<i>Aspergillus fumigatus</i> Hypoxia Adaptation Is Critical for the Establishment of Fungal Keratitis

Lightfoot, Jorge D.; Adams, Emily M.; Kamath, Manali M.; Wells, Becca L.; Fuller, Kevin K.

doi:10.1167/iovs.65.4.31

Cited by 3 publications

(1 citation statement)

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“…They also observed that during the progression of the infection, as the corneal edema progressed, the degree of hypoxia increased, and the endothelium decreased. Later, in the following stages, the accumulation of fungal metabolites together with leukocyte ROS/cytokines caused further loss of endothelial integrity and subsequent entry of fluid from the anterior chamber [66].…”

Section: Immune Mechanismsmentioning

confidence: 99%

Immune Mechanisms of Filamentous Fungal Keratitis

Mpakosi,

Kaliouli-Antonopoulou

2024

Cureus

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Filamentous fungal keratitis is a particularly serious eye infection that often results in ulceration, corneal perforation, and blindness. The cornea acts as a natural barrier against harmful agents due to the close connection of its epithelial cells. In addition, on its surface, there is a large number of substances with antiinflammatory and bactericidal properties, such as secretory IgA and mucin glycoproteins, and antimicrobial peptides (AMPs), such as human β-defensin 2 (HBD-2) and LL-37, which are especially increased in filamentous fungal keratitis.The interaction between pathogenic fungi and the host's immune mechanisms is a complex process: pathogen-associated molecular pattern (PAMP) molecules (chitin, β-glucan, and mannan) found in the fungal cell wall are recognized by pattern recognition receptors (PRRs) (toll-like receptors {TLRs}, C-type lectin receptors {CLRs}, nucleotide-binding oligomerization domain-like receptors {NLRs}, and scavenger receptors {SR}) found in host defense cells, triggering the secretion of various types of cytokines, such as interleukins (IL), tumor necrosis factors (TNFs), and chemokines, which recruit macrophages and neutrophils to migrate to the site of infection and activate inflammatory responses.In addition, the interaction of hyphae and corneal epithelial cells can activate cluster of differentiation (CD) 4+ T cells, CD8+ T cells, and B cells and induce secretion of T-helper (Th)-type cytokines 2 (IL-4 and IL-13) and IgG.

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Section: Immune Mechanismsmentioning

confidence: 99%

Immune Mechanisms of Filamentous Fungal Keratitis

Mpakosi,

Kaliouli-Antonopoulou

2024

Cureus

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The mammalian Ire1 inhibitor, 4µ8C, exhibits broad anti-Aspergillusactivityin vitroand in a treatment model of fungal keratitis

Kamath,

Adams,

Lightfoot

et al. 2024

Preprint

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ObjectiveThe fungal unfolded protein response consists of a two-component relay in which the ER-bound sensor, IreA, splices and activates the mRNA of the transcription factor, HacA. Previously, we demonstrated thathacAis essential forAspergillus fumigatusvirulence in a murine model of fungal keratitis (FK), suggesting the pathway could serve as a therapeutic target. Here we investigate the antifungal properties of known inhibitors of the mammalian Ire1 protein bothin vitroand in a treatment model of FK.MethodsThe antifungal activity of Ire1 inhibitors was tested against conidia of severalA. fumigatusisolates by a microbroth dilution assay and against fungal biofilm by XTT reduction. The influence of 4μ8C onhacAmRNA splicing inA. fumigatuswas assessed through gel electrophoresis and qRT-PCR of UPR regulatory genes. The toxicity and antifungal profile of 4μ8C in the cornea was assessed by applying drops to uninfected orA. fumigatus-infected corneas 3 times daily starting 4 hours post-inoculation. Corneas were evaluated daily through slit-lamp imaging and optical coherence tomography, or at endpoint through histology or fungal burden quantification via colony forming units.ResultsAmong six Ire1 inhibitors screened, the endonuclease inhibitor 4μ8C displayed the strongest antifungal profile with an apparent fungicidal action. The compound both blocked conidial germination and hyphal metabolism ofA. fumigatusAf293 in the same concentration range that blockedhacAsplicing and UPR gene induction (60-120 µM). Topical treatment of sham-inoculated corneas with 0.5 and 2.5 mM 4μ8C did not impact corneal clarity, but did transiently inhibit epithelialization of corneal ulcers. Relative to vehicle-treated Af293-infected corneas, treatment with 0.5 and 2.5 mM drug resulted in a 50% and >90% reduction in fungal load, respectively, the latter of which corresponded to an absence of clinical signs of infection or corneal pathology.ConclusionThein vitrodata suggest that 4μ8C displays antifungal activity againstA. fumigatusthrough the specific inhibition of IreA. Topical application of the compound to the murine cornea can furthermore block the establishment of infection, suggesting this class of drugs can be developed as novel antifungals that improve visual outcomes in FK patients.

