Jocelyn D. Glazier

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

Primarily, our objectives were to compare system A amino acid transporter activity in the microvillous plasma membrane (MVM) of placentas from normally grown (appropriate for gestational age, AGA) and intrauterine growth-restricted (IUGR) fetuses delivered during the third trimester, as a whole and in relation to the severity of IUGR. Ten AGA and 16 IUGR pregnancies were studied at the time of elective cesarean section performed between 28 and 40 wk of gestation. Severity of IUGR pregnancies was assessed primarily by Doppler velocimetry and fetal heart rate monitoring. Placental MVM vesicles were prepared, and system A activity in these was measured. The transporter activity was significantly lower in IUGR compared with AGA pregnancies. Within the IUGR group system A activity was only significantly lower, compared with AGA, in cases that presented with a reduction in umbilical blood flow. We conclude that placental MVM system A activity is lower in IUGR compared with AGA pregnancies delivered during the third trimester. System A activity is related to the severity of IUGR.

Amino acid (system A) transporter activity in microvillous membrane vesicles from the placentas of appropriate and small for gestational age babies

ABSTRACT: Although a number of causes of poor fetal growth are known, the involvement of placental transport proteins in the etiology of growth retardation is not understood. The aim of this study was to investigate the activity of the system A amino acid transporter and the Na+/H+ exchanger in vesicles isolated from the microvillous membrane of the syncytiotrophoblast of placentas of appropriate and small for gestational age babies. There were no biochemical differences between the membranes from the two groups of placentas, and there was no difference in the activity of the Na+/H+ exchanger. The initial rate of uptake of methylaminoisobutyric acid (a nonmetabolizable amino acid analogue) was 63% lower in vesicles from placentas of small for gestational age babies. Kinetic analysis of the system A transporter (utilized by methylaminoisobutyric acid) showed that the Vmax in the vesicles from placentas of small for gestational age babies (0.24 ± 0.03 nmol/mg protein/30 s, n = 5) was significantly lower than that in vesicles from placentas of appropriate for gestational age babies (0.64 ± 0.09 nmol/mg protein/30 s, n = 4, p < 0.001), whereas the Km was not different between the two groups. It is concluded that there is an abnormality of system A amino acid transporter function in placentas of small for gestational age babies.

Placental Phenotypes of Intrauterine Growth

The placenta is essential to nutrition before birth. Recent work has shown that a range of clearly defined alterations can be found in the placentas of infants with intrauterine growth restriction (IUGR). In the mouse, a placental specific knockout of a single imprinted gene, encoding IGF-2, results in one pattern of alterations in placenta structure and function which leads to IUGR. We speculate that the alterations in the human placenta can also be grouped into patterns, or phenotypes, that are associated with specific patterns of fetal growth. Identifying the placental phenotypes of different fetal growth patterns will improve the ability of clinicians to recognize high-risk patients, of laboratory scientists to disentangle the complexities of IUGR, and of public health teams to target interventions aimed at ameliorating the long-term adverse effects of inadequate intrauterine growth.

Expression of folate transporters in human placenta and implications for homocysteine metabolism

Poor folate status during pregnancy can lead to elevated maternal plasma levels of homocysteine (Hcy) with associated pregnancy complications and adverse neonatal outcomes, suggesting placental metabolism of Hcy might be an important determinant in influencing fetal development. The metabolic pathways for Hcy in placenta are not well defined. In this study we examined the gene expression of key enzymes involved in Hcy metabolism in first trimester and term human placenta to determine which metabolic pathways prevail. Expression of mRNA for methionine synthase and 5,10-methylene tetrahydrofolate reductase, enzymes involved in the methionine cycle and responsible for the re-methylation of Hcy to methionine, were expressed at similar levels between first trimester and term and in comparison to human liver as positive control. In contrast, cystathionine beta-synthase mRNA expression was markedly lower than that in liver at both gestational periods. Betaine-homocysteine methyltransferase mRNA was undetectable at either gestational age. These data suggest that re-methylation of Hcy using methyl donation from 5-methyltetrahydrofolate is the prevalent pathway, indicating a marked reliance on folate availability. This led to further investigations examining the expression and localisation of folate transporters in first trimester and term placenta. Folate receptor alpha (FRalpha) was highly polarised to the microvillous plasma membrane (MVM) of the syncytiotrophoblast at both gestational periods, a distribution shared by the proton-coupled folate transporter which co-localised with FRalpha. Reduced folate carrier was distributed to both MVM and basal syncytiotrophoblast plasma membranes at term suggesting a role at both loci, and in first trimester was localised to MVM as well as cytotrophoblast plasma membranes. These data support the concept that placental folate transport is established early in pregnancy, providing folate for utilisation in placental Hcy metabolism.

