Impact of Chronic Maternal Air Pollution Exposure (PM2.5) on Regulatory T Cell Number, Proliferation, and Suppressive Activity in Neonatal Umbilical Cord Blood: A Case-Control Study

Background Prenatal exposure to ambient fine particulate matter (PM _2.5 ) has been linked to a broad spectrum of adverse health outcomes; however, its impact on the establishment of neonatal immune tolerance remains inadequately characterized. Regulatory T cells (Tregs), defined by the CD4⁺CD25⁺FOXP3⁺ phenotype, play a pivotal role in maintaining immune homeostasis during early development. In this study, we investigated the effects of chronic maternal exposure to PM _2.5 on the frequency, proliferative capacity, cytokine secretion, and suppressive function of neonatal Tregs isolated from umbilical cord blood. Methods In a case-control study, pregnant women residing in Tehran, a highly polluted urban environment (n = 10), and Damavand, a clean rural environment (n = 10), were enrolled. Maternal exposure to ambient pollutants was quantified by measuring urinary concentrations of 6‑hydroxy‑nitropyrene and 8‑hydroxy‑pyrene using two-dimensional high-performance liquid chromatography coupled with tandem mass spectrometry (2D‑HPLC‑MS/MS). Umbilical cord blood mononuclear cells were subsequently isolated, and CD4⁺CD25⁺FOXP3⁺ regulatory T cells (Tregs) were phenotypically and functionally characterized by multicolor flow cytometry. Treg proliferative responses were assessed under interleukin-2 supplementation and anti-CD3/CD28 stimulation, while suppressive capacity was evaluated in co-culture assays with effector T cells. The secretion levels of IL-10 and TGF-β1 in the cell culture supernatants were quantitatively assessed using enzyme-linked immunosorbent assay (ELISA). Results Maternal urinary biomarker concentrations indicative of PM _2.5 exposure were significantly higher in women residing in Tehran compared with those from Damavand (p < 0.0001). The baseline frequencies of neonatal CD4⁺CD25⁺FOXP3⁺ regulatory T cells (Tregs) did not differ significantly between groups (p = 0.111). However, neonates from Tehran demonstrated reduced FOXP3 mean fluorescence intensity, impaired Treg proliferation on days 7, 12, and 14 of culture (p < 0.05), and diminished suppressive capacity at a 1:1 effector-to-Treg cell ratio (61% vs. 72%, p = 0.017). Regulatory T cells from infants exposed prenatally to higher pollution showed reduced IL-10 and TGF-β1 secretion, though differences were not statistically significant. Correlation analyses revealed positive associations between Treg frequency and IL‑10 production, as well as between suppressive capacity and FOXP3 expression, but these associations were observed only in neonates whose mothers resided in clean-air environments. Conclusion Chronic maternal exposure to PM _2.5 during pregnancy impairs critical functional attributes of neonatal regulatory T cells including proliferative capacity, cytokine production, and suppressive function despite unaltered baseline cell frequencies. These findings indicate that sustained prenatal exposure to ambient air pollution may disrupt the establishment of neonatal immune tolerance, thereby increasing susceptibility to immune‑mediated disorders in early life. Consequently, public health strategies aimed at mitigating maternal exposure to PM _2.5 are essential for safeguarding neonatal immune development and long-term health.