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Prophylactic Knockdown of the miR-183/96/182 Cluster Ameliorates Pseudomonas aeruginosa–Induced Keratitis

PURPOSE: Previously, we demonstrated that miR-183/96/182 cluster (miR-183C) knockout mice exhibit decreased severity of Pseudomonas aeruginosa (PA)-induced keratitis. This study tests the hypothesis that prophylactic knockdown of miR-183C ameliorates PA keratitis indicative of a therapeutic potentia...

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Autores principales: McClellan, Sharon, Pitchaikannu, Ahalya, Wright, Robert, Bessert, Denise, Iulianelli, Mason, Hazlett, Linda D., Xu, Shunbin
Formato: Online Artículo Texto
Lenguaje:English
Publicado: The Association for Research in Vision and Ophthalmology 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8684302/
https://www.ncbi.nlm.nih.gov/pubmed/34919120
http://dx.doi.org/10.1167/iovs.62.15.14
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author McClellan, Sharon
Pitchaikannu, Ahalya
Wright, Robert
Bessert, Denise
Iulianelli, Mason
Hazlett, Linda D.
Xu, Shunbin
author_facet McClellan, Sharon
Pitchaikannu, Ahalya
Wright, Robert
Bessert, Denise
Iulianelli, Mason
Hazlett, Linda D.
Xu, Shunbin
author_sort McClellan, Sharon
collection PubMed
description PURPOSE: Previously, we demonstrated that miR-183/96/182 cluster (miR-183C) knockout mice exhibit decreased severity of Pseudomonas aeruginosa (PA)-induced keratitis. This study tests the hypothesis that prophylactic knockdown of miR-183C ameliorates PA keratitis indicative of a therapeutic potential. METHODS: Eight-week-old miR-183C wild-type and C57BL/6J inbred mice were used. Locked nucleic acid–modified anti-miR-183C or negative control oligoribonucleotides with scrambled sequences (NC ORNs) were injected subconjunctivally 1 day before and then topically applied once daily for 5 days post-infection (dpi) (strain 19660). Corneal disease was graded at 1, 3, and 5 dpi. Corneas were harvested for RT-PCR, ELISA, immunofluorescence (IF), myeloperoxidase and plate count assays, and flow cytometry. Corneal nerve density was evaluated in flatmounted corneas by IF staining with anti-β-III tubulin antibody. RESULTS: Anti-miR-183C downregulated miR-183C in the cornea. It resulted in an increase in IL-1β at 1 dpi, which was decreased at 5 dpi; fewer polymorphonuclear leukocytes (PMNs) at 5 dpi; lower viable bacterial plate count at both 1 and 5 dpi; increased percentages of MHCII(+) macrophages (Mϕ) and dendritic cells (DCs), consistent with enhanced activation/maturation; and decreased severity of PA keratitis. Anti-miR-183C treatment in the cornea of naïve mice resulted in a transient reduction of corneal nerve density, which was fully recovered one week after the last anti-miR application. miR-183C targets repulsive axon-guidance receptor molecule Neuropilin 1, which may mediate the effect of anti-miR-183C on corneal nerve regression. CONCLUSIONS: Prophylactic miR-183C knockdown is protective against PA keratitis through its regulation of innate immunity, corneal innervation, and neuroimmune interactions.
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spelling pubmed-86843022022-01-06 Prophylactic Knockdown of the miR-183/96/182 Cluster Ameliorates Pseudomonas aeruginosa–Induced Keratitis McClellan, Sharon Pitchaikannu, Ahalya Wright, Robert Bessert, Denise Iulianelli, Mason Hazlett, Linda D. Xu, Shunbin Invest Ophthalmol Vis Sci Cornea PURPOSE: Previously, we demonstrated that miR-183/96/182 cluster (miR-183C) knockout mice exhibit decreased severity of Pseudomonas aeruginosa (PA)-induced keratitis. This study tests the hypothesis that prophylactic knockdown of miR-183C ameliorates PA keratitis indicative of a therapeutic potential. METHODS: Eight-week-old miR-183C wild-type and C57BL/6J inbred mice were used. Locked nucleic acid–modified anti-miR-183C or negative control oligoribonucleotides with scrambled sequences (NC ORNs) were injected subconjunctivally 1 day before and then topically applied once daily for 5 days post-infection (dpi) (strain 19660). Corneal disease was graded at 1, 3, and 5 dpi. Corneas were harvested for RT-PCR, ELISA, immunofluorescence (IF), myeloperoxidase and plate count assays, and flow cytometry. Corneal nerve density was evaluated in flatmounted corneas by IF staining with anti-β-III tubulin antibody. RESULTS: Anti-miR-183C downregulated miR-183C in the cornea. It resulted in an increase in IL-1β at 1 dpi, which was decreased at 5 dpi; fewer polymorphonuclear leukocytes (PMNs) at 5 dpi; lower viable bacterial plate count at both 1 and 5 dpi; increased percentages of MHCII(+) macrophages (Mϕ) and dendritic cells (DCs), consistent with enhanced activation/maturation; and decreased severity of PA keratitis. Anti-miR-183C treatment in the cornea of naïve mice resulted in a transient reduction of corneal nerve density, which was fully recovered one week after the last anti-miR application. miR-183C targets repulsive axon-guidance receptor molecule Neuropilin 1, which may mediate the effect of anti-miR-183C on corneal nerve regression. CONCLUSIONS: Prophylactic miR-183C knockdown is protective against PA keratitis through its regulation of innate immunity, corneal innervation, and neuroimmune interactions. The Association for Research in Vision and Ophthalmology 2021-12-17 /pmc/articles/PMC8684302/ /pubmed/34919120 http://dx.doi.org/10.1167/iovs.62.15.14 Text en Copyright 2021 The Authors https://creativecommons.org/licenses/by-nc-nd/4.0/This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.
spellingShingle Cornea
McClellan, Sharon
Pitchaikannu, Ahalya
Wright, Robert
Bessert, Denise
Iulianelli, Mason
Hazlett, Linda D.
Xu, Shunbin
Prophylactic Knockdown of the miR-183/96/182 Cluster Ameliorates Pseudomonas aeruginosa–Induced Keratitis
title Prophylactic Knockdown of the miR-183/96/182 Cluster Ameliorates Pseudomonas aeruginosa–Induced Keratitis
title_full Prophylactic Knockdown of the miR-183/96/182 Cluster Ameliorates Pseudomonas aeruginosa–Induced Keratitis
title_fullStr Prophylactic Knockdown of the miR-183/96/182 Cluster Ameliorates Pseudomonas aeruginosa–Induced Keratitis
title_full_unstemmed Prophylactic Knockdown of the miR-183/96/182 Cluster Ameliorates Pseudomonas aeruginosa–Induced Keratitis
title_short Prophylactic Knockdown of the miR-183/96/182 Cluster Ameliorates Pseudomonas aeruginosa–Induced Keratitis
title_sort prophylactic knockdown of the mir-183/96/182 cluster ameliorates pseudomonas aeruginosa–induced keratitis
topic Cornea
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8684302/
https://www.ncbi.nlm.nih.gov/pubmed/34919120
http://dx.doi.org/10.1167/iovs.62.15.14
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