Altered activity of the system A amino acid transporter in microvillous membrane vesicles from placentas of macrosomic babies born to diabetic women.

Fetal macrosomia (FM) is a well-recognized complication of diabetic pregnancy but it is not known whether placental transport mechanisms are altered. We therefore studied the activity of the system A amino acid transporter, the system L amino acid transporter, and the Na+/H+ exchanger in microvillous membrane vesicles from placentas of macrosomic babies born to diabetic women (FM group), from placentas of appropriately grown babies born to diabetic women (appropriate for gestational age group) and from placentas of appropriately grown babies of normal women (control group). Sodium-dependent uptake of [14C]-methylaminoisobutyric acid at 30 s (initial rate, a measure of system A activity) was 49% lower into FM vesicles than into control vesicles (P < 0.02); this effect was due to a decrease in Vmax of the transporter with no change in Km. There was no significant difference in system A activity between the appropriate for gestational age group and control or FM group. There was also no difference between system L transporter or Na+/H+ exchanger activity between the three groups. We conclude that the number of system A transporters per milligram of membrane protein in the placental microvillous membrane is selectively reduced in diabetic pregnancies associated with FM.

Placental transporter activity and expression in relation to fetal growth

The question of whether there are causative or compensatory changes in placental transport physiology affecting fetal growth is considered. Reductions in uterine and umbilical blood flow in growth retardation will reduce maternofetal exchange of lipophilic solutes, such as O2 and CO2, but will not have a major effect on the transfer of hydrophilic solutes. These solutes are transferred across the placenta by paracellular diffusion, transporter protein‐mediated transport and endocytosis‐exocytosis. Neither paracellular diffusion nor endocytosis‐exocytosis has been investigated in relation to fetal growth. The weight of evidence is that there is no change in the activity and expression of the syncytiotrophoblast GI UTI glucose transporter in fetal growth retardation. However, there is strong evidence that the activity of the system A amino acid transporter, per milligram of placental membrane protein, is altered in relation to fetal growth, but in a complex manner. There is also some weaker evidence that the activity of the Na(+)‐H+ exchanger, per milligram of placental membrane protein, is directly related to birth‐weight. There are no data for other solute transporters; a considerable amount of work still remains to be done in order to understand the relationship between placental function and fetal growth rate.

Placental-specific Igf2 knockout mice exhibit hypocalcemia and adaptive changes in placental calcium transport

Evidence is emerging that the ability of the placenta to supply nutrients to the developing fetus adapts according to fetal demand. To examine this adaptation further, we tested the hypothesis that placental maternofetal transport of calcium adapts according to fetal calcium requirements. We used a mouse model of fetal growth restriction, the placental-specific Igf2 knockout (P0) mouse, shown previously to transiently adapt placental System-A amino acid transporter activity relative to fetal growth. Fetal and placental weights in P0 mice were reduced when compared with WT at both embryonic day 17 (E17) and E19. Ionized calcium concentration [Ca2+] was significantly lower in P0 fetal blood compared with both WT and maternal blood at E17 and E19, reflecting a reversal of the fetomaternal [Ca2+] gradient. Fetal calcium content was reduced in P0 mice at E17 but not at E19. Unidirectional maternofetal calcium clearance (Ca K mf) was not different between WT and P0 at E17 but increased in P0 at E19. Expression of the intracellular calcium-binding protein calbindin-D9K, previously shown to be rate-limiting for calcium transport, was increased in P0 relative to WT placentas between E17 and E19. These data show an increased placental transport of calcium from E17 to E19 in P0 compared to WT. We suggest that this is an adaptation in response to the reduced fetal calcium accumulation earlier in gestation and speculate that the ability of the placenta to adapt its supply capacity according to fetal demand may stretch across other essential nutrients.

