Expression of Concentrative Nucleoside Transporters (SLC28A) in the Human Placenta: Effects of Gestation Age and Prototype Differentiation-Affecting Agents

Abstract

Equilibrative ( SLC29A) and concentrative ( SLC28A) nucleoside transporters contribute to proper placental development and mediate uptake of nucleosides/nucleoside-derived drugs. We analyzed placental expression of SLC28A mRNA during gestation. Moreover, we studied in choriocarcinoma-derived BeWo cells whether SLC29A and SLC28A mRNA levels can be modulated by activity of adenylyl cyclase, retinoic acid receptor activation, CpG islands methylation, or histone acetylation, using forskolin, all- trans-retinoic acid, 5-azacytidine, and sodium butyrate/sodium valproate, respectively. We found that expression of SLC28A1, SLC28A2, and SLC28A3 increases during gestation and reveals considerable interindividual variability. SLC28A2 was shown to be a dominant subtype in the first-trimester and term human placenta, while SLC28A1 exhibited negligible expression in the term placenta only. In BeWo cells, we detected mRNA of SLC28A2 and SLC28A3. Levels of the latter were affected by 5-azacytidine and all- trans-retinoic acid, while the former was modulated by sodium valproate (but not sodium butyrate), all- trans-retinoic acid, 5-azacytidine, and forskolin that caused 25-fold increase in SLC28A2 mRNA; we documented by analysis of syncytin-1 that the observed changes in SLC28A expression do not correlate with the morphological differentiation state of BeWo cells. Upregulated SLC28A2 mRNA was reflected in elevated uptake of [H]-adenosine, high-affinity substrate of concentrative nucleoside transporter 2. Using KT-5720 and inhibitors of phosphodiesterases, we subsequently confirmed importance of cAMP/protein kinase A pathway in SLC28A2 regulation. On the other hand, SLC29A genes exhibited constitutive expression and none of the tested compounds increased SLC28A1 expression to detectable levels. In conclusion, we provide the first evidence that methylation status and activation of retinoic acid receptor affect placental SLC28A2 and SLC28A3 transcription and substrates of concentrative nucleoside transporter 2 might be taken up in higher extent in placentas with overactivated cAMP/protein kinase A pathway and likely in the term placenta.

Equilibrative Nucleoside Transporter 1 (ENT1, SLC29A1) Facilitates Transfer of the Antiretroviral Drug Abacavir across the Placenta

“…The different effects of Na + depletion in BeWo cells and ex vivo human placental models can be explained by a three-orders-ofmagnitude higher expression of SLC28A2 (coding for CNT2) in BeWo cells when compared with the human placenta (Jiraskova et al, 2018). Moreover, the lack of CNT effect seems to be in accordance with the observation by Govindarajan et al (2007), who did not detect any protein expression of CNTs in the trophoblast layer of the term placenta.…”

“…Low hSLC29A2 placental expression may thus hamper the capability of our experimental models to detect abacavir interactions with ENT2. In contrast to findings for other placental transporters (e.g., ABCB1, ABCG2, and CNTs) (Gil et al, 2005;Meyer zu Schwabedissen et al, 2006;Jiraskova et al, 2018), the expression of hSLC29A1 and hSLC29A2 did not change in the course of gestation (Fig. 7A).…”

“…The presence of functional ENTs in BeWo cells has previously been confirmed by binding assays with NBMPR and by observing the effect of NBMPR on the accumulation of thymidine (Boumah et al, 1992;Mani et al, 1998;Karbanova et al, 2017). Recently, we have shown that our clone of BeWo expresses SLC28A2 and lower levels of SLC28A3 mRNA (Jiraskova et al, 2018). Here, we observed the effect of Na + depletion on [ 3 H]-adenosine (CNT2 and CNT3 substrate) but not [ 3 H]-thymidine (CNT1 and CNT3 substrate) (Molina-Arcas et al, 2009) (Fig.…”

“…Moreover, the lack of CNT effect seems to be in accordance with the observation by Govindarajan et al (2007), who did not detect any protein expression of CNTs in the trophoblast layer of the term placenta. Because the levels of CNTs increase over the course of gestation (Jiraskova et al, 2018), we do not hypothesize that CNTs are involved in maternal-placental transfer in earlier phases of gestation either.…”

“…The expression of drug transporters in the placenta frequently varies during gestation and differs among individuals (Gil et al, 2005;Mao, 2008;. This holds true also for SLC28A, exhibiting significant interindividual variability and higher expression in the term placenta compared with the first trimester (Jiraskova et al, 2018); however, such data are lacking for SLC29A.…”