Currently there is no test to accurately predict stillbirth. It is proposed that better identification of placental disease in utero may aid stillbirth prediction and prevention. Pregnancies complicated by reduced fetal movement (RFM) have increased risk of stillbirth. We hypothesised that RFM is a symptom of placental dysfunction associated with adverse pregnancy outcome (APO) and that this placental abnormality can be detected antenatally and used to identify fetuses at highest-risk of APO. We tested this hypothesis by: 1) comparison of ex vivo placental structure and function between APO RFM pregnancies and their normal outcome RFM counterparts, 2) comparison of in utero estimates of placental size, vascularity, vascular and endocrine functions obtained from placental ultrasound, Doppler waveform analysis and maternal circulating placentally-derived hormone concentrations, to their ex vivo correlates and 3) examination of the predictive potential of placental biomarkers at the time of RFM.Ex vivo placentas from APO RFM pregnancies, compared to normal outcome RFM counterparts, were smaller (diameter, area, weight and volume, p<0.0001), less vascular (vessel number and density, p≤0.002), with arteries that were less responsive to sodium nitroprusside (p<0.05), and with aberrant endocrine function (reduced tissue content and/or release of human chorionic gonadotrophin (hCG), human placental lactogen (hPL) and soluble fms-like Tyrosine Kinase-1 (sFlt-1), p<0.03). Placental volume (PV) ex vivo correlated with sonographic estimated PV (p<0.004), hPL, hCG and placental growth factor (PlGF) concentrations in the maternal circulation (p<0.03). Ex vivo villous vessel number and density correlated with Doppler impedance at the umbilical artery free-loop (UAD-F, p=0.02) and intraplacental arteries (p<0.0001) respectively, whilst UAD-F impedance correlated with arterial thromboxane sensitivity (p<0.04). Examination of placental structure and function at the time of presentation with RFM identified 15 independently-predictive biomarkers. Three potential predictive models, incorporating measures of placental size (PlGF), endocrine function (sFlt-1), arterial thromboxane sensitivity and villous vascularity (UAD-F), were proposed. Using these models, sensitivity for APO was improved from 8.9% with baseline care (assessment of fetal size and gestation) to up to 37.5% at a fixed specificity of 99% (p<0.05). This series of studies shows that antenatal placental examination is possible and improves identification of pregnancies at highest risk of stillbirth in a high-risk population by up to 29%. Therefore such tests merit further development to prospectively assess their ability to predict and prevent stillbirth itself.

Fetal growth restriction (FGR) is a common pregnancy complication resulting in increased neonatal mortality and morbidity. The aetiology of fetal growth restriction is not fully understood, but abnormalities in placental development are, leading to abnormalities in placental structure which are thought to affect supply of oxygen to the fetus. The source of fetal hypoxia is unknown due to the difficulty in obtaining oxygenation data in the context of pregnancy using existing techniques. There is also an absence of data relating to oxygenation in FGR pregnancies. Oxygen-Enhanced MRI (OE-MRI) and Blood Oxygen-Level Dependent (BOLD) MRI permit noninvasive acquisition of data related to changes in the concentration of dissolved oxygen (pO2) and changes in hemoglobin saturation (sO2) under air- and oxygen- breathing (hyperoxic challenge).The aim of this project was to determine whether MRI methods can provide information relating to placental oxygenation in normal and FGR-compromised pregnancy, to investigate fetal brain oxygenation and to assess the potential confound of placental perfusion changes under hyperoxic challenge. After optimization of sequences in non-pregnant volunteers, similar pO2 and sO2 increases under hyperoxic challenge were seen in normal and FGR pregnancy. This suggested placental oxygenation was similar and that fetal extraction of oxygen may be a likelier cause of fetal hypoxia. Normal fetal brain oxygenation was found not to increase under hyperoxic challenge, which may be due to hemodynamic adaptation to limit cerebral hyperoxygenation. Finally, the robustness of these oxygenation results was supported by the lack of placental perfusion changes observed under hyperoxia using Arterial Spin Labeling (ASL).In conclusion, MRI methods successfully provided information on placental and fetal oxygenation in normal and abnormal pregnancy, obtaining novel data informing the aetiology of FGR and the physiology of the fetal brain.

