Dectin-1 is essential for IL-1β production through JNK activation and apoptosis in Aspergillus fumigatus keratitis

Yuan, Kelan; Zhao, Guiqiu; Che, Chengye; Li, Cui; Lin, Jing; Zhu, Guoqiang; He, Kun

doi:10.1016/j.intimp.2017.09.008

Cited by 34 publications

(20 citation statements)

References 35 publications

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“…The immunofluorescence protocol used in this experiment is described in previous publications. 11 The primary antibody used was 10 μg/mL rat anti-mouse NIMP-R14 (Santa Cruz), and Alexa Fluor 488-conjugated goat anti-rat antibody (1:1000; CST) was used as a secondary antibody.…”

Section: Methodsmentioning

confidence: 99%

“…The PCR protocol used in this study is described in previous publications. 11 The primer pair sequences were as follows: hLOX-1 F-GCT CCT CCT GAG CGC AAG and R-CAT CTG CTG GAA GGT GGA CA, hIL-1β F-ATG CAC CTG TAC GAT CAC TGA and R-ACA AAG GAC ATG GAG AAC ACC, hβ-actin F-GCT CCT CCT GAG CGC AAG and R-CAT CTG CTG GAA GGT GGA CA, mLOX-1 F-AGG TCC TTG TCC ACA AGA CTG G and R-ACG CCC CTG GTC TTA AAG AAT TG, mIL-1β F-CGC AGC AGC ACA TCA ACA AGA GC and R-TGT CCT CAT CCT GGA AGG TCC ACG, mBcl-2 F-GGA CTT GAA GTG CCA TTG GT and R-AGC CCC TCT GTG ACA GCT TA, mBax F-CGA GCT GAT CAG AAC CAT CA and R-CTC AGC CCA TCT TCT TCC AG, mCyt-c F-AGT CCT TGG GCA CAG CAG TTG and R-GGC ACT GAG CAC ATT TCT GAA CA, mCaspase-9 F-AGT TCC CGG GTG CTG TCT AT and R-GCC ATG GTC TTT CTG CTC AC, mCaspase-8 F-CTC CGA AAA ATG AAG GAC AGA and R-CGT GGG ATA GGA TAC AGC AGA, mCaspase-3 F-TGG GCC TGA AAT ACC AAG TC and R-AAA TGA CCC CTT CAT CAC CA, and mβ-actin F-GAT TAC TGC TCT GGC TCC TAG C and R-GAC TCA TCG TAC TCC TGC TTG C.…”

Section: Methodsmentioning

confidence: 99%

“…The western blotting protocol that is used to assay corneas and cells is described in previous publications. 11 Membranes were incubated accordingly using the following antibodies: anti-LOX-1 (ProteinTech, Chicago, IL), anti-JNK (ProteinTech), anti-pJNK (CST), anti-mIL-1β (R&D, Minneapolis, MN), anti-hIL-1β (CST, Danvers, MA), anti-Bcl-2 (ProteinTech), anti-Bax (ProteinTech), anti-cytochrome c (ProteinTech), anti-cleaved-CASP9 (ProteinTech), anti-cleaved-CASP8 (ProteinTech), anti-cleaved-CASP3 (ProteinTech), and anti-β-actin (CST). The HRP-tagged secondary anti-bodies were purchased from CST.…”

Section: Methodsmentioning

confidence: 99%

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Fenretinide Inhibits Neutrophil Recruitment and IL-1β Production in Aspergillus fumigatus Keratitis

Zhao

Che

Liu

et al. 2018

Cornea

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Purpose:Fungal keratitis is a major cause of corneal ulcers, resulting in significant visual impairment and blindness. Fenretinide, a derivative of vitamin A, has been shown to suppress inflammation in a multitude of diseases. In this study, we aimed to characterize the effect of fenretinide in Aspergillus fumigatus keratitis of the eye in a mouse model.Methods:In vivo and in vitro experiments were performed in mouse models and THP-1 macrophage cell cultures infected with A. fumigatus, respectively. Experimental subjects were first pretreated with fenretinide, and then the effect of the compound was assessed with clinical evaluation, neutrophil staining, myeloperoxidase assay, quantitative polymerase chain reaction (qRT-PCR), and western blot.Results:We confirmed that fenretinide contributed to protection of corneal transparency during early mouse A. fumigatus keratitis by reducing neutrophil recruitment, decreasing myeloperoxidase (MPO) levels and increasing apoptosis. Compared with controls, fenretinide impaired proinflammatory cytokine interleukin 1 beta (IL-1β) production in response to A. fumigatus exposure with contributions by lectin-type oxidized LDL receptor 1 (LOX-1) and c-Jun N-terminal kinase (JNK).Conclusions:Together, these findings demonstrate that fenretinide may suppress inflammation through reduced neutrophil recruitment and inflammatory cytokine production in A. fumigatus keratitis.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Fenretinide Inhibits Neutrophil Recruitment and IL-1β Production in Aspergillus fumigatus Keratitis

