HDAC inhibitor-mediated epigenetic regulation of glaucoma-associated TGFβ2 in the trabecular meshwork

Jaclyn Y. Bermudez, Hannah C. Webber, Gaurang C. Patel, Xiangyang Liu, Yi Qiang Cheng, Abbot F. Clark, Weiming Mao

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

PURPOSE. Elevated intraocular pressure (IOP) in primary open-angle glaucoma (POAG) results from glaucomatous damage to the trabecular meshwork (TM). The glaucoma-associated factor TGFβ2 is increased in aqueous humor and TM of POAG patients. We hypothesize that histone acetylation has a role in dysregulated TGFβ2 expression. METHODS. Protein acetylation was compared between nonglaucomatous TM (NTM) and glaucomatous TM (GTM) cells using Western immunoblotting (WB). Nonglaucomatous TM cells were treated with 10 nM thailandepsin-A (TDP-A), a potent histone deacetylase inhibitor for 4 days. Total and nuclear proteins, RNA, and nuclear protein-DNA complexes were harvested for WB, quantitative PCR (qPCR), and chromatin immunoprecipitation (ChIP) assays, respectively. Paired bovine eyes were perfused with TDP-A versus DMSO, or TDP-A versus TDP-A plus the TGFβ pathway inhibitor LY364947 for 5 to 9 days. Intraocular pressure, TM, and perfusate proteins were compared. RESULTS. We found increased acetylated histone 3 and total protein acetylation in the GTM cells and TDP-A treated NTM cells. Chromatin immunoprecipitation assays showed that TDPA induced histone hyperacetylation associated with the TGFβ2 promoter. This change of acetylation significantly increased TGFβ2 mRNA and protein expression in NTM cells. In perfusion-cultured bovine eyes, TDP-A increased TGFβ2 in the perfusate as well as elevated IOP. Histologic and immunofluorescent analyses showed increased extracellular matrix and cytoskeletal proteins in the TM of TDP-A treated bovine eyes. Cotreatment with the TGFβ pathway inhibitor LY364947 blocked TDP-A–induced ocular hypertension. CONCLUSIONS. Our results suggest that histone acetylation has an important role in increased expression of the glaucoma-associated factor TGFβ2. Histone hyperacetylation may be the initiator of glaucomatous damage to the TM.

Original languageEnglish (US)
Pages (from-to)3698-3707
Number of pages10
JournalInvestigative Ophthalmology and Visual Science
Volume57
Issue number8
DOIs
StatePublished - Jul 1 2016
Externally publishedYes

Fingerprint

Trabecular Meshwork
Histone Deacetylase Inhibitors
Epigenomics
Glaucoma
Acetylation
Histones
Intraocular Pressure
Chromatin Immunoprecipitation
Nuclear Proteins
Proteins
Nuclear RNA
Ocular Hypertension
Cytoskeletal Proteins
Aqueous Humor
Extracellular Matrix Proteins
Dimethyl Sulfoxide
thailandepsin A
Perfusion
Western Blotting
Polymerase Chain Reaction

Keywords

  • Epigenetics
  • Glaucoma
  • Ocular hypertension
  • TGFb2
  • Trabecular meshwork

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

Cite this

HDAC inhibitor-mediated epigenetic regulation of glaucoma-associated TGFβ2 in the trabecular meshwork. / Bermudez, Jaclyn Y.; Webber, Hannah C.; Patel, Gaurang C.; Liu, Xiangyang; Cheng, Yi Qiang; Clark, Abbot F.; Mao, Weiming.

In: Investigative Ophthalmology and Visual Science, Vol. 57, No. 8, 01.07.2016, p. 3698-3707.

