Morphological and cellular effects of lead, cadmium, and perfluorooctanoic acid on Gallus gallus embryonic development: Insights into cephalic region vulnerabilities.

Lead (Pb), cadmium (Cd), and perfluorooctanoic acid (PFOA) are environmental pollutants with bioaccumulative properties, posing risks to embryonic development, particularly through cephalic region malformations. Despite their significance, few studies have investigated their effects at realistic concentrations or in mixtures. This study assessed the individual and combined effects of Pb, Cd, and PFOA on Gallus gallus embryos at relevant concentrations, focusing on the cephalic region at stages HH10 and HH14. Exposure to contaminant mixtures, particularly Pb + Cd, Pb + PFOA, and the full combination, resulted in a higher incidence of cranial malformations compared to single-compound exposures. Cd induced dose-dependent changes in cell morphology and significantly reduced proliferation in the head region. PFOA contributed additively to mixture toxicity, markedly impairing cell proliferation and modifying the expression of key adhesion molecules such as β-catenin and N-cadherin. Both embryonic stages exhibited aberrant cellular delamination from the neural tube, with HH10 embryos displaying heightened vulnerability. Systems biology analysis revealed distinct molecular responses to each contaminant, with Pb and Cd primarily affecting immune-related pathways, including complement activation, while PFOA operated through separate regulatory mechanisms. Integrative network analyses using STITCH 5 and the Comparative Toxicogenomics Database identified candidate genes potentially involved in developmental disruption. These findings underscore the complex interactions between pollutants, which may not be predicted solely by examining individual exposures and highlight the need for stricter environmental regulations to address the risks posed by contaminant mixtures.