Differentially expressed wound healing-related microRNAs in the human diabetic cornea.

MicroRNAs are powerful gene expression regulators, but their corneal repertoire and potential changes in corneal diseases remain unknown. Our purpose was to identify miRNAs altered in the human diabetic cornea by microarray analysis, and to examine their effects on wound healing in cultured telomera...

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Autores principales:	Vincent A Funari, Michael Winkler, Jordan Brown, Slobodan D Dimitrijevich, Alexander V Ljubimov, Mehrnoosh Saghizadeh
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2013
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/08866020531440b18efb714b45403f1a
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spelling	oai:doaj.org-article:08866020531440b18efb714b45403f1a2021-11-18T08:40:48ZDifferentially expressed wound healing-related microRNAs in the human diabetic cornea.1932-620310.1371/journal.pone.0084425https://doaj.org/article/08866020531440b18efb714b45403f1a2013-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/24376808/?tool=EBIhttps://doaj.org/toc/1932-6203MicroRNAs are powerful gene expression regulators, but their corneal repertoire and potential changes in corneal diseases remain unknown. Our purpose was to identify miRNAs altered in the human diabetic cornea by microarray analysis, and to examine their effects on wound healing in cultured telomerase-immortalized human corneal epithelial cells (HCEC) in vitro. Total RNA was extracted from age-matched human autopsy normal (n=6) and diabetic (n=6) central corneas, Flash Tag end-labeled, and hybridized to Affymetrix® GeneChip® miRNA Arrays. Select miRNAs associated with diabetic cornea were validated by quantitative RT-PCR (Q-PCR) and by in situ hybridization (ISH) in independent samples. HCEC were transfected with human pre-miR™miRNA precursors (h-miR) or their inhibitors (antagomirs) using Lipofectamine 2000. Confluent transfected cultures were scratch-wounded with P200 pipette tip. Wound closure was monitored by digital photography. Expression of signaling proteins was detected by immunostaining and Western blot. Using microarrays, 29 miRNAs were identified as differentially expressed in diabetic samples. Two miRNA candidates showing the highest fold increased in expression in the diabetic cornea were confirmed by Q-PCR and further characterized. HCEC transfection with h-miR-146a or h-miR-424 significantly retarded wound closure, but their respective antagomirs significantly enhanced wound healing vs. controls. Cells treated with h-miR-146a or h-miR-424 had decreased p-p38 and p-EGFR staining, but these increased over control levels close to the wound edge upon antagomir treatment. In conclusion, several miRNAs with increased expression in human diabetic central corneas were found. Two such miRNAs inhibited cultured corneal epithelial cell wound healing. Dysregulation of miRNA expression in human diabetic cornea may be an important mediator of abnormal wound healing.Vincent A FunariMichael WinklerJordan BrownSlobodan D DimitrijevichAlexander V LjubimovMehrnoosh SaghizadehPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 8, Iss 12, p e84425 (2013)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Vincent A Funari Michael Winkler Jordan Brown Slobodan D Dimitrijevich Alexander V Ljubimov Mehrnoosh Saghizadeh Differentially expressed wound healing-related microRNAs in the human diabetic cornea.
description	MicroRNAs are powerful gene expression regulators, but their corneal repertoire and potential changes in corneal diseases remain unknown. Our purpose was to identify miRNAs altered in the human diabetic cornea by microarray analysis, and to examine their effects on wound healing in cultured telomerase-immortalized human corneal epithelial cells (HCEC) in vitro. Total RNA was extracted from age-matched human autopsy normal (n=6) and diabetic (n=6) central corneas, Flash Tag end-labeled, and hybridized to Affymetrix® GeneChip® miRNA Arrays. Select miRNAs associated with diabetic cornea were validated by quantitative RT-PCR (Q-PCR) and by in situ hybridization (ISH) in independent samples. HCEC were transfected with human pre-miR™miRNA precursors (h-miR) or their inhibitors (antagomirs) using Lipofectamine 2000. Confluent transfected cultures were scratch-wounded with P200 pipette tip. Wound closure was monitored by digital photography. Expression of signaling proteins was detected by immunostaining and Western blot. Using microarrays, 29 miRNAs were identified as differentially expressed in diabetic samples. Two miRNA candidates showing the highest fold increased in expression in the diabetic cornea were confirmed by Q-PCR and further characterized. HCEC transfection with h-miR-146a or h-miR-424 significantly retarded wound closure, but their respective antagomirs significantly enhanced wound healing vs. controls. Cells treated with h-miR-146a or h-miR-424 had decreased p-p38 and p-EGFR staining, but these increased over control levels close to the wound edge upon antagomir treatment. In conclusion, several miRNAs with increased expression in human diabetic central corneas were found. Two such miRNAs inhibited cultured corneal epithelial cell wound healing. Dysregulation of miRNA expression in human diabetic cornea may be an important mediator of abnormal wound healing.
format	article
author	Vincent A Funari Michael Winkler Jordan Brown Slobodan D Dimitrijevich Alexander V Ljubimov Mehrnoosh Saghizadeh
author_facet	Vincent A Funari Michael Winkler Jordan Brown Slobodan D Dimitrijevich Alexander V Ljubimov Mehrnoosh Saghizadeh
author_sort	Vincent A Funari
title	Differentially expressed wound healing-related microRNAs in the human diabetic cornea.
title_short	Differentially expressed wound healing-related microRNAs in the human diabetic cornea.
title_full	Differentially expressed wound healing-related microRNAs in the human diabetic cornea.
title_fullStr	Differentially expressed wound healing-related microRNAs in the human diabetic cornea.
title_full_unstemmed	Differentially expressed wound healing-related microRNAs in the human diabetic cornea.
title_sort	differentially expressed wound healing-related micrornas in the human diabetic cornea.
publisher	Public Library of Science (PLoS)
publishDate	2013
url	https://doaj.org/article/08866020531440b18efb714b45403f1a
work_keys_str_mv	AT vincentafunari differentiallyexpressedwoundhealingrelatedmicrornasinthehumandiabeticcornea AT michaelwinkler differentiallyexpressedwoundhealingrelatedmicrornasinthehumandiabeticcornea AT jordanbrown differentiallyexpressedwoundhealingrelatedmicrornasinthehumandiabeticcornea AT slobodanddimitrijevich differentiallyexpressedwoundhealingrelatedmicrornasinthehumandiabeticcornea AT alexandervljubimov differentiallyexpressedwoundhealingrelatedmicrornasinthehumandiabeticcornea AT mehrnooshsaghizadeh differentiallyexpressedwoundhealingrelatedmicrornasinthehumandiabeticcornea
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Differentially expressed wound healing-related microRNAs in the human diabetic cornea.

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