Gaccioli Francesca, Aye Irving L M H, Roos Sara, Lager Susanne, Ramirez Vanessa I, Kanai Yoshikatsu, Powell Theresa L, Jansson Thomas
Department of Obstetrics and Gynaecology, University of Cambridge, Cambridge, UK.
Division of Basic Reproductive Sciences, Department of Obstetrics and Gynaecology, University of Colorado Denver Anschutz Medical Campus, Aurora, Denver, CO, USA.
Reprod Biol Endocrinol. 2015 Jun 9;13:57. doi: 10.1186/s12958-015-0054-8.
System L transporters LAT1 (SLC7A5) and LAT2 (SLC7A8) mediate the uptake of large, neutral amino acids in the human placenta. Many System L substrates are essential amino acids, thus representing crucial nutrients for the growing fetus. Both LAT isoforms are expressed in the human placenta, but the relative contribution of LAT1 and LAT2 to placental System L transport and their subcellular localisation are not well established. Moreover, the influence of maternal body mass index (BMI) on placental System L amino acid transport is poorly understood. Therefore the aims of this study were to determine: i) the relative contribution of the LAT isoforms to System L transport activity in primary human trophoblast (PHT) cells isolated from term placenta; ii) the subcellular localisation of LAT transporters in human placenta; and iii) placental expression and activity of System L transporters in response to maternal overweight/obesity.
System L mediated leucine uptake was measured in PHT cells after treatment with si-RNA targeting LAT1 and/or LAT2. The localisation of LAT isoforms was studied in isolated microvillous plasma membranes (MVM) and basal membranes (BM) by Western blot analysis. Results were confirmed by immunohistochemistry in sections of human term placenta. Expression and activity System L transporters was measured in isolated MVM from women with varying pre-pregnancy BMI.
Both LAT1 and LAT2 isoforms contribute to System L transport activity in primary trophoblast cells from human term placenta. LAT1 and LAT2 transporters are highly expressed in the MVM of the syncytiotrophoblast layer at term. LAT2 is also localised in the basal membrane and in endothelial cells lining the fetal capillaries. Measurements in isolated MVM vesicles indicate that System L transporter expression and activity is not influenced by maternal BMI.
LAT1 and LAT2 are present and functional in the syncytiotrophoblast MVM, whereas LAT2 is also expressed in the BM and in the fetal capillary endothelium. In contrast to placental System A and beta amino acid transporters, MVM System L activity is unaffected by maternal overweight/obesity.
L系统转运蛋白LAT1（SLC7A5）和LAT2（SLC7A8）介导人胎盘中大的中性氨基酸的摄取。许多L系统底物是必需氨基酸，因此是发育中胎儿的关键营养素。两种LAT亚型均在人胎盘中表达，但LAT1和LAT2对胎盘L系统转运的相对贡献及其亚细胞定位尚未明确。此外，母体体重指数（BMI）对胎盘L系统氨基酸转运的影响了解甚少。因此，本研究的目的是确定：i）LAT亚型对从足月胎盘分离的原代人滋养层（PHT）细胞中L系统转运活性的相对贡献；ii）LAT转运蛋白在人胎盘中的亚细胞定位；iii）胎盘L系统转运蛋白在母体超重/肥胖情况下的表达和活性。
在用靶向LAT1和/或LAT2的小干扰RNA处理后，测量PHT细胞中L系统介导的亮氨酸摄取。通过蛋白质印迹分析在分离的微绒毛质膜（MVM）和基底膜（BM）中研究LAT亚型的定位。结果在人足月胎盘切片中通过免疫组织化学得到证实。在来自不同孕前BMI的女性的分离的MVM中测量L系统转运蛋白的表达和活性。
LAT1和LAT2亚型均对人足月胎盘的原代滋养层细胞中的L系统转运活性有贡献。LAT1和LAT2转运蛋白在足月合体滋养层的MVM中高表达。LAT2也定位于基底膜和胎儿毛细血管内衬的内皮细胞中。在分离的MVM囊泡中的测量表明，L系统转运蛋白的表达和活性不受母体BMI的影响。
LAT1和LAT2存在于合体滋养层MVM中且具有功能，而LAT2也在基底膜和胎儿毛细血管内皮中表达。与胎盘A系统和β氨基酸转运蛋白相反，MVM的L系统活性不受母体超重/肥胖的影响。