The SNAT4 isoform of the system A amino acid transporter is functional in human placental microvillous plasma membrane

Placental system A activity is important for the supply of neutral amino acids needed for fetal growth. There are three system A isoforms: SNAT1, SNAT2 and SNAT4, but the contribution of each to system A‐mediated transport is unknown. Here, we have used immunohistochemistry to demonstrate that all three isoforms are present in the syncytiotrophoblast suggesting each plays a role in amino acid transport across the placenta. We next tested the hypothesis that the SNAT4 isoform is functional in microvillous plasma membrane vesicles (MVM) from normal human placenta using a method which exploits the unique property of SNAT4 to transport both cationic amino acids as well as the system A‐specific substrate MeAIB. The data show that SNAT4 contribution to system A‐specific amino acid transport across MVM is higher in first trimester placenta compared to term (approx. 70% and 33%, respectively, P < 0.01). Further experiments performed under more physiological conditions using intact placental villous fragments suggest a contribution of SNAT4 to system A activity in first trimester placenta but minimal contribution at term. In agreement, Western blotting revealed that SNAT4 protein expression is higher in first trimester MVM compared to term (P < 0.05). This study provides the first evidence of SNAT4 activity in human placenta and demonstrates the contribution of SNAT4 to system A‐mediated transport decreases between first trimester and term: our data lead us to speculate that at later stages of gestation SNAT1 and/or SNAT2 are more important for the supply of amino acids required for normal fetal growth.

Placental Malaria-Associated Inflammation Disturbs the Insulin-like Growth Factor Axis of Fetal Growth Regulation

BACKGROUND The pathogenetic mechanisms of fetal growth restriction associated with placental malaria are largely unknown. We sought to determine whether placental malaria and related inflammation were associated with disturbances in the insulin-like growth factor (IGF) axis, a major regulator of fetal growth. METHOD We measured IGF-1 and IGF-2 concentrations in plasma from 88 mother-neonate pairs at delivery and IGF binding proteins 1 and 3 (IGFBP-1 and IGFBP-3, respectively) in cord plasma from a cohort of Papua New Guinean women with and without placental malaria. Messenger RNA levels of IGF-1, IGF-2, and the IGF receptors were measured in matched placental biopsy specimens. RESULTS Compared with those for uninfected pregnancies, IGF-1 levels were reduced by 28% in plasma samples from women with placental Plasmodium falciparum infection and associated inflammation (P = .007) and by 25% in their neonates (P = .002). Levels of fetal IGFBP-1 were elevated in placental malaria with and without inflammation (P = .08 and P = .006, respectively) compared with uninfected controls. IGF-2 and IGFBP-3 plasma concentrations and placental IGF ligand and receptor messenger RNA transcript levels were similar across groups. CONCLUSION Placental malaria-associated inflammation disturbs maternal and fetal levels of IGFs, which regulate fetal growth. This may be one mechanism by which placental malaria leads to fetal growth restriction.

Review: Adaptation in placental nutrient supply to meet fetal growth demand: Implications for programming

This review considers the hypothesis that adaptations in blood flow, exchange surface area and transporter activity enable placental supply capacity to meet fetal demand and cause alterations in fetal composition which result in life-long programming of homeostatic set points. We consider the components of placental supply capacity and describe the predominant changes each of these could impose on solute and water exchange across the placenta. We next consider the evidence that adaptations in placental nutrient supply to meet the demands of fetal growth and development do occur. Evidence from human and mouse studies suggests that adaptations occur in regulation of blood flow through the fetoplacental circulation, in exchange barrier surface area and in transporter-mediated processes for amino acids and calcium. Crucially there appear to be differences in the gestational timing of these adaptations. Finally we suggest that each of these adaptations could have separate effects on the composition of the fetus. These could affect physiological set points in different ways and so programme the lifetime responses of the individual.