Fig 1.
Blood-retinal Barrier Breakdown in NaIO3-treated mice.
(A-D) Funduscopic imaging of mouse eyes 24 hours after injection of either PBS or NaIO3. In PBS-injected mice (A and B), the fluorescence from the AK-FLUOR dye demarcates the retinal and/or choroidal vasculature and distinguishes it from adjacent areas/structures. In NaIO3-injected mice (C and D), note the diffuse fluorescence resulting from increased outer BRB permeability and leakage of the dye. (E) Albumin leakage into the retina after injection of either PBS or NaIO3 was quantified using a modified Evans Blue protocol. Bars represent mean ± standard deviation (SD) for N ≥ 5 animals for all groups. A two-tailed t-test was used to assess significance between PBS-injected and NaIO3-injected mice (P = 0.01).
Table 1.
Incidence of K. pneumoniae and S. aureus EBE at 4 days postinfection in control and NaIO3-treated mice.
Table 2.
Incidence of K. pneumoniae and S. aureus EBE at 6 days postinfection in control and NaIO3-treated mice.
Fig 2.
Exoprotein-dependent alterations in ZO-1 immunoreactivity of cultured human RPE cells infected with S. aureus.
(A-H) Human ARPE-19 monolayers were infected with wild type S. aureus 8325–4 (A-C), an agr/sar-deficient mutant (E-G), or two ocular isolates of S. epidermidis (D and H), each at a concentration of 104 cfu/ml, MOI = 0.02. After 4, 6, or 8 hours postinfection, monolayers were stained with anti-ZO-1 and analyzed by immunofluorescence microscopy (10x magnification). (I) Quantitative analysis of ZO-1 staining demonstrates the exoprotein-dependency of ZO-1 disruption during S. aureus infection. The y-axes represent percent immunopositivity for anti-ZO-1 from 5 randomly-selected cells from each of N≥10 separate fields (S. aureus 8325–4 infected RPE cells versus S. aureus RN6390 agr/sar infected at 8 hours postinfection, *P<0.0001). (J) Alterations in the permeability of a cultured RPE barrier are dependent on S. aureus exoprotein production. Intact monolayers of human RPE cells in 0.4 micron transwells were infected with S. aureus 8325–4 or RN6390 agr/sar at a concentration of 104 cfu/ml (MOI = 0.01). After 4–8 hours of infection, diffusion of FITC-4-kDa-dextran across the monolayer was assessed by fluorescence spectrometry of media from the bottom chamber. After 8 hours, the fluorescence intensity in the bottom chamber media was significantly greater after infection with 8325–4 than after infection with the agr/sar-deficient strain (*P<0.0001). Values represent the mean concentration of the conjugate in the bottom chamber ± the SD (N≥3 at each time point) based on extrapolation from a standard curve of the fluorimetry of known FITC-dextran concentrations.