Expression and functional characterisation of System L amino acid transporters in the human term placenta

Reproductive Biology and Endocrinology

Thomas Jansson⁴  Theresa L. Powell²  Yoshikatsu Kanai⁵  Vanessa I. Ramirez³  Susanne Lager⁶  Sara Roos¹  Irving L. M. H. Aye⁴  Francesca Gaccioli⁶ 

[1] Institute of Biomedicine, The Sahlgrenska Academy at the University of Gothenburg, Gothenburg, Sweden;Section of Neonatology, Department of Pediatrics, University of Colorado Denver Anschutz Medical Campus, Aurora, Denver, CO, USA;Center for Pregnancy and Newborn Research, Department of Obstetrics and Gynecology, University of Texas Health Science Center, San Antonio, TX, USA;Division of Basic Reproductive Sciences, Department of Obstetrics and Gynaecology, University of Colorado Denver Anschutz Medical Campus, Aurora, Denver, CO, USA;Division of Bio-System Pharmacology, Department of Pharmacology, Osaka University Graduate School of Medicine, Osaka, Japan;Department of Obstetrics and Gynaecology, University of Cambridge, Cambridge, UK

关键词: Pregnancy;    Maternal obesity;    Leucine uptake;    Placental nutrient transport;    Trophoblast cells;   

Others  :  1216400
DOI  :  10.1186/s12958-015-0054-8

 received in 2014-10-26, accepted in 2015-05-28,  发布年份 2015

【 摘 要 】

Background

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.

Methods

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.

Results

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.

Conclusions

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.

【 授权许可】

   
2015 Gaccioli et al.

【 图 表 】

Fig. 1.

Fig. 2.

Fig. 3.

Fig. 4.

