Integration of transcriptome and DNA methylation reveals the mechanism of cilia-related genes in recurrent miscarriage

Abstract

Abstract Recurrent miscarriage (RM), a complex pregnancy disorder with largely undefined molecular mechanisms, has been associated with epigenetic abnormalities in chorionic tissue. This study aimed to elucidate methylation-dependent cilia-related genes (CRGs) implicated in RM. An integrative analysis combining RNA sequencing, public transcriptome data, and DNA methylation profiles was conducted to identify RM-related CRGs. Machine learning algorithms were applied to determine the most relevant candidates. Immune infiltration profiling, gene set enrichment analysis (GSEA), and competitive endogenous RNA (ceRNA) network construction were employed to clarify molecular pathways. RT-qPCR validation was performed using clinical samples. Fourteen methylation-regulated CRGs were identified, among which SLC1A2 and ZDHHC20 were confirmed as key candidates. GSEA indicated their association with spliceosome, cell cycle, and proteasome pathways. Immune analysis demonstrated decreased infiltration of activated CD4 + T cells, effector memory CD4 + T cells, and Th2 cells in RM, with SLC1A2 and ZDHHC20 expression positively correlated with these immune subsets. The ceRNA networks indicated that SLC1A2 and ZDHHC20 were modulated by 7 miRNAs and 19 lncRNAs, respectively. RT-qPCR results showed significant overexpression of SLC1A2, but not ZDHHC20, in RM chorionic tissue. Collectively, SLC1A2 represents a methylation-regulated CRG that links ciliary impairment, immune imbalance, and epigenetic modulation in RM, revealing a novel molecular axis and suggesting its diagnostic and therapeutic potential.