Rat-specific Expression of Placental MATE1 Contributes to Species Difference in Fetal Transfer of Metformin and 1-Methyl-4-phenylpyridinium

Metformin readily crosses the human placenta. However, the fetal-to-maternal plasma concentration ratio (F/M ratio) of metformin has been reported to be as low as 0.057 in rats. This study investigated the underlying mechanisms of these interspecies differences by examining the expression and function of candidate transporters, organic cation transporter 3 (OCT3/SLC22A3) and multidrug and toxin extrusion protein 1 (MATE1/SLC47A1), in the placenta across species. Expression analysis at gene and protein levels using RNA-sequencing and absolute quantification by LC-MS/MS revealed minimal species differences in placental OCT3 expression (less than 4-fold for mRNA and protein). In contrast, MATE1 showed almost exclusive expression in the rat placental labyrinth but minimal or undetectable levels were observed in samples prepared from human placental villi and mouse placental labyrinth. To evaluate the function of the placental MATE1, we compared the placental transfer of two MATE1/OCT3 substrates, metformin and 1-methyl-4-phenylpyridinium (MPP+), in pregnant rats and mice in the presence and absence of pyrimethamine, a MATE1 inhibitor. The pyrimethamine was dosed to reach the plasma concentration which specifically inhibits MATE1 based on the measured IC50 of pyrimethamine to rat Mate1 and Oct3, which were 0.015 μM and 100 μM, respectively. The F/M ratios of metformin and MPP+ in the absence of pyrimethamine were half or less in rats compared to mice. Notably, pre-administration of pyrimethamine increased the F/M ratios of metformin and MPP+ by approximately 50% in rats but not mice. These findings demonstrate that functional placental MATE1 expression is specific to rats, which, in part, contributes to the low fetal transfer of cationic compounds compared to other species. This is of significant concern, as it suggests that nonclinical developmental and reproductive toxicity studies of MATE1 substrate drugs in rats may underestimate fetal exposure and toxicity when extrapolated to humans.二甲双胍可轻松穿过人胎盘。不过有研究报道，大鼠体内二甲双胍的胎儿-母体血浆浓度比（F/M比）低至0.057。本研究通过检测不同物种胎盘中候选转运蛋白——有机阳离子转运蛋白3（OCT3/SLC22A3）与多药及毒素外排蛋白1（MATE1/SLC47A1）的表达与功能，探究此类种间差异的潜在机制。采用RNA测序（RNA-sequencing）与液相色谱-串联质谱法（LC-MS/MS）进行基因与蛋白水平的表达分析及绝对定量，结果显示胎盘OCT3的表达仅存在极小的种间差异（mRNA与蛋白水平差异均小于4倍）。与之相反，MATE1几乎仅在大鼠胎盘迷路层中表达，而在人胎盘绒毛及小鼠胎盘迷路层的样本中仅能检测到极微量或无法检测到其表达。为评估胎盘MATE1的功能，我们分别在妊娠大鼠与小鼠体内，给予与不给予MATE1抑制剂乙胺嘧啶的情况下，比较了两种MATE1/OCT3底物——二甲双胍与1-甲基-4-苯基吡啶鎓（MPP+）的胎盘转运情况。基于测得的乙胺嘧啶对大鼠Mate1与Oct3的半数抑制浓度（IC50）分别为0.015 μM与100 μM，我们调整乙胺嘧啶的给药剂量以使其血浆浓度可特异性抑制MATE1。未给予乙胺嘧啶时，大鼠体内二甲双胍与MPP+的F/M比仅为小鼠的一半或更低。值得注意的是，预先给予乙胺嘧啶可使大鼠体内二甲双胍与MPP+的F/M比升高约50%，但小鼠体内无此变化。上述研究结果表明，功能性胎盘MATE1的表达具有大鼠特异性，这在一定程度上导致相较于其他物种，阳离子化合物在大鼠体内的胎儿转运率更低。这一发现值得高度关注，因为这意味着以MATE1底物药物开展的大鼠非临床发育与生殖毒性研究，在将结果外推至人类时，可能会低估胎儿暴露量与毒性风险。