【 参考文献 】

• [1]Jansson T. Amino acid transporters in the human placenta. Pediatr Res. 2001; 49(2):141-7.
• [2]Cleal JK, Lewis RM. The mechanisms and regulation of placental amino acid transport to the human foetus. J Neuroendocrinol. 2008; 20(4):419-26.
• [3]Kanai Y, Segawa H, Miyamoto K, Uchino H, Takeda E, Endou H. Expression cloning and characterization of a transporter for large neutral amino acids activated by the heavy chain of 4F2 antigen (CD98). J Biol Chem. 1998; 273(37):23629-32.
• [4]Rajan DP, Kekuda R, Huang W, Devoe LD, Leibach FH, Prasad PD et al.. Cloning and functional characterization of a Na(+)-independent, broad-specific neutral amino acid transporter from mammalian intestine. Biochim Biophys Acta. 2000; 1463(1):6-14.
• [5]Kudo Y, Boyd CA. Human placental amino acid transporter genes: expression and function. Reproduction. 2002; 124(5):593-600.
• [6]Meier C, Ristic Z, Klauser S, Verrey F. Activation of system L heterodimeric amino acid exchangers by intracellular substrates. EMBO J. 2002; 21(4):580-9.
• [7]Verrey F. System L: heteromeric exchangers of large, neutral amino acids involved in directional transport. Pflugers Arch. 2003; 445(5):529-33.
• [8]Rossier G, Meier C, Bauch C, Summa V, Sordat B, Verrey F et al.. LAT2, a new basolateral 4F2hc/CD98-associated amino acid transporter of kidney and intestine. J Biol Chem. 1999; 274(49):34948-54.
• [9]Prasad PD, Wang H, Huang W, Kekuda R, Rajan DP, Leibach FH et al.. Human LAT1, a subunit of system L amino acid transporter: molecular cloning and transport function. Biochem Biophys Res Commun. 1999; 255(2):283-8.
• [10]Pineda M, Fernandez E, Torrents D, Estevez R, Lopez C, Camps M et al.. Identification of a membrane protein, LAT-2, that Co-expresses with 4F2 heavy chain, an L-type amino acid transport activity with broad specificity for small and large zwitterionic amino acids. J Biol Chem. 1999; 274(28):19738-44.
• [11]Okamoto Y, Sakata M, Ogura K, Yamamoto T, Yamaguchi M, Tasaka K et al.. Expression and regulation of 4F2hc and hLAT1 in human trophoblasts. Am J Physiol Cell Physiol. 2002; 282(1):C196-204.
• [12]Johnson LW, Smith CH. Neutral amino acid transport systems of microvillous membrane of human placenta. Am J Physiol. 1988; 254(6 Pt 1):C773-80.
• [13]Hoeltzli SD, Smith CH. Alanine transport systems in isolated basal plasma membrane of human placenta. Am J Physiol. 1989; 256(3 Pt 1):C630-7.
• [14]Kudo Y, Boyd CA. Characterization of amino acid transport systems in human placental basal membrane vesicles. Biochim Biophys Acta. 1990; 1021(2):169-74.
• [15]Kudo Y, Boyd CA. Characterisation of L-tryptophan transporters in human placenta: a comparison of brush border and basal membrane vesicles. J Physiol. 2001; 531(Pt 2):405-16.
• [16]Lewis RM, Glazier J, Greenwood SL, Bennett EJ, Godfrey KM, Jackson AA et al.. L-serine uptake by human placental microvillous membrane vesicles. Placenta. 2007; 28(5–6):445-52.
• [17]Cleal JK, Brownbill P, Godfrey KM, Jackson JM, Jackson AA, Sibley CP et al.. Modification of fetal plasma amino acid composition by placental amino acid exchangers in vitro. J Physiol. 2007; 582(Pt 2):871-82.
• [18]Gaccioli F, Lager S, Powell TL, Jansson T. Placental transport in response to altered maternal nutrition. J Dev Origins Health Dis. 2013; 4(02):101-15.
• [19]Jansson T, Scholtbach V, Powell TL. Placental transport of leucine and lysine is reduced in intrauterine growth restriction. Pediatr Res. 1998; 44(4):532-7.
• [20]Rosario FJ, Jansson N, Kanai Y, Prasad PD, Powell TL, Jansson T. Maternal protein restriction in the rat inhibits placental insulin, mTOR, and STAT3 signaling and down-regulates placental amino acid transporters. Endocrinology. 2011; 152(3):1119-29.
• [21]Jansson T, Ekstrand Y, Bjorn C, Wennergren M, Powell TL. Alterations in the activity of placental amino acid transporters in pregnancies complicated by diabetes. Diabetes. 2002; 51(7):2214-9.
• [22]Lager S, Aye IL, Gaccioli F, Ramirez VI, Jansson T, Powell TL. Labor inhibits placental mechanistic target of rapamycin complex 1 signaling. Placenta. 2014; 35(12):1007-12.
• [23]Cindrova-Davies T, Yung HW, Johns J, Spasic-Boskovic O, Korolchuk S, Jauniaux E et al.. Oxidative stress, gene expression, and protein changes induced in the human placenta during labor. Am J Pathol. 2007; 171(4):1168-79.
• [24]Lee KJ, Shim SH, Kang KM, Kang JH, Park DY, Kim SH et al.. Global gene expression changes induced in the human placenta during labor. Placenta. 2010; 31(8):698-704.
