Regulatory T cells (Tregs), interleukin 17-producing T helper cells (Th17 cells), and other immune cells play important roles in the maintenance of pregnancy, and their function is impacted by HIV infection. I hypothesized that FIV-infection may likewise alter placental T cell gene expression causing aberrant immune function and compromised pregnancy. The purpose of this project was to evaluate the expression placental immunomodulators related to Treg and Th17 cell function in the FIV-infected cat model. Gene expression was quantified in placenta and serum using quantitative PCR and ELISA, respectively. Abnormal expression of cytokines was frequently associated with infection and fetal nonviability, resulting in discordant relationships between cytokine pairs and the nuclear transcriptional regulators FoxP3 and RORã. The expression of IL-6 in the periphery mirrored that of the placenta, indicating a potential serological means to predict pregnancy outcome. In conclusion, aberrant placental immunomodulation accompanied failed pregnancy in the FIV-infected cat model.

<p>The pathophysiology of how a maternal infection induces fetal inflammation and subsequently premature birth is a growing area of research. The <em>ex-vivo</em> dual closed-loop placental perfusion model has been widely used to study placental physiology. To address the association between bacterial chorioamnionitis and fetal inflammation, TNF-α induction following lipopolysaccharide (LPS) challenge – a pyrogen of Gram-negative origin – was measured in the perfusion model. Preliminary analysis of perfusates unexpectedly revealed markedly elevated levels of TNF-α in control and LPS-treated groups indicating contamination of material(s) capable of activating innate immune responses.</p> <p>To identify source(s) driving high background TNF-α expression in perfusates, bovine serum albumin (BSA) – the chief component of the perfusion media – the perfusion system and the materno-feto-placental unit were independently examined. To validate a cleaning protocol effective in LPS removal, acid-base and oxidative depyrogenation techniques were also additionally assessed in the perfusion system.</p> <p>Using TNF-α as a surrogate marker of contamination, high background TNF-α expression in previously conducted placental perfusions were attributed to (1) LPS contaminated perfusion media and (2) LPS build up in the perfusion system. Additionally, results from depyrogenation experiments revealed both acid-base and oxidative techniques effectively reduced LPS buildup in the perfusion system to levels that were in accordance with FDA guidelines for medical equipment (< 0.5 EU/mL). Thus, to circumvent LPS-derived contamination placentas should be perfused using endotoxin-free perfusion media and the perfusion system should be cleaned with acid-base or oxidative depyrogenation techniques prior to its use.</p> / Master of Science (MSc)

Placental development can be disturbed by various factors, such as mutation of specific genes or maternal diabetes. Our previous work on interspecies hybrid placental dysplasia (IHPD) and two additional models of placental hyperplasia, cloned mice and Esx1 mutants, showed that many genes are deregulated in placental dysplasia. Two of these candidate placentation genes, Cpe and Lhx3, were further studied. We performed in situ hybridization to determine their spatio-temporal expression in the placentas and placental phenotypes were analyzed in mutant mice. Our results showed that the placental phenotype in Cpe mutant mice mimics some IHPD phenotypes. Deregulated expression of Cpe and Cpd, a functionally equivalent gene, prior to the manifestation of the IHPD phenotype, indicated that Cpe and Cpd are potentially causative genes in IHPD. Lhx3 mutants lacked any placental phenotype. Deletion of Lhx3 and Lhx4 together caused an inconsistent placental phenotype which did not affect placental lipid transport function or expression of Lhx3/Lhx4 target genes. Down regulation of Lhx3/Lhx4 did not rescue the placental phenotype of AT24 mice and hence could be excluded as causative genes in IHPD. Analysis of placental development in diabetic mice showed that severe maternal diabetes resulted in fetal intrauterine growth restriction (IUGR) without any change in placental weight and lipid transport function. The diabetic placentas however exhibited abnormal morphology. Gene expression profiling identified some genes that might contribute to diabetic pathology. In another study, it was found that the heterochromatin protein CBX1 is required for normal placentation, as deletion of the gene caused consistent spongiotrophoblast and labyrinthine phenotypes. Gene expression profiling and spatio-temporal expression analysis showed that several genes with known function in placental development were deregulated in the Cbx1 null placenta.