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第22744号 / 医博第4662号 / 新制||医||1046(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 髙折 晃史, 教授 竹内 理, 教授 近藤 玄 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Background: Pregnancy substance use is linked to low birth weight. However, less is known about relative contributions of various substances and whether effects are due to decreased gestational duration, restriction of fetal growth, or both. The study goal was to use causal modeling to evaluate the individual impact of alcohol, tobacco, cocaine, and marijuana on gestational duration and fetal growth. Methods: Participants were 3164 urban black women recruited at entry to prenatal care and followed to delivery, with all gestational dating ultrasound supported. Pregnancy substance use was assessed via self-report (alcohol, tobacco, cocaine, and marijuana). Results: Alcohol, cigarette, and cocaine use were all individually and negatively related to gestational age at delivery. However, only alcohol, cigarette, and marijuana use predicted fetal growth, with effects for alcohol and cigarette greater and more discrepant for older women. Overall, heavy cigarette smoking had the greatest individual impact on birth weight (up to 431 g). Heavy levels of use of all 4 substances by older women decreased birth weight by 26% (806 g). Conclusions: For perhaps the first time, reduced birth weight is apportioned both by type of substance and mechanism of effect. The use of alcohol and/or cigarettes was clearly more harmful to fetal growth than cocaine use. Findings demonstrate the need for continued emphasis on intervention efforts to address legal and illicit pregnancy substance use.

Introduction: Preeclampsia (PE) and fetal growth restriction (FGR) are common obstetrical complications, often with pathological features of maternal vascular malperfusion (MVM) in the placenta. Current placental clinical pathology methods involve a manual visual examination of histology sections, a practice that can be resource-intensive and demonstrate moderate-to-poor inter-pathologist agreement on diagnostic outcomes, dependant on the degree of pathologist sub-specialty training. Methods: This thesis aims to apply different machine learning (ML) feature extraction methods to classify digital images of placental histopathology specimens, collected from PE, FGR, PE + FGR, and healthy pregnancies, according to the presence or absence of MVM lesions. 166 digital images were captured from histological placental specimens, manually scored for MVM lesions (MVM- or MVM+) and used to develop various support vector machine (SVM) classifier models, differing in feature extraction methods. Classification performance of each model was assessed through accuracy, precision, and recall using confusion matrices. Results: SVM models demonstrated accuracies between 47-73% in MVM classification, with poorest performance observed on images with borderline MVM presence, as determined through manual observation. Data augmentation provided little to no improvement to the accuracies. Conclusion: The results are promising for the integration of ML methods into the placental histopathological examination process. Using this study as a proof-of-concept foundation will lead our group and others to carry ML models further in placental histopathology.

No description available.

京都大学 / 新制・課程博士 / 博士(医学) / 甲第24835号 / 医博第5003号 / 新制||医||1068(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 浅野, 雅秀, 教授 柳田, 素子, 教授 近藤, 玄 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

In human pregnancy, the feto-placental vessels are crucial for efficient materno-fetal transfer; hence they play a pivotal role in the pathogenesis of fetal growth restriction (FGR). We, as well as other research groups, have observed abnormalities in the FGR feto-placental vasculature, which, though inconclusive, were suggestive of a state of panhypovascularity. The goal of the work presented in this thesis was to investigate this. We hypothesised that the placenta may be panhypovascular in FGR due to failed angiogenesis; and enhancing angiogenesis in the placenta may improve fetal growth. Custom-designed techniques including advanced imaging, computer-aided analyses and tube-forming experiments were employed to compare feto-placental vessels and endothelial cells in placentas from normal and FGR-complicated pregnancies while aiming to answer two main research questions: (i) is the FGR placenta panhypovascular? (ii) can angiogenesis be induced or enhanced to improve placental vascularity?Findings include: (i) shorter arterial [p = 0.03 and 0.009 when data adjusted for placental surface area (PA) and weight (PW) respectively] and longer venous path [p = 0.05 and 0.03, adjusted for PA and PW respectively] in FGR placentas though no difference in the total number of arterial or venous branches, diameter, and tortuosity of the vessels compared to normal; (ii) altered angiogenic behaviour/response of FGR placental endothelial cells following in vitro pharmacological manipulation of WNT signalling; (iii) human placental endothelial cells are capable of regaining their angiogenic potential following withdrawal of WNT inhibition. These findings discount the hypothesis of panhypovascularity in FGR placentas, but identify additional, previously unreported, feto-placental vascular abnormalities associated with FGR. Also, the findings provide evidence that impairment of WNT signalling may play a role in defective angiogenesis and consequent dysvascularity in the FGR placenta. The evidence suggests the WNT pathway should be explored as a potential new target for therapeutic interventions to correct placental dysvascularity in FGR.

CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / avaliar os resultados perinatais em gestaÃÃes com centralizaÃÃo do fluxo fetal ao estudo dopplervelocimÃtrico e identificar os principais fatores prognÃsticos associados com o Ãbito neonatal. MÃtodos: estudo transversal a partir dos prontuÃrios das gestantes com diagnÃstico de centralizaÃÃo do fluxo fetal (CF), diÃstole zero (DZ) ou reversa (DR) acompanhadas no ServiÃo de Medicina Materno-Fetal da Maternidade-Escola Assis Chateaubriand â Universidade Federal do CearÃ. Foram analisadas 143 pacientes com gestaÃÃes Ãnicas, sem anomalias estruturais ou cromossÃmicas, apresentando idade gestacional superior a 22 semanas e com peso fetal igual ou acima de 500 gramas. Construiu-se curva ROC para idade gestacional e peso ao nascer (variÃveis independentes) e Ãbito neonatal (variÃvel dependente). Os resultados perinatais foram avaliados na populaÃÃo geral e em cada grupo (CF, DZ e DR), sendo posteriormente comparados entre si. Para a avaliaÃÃo estatÃstica, utilizou-se os testes: Shapiro-Wilk, Levene, t Student, Mann-Whitney, ANOVA, Kruskal Wallis, Exato de Fisher, Chi-quadrado de Pearson, RegressÃo LogÃstica e Multinomial. Todos foram considerados estatisticamente significantes quando p<0,05. Resultados: a maioria das gestantes (78,3%) apresentou algum distÃrbio hipertensivo associado à gravidez. A gestaÃÃo foi resolvida nas primeiras 24 horas apÃs o diagnÃstico dopplervelocimÃtrico na maioria dos casos (74,8%), sendo a via abdominal utilizada em 96,5% das vezes. No momento do parto, a idade gestacional mÃdia foi 33,6 semanas e o peso foi 1684g. Os recÃm-nascidos foram classificados como pequenos para idade gestacional em 69,6% e necessitaram de internamento em UTI em 63% dos casos. Os Ãndices de mortalidade perinatal para CF, DZ e DR foram, respectivamente, de 11,1, 31,1 e 70,6%. O peso do RN (Ãrea sob a curva ROC 0,934, p=0,000) e idade gestacional ao nascer (Ãrea 0,909, p=0,000) mostraram ser bons preditores de Ãbito neonatal. O ponto de corte calculado para o peso foi 1010g e para a idade gestacional foi 32,5 semanas. A incidÃncia do Ãndice de lÃquido amniÃtico (ILA) diminuÃdo nas gestaÃÃes com desfecho perinatal letal foi 41,2% e naquelas sem letalidade foi 41,3%. ConclusÃes: fetos com diagnÃstico de CF, DZ e DR apresentaram prognÃsticos progressivamente piores e estatisticamente diferentes entre si. A idade gestacional e peso ao nascer mostraram excelente correlaÃÃo com mortalidade neonatal. O ILA nÃo demonstrou associaÃÃo com taxa de letalidade. / evaluate the perinatal results in pregnancies with fetal brain sparing on the Doppler velocimetric study and identify the main prognostic factors associated with neonatal death. Methods: it is a transverse study from the charts of pregnant women with diagnosis of brain sparing, absent or reversed end-diastolic flow in the umbilical artery, followed at the Service of Maternal-Fetal Medicine of Maternidade-Escola Assis Chateaubriand â Universidade Federal do CearÃ. There were analyzed 143 patients with single pregnancies, without structural or chromosomal anomalies, presenting gestational age above 22 weeks and fetal weight equal or above 500 grams. ROC curve was constructed for gestational age and weight at birth (independent variables) and neonatal death (dependent variable). The perinatal results were evaluated on the general population and on each group (brain sparing, absent and reversed end-diastolic flow), later compared with each other. For the statistical analisys it was utilized the tests: Shapiro-Wilk, Levene, t Student, Mann-Whitney, ANOVA, Kruskal Wallis, Fisher, Chi-square, Logistical and Multinomial Regression. All were considered statistically significant when p < 0.05. Results: the majority of pregnant women (78.3%) presented some hypertensive disturb associated to the pregnancy. The pregnancy was resolved in the first 24 hours after Doppler velocimetric diagnosis on most cases (74.8%), being the abdominal access utilized in 96.5% of the times. At the moment of delivery, the average gestational age was 33.6 weeks and the weight was 1684g. The newborns were classified as small for gestational age in 69.6% and needed ICU admission in 63% of the cases. The indexes of perinatal mortality for brain sparing, absent and reversed end-diastolic flow were respectively 11.1, 31.1 and 70.6%. The weight of the newborn (area bellow the ROC curve 0.934, p=0.000) and gestational age at birth (area 0.909, p=0.000) have shown to be good predictors of neonatal death. The cutoff point calculated for the weight was 1010g and for the gestational age was 32.5 weeks. The incidence of diminished amniotic fluid index (AFI) in the pregnancies with lethal perinatal outcome was 41.2% and in those without lethality was 41.3%. Conclusions: fetuses with diagnosis of brain sparing, absent and reversed end-diastolic flow presented progressively worse and statistically different with each other prognosis. The gestational age and weight at birth showed excellent correlation with neonatal mortality. The AFI did not demonstrate association with lethality rate.

