Folate Transporters in Placentas from Preterm Newborns and Their Relation to Cord Blood Folate and Vitamin B12 Levels

RESEARCH ARTICLE Folate Transporters in Placentas from Preterm Newborns and Their Relation to Cord Blood Folate and Vitamin B12 Levels Erika Castaño1, Lorena Caviedes1, Sandra Hirsch1, Miguel Llanos1, Germán Iñiguez2, Ana Marı́a Ronco1* 1 Laboratory of Nutrition and Metabolic Regulation, Human Nutrition Unit, Institute of Nutrition and Food Technology, Dr. Fernando Monckeberg Barros (INTA), University of Chile, Santiago, Chile, 2 Mother and Child Research Institute, Division of Medical Sciences, School of Medicine, University of Chile, Santiago, Chile a1111111111 a1111111111 a1111111111 a1111111111 a1111111111 OPEN ACCESS Citation: Castaño E, Caviedes L, Hirsch S, Llanos M, Iñiguez G, Ronco AM (2017) Folate Transporters in Placentas from Preterm Newborns and Their Relation to Cord Blood Folate and Vitamin B12 Levels. PLoS ONE 12(1): e0170389. doi:10.1371/journal.pone.0170389 Editor: Colette Kanellopoulos-Langevin, Xavier Bichat Medical School, INSERM-CNRS - Université Paris Diderot, FRANCE Received: April 28, 2016 Accepted: January 4, 2017 Published: January 19, 2017 Copyright: © 2017 Castaño et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. * Abstract Folate deficiency during pregnancy has been related to low birth weight, preterm (PT) birth and other health risks in the offspring; however, it is unknown whether prematurity is related to low folate transport through the placenta due to altered expression of specific folate transporters. We determined placental expression (mRNA and protein concentrations by RTqPCR and WB respectively) of specific folate transporters: RFC, PCFT/HCP1 and FOLR1 in chorionic (fetal) and basal (maternal) plates of placentas of PT pregnancies (PT, 32–36 weeks, n = 51). Term placentas were used as controls (T, 37–41 weeks, n = 47). Folates and vitamin B12 levels were measured by electrochemiluminescence in umbilical cord blood of newborns. FOLR1 mRNA expression was lower and protein concentration higher in PT placentas (both plates) relative to the control group (p <0.05). In addition, gestational age was positively correlated with mRNA expression (Rho = 0.7), and negatively with protein concentration (Rho = -0.7 for chorionic and -0.43 for basal plate). PCFT/HCP1 mRNA was lower in PT placentas, without changes in protein levels. RFC did not differ in PT placentas compared to controls. PT newborns presented higher cord blood folate level (p = 0.049) along with lower vitamin B12 concentration compared to controls (p = 0.037).In conclusion, placental FOLR1 mRNA was positively associated with gestational age. Conversely, FOLR1 protein concentrations along with folate/vitamin B12 ratio in cord blood were negatively associated with gestational age. Placental FOLR1 is likely the main placental folate transporter to the fetus in newborns. Data Availability Statement: All data underlying our findings are presently available within the manuscript. Funding: This study was funded by Fondo Nacional de Ciencia y Tecnologı́a (FONDECYT # 1130188 to AMR), Gobierno de Chile. Competing Interests: The authors have declared that no competing interests exist. Introduction Folates are water-soluble vitamins of the B complex that are naturally present in foods as reduced forms or glutamate chains. The synthetic form, folic acid (FA), is fully oxidized and more stable; it is frequently used in fortified foods or as supplements [1,2]. Folates are needed for fetal growth and placental development, since they activate cell growth and biosynthetic PLOS ONE | DOI:10.1371/journal.pone.0170389 January 19, 2017 1 / 14 Folate Transporters in Placentas of Preterm Newborns processes that are essential during pregnancy [3]. For these reasons maternal requirements during pregnancy are 50% greater than adult requirements (400 μg/day) [3,4]. Folate deficiency has been associated with preterm births (PT) [1], defined as <37 week gestation at birth; these are a major public health concern [5], with reported rates ranging from 5% to 20% worldwide [6]. Deficient maternal folate intake [1] and decreases in folate bioavailability associated with reduced folate transport to the fetus or the presence of FRα (folate receptor alpha) autoantibodies (FRAbs) [7] are included in the etiology of PT. Also, folate deficiency may induce low birth weight and other altered birth outcomes such as neural tube defects (NTD) [3,4,8]. Folates and FA reduce the risk of NTD [8], thus several countries have implemented food fortification programs with FA to increase the consumption of FA during pregnancy [9]. Chile started FA fortification in 2001 with 2.4 mg of FA/1000 g in wheat flour [10], which led to a 43% decrease in the prevalence of NTD two years later [11]. This greater supply of FA along with the greater bioavailability of FA (near 100%), twice that of natural folate [4], have dramatically increased FA consumption levels with unknown long-term effects on offspring. It is important to note that FA has a tolerable upper intake limit (UL) value of 1000 μg/day [4], mainly because higher levels may mask vitamin B12 deficiency [12,13]. Vitamin B12, which is present mainly in foods of animal origin [14], is also essential during pregnancy due to its role in folate metabolism, normal cell growth, prevention of birth defects and neurocognitive development [15,16]. The maternal requirement of vitamin B12 during pregnancy is slightly higher than in adults (from 2.4 to 2.6 μg/day) [4]. Due to the key role played by these vitamins (folates and vitamin B12) in the metabolism of methionine and methylation reactions, a deficiency, excess or imbalance in the supply of some of them during pregnancy could create an adverse intrauterine environment, and consequently induce fetal programming through epigenetic mechanisms [17,18]. Some studies have reported lower fetal growth due to a high folate/vitamin B12 ratio [19]. However, more studies are needed to confirm these findings. The delivery of nutrients from mother to fetus only occurs through the placenta [20]. Three specific folate transport mechanisms operate in the human placenta, including folate receptor (FOLR) in its -α (FOLR1) and β (FOLR2) forms [21], the reduced folate carrier (RFC) and the proton-coupled folate transporter (PCFT/HCP1) [22–24]. The transfer of folate from maternal circulation to the fetus is the result of the coordinated action of these transporters, although there is published evidence that FRα is the primary transporter for maternal folate to the fetus [25]. Involvement of members of the ABC superfamily of transporters in FA cellular homeostasis in the human placenta has also been reported [23]. Since altered folate levels have been linked to preterm pregnancies, the aim of this study was to investigate whether preterm conditions prese (...truncated)