Disruption of Estrogenic and Androgenic Bioactivities in Human Fetuses Exposed to Maternal Smoking

Abstract Endocrine disruptors (EDs) interfere with hormonal signalling and, given that multiple developmental processes are hormone-driven, the prenatal period is a window of increased sensitivity. Maternal smoking is a real-life model of in utero exposure to a complex mixture of EDs. Cigarette smoke contains of >7,000 pollutants, including polycyclic aromatic hydrocarbons (PAHs), which are AhR ligands and cross-talk with the estrogen receptor (ER) system. Prenatal exposure to cigarette smoke is associated with adverse outcomes, including intrauterine growth restriction and increased risk of metabolic syndrome later in life. We aimed to evaluate ED effects associated with smoke exposure in human fetuses. Fetal tissues (plasma, n=48; placenta, n=30; liver, n=29) from elective terminations of normally progressing pregnancies, ranging from 10 to 20 gestation weeks, were collected (SAFeR and FEGO studies: REC 15/NS/0123, REC 04/S0802/21). PAHs and PAH-like compounds were extracted from placenta and fetal liver. Bioactivity levels in plasma, placenta and liver extracts were determined using ER and androgen receptor (AR) transactivation reporter gene assays. PAH burden was evaluated using the AhR-responsive DRhp-CALUX assay. Smoke exposure was associated with a 1.3-fold increase in plasma estrogenic activity. The developmental trajectory of androgenic activity was altered in plasma of smoke-exposed fetuses, with significant anti-androgenic activity in older fetuses (>16 weeks of gestation). In males, plasma androgenic activity was positively associated with testes weight and anogenital distance. In contrast, placentas from smoking mothers had significantly increased androgenic potential. Furthermore, AhR-like activity was 2.9-fold higher in smoke-exposed placentas compared to controls, and 2.3-fold higher in female compared to male fetal livers. Overall, all bioactivity levels were higher in placentas compared to fetal liver. Prenatal exposure to cigarette smoke is associated with higher placental AhR activation, indicative of increased xenotoxicants burden. We also report that smoke-exposed fetuses showed increased circulating estrogenic activity and disrupted androgenic potential, across 10-20 weeks of gestation, in both fetal plasma and placenta. This demonstrates that EDs present in cigarette smoke are able to interfere with hormonal signalling and alter dynamic endocrine activity profiles, which are critical to ensure appropriate, sex-specific, development. These ED effects are likely to disturb placental function and reprogramme fetal development and thus impacting on life-long health.