A gestação confere ao organismo materno uma série de mudanças e desafios, envolvendo especialmente seu sistema imunológico. O feto, do ponto de vista imunológico, é comparado a um enxerto semi-alogenêico, pois expressa moléculas do complexo de histocompatibilidade principal (MHC) paterno, considerado não próprio ao organismo materno, capaz de desencadear uma resposta imunológica no ambiente intrauterino. Porém, durante todo o período gestacional, o organismo materno reconhece o embrião sem uma resposta imunológica contra sua permanência no útero, estabelecendo-se uma tolerância materno-fetal. Vários mecanismos contribuem para esse estado tolerogênico, como a atividade da enzima indoleamina 2,3-dioxigenase (IDO). A IDO promove o catabolismo do aminoácido triptofano, levando as células T à apoptose, devido à carência deste aminoácido e pela ação de seus catabólitos no micro ambiente placentário. Diversos tipos celulares estão presentes na interface materno fetal e vários deles podem potencialmente expressar a IDO. Em ratas Wistar, sabe-se que diversas proteínas, principalmente o interferon γ e, recentemente, a progesterona, podem aumentar a expressão desta enzima; contudo, ainda não são conhecidos quais tipos celulares são efetivamente influenciados por estas moléculas. Desta forma, este trabalho buscou suprir esta lacuna, por meio da identificação das células IDO positivas pela imunofenotipagem e citometria de fluxo. De acordo com as analises realizadas, os resultados deste trabalho mostraram que todos os tipos celulares fenotipados foram capazes de expressar a enzima. Contudo somente alguns grupos específicos de células presentes no ambiente uterino placentário sofrem influência das proteínas supracitadas, principalmente as células dendríticas, e os linfócitos CD4 que elevam a expressão de IDO na presença de progesterona e IFN-γ. Tais achados sugerem que a ação da enzima em grupos celulares específicos poderiam constituir meios pelo qual o organismo regula o sistema imunológico no ambiente uterino-placentário, em que é capaz de impedir o desenvolvimento de células imunológicas potencialmente ativas, colaborando com a formação de um ambiente tolerogênico favorável para o desenvolvimento embrionário. / Pregnancy causes several changes and challenges to the maternal body, especially involving the immune system. The fetus, on the immunological point of view, is compared to a semialogeneic graft, expressed as molecules of the paternal major histocompatibility complex (MHC) considered not self to the maternal organism, capable of triggering an immune response in the intrauterine environment. However, throughout the gestational period, maternal organism tolerates the embryo without an immune response against its permanence in the uterus, establishing maternal-fetal tolerance. Several mechanisms contribute to this tolerogenic state, as the activity of the enzyme indoleamine 2,3-dioxygenase (IDO). The IDO promotes the catabolism of tryptophan, inducing T cells apoptosis due to the deprivation of this amino acid and by the action of its catabolites in the placental microenvironment. Several cell types are present in the maternal-fetal interface and many of them can potentially express IDO. In Wistar rats, it is known that interferon γ and recently, progesterone, may increase the expression of this enzyme; however, which cell types are actually influenced by these molecules is still unknown. Thus, this study intends to fill this gap, by identifying IDO positive cells by immunophenotyping and flow cytometry. According to the analysis carried out, the results of this study showed that all phenotyped cell types were able to express the enzyme, however only some specific groups present in placental uterine environment are influenced by the factors above mentioned, especially dendritic cells and lymphocytes CD4 increase the IDO expression at the presence of progesterone and IFN-γ. These findings suggest that the action of the enzyme in specific cell groups may be one of the means by which the body regulates the immune system uterine-placental environment, which is able to prevent the development of potentially active immune cells, contributing to the formation tolerogenic environment favorable for embryonic development.

