Maternal placental mitochondrial dysfunction, oxidative stress, and autophagy by regulating ferroptosis in pregnant ewe model.

Elevated prenatal testosterone (T) induces placental insufficiency and fetal growth restriction (FGR) in sheep, a process hypothesized to involve oxidative stress (OS), mitochondrial dysfunction, autophagy, and ferroptosis. While ferroptosis is recognized as a significant contributor to placental pathophysiology, its specific role in T-mediated ovine placental dysfunction required further investigation. To address this, an in vivo study was conducted wherein pregnant Hu sheep received intramuscular injections of 100 mg T propionate or a control vehicle twice weekly from gestational day (GD) 60-130. Complementarily, in vitro experiments utilized dihydrotestosterone (DHT)-exposed ovine trophoblast cells (OTCs), which were further treated with the ferroptosis activator Erastin or inhibitor Ferrostatin-1 (Fer-1) to directly probe the functional impact of ferroptosis. Our results demonstrated that T administration in vivo recapitulated the pathological phenotype, triggering placental OS, mitochondrial dysfunction, ferroptosis, autophagy, and culminating in FGR. Consistent with these findings, DHT exposure in OTCs induced a similar suite of cellular stresses, including OS, mitochondrial impairment, ferroptosis, and autophagy. Crucially, the inhibition of ferroptosis with Fer-1 in DHT-treated OTCs was found to attenuate these detrimental effects, notably alleviating OS, iron overload, mitochondrial dysfunction, and autophagic activity. Conversely, the co-administration of the ferroptosis inducer Erastin effectively abolished the protective changes conferred by Fer-1, thereby substantiating a central role for ferroptosis in the cascade of T-induced placental dysfunction.