Title page, table of contents and abstract only. The complete thesis in print form is available from the University of Adelaide Library. / A range of epidemiological, clinical and experimental studies have demonstrated that exposure of an embryo to a suboptimal environment in vivo or ex vivo during early embryo development is associated with altered development of cardiovascular, neuroendocrine and metabolic disorders in adult life. A number of perturbations during early embryo development result in developmental adaptations by the embryo to ensure immediate survival, whilst programming the embryo for altered fetal and placental development, resulting in the eventual onset of adult disease. It has been previously shown that maternal nutrient restriction during the periconceptional period results in a hyperactivation of the pituitary - adrenal axis and increased mean arterial blood pressure in twin but not singleton pregnancies. It was therefore the first aim of this thesis to interrogate the impact of maternal undernutrition during the periconceptional period (defined as from at least 45 days prior until 7 days after conception) on fetal and placental development during early pregnancy at - day 55 of pregnancy, which coincides with the period of maximal placental growth. In Chapter 2, it has been demonstrated that there are important relationships between maternal weight gain during the periconceptional period and feto-placental growth during the first - 55 days of pregnancy and that periconceptional undernutrition has a differential effect on these relationships in singleton and twin pregnancies. In singleton pregnancies, periconceptional undernutrition disrupts the relationship between maternal weight gain during the periconceptional period and utero-placental growth and in twin pregnancies, periconceptional undernutrition results in the emergence of an inverse relationship between maternal weight gain during early pregnancy and uteroplacental growth and in a dependence of fetal growth on placental growth. (Chapter 2) In order to investigate the origins of the physiological adaptations that lead to the development of hyperactivation of the pituitary - adrenal axis and increased mean arterial blood pressure in late gestational fetuses after exposure as an embryo to periconceptional undernutrition, we investigated the development and steroidogenic capacity of the fetal adrenal gland and development of the fetal heart and kidney at - 55 days gestation (Chapter 3 and 4). The relative weight of the fetal adrenal and adrenal IGF-1, IGF-1 R, IGF-2, IGF-2R and CYP 17 mRNA expression were lower in twin compared to singleton fetuses. There was evidence that in control singletons, IGF-2R expression plays an important role in the regulation of adrenal growth and CYP 17 mRNA expression during early pregnancy. In control twins, however, whilst there was a significant positive relationship between adrenal CYP 17 and IGF-2 mRNA expression, adrenal weight was directly related to the level of adrenal IGF-1 mRNA expression. There was no effect of periconceptional undernutrition on the level of expression of any of the placental or adrenal genes in the study. In PCUN ewes, carrying singletons, however, there was a loss of the relationships between either adrenal IGF-2, IGF-2R and IGF-1 mRNA expression and adrenal growth and CYP 17 expression which were present in control singletons. Similarly in ewes carrying twins, maternal undernutrition during the periconceptional period resulted in the loss of the relationships between adrenal growth and IGF-1 expression and between _ adrenal CYP 17 and IGF-2 expression which were present in control twin fetuses. Whilst there was no effect of fetal number on fetal heart growth at - d55 in twin fetuses, there was a direct relationship between relative fetal heart and adrenal