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SFRP1 promoter methylation and expression in human trabecular meshwork cells
Weiming Mao1, Jeffrey S Rubin, Nancy Anoruo
1North Texas Eye Research Institute, Department of Cell Biology & Anatomy, University of North Texas Health Science Center, CBH440, Fort Worth, TX 76107, USA. weiming.mao@unthsc.edu
Experimental Eye Research
|January 18, 2012
Summary
Elevated secreted frizzled-related protein 1 (SFRP1) in glaucoma may not stem from promoter methylation. This study found SFRP1 is highly expressed in glaucomatous human trabecular meshwork (HTM) cells, but their promoters remain unmethylated.
Area of Science:
- Ophthalmology
- Molecular Biology
- Genetics
Background:
- Primary open angle glaucoma (POAG) involves impaired trabecular meshwork (TM) function, leading to elevated intraocular pressure (IOP) and optic neuropathy.
- Secreted frizzled-related protein 1 (SFRP1), a Wnt signaling inhibitor, is elevated in glaucomatous TM (GTM) and linked to increased IOP.
Purpose of the Study:
- To investigate whether differential promoter methylation contributes to elevated SFRP1 expression in GTM cells.
- To determine if DNA methylation inhibitors impact SFRP1 expression in human TM (HTM) cells.
Main Methods:
- Cultured normal TM (NTM) and GTM cell strains.
- Quantified SFRP1 expression using quantitative PCR (qPCR).
- Assessed SFRP1 promoter methylation status via bisulfite conversion and methylation-sensitive high-resolution melting analysis (MS-HRM).
- Treated TM cells with 5-aza-2'-deoxycytidine (AZA-dC) to assess the effect of DNA methylation inhibitors on SFRP1 expression.
Main Results:
- GTM cell strains exhibited significantly higher SFRP1 expression compared to NTM cell strains.
- SFRP1 promoter regions were unmethylated in all tested TM cell strains.
- AZA-dC treatment did not alter SFRP1 expression in TM cells, unlike in NCI-H460 cancer cells (positive control).
Conclusions:
- Differential expression of SFRP1 in HTM cells is not regulated by differential promoter methylation.
- Alternative epigenetic or regulatory mechanisms may be responsible for elevated SFRP1 in glaucomatous TM.