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The mammalian Ire1 inhibitor, 4µ8C, exhibits broad anti-Aspergillus activity in vitro and in a treatment model of fungal keratitis

Kamath,

Adams,

Lightfoot

et al. 2024

Front. Cell. Infect. Microbiol.

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ObjectiveThe fungal unfolded protein response consists of a two-component relay in which the ER-bound sensor, IreA, splices and activates the mRNA of the transcription factor, HacA. Previously, we demonstrated that hacA is essential for Aspergillus fumigatus virulence in a murine model of fungal keratitis (FK), suggesting the pathway could serve as a therapeutic target. Here we investigate the antifungal properties of known inhibitors of the mammalian Ire1 protein both in vitro and in a treatment model of FK.MethodsThe antifungal activity of Ire1 inhibitors was tested against conidia of several A. fumigatus isolates by a broth microdilution assay and against fungal biofilm by XTT reduction. The influence of 4μ8C on hacA mRNA splicing in A. fumigatus was assessed through gel electrophoresis and qRT-PCR of UPR regulatory genes. The toxicity and antifungal profile of 4μ8C in the cornea was assessed by applying drops to uninfected or A. fumigatus-infected corneas 3 times daily starting 4 hours post-inoculation. Corneas were evaluated daily through slit-lamp imaging and optical coherence tomography, or at endpoint through histology or fungal burden quantification via colony forming units.ResultsAmong six Ire1 inhibitors screened, the endonuclease inhibitor 4μ8C displayed the strongest antifungal profile with an apparent fungicidal action. The compound both blocked conidial germination and hyphal metabolism of A. fumigatus Af293 in the same concentration range that blocked hacA splicing and UPR gene induction (60-120 µM). Topical treatment of sham-inoculated corneas with 0.5 and 2.5 mM 4μ8C did not impact corneal clarity, but did transiently inhibit epithelialization of corneal ulcers. Relative to vehicle-treated Af293-infected corneas, treatment with 0.5 and 2.5 mM drug resulted in a 50% and >90% reduction in fungal load, respectively, the latter of which corresponded to an absence of clinical signs of infection or corneal pathology.ConclusionThe in vitro data suggest that 4μ8C displays antifungal activity against A. fumigatus through the specific inhibition of IreA. Topical application of the compound to the murine cornea can furthermore block the establishment of infection, suggesting this class of drugs can be developed as novel antifungals that improve visual outcomes in FK patients.

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Aspergillus fumigatus Hypoxia Adaptation Is Critical for the Establishment of Fungal Keratitis

Cited by 3 publications

References 85 publications

Immune Mechanisms of Filamentous Fungal Keratitis

Immune Mechanisms of Filamentous Fungal Keratitis

The mammalian Ire1 inhibitor, 4µ8C, exhibits broad anti-Aspergillusactivityin vitroand in a treatment model of fungal keratitis

The mammalian Ire1 inhibitor, 4µ8C, exhibits broad anti-Aspergillus activity in vitro and in a treatment model of fungal keratitis

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