The outer surface of the human placenta, the syncytiotrophoblast, results from the fusion of many cytotrophoblast cells such that many nuclei are contained in this layer. It is possible for these nuclei to cluster forming syncytial nuclear aggregates (SNAs). SNAs have been linked to pathology with increased numbers and earlier formation of SNAs in preeclampsia and fetal growth restriction (FGR). SNAs can be grouped into subtypes including bridges, knots and sprouts, dependent on morphology and attachment to surrounding placental villi. Little is known about SNA formation, but the pyknotic appearance of nuclei within SNAs has led to development of a hypothesis that SNAs are the terminal point of nuclear turnover in the syncytiotrophoblast. Some cytoskeletal proteins have been associated with SNAs indicating their potential involvement in SNA formation. This project aimed to uncover differences between SNA subtypes, whether the degenerate nuclear morphology represents apoptosis and to understand which mechanisms drive nuclear collection into SNAs. Experimental approaches included a review of an electron micrograph archive and application of immunohistochemical techniques to ex vivo placental tissue. A long-term explant model was developed to examine SNA development in vitro; these experiments were further explored using an isolated primary cytotrophoblast model. Nuclei within SNAs were more frequently pyknotic and less frequently eukaryotic than nuclei dispersed in the syncytiotrophoblast. However, few SNAs were positive for the cytokeratin-M30 neoepitope, a caspase dependent breakdown product of cytokeratin-18 and no subtype of SNA showed greater M30 staining than general areas of syncytiotrophoblast. There were increased syncytial knots and decreased syncytial bridges in placentas from women with preeclampsia compared to controls and FGR. While cytoskeletal proteins are seen surrounding SNAs, inhibition of actin and tubulin had no effect on SNA turnover or stability. Very limited nuclear movement was recorded from in vitro culture indicating that syncytiotrophoblast nuclei move far less than had been expected. These data suggest that cell death was not prominent within SNAs but different prevalence of subtypes were present in preeclampsia indicating that SNAs might represent larger changes in placenta structure. As nuclei moved less and SNAs were more static than expected it is suggests that SNAs are more stable than previously thought. Overall, the hypothesis that SNAs are highly active in preeclampsia is questioned and new hypotheses of the role of SNAs are considered in the light of these experimental findings, including whether they form by chance and represent changes in cell turnover of the syncytiotrophoblast.