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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...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
The Association for Research in Vision and Ophthalmology
2021
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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. |
format | Online Article Text |
id | pubmed-8684302 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | The Association for Research in Vision and Ophthalmology |
record_format | MEDLINE/PubMed |
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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