Maternal 3,3’-Diiodothyronine Sulfate Formation from Guinea Pig Placenta Perfused with 3,3’,5-Triodothyronine

Objective: Serum 3, 3’,5-triiodothyronine (T3) remains low in near-term fetus to prevent the growing fetus from undue exposure to its active catabolic effect in mammals. The present study was undertaken to gain insight in the role of placenta in T3 metabolism, fetal to maternal transfer of T3, and its metabolites by in situ placenta perfusion with outer-ring labeled [125I]-T3 in pregnant guinea pig, a species showing increased sulfated 3, 3’-diiodothyronine (T2S) levels in maternal serum in late pregnancy (term = 65 days), similarly to humans in pregnancy. Materials and Methods: One-pass placenta perfusions performed on pregnant guinea pigs were studied between 58 - 65 days of gestation. In two separate experiments, the umbilical artery of the guinea pig placenta was perfused in situ at 37°C with outer-ring labeled [125I]-T3. Maternal sera and umbilical effluents were obtained for analysis at the end of a 60-minute perfusion, when the steady-state levels of radioactivity were reached in the placenta effluent after 30-minute. Results: Sulfated [125I]-T2S was readily detected in the maternal serum as the major metabolite of T3 following the perfusion of placenta with [125I]-T3, suggesting that placental inner-ring deiodinase and sulfotransferase may play an important role in fetal T3 homeostasis and in the fetal to maternal transfer of sulfated iodothyronine metabolites. Conclusions: The expression of type 3 deiodinase (D3) and thyroid hormone sulfotransferase activity in placenta may play an important role to protect developing organs against undue exposure to active thyroid hormone in late gestation in the fetus. The combined activities of D3 and sulfotransferase promoted a placental transfer of T2S into maternal circulation. The maternal circulation of T2S is fetal T3 in origin and its role as a fetal thyroid function biomarker deserves further evaluations and studies.

Nanoparticle-mediated transgene expression of insulin-like growth factor 1 in the guinea pig placenta differentially affects fetal liver gene expression depending on maternal nutrient status

Fetal growth restriction (FGR) occurs in up to 10% of pregnancies and is a leading cause of infant morbidity and mortality. Additionally, FGR has been implicated in contributing to the development of long-term health outcomes including increasing the risk for future cardiovascular and endocrine diseases. Currently, there is limited preventative strategies and no effective treatment options for FGR. To address this need, we are developing a therapeutic targeting the placenta to increase expression of human insulin-like growth factor 1 (hIGF-1) and enhance placental development and function, with the goal of correcting fetal growth trajectories. Methods Initially, an ultrasound-guided, transcutaneous, intra-placental injection of a non-viral, Texas-red conjugated polymer-based nanoparticle containing a plasmid with the green fluorescent protein (GFP) gene under the control of the placenta-specific promotors Plac1 or Cyp19a1 was performed to determine nanoparticle uptake and transgene expression in the guinea pig placenta. Subsequently, using the established maternal nutrient restriction (MNR) guinea pig model of FGR, placentas were treated with an unconjugated nanoparticle containing a plasmid with the hIGF-1 gene under the Cyp19a1 promotor at mid-pregnancy (gestational day (GD) 30-33). Five days after treatment placentas and fetal liver tissue was collected, weighed, and fixed for histology or snap-frozen for qPCR analysis of mRNA expression. Results Histological analysis of Texas-red and GFP fluorescence in placenta and fetal liver tissue confirmed nanoparticle uptake and transgene expression and that nanoparticle was unable to cross the placenta to fetal circulation. In situ hybridization for plasmid-specific mRNA confirmed sustained hIGF-1 expression five days after treatment. MNR resulted in 20-25% decreased fetal weight at mid-pregnancy (P<0.001) that was not changed with nanoparticle treatment (P>0.05). There was no effect of nanoparticle treatment on the volume densities of trophoblasts or fetal capillaries in the placenta (P>0.05 for both). However, treatment did reduce the interhaemal distance between the maternal blood space and fetal circulation in the MNR placentas compared to sham treated MNR placentas (P<0.001). In the fetuses, placental nanoparticle treatment increased circulating glucose by 38-50% (P<0.001) and was associated with differential changes to fetal liver mRNA expression of genes associated with gluconeogenesis. Gene expression changes were dependent on if the fetus was growth restricted or not; nanoparticle treatment: down-regulated gluconeogenesis gene expression in the normal growing fetuses but increased expression in the FGR fetuses. Conclusions The current study shows that treatment of the guinea pig placenta with a polymer-based nanoparticle causes expression of hIGF-1 and ultimately increases fetal glucose concentrations within five days of treatment. Furthermore, the data shows that the placenta and fetal liver respond differently to nanoparticle treatment depending on fetal growth conditions.

Molecular Characterization of Microtubule Affinity-Regulating Kinase4 from Sus scrofa and Promotion of Lipogenesis in Primary Porcine Placental Trophoblasts

This study aimed to characterize the full-length cDNA of MARK4 in Sus scrofa, and evaluated its potential role in the regulation of lipid accumulation in pig placental trophoblasts and analyzed signaling pathways involved, thereby providing insights into mechanisms for placental lipotoxicity induced by excessive back-fat during pregnancy of sows. The cDNA obtained with 5′ and 3′ RACE amplification covered 3216 bp with an open reading frame of 2259 bp encoding 752 amino acids. Multiple alignments and phylogenetic analysis revealed MARK4 protein of Sus scrofa had a high homology (95%–99%) to that of other higher vertebrates. After transfection, enhanced MARK4 significantly promoted lipogenesis in pig trophoblasts, as evidenced by accelerated lipid accumulation and consistently increased mRNA expressions of lipogenic genes DGAT1, LPIN1, LPIN3, LPL, PPARδ and SREBP-1c. Meanwhile, PPARγ remarkably inhibited the stimulating effect of MARK4 on non-receptor-mediated lipid accumulation in trophoblasts. Further analyses revealed WNT signaling enhanced lipid accumulation and activation of MARK4 in pig trophoblast cells. Finally, we demonstrated that WNT/β-catenin signal pathway is involved in MARK4 activated lipogenesis. These results suggest that MARK4 promotes lipid accumulation in porcine placental trophoblasts and can be considered as a potential regulator of lipotoxicity associated with maternal obesity in the pig placenta.