli into the CSF spaces. Here, we used a human CP papilloma (HIBCPP) cell line that was previously established as valid model for the study of the BCSFB. We show that E. coli invades HIBCPP cells in a polar fashion preferentially from the physiologically relevant basolateral side. Moreover, we demonstrate that deletion of outer membrane protein A, ibeA or neuDB genes results in decreased cell infection, while absence of fimH enhances invasion, although causing reduced adhesion to the apical side of HIBCPP cells. Our findings suggest that the BCSFB might constitute an entry point for E. coli into the central nervous system, and HIBCPP cells are a valuable tool for investigating E. coli entry of the BCSFB.

Figures

Figure 1.
Figure 1.
Escherichia coli induced changes to TEER in HIBCPP standard and inverted cultures over time. Changes in TEER after infection of HIBCPP cells with E. coli at an (a) MOI of 10 in standard cultures, (b) MOI of 10 in inverted cultures, (c) MOI 1 in standard cultures and (d) MOI of 1 in inverted cultures. Values are given as % of the initial TEER prior to infection (0 h value); initial TEER was between 300 and 660 Ω × cm2 (SD +/– 129 Ω × cm2). All conditions were carried out three times in triplicates (n = 9).
Figure 2.
Figure 2.
Immunofluorescence and electron microscopy of E. coli RS218-infected HIBCPP cells. Shown is invasion of HIBCPP cells by E. coli RS218 in inverted cell cultures. Assays were carried out after a 1-h incubation period with E. coli with an initial MOI of 10. (a, b) Apotome-generated images of HIBCPP monolayers with staining of cell nuclei (blue), actin filaments (purple) and double-immunofluorescent staining of E. coli RS218 bacteria (intracellular, green; extracellular, yellow). Panels show an en face view (xy axis) with cross-sections through multiple optical layers (z axis) above the image and to its right. (a) Shown are cells containing intracellular bacteria (green arrows), whereas some bacteria adhered to the cell surface, presumably after cell transversal (red arrows). (b) Intracellular E. coli is labelled with green arrows. A red arrow indicates extracellular bacteria at the basolateral cell side, not yet having entered the cell close to bacteria located intracellularly after successful cell invasion. (cf) Ultrathin section transmission electron microscopic images show E. coli adhered to HIBCPP cells at the apical side to microvilli (c, d), and E. coli within membrane-bound vacuoles (labelled with arrowheads) inside HIBCPP cells (e, f). Multiple images showed vesicles containing more than one bacterium (e).
Figure 3.
Figure 3.
Escherichia coli adhesion to and invasion of HIBCPP cells from the apical (standard culture) and the basolateral side (inverted culture). Apical adhesion of E. coli to (a) and apical and basolateral invasion of (b) HIBCPP cell cultures in standard and inverted cell cultures, respectively. Assays were carried out after a 1-h incubation period with E. coli with an initial MOI of 10. Values are given as % of bacteria present after 1 h (derived from parallel growth curves). All assays were performed in triplicates and repeated at least five times (n = 15). *, significant, P < 0.05; **, highly significant, P < 0.01; ***, extremely significant, P < 0.001. TEER values remain stable when the HIPPC cells were challenged in the standard culture from the apical side (c) or in the inverted culture from the basolateral side (d) with the indicated E. coli strains with an MOI of 10 for 1 h.

Similar articles

See all similar articles

Cited by 3 articles

MeSH terms

Feedback