The transfer of ethyl glucuronide across the dually perfused human placenta
Introduction
Determining the extent of fetal exposure to alcohol during pregnancy can help identify the risk of Fetal Alcohol Spectrum Disorder (FASD) in newborns. FASD is the leading preventable cause of malformations, intellectual disabilities, and neurobehavioral disorders in the Western world [1]. Left untreated, individuals with FASD can develop secondary characteristics, such as mental health issues, addictions, and trouble with the law [2]. As such, it is of vital importance for healthcare providers to adequately screen for maternal use of and fetal exposure to alcohol to help identify high-risk cases.
While confirmed alcohol use during pregnancy is the best indicator of fetal risk for FASD, many women under-report their consumption due to fear of stigmatization and blame [3]. For this reason, ethanol metabolites such as fatty acid ethyl esters (FAEE) are often used as objective biomarkers of fetal alcohol exposure. FAEEs have been detected in maternal hair and neonatal meconium, which begins to form at 12 weeks gestation. They were also shown to not cross the human placenta and therefore are biomarkers of direct fetal exposure to ethanol [4]. Despite these findings, there are several sources of false results with FAEE analysis, such as prenatal vitamin use, olive oil use, and contamination of meconium with post-natal stool [5], [6]. These sources of false results warrant investigation into other biomarkers that could supplement FAEE analysis.
Similar to FAEE, ethyl glucuronide (EtG) is a minor, direct, non-oxidative metabolite of ethanol. EtG is formed by the net addition of UDP-glucuronic acid to ethanol, a reaction catalyzed by the UDP-glucuronosyl transferase (UGT) family [7]. Compared to ethanol, EtG is stable within the body for longer periods and can therefore provide a larger window of alcohol consumption. With respect to pregnant patients, EtG has previously been detected in maternal hair [8], neonatal meconium [3], [9], [10], [11], as well as first trimester placental tissue and fetal remains [12]. Recently, a cut-off of 2 nmol/g has been established in meconium to differentiate excessive alcohol consumption during pregnancy [10]. These findings suggest that EtG could be a useful supplemental biomarker to FAEEs to help improve screening accuracy.
Whether EtG detected in these matrices is of maternal or fetal origin needs to be determined, since the EtG cut-off in meconium may depend on the relative contributions of these factors. While EtG is extensively formed in the adult liver [7], no studies have investigated fetal hepatic ethanol glucuronidation. Several reports have indicated that, compared to the adult liver, glucuronidation is greatly reduced in the fetus and only begins to develop postpartum [13], [14]. If, like FAEEs, EtG does not cross the placenta, then EtG detected in meconium is solely of fetal origin. However, it is much more likely that there is transfer, in which case EtG is primarily of maternal origin with a potentially minor fetal contribution. To help clarify the utility of EtG in alcohol screens, the objective of this study was to determine if physiologically relevant levels of EtG cross the human placenta.
Section snippets
Ex vivo perfusion of a single placental cotyledon
The ex vivo placental perfusion model has been previously explained in detail [15] and adapted in our laboratory [16], [17]. Term placentae were obtained from scheduled elective Caeserian sections at the obstetrics ward at St. Michael's Hospital in Toronto, Ontario, Canada. Research ethics board approval was obtained from the hospital and mothers gave written consent prior to delivery. Immediately after delivery, placentae were transported to the on-site perfusion laboratory in ice-cold
Results
A total of 4 cotyledons from different placentae were perfused with 1 μg/mL EtG and mean lobule weight of the 4 perfused lobules was 19.03 ± 1.29 g. Maternal and fetal flow rates were 13.65 ± 0.68 and 2.11 ± 0.06 mL/min, respectively. Parameters of placental integrity and viability are presented in Table 1. None of the parameters were statistically different between the control and experimental phases. Antipyrine equilibrated between the two circulations after 3 h with a final mean F:M ratio of
Discussion
The results of this perfusion study show that EtG crosses the term human placenta slowly, reaching an F:M ratio of 0.29 after 3 h of perfusion. Glucuronide metabolites have previously been shown to cross the placenta, resulting in significant fetal concentrations [24], [25]. In pregnant baboons injected with morphine-3-glucuronide, F:M ratios of 0.7–0.9 were measured after 24 h, by which point the metabolite was presumably at steady state [25]. The final F:M ratio of 0.29 is likely not
Acknowledgments
The authors would like to thank the generous staff at St. Michael's Hospital for facilitating sample collection. This study is supported by a grant from the Canadian Institutes for Health Research and the Research Leadership for Better Pharmacotherapy During Pregnancy and Lactation. JNM is supported by an Ontario Graduate Scholarship. GK is the holder of the Ivey Chair in Molecular Toxicology, The Department of Medicine, University of Western Ontario.
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