weights, which was present in both the PCUN and control groups. There was also a significant inverse relationship between maternal weight at conception and relative fetal heart weight in PCUN twin, but not PCUN singleton or control fetuses (Chapter 3). In control pregnancies maternal weight gain during the periconceptional period is inversely related to the relative weight of the fetal kidney at -55d pregnancy. In this group, relative kidney weight was also directly related to renal IGF-1 mRNA expression. In control twins maternal weight gain was inversely related to fetal kidney weight and this effect was ablated when the effects of maternal cortisol was controlled for in the analysis. In the PCUN group, whilst there was an inverse relationship between maternal weight gain during the periconceptional period and relative kidney weight, it was not possible to separate the independent effects of maternal weight loss during the periconceptional period and the subsequent weight gain during the period of refeeding. Renal IGF-1 mRNA expression was higher and renal lGF-1 R and 2R expression were lower in twin fetuses compared to singletons. After exposure to PCUN, renal IGF-1 expression was also higher than in control pregnancies independent of the fetal number (Chapter 4). Superovulation, artificial insemination, embryo transfer and in vitro embryo culture are used in a range of assisted reproductive technologies, and it has been demonstrated that varying the composition of the culture media can result in a change in pre and postnatal development. Culture of sheep embryos in media containing serum is associated with fetal overgrowth which is phenotypic of the Large Offspring Syndrome. It is not known how the combination of superovulation, artificial insemination and embryo transfer alone impacts fetoplacental development in late gestation of the sheep. There have been no studies, however, examining the differential impact of superovulation, artificial insemination and embryo transfer with or without in vitro embryo culture in the absence or presence of human serum on feto-placental development in Singleton and twin pregnancies (Chapter 5). I have therefore tested the hypothesis that superovulation, artificial insemination and embryo transfer with or without in vitro embryo culture in the presence or absence of human serum differentially alters the growth of the placenta, fetus and fetal organs during late gestation when compared to naturally conceived controls and that these effects are different in singleton and twin pregnancies. The fetal weight, CRL and abdominal circumference were significantly larger in IVCHS singleton fetuses. A novel finding in this study was lower fetal weights of twin fetuses in the ET and IVCNS groups compared to NM control twin fetuses. In addition, placental weights were lighter in twin fetuses in the ET, IVCNS and IVCHS treatment groups and this is partially due to a failure to initiate compensatory growth of placentomes in twin pregnancies (Chapter 5). The results of this thesis therefore highlight the complex interactions between the periconceptional environment (in vivo or ex vivo) and embryo or fetal number on the programming fetal and placental development. Maternal undernutrition during the periconceptional period and superovulation, artificial insemination and embryo transfer with or without in vitro culture in the absence or presence of serum alters fetal development, and I have demonstrated that these changes in fetal growth can be explained by changes in placental growth trajectory. Furthermore, a novel finding of this study is that perturbations of the periconceptional environment affect feto-placental development differently in singleton and twin pregnancies. / Thesis (Ph.D.) -- University of Adelaide, School of Molecular and Biomedical Science, 2006