Zhao

Che

Liu

et al. 2018

Cornea

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“…Mice were anaesthetized with 8% chloral hydrate, and then one eye was randomly selected from each mouse. A. fumigatus conidia (2 µ L, concentration 0.5 × 10 5 µ L) were released into the corneal stroma of the mouse [16], and the mice were subsequently examined by a slit lamp microscope for corneal opacification and ulceration. Corneas were harvested for use in western blot assay at 1/2, 1, 2, 3, 5, 7, 10, and 14 days after infection.…”

Section: Methodsmentioning

confidence: 99%

ATF4 Involvement in TLR4 and LOX-1-Induced Host Inflammatory Response to Aspergillus fumigatus Keratitis

Zhang

Meng

Liu

et al. 2018

Journal of Ophthalmology

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Purpose Activating transcription factor 4 (ATF4) is induced by various stressors. Here, we investigated the expression of ATF4 in the host inflammatory response to Aspergillus fumigatus (A. fumigatus) keratitis. Methods A. fumigatus keratitis mouse models developed by intrastromal injection as well as corneal epithelium scratching were examined daily with a slit lamp microscope for corneal opacification and ulceration. Subsequent in vitro experimentation was carried out in human corneal epithelial cells (HCECs) as well as THP-1 macrophages infected with A. fumigatus. Inhibitors, including CLI-095, Poly (I), SCH772984, and SP600125, were used to assess the role of proteins like toll-like receptor 4 (TLR4), lectin-type oxidized LDL receptor 1 (LOX-1), extracellular signal-regulated kinases (ERK1/2), and c-Jun N-terminal kinase (JNK) in ATF4 expression as a response to A. fumigatus infection. This assessment was made in both mouse models and HCECs using western blot. Results Compared to the controls, ATF4 was increased in corneas from two kinds of A. fumigatus keratitis models at 3 days after infection. ATF4 expression was upregulated with A. fumigatus conidia both in HCECs and THP-1 macrophages 16 hours after stimulation. Furthermore, ATF4 expression in response to A. fumigatus infection was shown to be dependent on TLR4 and LOX-1 expression, and ERK1/2 and JNK contributed to the expression of ATF4 in response to A. fumigatus. Conclusion Our results clearly indicate that ATF4 was involved in the host antifungal immune response to A. fumigatus keratitis; expression was found to be dependent on TLR4, LOX-1 expression, and MAPKs pathway.

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“…Then, a 33G Hamilton syringe is inserted into the tunnel to deliver a 1-2 µL inoculum directly into the underlying stroma. Regardless of fungal species, 10 5 total conidia is commonly used inoculum, although this can vary by a log-fold either way [1,32,[47][48][49][50][51][52].…”

Section: Intrastromal Injectionmentioning

confidence: 99%

Experimental Models for Fungal Keratitis: An Overview of Principles and Protocols

Montgomery

Fuller

2020

Cells

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Fungal keratitis is a potentially blinding infection of the cornea that afflicts diverse patient populations worldwide. The development of better treatment options requires a more thorough understanding of both microbial and host determinants of pathology, and a spectrum of experimental models have been developed toward this end. In vivo (animal) models most accurately capture complex pathological outcomes, but protocols may be challenging to implement and vary widely across research groups. In vitro models allow for the molecular dissection of specific host cell–fungal interactions, but they do so without the appropriate environmental/structural context; ex vivo (corneal explant) models provide the benefits of intact corneal tissue, but they do not provide certain pathological features, such as inflammation. In this review, we endeavor to outline the key features of these experimental models as well as describe key technical variations that could impact study design and outcomes.

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Dectin-1 is essential for IL-1β production through JNK activation and apoptosis in Aspergillus fumigatus keratitis

Cited by 34 publications

References 35 publications

Fenretinide Inhibits Neutrophil Recruitment and IL-1β Production in Aspergillus fumigatus Keratitis

Fenretinide Inhibits Neutrophil Recruitment and IL-1β Production in Aspergillus fumigatus Keratitis

ATF4 Involvement in TLR4 and LOX-1-Induced Host Inflammatory Response to Aspergillus fumigatus Keratitis

Experimental Models for Fungal Keratitis: An Overview of Principles and Protocols

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