• [25]Kliman HJ, Nestler JE, Sermasi E, Sanger JM, Strauss JF. Purification, characterization, and in vitro differentiation of cytotrophoblasts from human term placentae. Endocrinology. 1986; 118(4):1567-82.
• [26]Aye IL, Jansson T, Powell TL. Interleukin-1beta inhibits insulin signaling and prevents insulin-stimulated system A amino acid transport in primary human trophoblasts. Mol Cell Endocrinol. 2013; 381(1–2):46-55.
• [27]Meller M, Vadachkoria S, Luthy DA, Williams MA. Evaluation of housekeeping genes in placental comparative expression studies. Placenta. 2005; 26(8–9):601-7.
• [28]Roos S, Lagerlof O, Wennergren M, Powell TL, Jansson T. Regulation of amino acid transporters by glucose and growth factors in cultured primary human trophoblast cells is mediated by mTOR signaling. Am J Physiol Cell Physiol. 2009; 297(3):C723-31.
• [29]Lowry OH, Rosebrough NJ, Farr AL, Randall RJ. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951; 193(1):265-75.
• [30]Ericsson A, Hamark B, Powell TL, Jansson T. Glucose transporter isoform 4 is expressed in the syncytiotrophoblast of first trimester human placenta. Hum Reprod. 2005; 20(2):521-30.
• [31]Park SY, Kim JK, Kim IJ, Choi BK, Jung KY, Lee S et al.. Reabsorption of neutral amino acids mediated by amino acid transporter LAT2 and TAT1 in the basolateral membrane of proximal tubule. Arch Pharm Res. 2005; 28(4):421-32.
• [32]Yanagida O, Kanai Y, Chairoungdua A, Kim DK, Segawa H, Nii T et al.. Human L-type amino acid transporter 1 (LAT1): characterization of function and expression in tumor cell lines. Biochim Biophys Acta. 2001; 1514(2):291-302.
• [33]Altman A, Cardenas JM, Houghten RA, Dixon FJ, Theofilopoulos AN. Antibodies of predetermined specificity against chemically synthesized peptides of human interleukin 2. Proc Natl Acad Sci U S A. 1984; 81(7):2176-80.
• [34]Hisano S, Haga H, Miyamoto K, Takeda E, Fukui Y. The basic amino acid transporter (rBAT)-like immunoreactivity in paraventricular and supraoptic magnocellular neurons of the rat hypothalamus. Brain Res. 1996; 710(1–2):299-302.
• [35]Illsley NP, Wang ZQ, Gray A, Sellers MC, Jacobs MM. Simultaneous preparation of paired, syncytial, microvillous and basal membranes from human placenta. Biochim Biophys Acta. 1990; 1029(2):218-26.
• [36]Acosta O, Ramirez VI, Lager S, Gaccioli F, Dudley DJ, Powell TL et al.. Increased glucose and placental GLUT-1 in large infants of obese nondiabetic mothers. Am J Obstet Gynecol. 2014.
• [37]Bowers GN, McComb RB. A continuous spectrophotometric method for measuring the activity of serum alkaline phosphatase. Clin Chem. 1966; 12(2):70-89.
• [38]Jansson N, Rosario FJ, Gaccioli F, Lager S, Jones HN, Roos S et al.. Activation of placental mTOR signaling and amino acid transporters in obese women giving birth to large babies. J Clin Endocrinol Metab. 2013; 98(1):105-13.
• [39]Gaccioli F, White V, Capobianco E, Powell TL, Jawerbaum A, Jansson T. Maternal overweight induced by a diet with high content of saturated fat activates placental mTOR and eIF2 alpha signaling and increases fetal growth in rats. Biol Reprod. 2013; 89(4):96.
• [40]Sebire NJ, Jolly M, Harris JP, Wadsworth J, Joffe M, Beard RW et al.. Maternal obesity and pregnancy outcome: a study of 287,213 pregnancies in London. Int J Obes Relat Metab Disord. 2001; 25(8):1175-82.
• [41]Araujo JR, Correia-Branco A, Ramalho C, Goncalves P, Pinho MJ, Keating E et al.. L-methionine placental uptake: characterization and modulation in gestational diabetes mellitus. Reprod Sci. 2013; 20(12):1492-507.
• [42]Farmer DR, Nelson DM. A fibrin matrix modulates the proliferation, hormone secretion and morphologic differentiation of cultured human placental trophoblast. Placenta. 1992; 13(2):163-77.
• [43]Esterman A, Greco MA, Mitani Y, Finlay TH, Ismail-Beigi F, Dancis J. The effect of hypoxia on human trophoblast in culture: morphology, glucose transport and metabolism. Placenta. 1997; 18(2–3):129-36.
• [44]Morrish DW, Dakour J, Li H, Xiao J, Miller R, Sherburne R et al.. In vitro cultured human term cytotrophoblast: a model for normal primary epithelial cells demonstrating a spontaneous differentiation programme that requires EGF for extensive development of syncytium. Placenta. 1997; 18(7):577-85.
• [45]Cleal JK, Glazier JD, Ntani G, Crozier SR, Day PE, Harvey NC et al.. Facilitated transporters mediate net efflux of amino acids to the fetus across the basal membrane of the placental syncytiotrophoblast. J Physiol. 2011; 589(Pt 4):987-97.
• [46]Segawa H, Fukasawa Y, Miyamoto K, Takeda E, Endou H, Kanai Y. Identification and functional characterization of a Na + −independent neutral amino acid transporter with broad substrate selectivity. J Biol Chem. 1999; 274(28):19745-51.
