Identification and Characterization of Ovarian Extrachromosomal Circular Deoxyribonucleic Acid in Rats with Polycystic Ovary Syndrome

Objective: To characterize the expression profile of extrachromosomal circular deoxyribonucleic acid in polycystic ovary syndrome ovaries and elucidate their potential functions and interactions with non-coding ribonucleic acids and circular ribonucleic acids. Design: An experimental study using a polycystic ovary syndrome rat model combined with high-throughput sequencing and bioinformatics analysis. Subjects: Female Sprague-Dawley rats randomly assigned to control and polycystic ovary syndrome groups. Exposure: Polycystic ovary syndrome was induced via subcutaneous injection of dehydroepiandrosterone for 21 consecutive days. Main Outcome Measures: Follicular development assessed by hematoxylin-eosin staining; serum sex hormones measured by enzyme-linked immunosorbent assay; estrous cycles monitored by light microscopy; extrachromosomal circular deoxyribonucleic acid profiles determined by circle sequencing; host gene functions and pathways analyzed by gene ontology and pathway enrichment analysis; extrachromosomal circular deoxyribonucleic acid presence confirmed by transmission electron microscopy; differential expression validated by gel electrophoresis. Results: Polycystic ovary syndrome rats exhibited abnormal follicular development, disrupted sex hormone levels, and irregular estrous cycles. A total of 15,207 extrachromosomal circular deoxyribonucleic acid molecules representing 13,762 distinct types were identified, with significantly lower abundance in the polycystic ovary syndrome group. Among 263 differentially expressed extrachromosomal circular deoxyribonucleic acid molecules, all were small circular deoxyribonucleic acids. Length distribution showed a unimodal pattern in controls peaking at 368 base pairs versus a bimodal pattern in polycystic ovary syndrome with peaks at 212 base pairs and 359 base pairs. Extrachromosomal circular deoxyribonucleic acid originated widely from various chromosomes without significant intergroup differences, predominantly from gene-enriched regions with 4-20 base pair repeats at junctions. No significant guanine-cytosine content differences were observed. Host genes partially or completely overlapped with non-coding ribonucleic acids, transposable elements, and circular ribonucleic acids. Functional analysis revealed enrichment in synaptic structure, signal transmission, iron ion homeostasis, and cell junction formation. Pathway analysis indicated T cell receptor signaling pathway enrichment. Transmission electron microscopy confirmed ovarian extrachromosomal circular deoxyribonucleic acid presence, and gel electrophoresis validated differential expression. Conclusion: This study first reveals the extrachromosomal circular deoxyribonucleic acid expression profile in polycystic ovary syndrome ovaries, clarifies their functions, and elucidates synergistic mechanisms with non-coding ribonucleic acids and circular ribonucleic acids, providing novel insights into polycystic ovary syndrome pathogenesis and potential diagnostic biomarkers and therapeutic targets.