Intrauterine growth restriction is associated with alterations in placental lipoprotein receptors and maternal lipoprotein composition

Among other factors, fetal growth requires maternal supply of cholesterol. Cellular cholesterol uptake is mainly mediated by the LDL receptor (LDL-R) and the scavenger receptor family. We hypothesized that expression levels of key receptors of these families were regulated differently in placentas from IUGR pregnancies with varying degrees of severity. Third-trimester placentas from IUGR pregnancies with (IUGR-S) and without (IUGR-M) fetal hemodynamic changes and from control (AGA) pregnancies were studied. LDL-R, LDL-Rrelated protein (LRP-1), and scavenger receptor class B type I (SR-BI) mRNA and protein levels were measured. Cholesterol concentration and composition of lipoproteins were analyzed enzymatically and by lipid electrophoresis, respectively, in maternal and umbilical cord blood. LDL-R mRNA levels in IUGR-M were similar to AGA but lower (P ! 0.05) in IUGR-S. In contrast, LDL-R protein was twofold (IUGR-M) and 1.8-fold (IUGR-S) higher (P ! 0.05) than in the AGA group. LRP-1 mRNA and protein levels were not altered in the IUGR cases. SR-BI mRNA was unchanged in IUGR, but protein levels were lower (P ! 0.05) in IUGR-S than in the other groups. Maternal plasma concentrations of LDL cholesterol were higher (P ! 0.05) in the AGA group (188.5 " 23.6 mg/dl) than in the IUGR-S group (154.2 " 26.1). Electrophoretic mobility of the LDL fraction in maternal plasma demonstrated significant changes in migration toward higher values (AGA 0.95 " 0.06, IUGR-M 1.12 " 0.11, P ! 0.001; IUGR-S 1.28 " 0.20, P # 0.002). We conclude that LDL-R and SR-BI levels are altered in IUGR pregnancies. These differences were associated with changes in LDL, but not HDL, mobility and cholesterol concentration in maternal circulation

Inhibition of diacylglycerol lipase beta modulates lipid and endocannabinoid levels in the ex vivo human placenta

IntroductionLipids and fatty acids are key components in metabolic processes of the human placenta, thereby contributing to the development of the fetus. Placental dyslipidemia and aberrant activity of lipases have been linked to diverse pregnancy associated complications, such as preeclampsia and preterm birth. The serine hydrolases, diacylglycerol lipase alpha and beta (DAGL alpha, DAGL beta) catalyze the degradation of diacylglycerols, leading to the formation of monoacylglycerols (MAG), including one main endocannabinoid 2-arachidonoylglycerol (2-AG). The major role of DAGL in the biosynthesis of 2-AG is evident from various studies in mice but has not been investigated in the human placenta. Here, we report the use of the small molecule inhibitor DH376, in combination with the ex vivo placental perfusion system, activity-based protein profiling (ABPP) and lipidomics, to determine the impact of acute DAGL inhibition on placental lipid networks. MethodsDAGL alpha and DAGL beta mRNA expression was detected by RT-qPCR and in situ hybridization in term placentas. Immunohistochemistry staining for CK7, CD163 and VWF was applied to localize DAGL beta transcripts to different cell types of the placenta. DAGL beta activity was determined by in- gel and MS-based activity-based protein profiling (ABPP) and validated by addition of the enzyme inhibitors LEI-105 and DH376. Enzyme kinetics were measured by EnzChek (TM) lipase substrate assay. Ex vivo placental perfusion experiments were performed +/- DH376 [1 mu M] and changes in tissue lipid and fatty acid profiles were measured by LC-MS. Additionally, free fatty acid levels of the maternal and fetal circulations were determined. ResultsWe demonstrate that mRNA expression of DAGL beta prevails in placental tissue, compared to DAGL alpha (p <= 0.0001) and that DAGL beta is mainly located to CK7 positive trophoblasts (p <= 0.0001). Although few DAGL alpha transcripts were identified, no active enzyme was detected applying in-gel or MS-based ABPP, which underlined that DAGL beta is the principal DAGL in the placenta. DAGL beta dependent substrate hydrolysis in placental membrane lysates was determined by the application of LEI-105 and DH376. Ex vivo pharmacological inhibition of DAGL beta by DH376 led to reduced MAG tissue levels (p <= 0.01), including 2-AG (p <= 0.0001). We further provide an activity landscape of serine hydrolases, showing a broad spectrum of metabolically active enzymes in the human placenta. DiscussionOur results emphasize the role of DAGL beta activity in the human placenta by determining the biosynthesis of 2-AG. Thus, this study highlights the special importance of intra-cellular lipases in lipid network regulation. Together, the activity of these specific enzymes may contribute to the lipid signaling at the maternal-fetal interface, with implications for function of the placenta in normal and compromised pregnancies.Molecular Physiolog

