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Updated: Jul 23, 2025
A 5-mC Dot Blot Assay Quantifying the DNA Methylation Level of Chondrocyte Dedifferentiation In Vitro
Published on: May 17, 2017
The identification of hub-methylated differentially expressed genes in osteoarthritis patients is based on epigenomic
Zhen-Chen Chu1,2, Ting Cong2,3, Jian-Yu Zhao1
1Department of Orthopedics, First Affiliated Hospital of Dalian Medical University, Dalian, Liaoning, China.
Introduction:
Osteoarthritis (OA) refers to a commonly seen degenerative joint disorder and a major global public health burden. According to the existing literature, osteoarthritis is related to epigenetic changes, which are important for diagnosing and treating the disease early. Through early targeted treatment, costly treatments and poor prognosis caused by advanced osteoarthritis can be avoided.
Methods:
This study combined gene differential expression analysis and weighted gene co-expression network analysis (WGCNA) of the transcriptome with epigenome microarray data to discover the hub gene of OA. We obtained 2 microarray datasets (GSE114007, GSE73626) in Gene Expression Omnibus (GEO). The R software was utilized for identifying differentially expressed genes (DEGs) and differentially methylated genes (DMGs). By using WGCNA to analyze the relationships between modules and phenotypes, it was discovered that the blue module (MEBlue) has the strongest phenotypic connection with OA (cor = 0.92, p = 4e-16). The hub genes for OA, also known as the hub methylated differentially expressed genes, were identified by matching the MEblue module to differentially methylated differentially expressed genes. Furthermore, this study used Gene set variation analysis (GSVA) to identify specific signal pathways associated with hub genes. qRT-PCR and western blotting assays were used to confirm the expression levels of the hub genes in OA patients and healthy controls.
Results:
Three hub genes were discovered: HTRA1, P2RY6, and RCAN1. GSVA analysis showed that high HTRA1 expression was mainly enriched in epithelial-mesenchymal transition and apical junction; high expression of P2RY6 was mainly enriched in the peroxisome, coagulation, and epithelial-mesenchymal transition; and high expression of RCAN1 was mainly enriched in epithelial-mesenchymal-transition, TGF-β-signaling, and glycolysis. The results of the RT-qPCR and WB assay were consistent with the findings.
Discussion:
The three genes tested may cause articular cartilage degeneration by inducing chondrocyte hypertrophy, regulating extracellular matrix accumulation, and improving macrophage pro-inflammatory response, resulting in the onset and progression of osteoarthritis. They can provide new ideas for targeted treatment of osteoarthritis.
