Apolipoprotein D as a Potential Biomarker and Construction of a Transcriptional Regulatory-Immune Network Associated with Osteoarthritis by Weighted Gene Coexpression Network Analysis

Abstract

Objective Synovial inflammation influences the progression of osteoarthritis (OA). Herein, we aimed to identify potential biomarkers and analyze transcriptional regulatory-immune mechanism of synovitis in OA using weighted gene coexpression network analysis (WGCNA). Design A data set of OA synovium samples (GSE55235) was analyzed based on WGCNA. The most significant module with OA was identified and function annotation of the module was performed, following which the hub genes of the module were identified using Pearson correlation and a protein-protein interaction network was constructed. A transcriptional regulatory network of hub genes was constructed using the TRRUST database. The immune cell infiltration of OA samples was evaluated using the single-sample Gene Set Enrichment Analysis (ssGSEA) method. The hub genes coexpressed in multiple tissues were then screened out using data sets of synovium, cartilage, chondrocyte, subchondral bone, and synovial fluid samples. Finally, transcriptional factors and coexpressed hub genes were validated via experiments. Results The turquoise module of GSE55235 was identified via WGCNA. Functional annotation analysis showed that “mineral absorption” and “FoxO signaling pathway” were mostly enriched in the module. JUN, EGR1, FOSB, and KLF4 acted as central nodes in protein-protein interaction network and transcription factors to connect several target genes. “Activated B cell,” “activated CD4T cell,” “eosinophil,” “neutrophil,” and “type 17 T helper cell” showed high immune infiltration, while FOSB, KLF6, and MYBL2 showed significant negative correlation with type 17 T helper cell. Conclusions Our results suggest that the expression level of apolipoprotein D (APOD) was correlated with OA. Furthermore, transcriptional regulatory-immune network was constructed, which may contribute to OA therapy.

Identification of susceptibility modules and hub genes of osteoarthritis by WGCNA analysis

“…Currently, with the rapid development of computational algorithms and gene chip technology, many researchers have focused on exploring biomarkers of numerous diseases by application of multiple bioinformatic approaches, which enables in-depth analysis of the whole genome and accelerates the progression of disease study. However, many studies are confronted with limited databases (Gao et al, 2019;Cao et al, 2021;Qin et al, 2021), lack of experimental validation (Gao et al, 2019;Zhang et al, 2020b;Cao et al, 2021), controversial methodology validation (Qin et al, 2021;Li et al, 2022), and unconvincing outcomes. Frontiers in Genetics frontiersin.org chondrocytes (Li et al, 2022).…”

“…Frontiers in Genetics frontiersin.org chondrocytes (Li et al, 2022). Other research groups also only chose one dataset for WGCNA analysis, and many of them lacked validation experiments (Gao et al, 2019;Zhang et al, 2020b;Cao et al, 2021;Wang et al, 2022) or lacked synoviocyte validation data (Qin et al, 2021;Li et al, 2022). None of them verified and evaluated the diagnostic efficacy of candidate markers by mathematical models.…”

“…Qin et al (2021) chose only one dataset (GSE55235) for WGCNA analysis and validated their findings in chondrocytes stimulated by IL-1β. First, in terms of precision, their limited dataset may lead to sampling error, and the conclusions may cause bias.…”

APOD acts on fibroblast‐like synoviocyte and chondrocyte to alleviate the process of osteoarthritis in vitro

“…In our previous study, we analyzed samples from patients with OA by weighted correlation network analysis (WGCNA) to identify significant modules, along with function annotation of module genes and hub genes. After validation through enzyme‐linked immunosorbent assay (ELISA) and quantitative reverse‐transcription polymerase chain reaction (RT‐qPCR), apolipoprotein D (APOD) was validated as a hub gene coexpressed and downregulated in OA cartilage and synovium 9 . APOD is a secreted glycosylated protein and an atypical lipoprotein, which can bind to several small molecules, such as arachidonic acid, steroids, and cholesterol 10 .…”

“…After validation through enzyme-linked immunosorbent assay (ELISA) and quantitative reverse-transcription polymerase chain reaction (RT-qPCR), apolipoprotein D (APOD) was validated as a hub gene coexpressed and downregulated in OA cartilage and synovium. 9 APOD is a secreted glycosylated protein and an atypical lipoprotein, which can bind to several small molecules, such as arachidonic acid, steroids, and cholesterol. 10 Studies have reported the important functions of APOD, such as oxidation resistance, anti-inflammation, and stress resistance.…”