CD39 abrogates platelet-derived factors induced IL-1β expression in the human placenta

Tissue insults in response to inflammation, hypoxia and ischemia are accompanied by the release of ATP into the extracellular space. There, ATP modulates several pathological processes, including chemotaxis, inflammasome induction and platelet activation. ATP hydrolysis is significantly enhanced in human pregnancy, suggesting that increased conversion of extracellular ATP is an important anti-inflammatory process in preventing exaggerated inflammation, platelet activation and hemostasis in gestation. Extracellular ATP is converted into AMP, and subsequently into adenosine by the two major nucleotide-metabolizing enzymes CD39 and CD73. Here, we aimed to elucidate developmental changes of placental CD39 and CD73 over gestation, compared their expression in placental tissue from patients with preeclampsia and healthy controls, and analyzed their regulation in response to platelet-derived factors and different oxygen conditions in placental explants as well as the trophoblast cell line BeWo. Linear regression analysis showed a significant increase in placental CD39 expression, while at the same time CD73 levels declined at term of pregnancy. Neither maternal smoking during first trimester, fetal sex, maternal age, nor maternal BMI revealed any effects on placental CD39 and CD73 expression. Immunohistochemistry detected both, CD39 and CD73, predominantly in the syncytiotrophoblast layer. Placental CD39 and CD73 expression were significantly increased in pregnancies complicated with preeclampsia, when compared to controls. Cultivation of placental explants under different oxygen conditions had no effect on the ectonucleotidases, whereas presence of platelet releasate from pregnant women led to deregulated CD39 expression. Overexpression of recombinant human CD39 in BeWo cells decreased extracellular ATP levels after culture in presence of platelet-derived factors. Moreover, platelet-derived factors-induced upregulation of the pro-inflammatory cytokine, interleukin-1β, was abolished by CD39 overexpression. Our study shows that placental CD39 is upregulated in preeclampsia, suggesting an increasing demand for extracellular ATP hydrolysis at the utero-placental interface. Increased placental CD39 in response to platelet-derived factors may lead to enhanced conversion of extracellular ATP levels, which in turn could represent an important anti-coagulant defense mechanism of the placenta

The effect of maternal undernutrition on the rat placental transcriptome: protein restriction up-regulates cholesterol transport

Fetal exposure to a maternal low protein diet during rat pregnancy is associated with hypertension, renal dysfunction and metabolic disturbance in adult life. These effects are present when dietary manipulations target only the first half of pregnancy. It was hypothesised that early gestation protein restriction would impact upon placental gene expression and that this may give clues to the mechanism which links maternal diet to later consequences. Pregnant rats were fed control or a low protein diet from conception to day 13 gestation. Placentas were collected and RNA Sequencing performed using the Illumina platform. Protein restriction down-regulated 67 genes and up-regulated 24 genes in the placenta. Ingenuity pathway analysis showed significant enrichment in pathways related to cholesterol and lipoprotein transport and metabolism, including atherosclerosis signalling, clathrin-mediated endocytosis, LXR/RXR and FXR/RXR activation. Genes at the centre of these processes included the apolipoproteins ApoB, ApoA2 and ApoC2, microsomal triglyceride transfer protein (Mttp), the clathrin-endocytosis receptor cubilin, the transcription factor retinol binding protein 4 (Rbp4) and transerythrin (Ttr; a retinol and thyroid hormone transporter). Real-time PCR measurements largely confirmed the findings of RNASeq and indicated that the impact of protein restriction was often striking (cubilin up-regulated 32-fold, apoC2 up-regulated 17.6-fold). The findings show that gene expression in specific pathways is modulated by maternal protein restriction in the day-13 rat placenta. Changes in cholesterol transport may contribute to altered tissue development in the fetus and hence programme risk of disease in later life

Maternal Angiotensin Increases Placental Leptin in Early Gestation via an Alternative Renin-Angiotensin System Pathway: Suggesting a Link to Preeclampsia.

