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Updated: Jan 19, 2026

Continuous Fluorescence-Based Endonuclease-Coupled DNA Methylation Assay to Screen for DNA Methyltransferase Inhibitors
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Continuous Fluorescence-Based Endonuclease-Coupled DNA Methylation Assay to Screen for DNA Methyltransferase Inhibitors

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DNA methyltransferase inhibitor suppresses fibrogenetic changes in human conjunctival fibroblasts

Hitomi Yonemura1, Akiko Futakuchi1, Miyuki Inoue-Mochita1

  • 1Department of Ophthalmology, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan.

Molecular Vision
|September 17, 2019
PubMed
Abstract

Insights

A DNA methyltransferase inhibitor partially suppressed fibrogenetic changes in human conjunctival fibroblasts. This suggests an indirect inhibitory effect on the COL1A2 promoter, impacting collagen production.

Area of Science:

  • Ophthalmology
  • Cell Biology
  • Molecular Biology

Background:

  • Fibrogenesis, the process of forming scar tissue, is a significant concern in ocular surface diseases.
  • Human conjunctival fibroblasts (HConF) play a crucial role in extracellular matrix production and fibrotic responses.
  • DNA methyltransferase inhibitors are being explored for their potential to modulate cellular processes.

Purpose of the Study:

  • To investigate the impact of a DNA methyltransferase inhibitor on fibrogenetic changes in human conjunctival fibroblasts (HConF).
  • To determine if 5-aza-2'-deoxycytidine (5-Aza-dC) can inhibit transforming growth factor (TGF)-β2-induced fibrogenesis in HConF.

Main Methods:

  • HConF were treated with 5-aza-2'-deoxycytidine (5-Aza-dC) followed by transforming growth factor (TGF)-β2 stimulation.
  • Expression levels of α-smooth muscle actin (α-SMA) and extracellular matrix proteins were assessed using western blotting.
  • COL1A2 promoter activity was evaluated using a luciferase reporter gene assay.

Main Results:

  • 5-Aza-dC pretreatment dose-dependently suppressed TGF-β2-induced α-SMA upregulation.
  • Type I collagen upregulation was inhibited by 10 μM 5-Aza-dC.
  • 5-Aza-dC inhibited COL1A2 promoter activity but did not affect fibronectin or phosphorylated Smad3 expression.

Conclusions:

  • DNA methyltransferase inhibition partially suppressed fibrogenetic changes in HConF.
  • The findings suggest an indirect inhibitory mechanism of 5-Aza-dC on the COL1A2 promoter in HConF.
  • This study provides insights into potential therapeutic strategies targeting fibrotic pathways in the conjunctiva.

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