The m6A/m1A/m5C-Related Methylation Modification Patterns and Immune Landscapes in Rheumatoid Arthritis and Osteoarthritis Revealed by Microarray and Single-Cell Transcriptome

Abstract

Purpose

The goal of this study was to explore the expression characteristics of RNA modification-related genes, reveal immune landscapes and identify novel potential diagnostic biomarkers in osteoarthritis (OA) and rheumatoid arthritis (RA) patients.

Patients and Methods

RNA microarray and single-cell sequencing (scRNA-seq) data were downloaded from gene expression omnibus (GEO) database. Differentially expressed RNA modification-related genes were identified and then functionally annotated. Univariate logistic regression and lasso regression analysis were used to identify primary disease genes for OA and RA. Validation was done using scRNA-seq analysis and immunohistochemistry (IHC) in human knee synovial tissues and a murine destabilization of the medial meniscus (DMM) model. Through WGCNA analysis, genes associated with cell pyroptosis or autophagy in OA and RA were identified, which were then combined with differentially expressed RNA modification-related genes to construct a PPI interaction network. Furthermore, hub genes were selected for ceRNA interaction network analysis, correlation analysis with OA and RA molecular subtypes, as well as correlation analysis with 22 immune cells.

Results

Six RNA modification-related genes (ADAMDEC1, IGHM, OGN, TNFRSF11B, SCARA3 and PTN) were identified as potential OA and RA pathogenesis biomarkers. Their expression was validated in human knee synovial tissues and a murine DMM model. Functional enrichment of differentially expressed RNA modification-related genes between RA and OA was analyzed using GO, KEGG, GSEA, and GSVA. Based on WGCNA and PPI analysis, the six hub genes related to pyroptosis and RNA modification (CXCL10, CXCL9, CCR7, CCL5, CXCL1, and CCR2) were identified as central nodes for ceRNA interaction, correlation with OA and RA molecular subtypes, and association with 22 immune cells.

Conclusion

Our research revealed the significance of RNA modification-related genes in the development of OA and RA pathogenesis, thereby providing a novel research direction for understanding the mechanisms, diagnosis, and treatment of OA and RA.

Keywords:

• m6A
• m5C
• m1A
• osteoarthritis
• rheumatoid arthritis
• single-cell sequencing

Data Sharing Statement

The data supporting the results of the study is available from the GEO database (https://www.ncbi.nlm.nih.gov/geo/) and and the name of the database and accession number(s) can be found in the article. The code used for our analysis was uploaded to GitHub (https://github.com/JEASTYJJ/Code.git).

Ethics Approval Statement

The study adhered to the guidelines set forth by the Declaration of Helsinki. Ethical approval for research involving human participants was granted by the Biomedical Research Ethics Committee at HongHui Hospital, affiliated with Xi'an Jiaotong University (No. 202211012, November 28, 2022). The participants provided their written informed consent to participate in this study. All animal experiments were conducted in strict accordance with the ARRIVE guidelines and adhered to the “3Rs” principles of animal welfare, emphasizing Replacement, Reduction, and Refinement. The study protocols were duly approved by the Animal Care and Use Committee of Xi’an Jiaotong University (No. 2022-1634, December 1, 2022).

Acknowledgments

We are grateful to the patients for their contributions to this study.

Disclosure

The authors report no conflicts of interest in this work.

Additional information

Funding

This work was supported by funding from the National Natural Science Foundation of China (No. 82072432).