Hsa_circ_0065149 mediates the ceRNA network pathway: Research on the mechanism regulating myasthenia gravis.

CircRNAs have been linked to the pathogenesis and progression of myasthenia gravis (MG), but research on this association remains limited. This study aims to provide a new perspective for understanding the molecular pathogenesis of MG. We obtained RNA-seq data from patients with acetylcholine receptor (AChR)-positive MG and healthy individuals from the Gene Expression Omnibus (GEO) database. A bioinformatics analysis was performed to identify differentially expressed circRNAs related to MG; the circRNA that was most consistently identified in most databases as a key factor involved in MG, hsa_circ_0065149, was chosen for further analysis. AChR localized at the postsynaptic membrane of the neuromuscular junction (NMJ) plays a pivotal role in mediating synaptic signal transmission. The CHRNA1 gene, which encodes the α1 subunit of AChR and is critical for its function-was subsequently selected as the target mRNA in our investigation, whereas our prediction pipeline identified miR-330-5p as the target. For functional assays, various constructs (e.g., hsa_circ_0065149-siRNA) were transfected as described in the Methods section. The qRT‒PCR results indicated that hsa_circ_0065149 knockdown significantly upregulated miR-330-5p and downregulated CHRNA1, while the inhibition of miR-330-5p significantly upregulated both hsa_circ_0065149 and CHRNA1. We also performed functional rescue experiments to confirm the regulatory hierarchical relationships among these factors. A dual-luciferase reporter assay confirmed that hsa_circ_0065149 and CHRNA1 are direct targets of miR-330-5p. These results suggest that hsa_circ_0065149 regulates MG by mediating the ceRNA network pathway, and we reasonably hypothesize that it may further contribute to the pathological process of MG by impairing signal transmission in the NMJ.