• [47]Panitchob N, Widdows KL, Crocker IP, Hanson MA, Johnstone ED, Please CP et al.. Computational modelling of amino acid exchange and facilitated transport in placental membrane vesicles. J Theor Biol. 2015; 365:352-64.
• [48]Diaz P, Powell TL, Jansson T. The role of placental nutrient sensing in maternal-fetal resource allocation. Biol Reprod. 2014; 91(4):82.
• [49]Mahendran D, Donnai P, Glazier JD, D’Souza SW, Boyd RD, Sibley CP. Amino acid (system A) transporter activity in microvillous membrane vesicles from the placentas of appropriate and small for gestational age babies. Pediatr Res. 1993; 34(5):661-5.
• [50]Norberg S, Powell TL, Jansson T. Intrauterine growth restriction is associated with a reduced activity of placental taurine transporters. Pediatr Res. 1998; 44(2):233-8.
• [51]Glazier JD, Cetin I, Perugino G, Ronzoni S, Grey AM, Mahendran D et al.. Association between the activity of the system A amino acid transporter in the microvillous plasma membrane of the human placenta and severity of fetal compromise in intrauterine growth restriction. Pediatr Res. 1997; 42(4):514-9.
• [52]Paolini CL, Marconi AM, Ronzoni S, Di Noio M, Fennessey PV, Pardi G et al.. Placental transport of leucine, phenylalanine, glycine, and proline in intrauterine growth-restricted pregnancies. J Clin Endocrinol Metab. 2001; 86(11):5427-32.
• [53]Kuruvilla AG, D’Souza SW, Glazier JD, Mahendran D, Maresh MJ, Sibley CP. Altered activity of the system A amino acid transporter in microvillous membrane vesicles from placentas of macrosomic babies born to diabetic women. J Clin Invest. 1994; 94(2):689-95.
• [54]Ditchfield AM, Desforges M, Mills TA, Glazier JD, Wareing M, Mynett K, et al. Maternal obesity is associated with a reduction in placental taurine transporter activity. Int J Obes (2005). 2014. doi:10.1038/ijo.2014.212
• [55]Farley DM, Choi J, Dudley DJ, Li C, Jenkins SL, Myatt L et al.. Placental amino acid transport and placental leptin resistance in pregnancies complicated by maternal obesity. Placenta. 2010; 31(8):718-24.
• [56]Carter MF, Powell TL, Li C, Myatt L, Dudley D, Nathanielsz P et al.. Fetal serum folate concentrations and placental folate transport in obese women. Am J Obstet Gynecol. 2011; 205(1):83.
• [57]Colomiere M, Permezel M, Riley C, Desoye G, Lappas M. Defective insulin signaling in placenta from pregnancies complicated by gestational diabetes mellitus. Eur J Endocrinol. 2009; 160(4):567-78.
• [58]Dube E, Gravel A, Martin C, Desparois G, Moussa I, Ethier-Chiasson M et al.. Modulation of fatty acid transport and metabolism by maternal obesity in the human full-term placenta. Biol Reprod. 2012; 87(1):14.
• [59]Chu SY, Callaghan WM, Kim SY, Schmid CH, Lau J, England LJ et al.. Maternal obesity and risk of gestational diabetes mellitus. Diabetes Care. 2007; 30(8):2070-6.
• [60]Catalano PM. The impact of gestational diabetes and maternal obesity on the mother and her offspring. J Dev Orig Health Dis. 2010; 1(4):208-15.
• [61]Leddy MA, Power ML, Schulkin J. The impact of maternal obesity on maternal and fetal health. Rev Obstet Gynaecol. 2008; 1(4):170-8.
• [62]Knight KM, Pressman EK, Hackney DN, Thornburg LL. Perinatal outcomes in type 2 diabetic patients compared with non-diabetic patients matched by body mass index. J Matern Fetal Neonatal Med. 2012; 25(6):611-5.
• [63]Wahabi HA, Fayed AA, Alzeidan RA, Mandil AA. The independent effects of maternal obesity and gestational diabetes on the pregnancy outcomes. BMC Endocr Disord. 2014; 14:47. BioMed Central Full Text
• [64]Bystrom M, Liu A, Quinton AE, Champion BL, Mann K, Peek M et al.. Gestational diabetes independently increases birth length and augments the effects of maternal BMI on birth weight: a retrospective cohort study. Frontiers Pediatr. 2014; 2:112.
• [65]Uebel K, Pusch K, Gedrich K, Schneider KT, Hauner H, Bader BL. Effect of maternal obesity with and without gestational diabetes on offspring subcutaneous and preperitoneal adipose tissue development from birth up to year-1. BMC Pregnancy Childbirth. 2014; 14:138. BioMed Central Full Text
• [66]Boyle KE, Hwang H, Janssen RC, DeVente JM, Barbour LA, Hernandez TL et al.. Gestational diabetes is characterized by reduced mitochondrial protein expression and altered calcium signaling proteins in skeletal muscle. PLoS One. 2014; 9(9):e106872.
• [67]Hastie R, Lappas M. The effect of pre-existing maternal obesity and diabetes on placental mitochondrial content and electron transport chain activity. Placenta. 2014; 35(9):673-83.