Research output: Contribution to journalArticle

Bermudez, Jaclyn Y. ; Webber, Hannah C. ; Patel, Gaurang C. ; Liu, Xiangyang ; Cheng, Yi Qiang ; Clark, Abbot F. ; Mao, Weiming. / HDAC inhibitor-mediated epigenetic regulation of glaucoma-associated TGFβ2 in the trabecular meshwork. In: Investigative Ophthalmology and Visual Science. 2016 ; Vol. 57, No. 8. pp. 3698-3707.
@article{f650f1246587402fac71e14d150db246,
title = "HDAC inhibitor-mediated epigenetic regulation of glaucoma-associated TGFβ2 in the trabecular meshwork",
abstract = "PURPOSE. Elevated intraocular pressure (IOP) in primary open-angle glaucoma (POAG) results from glaucomatous damage to the trabecular meshwork (TM). The glaucoma-associated factor TGFβ2 is increased in aqueous humor and TM of POAG patients. We hypothesize that histone acetylation has a role in dysregulated TGFβ2 expression. METHODS. Protein acetylation was compared between nonglaucomatous TM (NTM) and glaucomatous TM (GTM) cells using Western immunoblotting (WB). Nonglaucomatous TM cells were treated with 10 nM thailandepsin-A (TDP-A), a potent histone deacetylase inhibitor for 4 days. Total and nuclear proteins, RNA, and nuclear protein-DNA complexes were harvested for WB, quantitative PCR (qPCR), and chromatin immunoprecipitation (ChIP) assays, respectively. Paired bovine eyes were perfused with TDP-A versus DMSO, or TDP-A versus TDP-A plus the TGFβ pathway inhibitor LY364947 for 5 to 9 days. Intraocular pressure, TM, and perfusate proteins were compared. RESULTS. We found increased acetylated histone 3 and total protein acetylation in the GTM cells and TDP-A treated NTM cells. Chromatin immunoprecipitation assays showed that TDPA induced histone hyperacetylation associated with the TGFβ2 promoter. This change of acetylation significantly increased TGFβ2 mRNA and protein expression in NTM cells. In perfusion-cultured bovine eyes, TDP-A increased TGFβ2 in the perfusate as well as elevated IOP. Histologic and immunofluorescent analyses showed increased extracellular matrix and cytoskeletal proteins in the TM of TDP-A treated bovine eyes. Cotreatment with the TGFβ pathway inhibitor LY364947 blocked TDP-A–induced ocular hypertension. CONCLUSIONS. Our results suggest that histone acetylation has an important role in increased expression of the glaucoma-associated factor TGFβ2. Histone hyperacetylation may be the initiator of glaucomatous damage to the TM.",
keywords = "Epigenetics, Glaucoma, Ocular hypertension, TGFb2, Trabecular meshwork",
author = "Bermudez, {Jaclyn Y.} and Webber, {Hannah C.} and Patel, {Gaurang C.} and Xiangyang Liu and Cheng, {Yi Qiang} and Clark, {Abbot F.} and Weiming Mao",
year = "2016",
month = "7",
day = "1",
doi = "10.1167/iovs16-19446",
language = "English (US)",
volume = "57",
pages = "3698--3707",
journal = "Investigative Ophthalmology and Visual Science",
issn = "0146-0404",
publisher = "Association for Research in Vision and Ophthalmology Inc.",
number = "8",

}

TY - JOUR

T1 - HDAC inhibitor-mediated epigenetic regulation of glaucoma-associated TGFβ2 in the trabecular meshwork

AU - Bermudez, Jaclyn Y.

AU - Webber, Hannah C.

AU - Patel, Gaurang C.

AU - Liu, Xiangyang

AU - Cheng, Yi Qiang

AU - Clark, Abbot F.

