Proteomic Dynamics of Ovine Allantoic and Amniotic Fluids During Gestation

Gestational success in ruminants depends on the maintenance of a dynamic maternal-foetal microenvironment, resulting from complex interactions among the conceptus, extraembryonic membranes, maternal tissues, and gestational fluids. This study performed a comparative and longitudinal proteomic characterization of allantoic and amniotic fluids throughout pregnancy in sheep, aiming to elucidate the molecular mechanisms involved in maternal-foetal communication and embryonic development. Samples of allantoic and amniotic fluids aseptically collected from extraembryonic compartments after uterine opening following animal slaughter and allocated into three gestational groups according to foetal development: early gestation (0-50 days; n = 6), mid gestation (50-100 days; n = 3) and late gestation (100-150 days; n = 5). After centrifugation and protein quantification using the Bradford method, standardized aliquots were subjected to in-solution protein digestion and analysed by nano-LC-ESI-Q-TOF mass spectrometry. A total of 201 proteins were identified in the allantoic fluid and 159 proteins in the amniotic fluid, with five proteins showing significant differences across gestational stages. In the allantoic fluid, differentially abundant proteins included alpha-2-macroglobulin, gelsolin, basement membrane-specific heparan sulphate proteoglycan core protein and alpha-globin chain. In the amniotic fluid, changes were observed in peroxiredoxin-2, mucin-5 AC, fetuin-B, cystatin domain-containing protein and globin domain-containing protein. Multivariate analyses revealed progressive remodelling of the foetal extraembryonic microenvironment, associated with proteolytic control, tissue remodelling, redox homeostasis, epithelial protection and metabolic processes. Collectively, these findings reinforce the active role of foetal extraembryonic fluids in gestational physiology and highlight their potential as sources of biomarkers associated with reproductive success in sheep.