Therapeutic development of group B Streptococcus meningitis by targeting a host cell signaling network involving EGFR (2020)
Figure 1

Fig. 1. Group B Streptococcus (GBS) exploits host EGFR for penetration of the blood-brain barrier.

(A) Protein-Protein Interaction enrichment analysis of differentially expressed host factors in HBMEC with and without GBS infection revealing signaling network involving EGFR.

(B) Wild type mice received strain GFP-K79 via the tail vein and 1 hr later, animals were perfused and the meninges, cortex and choroid plexus were obtained for demonstration of intravenously injected bacteria. The arrows represent bacteria co-localized with capillaries (e.g., within capillaries) and arrowheads represent bacteria outside the capillaries (e.g., exited from the capillaries). A few bacteria were successfully passed through the meningeal and cortex capillaries, but no bacteria were demonstrated in the choroid plexus at this time. Scale bar = 100 μm. The completed figures with control and 12 h infection group were shown in Fig. EV2A, B and C.

(C) Sulfo-NHS-biotin was administered via intraperitoneal injection 10 min before perfusion for assessing the blood-brain barrier permeability, i.e., extravasation is indicative of leakage from the intravascular lumen. Sulfo-NHS-Biotin was confined to the capillaries of the meninges and cortex (as shown with arrows), and there was no evidence of extravasation. Scale bar = 100 μm. The completed figures with control and 12 h infection group were shown in Fig. EV2 D and E.

(D) Relative invasion frequency of GBS strain K79 in HBMEC with or without EGFR inhibitor (gefitinib). Data represent the means ± SEM from three independent experiments, with statistical analysis by Student's t-test, **p = 0.0020, ***p = 8.6E-06 (from left to right).

(E) Bacterial counts recovered from the blood and brain in wild type mice receiving vehicle control (n = 5) or gefitinib (10 mg/kg) (n = 6), infected with strain K79 for 1 h. Data represent the means ± SEM with Wilcoxon rank sum test, **p = 0.0038.

(F) Tyrosine phosphorylation of EGFR in HBMEC in response to GBS strain K79 infection at 15 min and 60 min post infection.

(G) EGFR protein expression in EGFR knockout HBMEC using CRISPR/Cas9 (left panel) and relative invasion frequency of 8 meningitis isolates of GBS strains in EGFR knockout and control HBMEC (right panel). Data represent the means ± SEM from three independent experiments, with statistical analysis by Student's t-test. From left to right, p value of K79 is 0.0059, p value of K160 is 0.00024, p value of K161 is 0.00080, p value of K181 is 0.00042, p value of K226 is 0.00011, p value of K237 is 0.00072, p value of K238 is 0.0093, p value of P539 is 0.00013 (right panel).

(H) GBS strain K79 traversal of HBMEC monolayer was significantly decreased in EGFR knockout HBMEC compare to control HBMEC. Data represent the means ± SEM from three independent experiments, with statistical analysis by Student's t-test, **p = 0.0027.

(I) Bacterial counts recovered from the blood and brain of EGFR conditional knockout (n = 5) and control mice (n = 5) 1 h after intravenous inoculation with strain K79. Data represent the means ± SEM with statistical analysis by Student's t-test, ***p = 0.00026.