Title page, table of contents and abstract only. The complete thesis in print form is available from the University of Adelaide Library. / A range of epidemiological, clinical and experimental studies have demonstrated that exposure of an embryo to a suboptimal environment in vivo or ex vivo during early embryo development is associated with altered development of cardiovascular, neuroendocrine and metabolic disorders in adult life. A number of perturbations during early embryo development result in developmental adaptations by the embryo to ensure immediate survival, whilst programming the embryo for altered fetal and placental development, resulting in the eventual onset of adult disease. It has been previously shown that maternal nutrient restriction during the periconceptional period results in a hyperactivation of the pituitary - adrenal axis and increased mean arterial blood pressure in twin but not singleton pregnancies. It was therefore the first aim of this thesis to interrogate the impact of maternal undernutrition during the periconceptional period (defined as from at least 45 days prior until 7 days after conception) on fetal and placental development during early pregnancy at - day 55 of pregnancy, which coincides with the period of maximal placental growth. In Chapter 2, it has been demonstrated that there are important relationships between maternal weight gain during the periconceptional period and feto-placental growth during the first - 55 days of pregnancy and that periconceptional undernutrition has a differential effect on these relationships in singleton and twin pregnancies. In singleton pregnancies, periconceptional undernutrition disrupts the relationship between maternal weight gain during the periconceptional period and utero-placental growth and in twin pregnancies, periconceptional undernutrition results in the emergence of an inverse relationship between maternal weight gain during early pregnancy and uteroplacental growth and in a dependence of fetal growth on placental growth. (Chapter 2) In order to investigate the origins of the physiological adaptations that lead to the development of hyperactivation of the pituitary - adrenal axis and increased mean arterial blood pressure in late gestational fetuses after exposure as an embryo to periconceptional undernutrition, we investigated the development and steroidogenic capacity of the fetal adrenal gland and development of the fetal heart and kidney at - 55 days gestation (Chapter 3 and 4). The relative weight of the fetal adrenal and adrenal IGF-1, IGF-1 R, IGF-2, IGF-2R and CYP 17 mRNA expression were lower in twin compared to singleton fetuses. There was evidence that in control singletons, IGF-2R expression plays an important role in the regulation of adrenal growth and CYP 17 mRNA expression during early pregnancy. In control twins, however, whilst there was a significant positive relationship between adrenal CYP 17 and IGF-2 mRNA expression, adrenal weight was directly related to the level of adrenal IGF-1 mRNA expression. There was no effect of periconceptional undernutrition on the level of expression of any of the placental or adrenal genes in the study. In PCUN ewes, carrying singletons, however, there was a loss of the relationships between either adrenal IGF-2, IGF-2R and IGF-1 mRNA expression and adrenal growth and CYP 17 expression which were present in control singletons. Similarly in ewes carrying twins, maternal undernutrition during the periconceptional period resulted in the loss of the relationships between adrenal growth and IGF-1 expression and between _ adrenal CYP 17 and IGF-2 expression which were present in control twin fetuses. Whilst there was no effect of fetal number on fetal heart growth at - d55 in twin fetuses, there was a direct relationship between relative fetal heart and adrenal weights, which was present in both the PCUN and control groups. There was also a significant inverse relationship between maternal weight at conception and relative fetal heart weight in PCUN twin, but not PCUN singleton or control fetuses (Chapter 3). In control pregnancies maternal weight gain during the periconceptional period is inversely related to the relative weight of the fetal kidney at -55d pregnancy. In this group, relative kidney weight was also directly related to renal IGF-1 mRNA expression. In control twins maternal weight gain was inversely related to fetal kidney weight and this effect was ablated when the effects of maternal cortisol was controlled for in the analysis. In the PCUN group, whilst there was an inverse relationship between maternal weight gain during the periconceptional period and relative kidney weight, it was not possible to separate the independent effects of maternal weight loss during the periconceptional period and the subsequent weight gain during the period of refeeding. Renal IGF-1 mRNA expression was higher and renal lGF-1 R and 2R expression were lower in twin fetuses compared to singletons. After exposure to PCUN, renal IGF-1 expression was also higher than in control pregnancies independent of the fetal number (Chapter 4). Superovulation, artificial insemination, embryo transfer and in vitro embryo culture are used in a range of assisted reproductive technologies, and it has been demonstrated that varying the composition of the culture media can result in a change in pre and postnatal development. Culture of sheep embryos in media containing serum is associated with fetal overgrowth which is phenotypic of the Large Offspring Syndrome. It is not known how the combination of superovulation, artificial insemination and embryo transfer alone impacts fetoplacental development in late gestation of the sheep. There have been no studies, however, examining the differential impact of superovulation, artificial insemination and embryo transfer with or without in vitro embryo culture in the absence or presence of human serum on feto-placental development in Singleton and twin pregnancies (Chapter 5). I have therefore tested the hypothesis that superovulation, artificial insemination and embryo transfer with or without in vitro embryo culture in the presence or absence of human serum differentially alters the growth of the placenta, fetus and fetal organs during late gestation when compared to naturally conceived controls and that these effects are different in singleton and twin pregnancies. The fetal weight, CRL and abdominal circumference were significantly larger in IVCHS singleton fetuses. A novel finding in this study was lower fetal weights of twin fetuses in the ET and IVCNS groups compared to NM control twin fetuses. In addition, placental weights were lighter in twin fetuses in the ET, IVCNS and IVCHS treatment groups and this is partially due to a failure to initiate compensatory growth of placentomes in twin pregnancies (Chapter 5). The results of this thesis therefore highlight the complex interactions between the periconceptional environment (in vivo or ex vivo) and embryo or fetal number on the programming fetal and placental development. Maternal undernutrition during the periconceptional period and superovulation, artificial insemination and embryo transfer with or without in vitro culture in the absence or presence of serum alters fetal development, and I have demonstrated that these changes in fetal growth can be explained by changes in placental growth trajectory. Furthermore, a novel finding of this study is that perturbations of the periconceptional environment affect feto-placental development differently in singleton and twin pregnancies. / Thesis (Ph.D.) -- University of Adelaide, School of Molecular and Biomedical Science, 2006

