Whole transcriptome analysis of alternative ribonucleic acid splicing events in severe and moderate forms of fetal growth retardation

Background. Fetal growth retardation (FGR) remains one of the leading causes of perinatal morbidity and a major risk factor for longterm adverse health outcomes in children, including an increased likelihood of neurological, metabolic, and cardiovascular disorders. Despite extensive research interest, the molecular mechanisms underlying FGR are still insufficiently understood. In particular, little is known about the role of post-transcriptional regulation in the development of this condition. Alternative splicing is of special interest. It determines transcriptome diversity and expands the functional capacities of cells. Through this mechanism, cells gain the ability to adapt to pathological stimuli. At the same time, it influences their susceptibility to disease, including obstetric complications.

Aim: To characterize alternative splicing profiles in placental decidual cells (DCs) that determine the severity of fetal growth retardation. Material and Methods. The study was conducted on placental tissue samples from patients with moderate and severe forms of FGR. Whole-transcriptome analysis was performed on decidual cells isolated by laser microdissection from stained thin tissue sections. Whole-genome ribonucleic acid (RNA) sequencing was performed using the SMARTer Stranded Total RNA-Seq kit v2 (Takara BIO). Alternative splicing events were analyzed with the MAJIQ package under a Linux operating system.

Results. In the analyzed samples, 13,688 alternative splicing (AS) events were detected across 4,002 genes expressed in decidual cells. More than 52% of these events were identical between both groups. In severe FGR, both annotated and de novo events demonstrated a statistically significant decrease in the frequency of the alternative first exon (χ2 = 8.48, p = 0.004; χ2 = 6.15, p = 0.014, respectively). The alternatively spliced genes specific to severe FGR were involved in the following biological processes: catalytic activity acting on nucleic acids (pFDR = 0.020), regulation of GTPase activity (pFDR = 0.021), regulation of nucleoside triphosphatase activity (pFDR = 0.021), and peptide N-acetyltransferase activity (pFDR = 0.028). Comparison of the moderate and severe FGR groups identified 84 differentially spliced genes (0.200 < deltaPSI < 0.648; p < 0.05). These genes were significantly associated with biological and signaling pathways including multiple types of DNA repair, the ligand-gated ion channel pathway, vesicular transport to the plasma membrane, regulation of mRNA metabolism, peroxisome organization, lysosome organization, morphogenesis, the SMAD signaling pathway, sulfur metabolism, and ATP-dependent chromatin remodeling.

Conclusion. The data indicate a specific set of AS-related molecular changes characteristic of FGR, regardless of its severity. AS patterns unique to severe FGR are associated with disruptions of fundamental cellular regulatory systems. Functional annotation of differentially spliced genes suggests that AS affects post-transcriptional control, cellular architecture, and intercellular signaling interactions in severe FGR.

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