Disruption of staphylococcal aggregation protects against lethal lung injury
Jaime L. Hook, … , Sunita Bhattacharya, Jahar Bhattacharya
Jaime L. Hook, … , Sunita Bhattacharya, Jahar Bhattacharya
Published February 12, 2018
Citation Information: J Clin Invest. 2018;128(3):1074-1086. https://doi.org/10.1172/JCI95823.
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Research Article Cell biology Pulmonology

Disruption of staphylococcal aggregation protects against lethal lung injury

Abstract

Infection by Staphylococcus aureus strain USA300 causes tissue injury, multiorgan failure, and high mortality. However, the mechanisms by which the bacteria adhere to, then stabilize on, mucosal surfaces before causing injury remain unclear. We addressed these issues through the first real-time determinations of USA300-alveolar interactions in live lungs. We found that within minutes, inhaled USA300 established stable, self-associated microaggregates in niches at curved, but not at flat, regions of the alveolar wall. The microaggregates released α-hemolysin toxin, causing localized alveolar injury, as indicated by epithelial dye loss, mitochondrial depolarization, and cytosolic Ca2+ increase. Spread of cytosolic Ca2+ through intercellular gap junctions to adjoining, uninfected alveoli caused pulmonary edema. Systemic pretreatment with vancomycin, a USA300-cidal antibiotic, failed to protect mice infected with inhaled WT USA300. However, vancomycin pretreatment markedly abrogated mortality in mice infected with mutant USA300 that lacked the aggregation-promoting factor PhnD. We interpret USA300-induced mortality as having resulted from rapid bacterial aggregation in alveolar niches. These findings indicate, for the first time to our knowledge, that alveolar microanatomy is critical in promoting the aggregation and, hence, in causing USA300-induced alveolar injury. We propose that in addition to antibiotics, strategies for bacterial disaggregation may constitute novel therapy against USA300-induced lung injury.

Authors

Jaime L. Hook, Mohammad N. Islam, Dane Parker, Alice S. Prince, Sunita Bhattacharya, Jahar Bhattacharya

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Figure 5

USA300 MAs rapidly induce Hla- and Ca2+-dependent loss of alveolar function.

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USA300 MAs rapidly induce Hla- and Ca2+-dependent loss of alveolar funct...
Confocal images and quantifications show fluorescence in live alveoli. All bars: mean ± SEM; *P < 0.05 as indicated using ANOVA with Bonferroni correction. (A–G) A shows an alveolus (alv) with epithelial tetramethylrhodamine, ethyl ester (TMRE), fluorescence. B and C show the marked region (dashed rectangle) at high magnification, before (B) and 1 hour after (C) microinstillation of WT USA300 (WT). D and E are high-magnification regions of other alveoli that received Hla-deficient USA300 (hla–) or BAPTA pretreatment, respectively. Bacterial fluorescence was digitally removed in C–E. Dashed regions indicate MA locations. Scale bars: 30 (A) and 5 (B–E) μm. Plots (F) and bars (G) show effects of the indicated treatments on fluorescence at MA-associated epithelial sites. Plots are representative for individual sites. Bars show group data 1 hour after the indicated microinstillations; n = 3 lungs (3 sites quantified per lung). (H–M) H shows fluorescence of alveolar type 2 (AT2) cell lamellar bodies (LBs) loaded with LysoTracker Red (LTR). We microinstilled alveoli as indicated, transiently hyperinflated the lung, then obtained images I–K. LB fluorescence loss indicates hyperinflation-induced surfactant secretion. Bacterial fluorescence was digitally removed in J and K. Dashed regions indicate MA locations. Scale bars: 5 μm. Plots (L) are representative for individual cells. Bars (M) show group data 1 hour after the indicated microinstillations; n = 3 lungs (5 cells quantified per lung). (N–Q) Alveoli were pretreated with vehicle (N and O) or BAPTA (P), then microinstilled as indicated. After 6 hours, we gave intravascular 20-kDa dextran (green), then obtained the images. MAs are indicated in red. Arrows indicate dextran-filled airspaces. FD20, FITC-labeled dextran. Scale bars: 50 μm. Bars (Q): n = 3 lungs (10 alveoli quantified per lung).