Pentamidine is used to treat Pneumocystis carinii pneumonia. The incidence of this infection in pregnancy has paralleled the increasing incidence of acquired immunodeficiency syndrome in pregnancy. Using the in vitro bidirectionally perfused human placenta, we studied the transfer of pentamidine across the placenta. Pentamidine was added to the maternal circulation at therapeutic concentrations (2 wg/ml). No transfer of pentamidine was detectable with a newly devised high-performance liquid chromatography method sensitive to 0.05 wg/ml of pentamidine. Increasing the pentamidine concentration tenfold produced a low level of transfer to the fetal circuit. Fetal concentrations were far below maternal perfusate concentrations. Placental tissue levels were higher than media levels. These data are suggestive of minimal drug transfer to the fetus and significant concentration of the drug in placental tissue. (Am J Obstet Gynecol 1989;160:759-61.)

No description available.

The genetic architecture of placental abruption (PA) remains poorly understood. We examined variations in SNPs of circadian clock-related genes in placenta with PA risk. We also explored placental and maternal genomic contributions to PA risk. Placental genomic DNA samples were isolated from 280 PA cases and 244 controls. Genotyping was performed using the Illumina Cardio-MetaboChip. We examined 116 SNPs in 13 genes known to moderate circadian rhythms. Logistic regression models were fit to estimate odds ratios (ORs). The combined effect of multiple SNPs on PA risk was estimated using a weighted genetic risk score. We examined independent and joint associations of wGRS derived from placental and maternal genomes with PA. Seven SNPs in five genes (ARNTL2, CRY2, DEC1, PER3 and RORA), in the placental genome, were associated with PA risk. Each copy of the minor allele (G) of a SNP in the RORA gene (rs2899663) was associated with a 30% reduced odds of PA (95% CI 0.52-0.95). The odds of PA increased with increasing placental-wGRS (P_trend<0.001). The ORs were 1.00, 2.16, 3.24 and 4.48 across quartiles. Associations persisted after the maternal-wGRS was included in the model. There was evidence of an additive contribution of placental and maternal genetic contributions to PA risk. Participants with placental- and maternal-wGRS in the highest quartile, compared with those in the lowest quartile, had a 15.57-fold (95% CI 3.34- 72.60) increased odds of PA. Placental variants in circadian clock-related genes are associated with PA risk; and the association persists after control of genetic variants in the maternal genome

Preeclampsia (PE) remains one of the most enigmatic and pervasive conditions developed during pregnancy and is a leading cause of maternal and fetal morbidity and mortality throughout the world. Afflicting nearly 5-8% of pregnancies in the Unites States, PE is most commonly characterized by an increase in blood pressure and high protein excretion near or after the 20th week of gestation. Unfortunately, few effective treatments are available, and the only "cure" is delivery. While the molecular pathogenesis of PE remains undefined, an interruption in placental blood flow, or placental ischemia, is widely observed as a primary contributor to the syndrome progression. Furthermore, to investigate the role of both pharmacological and non-pharmacological modalities to prevent placental ischemia induced hypertension, we employed a robust model of reduced utero-placental perfusion pressure (RUPP) in the pregnant rat. First, in Chapter IV, exercise initiated during gestation was not effective in the prevention of RUPP-induced hypertension, whereas exercise training prior to and continued through gestation prevented the increase in blood pressure. Though the molecular contributions to this effect are undefined, the effects appear to be independent of angiogenic balance restoration. Finally, in Chapter V, administration of 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside (AICAR) was explored as a novel pharmacological modality to prevent the onset of hypertension and endothelial dysfunction in the RUPP model. As hypothesized, AICAR ameliorated the RUPP-induced hypertension, and the anti-hypertensive effect in the RUPP appears to be dependent on the restoration of angiogenic balance in the maternal plasma. This dissertation includes previously published and unpublished co-authored material.

Gestational diabetes, a common medical complication in pregnancy, may lead to severe fetal consequences if left untreated. A major concern with the use of antidiabetic drugs in pregnancy is the potential for placental transfer and fetal toxicity. The presence of endocytic pathways and several ABC transporter proteins has been demonstrated in the human placenta and are believed to play an important role in determining fetal exposure to drugs used in pregnancy. The objective of this thesis is to investigate the safety and transfer of the oral hypoglycemic agent, glyburide, and the new long acting insulin analog, insulin glargine, across the human placenta. The oral antidiabetic, glyburide, has been shown to be actively effluxed across the placenta in the fetal to maternal direction. The transport of glyburide in the presence of a breast cancer resistance protein (BCRP) inhibitor was investigated in the dually perfused human placenta model. The results of the perfusion studies indicate that BCRP plays a role in protecting the fetus from the accumulation of glyburide. Subsequently, cellular studies were carried out to determine the effect of the naturally occurring single nucleotide polymorphism within the coding region of BCRP (C421A/Q141K) on glyburide transport. Results suggest that glyburide transport may be reduced in the presence of the Q141K polymorphism. While insulin remains as the gold standard, the potential for maternal hypoglycemia with insulin injection has resulted in the development of insulin analogs. Insulin glargine, a human insulin analog, has a long half life with no pronounced peak when compared to currently used NPH insulin. Human placental perfusion experiments examining the extent and rate of transfer of insulin glargine across the placenta demonstrated that, at therapeutic concentrations, insulin glargine does not cross the placenta to a measurable extent. To further determine the fetal safety of insulin glargine therapy compared with NPH insulin, a systematic review and meta-analysis were performed. No evidence was identified for increased adverse fetal outcomes with the use of insulin glargine during pregnancy. Overall, the results of this research serve to provide improved treatment options for women with diabetes in pregnancy.