Effects of single and repetitive valproic acid administration on the gene expression of placental transporters in pregnant rats: An analysis by gestational period
Introduction
About 0.3–0.7 % of pregnant women have epilepsy [1]. Seizure control needs to be maintained by continuous pharmacotherapy throughout pregnancy. Generally, medication use during pregnancy is a fetal risk factor, needing consideration in gestational pharmacotherapy.
Valproic acid (VPA) is a well-established and frequently used antiepileptic drug. However, it increases the risk of fetal malformations, autism spectrum disorders, and cognitive defects [[2], [3], [4],5]. VPA treatment should be avoided in women of childbearing age; however, it is prescribed to pregnant women when required [6,7]. Hence, the effect of VPA on the fetus and related organs needs to be evaluated.
The placenta is a crucial temporary organ, which is in contact with both maternal and fetal blood. ATP‐binding cassette (ABC) and solute carrier (SLC) transporters play a role in nutrient, metabolic waste, and xenobiotic exchange between mother and fetus [8,9]. Hence, the expression levels of placental transporters are one of the factors affecting the intrauterine environment and fetal growth. Since the expression of placental transporters changes across gestation, their evaluation at each gestational stage is important. Additionally, transporters can be affected in a sex-specific manner, in some cases [10].
VPA has been reported to inhibit histone deacetylases (HDACs), which remove acetyl groups from histones. Four classes of HDACs have been identified, and VPA acts predominantly on Class I HDACs, including HDAC 1–3 and 8 [11,12]. Therefore, VPA likely affects the expression of various genes. So far, there are several reports that have investigated the effects of VPA on placental transporters. An in vitro study indicated that VPA treatment altered the expression and function of breast cancer resistance protein (BCRP) [13]. Moreover, we previously showed that VPA exposure induces increased mRNA levels of folate receptor alpha (FRα) and proton-coupled folate transporter in BeWo and JEG-3 cell lines, derived from human choriocarcinoma [14]. Additionally, perfusion with VPA reportedly has reduced FRα and glucose transporter (GLUT) 1 in ex vivo human placentas [15,16]. However, the perfusion time was short and variability among samples was high in the model. In particular, placental structure, function, and transporters are dramatically altered. Therefore, an in vivo study is well suited to sequentially investigate, throughout gestation, and assess the influence of fetal sex on VPA-mediated changes. Although a previous in vivo study indicated that the expression of l-type amino acid transporter (LAT) 1, organic anion transporting polypeptide (OATP) 4a1, and reduced folate carrier were lower in placentas from VPA-treated mid-pregnant mice [17], transporter expression level changes were not recorded.
This study aimed to comprehensively reveal the effects of both single and repetitive VPA treatment on placental transporters, using pregnant rats. We analyzed the expression of 8 ABC transporters, including multiple drug-resistant (MDR) 1a and 1b (Abcb1a, 1b), multidrug resistance-associated proteins (MRPs) 1–5 (Abcc1–5), and BCRP (Abcg2), as well as 10 SLC transporters, including LAT1 and 2 (Slc7a5 and Slc7a8), monocarboxylate transporter (MCT) 4 (Slc16a3), organic cation transporter (OCT) 3 (Slc22a3), organic cation/carnitine transporter (OCTN) 1 and 2 (Slc22a4, 5), equilibrative nucleoside transporter (ENT) 1 (Slc29a1), and OATP2a1, 2b1, and 4a1 (Slco2a1, 2b1, 4a1), by gestational age. Furthermore, the results were secondarily analyzed by fetal sex.
Section snippets
Chemicals and reagents
Valproic acid sodium salt (VPA Na) and valproic acid were purchased from Sigma-Aldrich (St. Louis, MO, USA). Deuterium-labeled valproic acid (VPA-d6) and 2-propyl-4-pentenoic acid (4-ene-VPA) were purchased from Toronto Research Chemicals (Toronto, Canada).
Animals
Pregnant female Wistar rats (12–13 weeks old) were provided by CLEA Japan (Tokyo, Japan). The existence of a vaginal plug indicated the first day of gestation (Gestational day 0; G0). G9–13 and G16–20 were considered as mid- and late
Effects of single VPA Na administration on placental transporters at G13 and G20
First, we confirmed the presence of VPA and 4-ene-VPA in the plasma from the pregnant rats 30 min (peak) post oral VPA Na administration. In a previous study in pregnant rats, the VPA level peaked at 0.5–0.9 h post-oral VPA Na administration [23]. The plasma levels of VPA and its toxic active metabolite, 4-ene-VPA, 30 min post-administration were approximately 200 and 1.5 μg/mL, respectively (Table 1); this level was similar to that reported in a previous study [23]. After 24 h, their
Discussion
VPA use during pregnancy poses risks to the fetus such as malformations, autism spectrum disorders, and cognitive defects [[2], [3], [4], [5]]. Therefore, evaluation of its effects on the placenta, which supports fetal development and health maintenance in pregnancy, is important. In the present study, we comprehensively evaluated the effects of VPA administration on the expression of rat placental transporters.
In the study, we determined the experimental dose to observe fetal effects and not
Declaration of Competing Interest
The authors declare no conflicts of interest.
Acknowledgment
None
Funding
This work was supported by a grant from the Japan Society for the Promotion of Science (JSPS) KAKENHI (grant number, 18K1497208) (provided to A.F.).
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