Various studies found an association of different renin-angiotensin system (RAS) components with gestational duration and preterm birth, as well as with preeclampsia. Approximately 25% of first-time pregnant women develop a mild to severe hypertension in pregnancy or even preeclampsia. Based on recently published single-cell RNA-sequencing, we hypothesized an alternative RAS function in placenta and furthermore, an implication in hypertensive disorders in pregnancy. Placental RAS expression and localization was analyzed via quantitative polymerase chain reaction and in situ mRNA padlock probes. Tissue was collected from first-trimester elective termination (n=198), from healthy third-trimester controls (n=54), from early-onset preeclamptic (n=54) and age-matched controls (n=29), as well as first-trimester placentae from women with a high uterine artery resistance index (high-risk for preeclampsia, n=9) and controls (n=8). Serum levels of Ang (angiotensin) I to IV from women before and after conception were measured via mass spectrometry (n=10). Placental explants were cultured in 2.5% oxygen with Ang II, candesartan, and leptin. Seahorse XF96 MitoStress assays assessed trophoblast metabolism. Here, we show that maternal angiotensin acts on placental LNPEP (leucine aminopeptidase), that is, angiotensin IV-receptor and fetal angiotensin on placental AGTR1 (angiotensin II receptor type 1). Maternal circulating RAS shifts towards Ang IV in pregnancy. Ang IV decreases trophoblastic mitochondrial respiration and increases placental leptin via placental LNPEP. Lower placental LNPEP in preeclampsia and in first-trimester patients at high-risk for preeclampsia suggests a new alternative route in maternal RAS signaling and may contribute to hypertension and disease in pregnancy. The study shows how hypertensive disorders in pregnancy may be connected metabolic alterations that finally seem to contribute to the multifactorial disease in pregnancy, preeclampsia

Synthesis, structure and luminescence of Er3+-doped Y3Ga5O12 nano-garnets

A novel Y3(1-x)Er3xGa5O12 nanocrystalline garnet has been synthesized by a sol-gel technique and a complete structural, morphological, vibrational, and optical characterization has been carried out in order to correlate the local structure of the Er3+ ions with their optical properties. The synthesized nanocrystals are found in a single-phase garnet structure with an average grain size of around 60 nm. The good crystalline quality of the garnet structure is confirmed by FTIR and Raman measurements, since the phonon modes of the nano-garnet are similar to those found in the single crystal garnet. Under blue laser excitation, intense green and red visible and 1.5 mu m infrared luminescences are observed, whose relative intensities are very sensitive to the Er3+ concentration. The dynamics of these emissions under pulsed laser excitations are analyzed in the framework of different energy transfer interactions. Intense visible upconverted luminescence can be clearly observed by the naked eye for all synthesized Er3+-doped Y3Ga5O12 nano-garnets under a cw 790 nm laser excitation. The power dependency and the dynamics of the upconverted luminescence confirm the existence of different two-photon upconversion processes for the green and red emissions that strongly depend on the Er3+ concentration, showing the potential of these nano-garnets as excellent candidates for developing new optical devices.This work has been partially supported by Ministerio de Ciencia e Innovacion of Spain (MICCIN) under The National Program of Materials (MAT2010-21270-C04-02; -03; -04), The Consolider-Ingenio 2010 Program (MALTA CSD2007-0045), and The National Infrastructure Program, by Ministerio de Economia y Competitividad of Spain (MINECO) within The Indo-Spanish Joint Programme of Cooperation in Science and Technology (PRI-PIBIN-2011-1153/DST-INT-Spain-P-38-11), and by the EU-FEDER funds (UCAN08-4E-008). S.F. Leon-Luis and V. Monteseguro wish to thank MICINN for the FPI grants (BES-2008-003353 and BES-2011-044596). Dr V. Venkatramu is grateful to DAE-BRNS, Government of India for the award of DAE Research Award for Young Scientists (no. 2010/20/34/5/BRNS/2223).Venkatramu, V.; León-Luis, SF.; Rodriguez-Mendoza, UR.; Monteseguro, V.; Manjón, FJ.; Lozano-Gorrín, AD.; Valiente, R.... (2012). Synthesis, structure and luminescence of Er3+-doped Y3Ga5O12 nano-garnets. Journal of Materials Chemistry. 22:13788-13799. doi:10.1039/c2jm31386cS13788137992