Infection of Human Lung Tissue Explants (HLTEs) with Legionella pneumophila

Scheithauer, Lina; Steinert, Michael

doi:10.1007/978-1-4939-9048-1_21

Cited by 2 publications

(4 citation statements)

References 14 publications

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“…The HLTE infection assay was performed as previously described (Hoppe et al, 2017; Jäger et al, 2014; Scheithauer & Steinert, 2019). Tumour‐free pulmonary tissue samples obtained from patients undergoing lobe‐ or pneumectomy were examined by frozen section analysis.…”

Section: Methodsmentioning

confidence: 99%

Zinc metalloprotease ProA of Legionella pneumophila increases alveolar septal thickness in human lung tissue explants by collagen IV degradation

Scheithauer

Thiem

Schmelz

et al. 2021

Cellular Microbiology

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ProA is a secreted zinc metalloprotease of Legionella pneumophila causing lung damage in animal models of Legionnaires' disease. Here we demonstrate that ProA promotes infection of human lung tissue explants (HLTEs) and dissect the contribution to cell type specific replication and extracellular virulence mechanisms. For the first time, we reveal that co‐incubation of HLTEs with purified ProA causes a significant increase of the alveolar septal thickness. This destruction of connective tissue fibres was further substantiated by collagen IV degradation assays. The moderate attenuation of a proA‐negative mutant in A549 epithelial cells and THP‐1 macrophages suggests that effects of ProA in tissue mainly result from extracellular activity. Correspondingly, ProA contributes to dissemination and serum resistance of the pathogen, which further expands the versatile substrate spectrum of this thermolysin‐like protease. The crystal structure of ProA at 1.48 Å resolution showed high congruence to pseudolysin of Pseudomonas aeruginosa, but revealed deviations in flexible loops, the substrate binding pocket S1′ and the repertoire of cofactors, by which ProA can be distinguished from respective homologues. In sum, this work specified virulence features of ProA at different organisational levels by zooming in from histopathological effects in human lung tissue to atomic details of the protease substrate determination.

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Section: Methodsmentioning

confidence: 99%

Zinc metalloprotease ProA of Legionella pneumophila increases alveolar septal thickness in human lung tissue explants by collagen IV degradation

Scheithauer

Thiem

Schmelz

et al. 2021

Cellular Microbiology

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“…Additionally, the protease is associated with lung lesions and tissue necrosis, and can hinder different types of immune cells [41][42][43]. In a human lung tissue explant model, we earlier demonstrated how effectively L. pneumophila can use the lung as a habitat and lead to substantial tissue destruction [44][45][46]. In this case, ProA massively contributed to the observed damage during infections and induced a phenotype with significantly increased alveolar septal thickness [40].…”

Section: Introductionmentioning

confidence: 94%

“…Tumor-free human lung tissue explants (HLTEs) were obtained from surgery patients and infected as described previously [44][45][46]. RPMI 1640 with 10% (v/v) FBS, 20 mM HEPES and 1 mM sodium pyruvate was inoculated with L. pneumophila Corby from stationary phase cultures at a density of 10 7 bacteria/mL.…”

Section: Collection Of Human Lung Tissue Explant Supernatantmentioning

confidence: 99%

“…For analyzing the ability of L. pneumophila to evade the TLR5-dependent immune response on tissue level, we infected human lung tissue explants (HLTEs) with different Legionella mutant strains. This model was previously demonstrated to be capable of revealing intra-and extracellular aspects of the host-pathogen interaction during Legionnaires' disease via bacterial growth kinetics, transmigration and localization as well as histopathological alterations of the lung tissue architecture [44][45][46]. Therefore, it is well suitable to compare pro-inflammatory host cell responses in infections with the wild type (WT), a proA-and a flaA-negative mutant (Figure 4).…”

Section: Proa Diminishes Extracellular Flaa Stimuli During Human Lung...mentioning

confidence: 99%

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Zinc Metalloprotease ProA from Legionella pneumophila Inhibits the Pro-Inflammatory Host Response by Degradation of Bacterial Flagellin

et al. 2022

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The environmental bacterium Legionella pneumophila is an intracellular pathogen of various protozoan hosts and able to cause Legionnaires’ disease, a severe pneumonia in humans. By encoding a wide selection of virulence factors, the infectious agent possesses several strategies to manipulate its host cells and evade immune detection. In the present study, we demonstrate that the L. pneumophila zinc metalloprotease ProA functions as a modulator of flagellin-mediated TLR5 stimulation and subsequent activation of the pro-inflammatory NF-κB pathway. We found ProA to be capable of directly degrading immunogenic FlaA monomers but not the polymeric form of bacterial flagella. These results indicate a role of the protease in antagonizing immune stimulation, which was further substantiated in HEK-BlueTM hTLR5 Detection assays. Addition of purified proteins, bacterial suspensions of L. pneumophila mutant strains as well as supernatants of human lung tissue explant infection to this reporter cell line demonstrated that ProA specifically decreases the TLR5 response via FlaA degradation. Conclusively, the zinc metalloprotease ProA serves as a powerful regulator of exogenous flagellin and presumably creates an important advantage for L. pneumophila proliferation in mammalian hosts by promoting immune evasion.

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Infection of Human Lung Tissue Explants (HLTEs) with Legionella pneumophila

Cited by 2 publications

References 14 publications

Zinc metalloprotease ProA of Legionella pneumophila increases alveolar septal thickness in human lung tissue explants by collagen IV degradation

Zinc metalloprotease ProA of Legionella pneumophila increases alveolar septal thickness in human lung tissue explants by collagen IV degradation

Zinc Metalloprotease ProA from Legionella pneumophila Inhibits the Pro-Inflammatory Host Response by Degradation of Bacterial Flagellin

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