Maternal Exposure to Wood-Smoke-Derived PM2.5 Is Associated with Delayed Fetal Neurocranial Intramembranous Ossification in a Rat Model

Maternal exposure to airborne particulate matter smaller than 2.5 μm (PM2.5) has been associated with adverse fetal outcomes, although its effects on intramembranous ossification remain poorly understood. This study evaluated the impact of gestational and pregestational exposure to wood-smoke-derived PM2.5 on fetal neurocranial ossification in Sprague–Dawley rats. Females were allocated to four exposure conditions combining filtered air (FA) and non-filtered air (NFA): FA/FA, FA/NFA, NFA/FA, and NFA/NFA. Fetuses were collected at gestational day 21 and analyzed using fetal morphometry, radiography, micro-computed tomography, whole-mount alizarin red skeletal staining, histology, and immunohistochemistry for HIF-1α, COL-1, BMP-2, FGF-R1, and TGF-β. Continuous exposure (NFA/NFA) was associated with reduced fetal weight, shorter crown–rump length, impaired craniofacial mineralization, widened cranial sutural regions, and reduced mineral density, particularly in the occipital and interparietal bones. Histologically, exposed fetuses exhibited abundant osteoid, reduced osteocyte incorporation, and diffuse osteoblastic distribution, consistent with delayed osteogenic maturation. Immunohistochemistry showed increased HIF-1α immunoreactivity, altered TGF-β regulation, and reduced COL-1 expression in continuously exposed fetuses, whereas BMP-2 and FGF-R1 showed no significant changes. These findings suggest that maternal exposure to wood-smoke-derived PM2.5 is associated with delayed fetal neurocranial intramembranous ossification, particularly under continuous exposure. The observed immunohistochemical profile, elevated HIF-1α, reduced COL-I, and altered TGF-β, is consistent with a hypoxia-associated imbalance between extracellular matrix deposition and mineral maturation; however, the underlying mechanistic pathway was not directly functionally tested and should be regarded as a biologically plausible inferential model requiring further experimental validation.