AU - Mao, Weiming

PY - 2016/7/1

Y1 - 2016/7/1

N2 - PURPOSE. Elevated intraocular pressure (IOP) in primary open-angle glaucoma (POAG) results from glaucomatous damage to the trabecular meshwork (TM). The glaucoma-associated factor TGFβ2 is increased in aqueous humor and TM of POAG patients. We hypothesize that histone acetylation has a role in dysregulated TGFβ2 expression. METHODS. Protein acetylation was compared between nonglaucomatous TM (NTM) and glaucomatous TM (GTM) cells using Western immunoblotting (WB). Nonglaucomatous TM cells were treated with 10 nM thailandepsin-A (TDP-A), a potent histone deacetylase inhibitor for 4 days. Total and nuclear proteins, RNA, and nuclear protein-DNA complexes were harvested for WB, quantitative PCR (qPCR), and chromatin immunoprecipitation (ChIP) assays, respectively. Paired bovine eyes were perfused with TDP-A versus DMSO, or TDP-A versus TDP-A plus the TGFβ pathway inhibitor LY364947 for 5 to 9 days. Intraocular pressure, TM, and perfusate proteins were compared. RESULTS. We found increased acetylated histone 3 and total protein acetylation in the GTM cells and TDP-A treated NTM cells. Chromatin immunoprecipitation assays showed that TDPA induced histone hyperacetylation associated with the TGFβ2 promoter. This change of acetylation significantly increased TGFβ2 mRNA and protein expression in NTM cells. In perfusion-cultured bovine eyes, TDP-A increased TGFβ2 in the perfusate as well as elevated IOP. Histologic and immunofluorescent analyses showed increased extracellular matrix and cytoskeletal proteins in the TM of TDP-A treated bovine eyes. Cotreatment with the TGFβ pathway inhibitor LY364947 blocked TDP-A–induced ocular hypertension. CONCLUSIONS. Our results suggest that histone acetylation has an important role in increased expression of the glaucoma-associated factor TGFβ2. Histone hyperacetylation may be the initiator of glaucomatous damage to the TM.

AB - PURPOSE. Elevated intraocular pressure (IOP) in primary open-angle glaucoma (POAG) results from glaucomatous damage to the trabecular meshwork (TM). The glaucoma-associated factor TGFβ2 is increased in aqueous humor and TM of POAG patients. We hypothesize that histone acetylation has a role in dysregulated TGFβ2 expression. METHODS. Protein acetylation was compared between nonglaucomatous TM (NTM) and glaucomatous TM (GTM) cells using Western immunoblotting (WB). Nonglaucomatous TM cells were treated with 10 nM thailandepsin-A (TDP-A), a potent histone deacetylase inhibitor for 4 days. Total and nuclear proteins, RNA, and nuclear protein-DNA complexes were harvested for WB, quantitative PCR (qPCR), and chromatin immunoprecipitation (ChIP) assays, respectively. Paired bovine eyes were perfused with TDP-A versus DMSO, or TDP-A versus TDP-A plus the TGFβ pathway inhibitor LY364947 for 5 to 9 days. Intraocular pressure, TM, and perfusate proteins were compared. RESULTS. We found increased acetylated histone 3 and total protein acetylation in the GTM cells and TDP-A treated NTM cells. Chromatin immunoprecipitation assays showed that TDPA induced histone hyperacetylation associated with the TGFβ2 promoter. This change of acetylation significantly increased TGFβ2 mRNA and protein expression in NTM cells. In perfusion-cultured bovine eyes, TDP-A increased TGFβ2 in the perfusate as well as elevated IOP. Histologic and immunofluorescent analyses showed increased extracellular matrix and cytoskeletal proteins in the TM of TDP-A treated bovine eyes. Cotreatment with the TGFβ pathway inhibitor LY364947 blocked TDP-A–induced ocular hypertension. CONCLUSIONS. Our results suggest that histone acetylation has an important role in increased expression of the glaucoma-associated factor TGFβ2. Histone hyperacetylation may be the initiator of glaucomatous damage to the TM.

KW - Epigenetics

KW - Glaucoma

KW - Ocular hypertension

KW - TGFb2

KW - Trabecular meshwork

UR - http://www.scopus.com/inward/record.url?scp=84978530248&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84978530248&partnerID=8YFLogxK

U2 - 10.1167/iovs16-19446

DO - 10.1167/iovs16-19446

M3 - Article

VL - 57

SP - 3698

EP - 3707

JO - Investigative Ophthalmology and Visual Science

JF - Investigative Ophthalmology and Visual Science

SN - 0146-0404

IS - 8

ER -