FOXP3 isoform expression (Full-length and Delta 2) & immunoregulatory markers (Tregs, IL-6, IL-10) in recurrent pregnancy loss: A South-Indian study.

Recurrent Pregnancy Loss (RPL), a complex reproductive disorder, with 50% of cases lacking a definitive cause. Immune dysregulation, particularly involving regulatory T cells (Tregs) and their master transcription factor FOXP3, is increasingly recognized as a critical contributor. FOXP3 exists in functionally distinct splice variants-full-length (FOXP3-FL) and exon 2-deleted (FOXP3-D2) whose roles in RPL remain poorly understood. To investigate FOXP3-FL and FOXP3-D2 expression, Treg populations, and serum IL-6 and IL-10 levels in second-trimester pregnant women with and without RPL. Sixty pregnant women were enrolled, equally divided into RPL and non-RPL (NRPL) groups. Treg populations and cytokine levels (IL-6 and IL-10) were assessed in all participants using flow cytometry and ELISA, respectively. FOXP3 isoform expression was further validated by Western blot analysis in a subset of 30 samples, including 15 NRPL and 15 RPL cases. Statistical comparisons included subgroup analysis (2 losses vs. >2 losses), with p < 0.05 considered significant. Compared to NRPL, RPL women had significantly higher age, BMI, and age at menarche, with reduced Treg percentage, FOXP3-FL, and IL-10 levels, and elevated FOXP3-D2 and IL-6. ROC analysis indicated strong diagnostic accuracy for Tregs, IL-6, and FOXP3-D2. Subgroup analysis revealed progressive FOXP3-D2 increase and FOXP3-FL decrease with the severity. The IL-6/IL-10 ratio was significantly higher in RPL, indicating a pro-inflammatory shift independent of loss severity. This is the first report of altered FOXP3 isoform expression in RPL, revealing a disrupted FOXP3-FL/D2 ratio that may underlie impaired Treg function and immune tolerance failure. FOXP3 isoforms, alongside cytokine markers, hold promise as novel immunological biomarkers for RPL diagnosis and risk stratification.