A poluição atmosférica de centros urbanizados e industrializados é associada a diversos efeitos nocivos, incluindo alterações na fertilidade masculina. A toxicidade da poluição atmosférica da cidade de São Paulo já foi demonstrada em estudos experimentais e especula-se que seus efeitos sejam mediados por estresse oxidativo, porém, os mecanismos envolvidos ainda não são totalmente conhecidos. O objetivo deste estudo foi avaliar os efeitos da exposição à poluição atmosférica da cidade de São Paulo sobre os testículos de camundongos. Para isso, os animais foram expostos à fração da poluição atmosférica mais implicada em danos à saúde, o material particulado inalável fino, que é uma complexa mistura de substâncias com 2,5 ?m ou menos de diâmetro aerodinâmico (MP2,5). Um grupo de camundongos foi exposto ao MP2,5 durante o período gestacional e após o nascimento, no período entre o pós-desmame e a idade adulta (grupo pré-natal e pós-desmame - GPNPD); um segundo grupo de camundongos foi exposto ao MP2,5 apenas durante o período gestacional (grupo pré-natal - GPN); o terceiro grupo foi exposto ao MP2,5 apenas após o nascimento, depois do desmame até a idade adulta (grupo pós-desmame - GPD); e o quarto grupo foi o controle (GC), formado por camundongos que foram expostos ao ar filtrado, tanto durante o período gestacional quanto após o nascimento até a idade adulta. Os efeitos do MP2,5 foram avaliados nos testículos de animais adultos, com 90 dias de vida. Técnicas estereológicas foram aplicadas para analisar aspectos estruturais do testículo e o escore de Johnsen foi utilizado para avaliar qualitativamente a espermatogênese. A expressão gênica foi realizada pela técnica de microarranjos de DNA. Na comparação com o GC, os animais do GPNPD apresentaram peso e volume dos testículos, área de superfície dos túbulos seminíferos, volume do epitélio seminífero e peso dos epidídimos maiores, bem como menor qualidade da espermatogênese, com p < 0,05 em todos esses parâmetros. O GPD teve comportamento semelhante, porém, apenas o volume do epitélio seminífero e a qualidade da espermatogênese diferiram significativamente do GC. O volume do epitélio seminífero do GPN foi menor que o do GC e esse foi o único parâmetro com diferença estatística em relação ao GC. A análise de expressão gênica evidenciou alteração de expressão de genes envolvidos na espermatogênese e na esteroidogênese, além de vários outros genes envolvidos nas vias de resposta imune, estresse oxidativo, dano de DNA e apoptose. Os resultados deste estudo mostraram que a exposição ao MP2,5 da cidade de São Paulo foi capaz de causar alterações nas funções testiculares de camundongos / Air pollution in industrial and urban centers is related to negative effects in living organisms, including alterations in male fertility. Experimental studies had already showed toxicity aspects related to the air pollution of the city of Sao Paulo. The hypothesis surrounding its toxicity is based on oxidative stress, but the mechanisms are not yet completed understood. The objective of this study was to evaluate the effects of the exposure to air pollution of the city of Sao Paulo on mice testes. To do so, animals were exposed to the most deleterious fraction of the air pollution, the fine inhalable particulate matter, which consists of a complex mixture of particles sized 2,5 ?m or less of aerodynamic diameter (PM2,5). A group of mice was exposed to PM2,5 during gestational period and after birth, from the weaning day until adulthood (pré-natal and post-weaning group - PNPWG); another group was exposed to PM2,5 during gestational period only (pré-natal group - PNG); a third group was exposed to PM2,5 after birth, from the weaning day until adulthood only (post-weaning group - PWG); and finally, a fourth group of mice was exposed to filtered air during gestational period and from post-weaning day until adulthood (control group - CG). The analyses were performed on testes from adult animals. Stereological techniques were used to analyze structures of the testes and Johnsen\'s score to evaluate spermatogenesis in a qualitative way. DNA microarray were used to evaluate gene expression. Significant differences (p < 0,05) between the PNPWG and the CG were found for testis weight and volume, surface area of the seminiferous tubules, volume of the seminiferous epithelium and epididymis weight. For all these parameters the mean values of the PNPWG were higher. The quality of spermatogenesis in the PNPWG was also significant differently from CG, showing a worse quality of spermatogenesis. The PWG had similar results, however the only parameters with significant difference from the CG were the volume of the seminiferous epithelium and the quality of spermatogenesis. The volume of the seminiferous epithelium from the PNG showed significant lower mean compared to the CG. The gene expression analysis showed differential expression of genes related to spermatogenesis and steroidogenesis. Also, altered expression was found for genes related to oxidative stress, immune response, DNA damage and apoptosis ways. The results of the present study showed that the exposition to the PM2,5 of the city of Sao Paulo was capable of elicit testes alterations in mice

O presente estudo analisou a hipótese de que as alterações hemodinâmicas fetais estão associadas à disfunção miocárdica fetal em gestações complicadas pela insuficiência placentária. Os objetivos do estudo foram correlacionar os valores do peptídeo natriurético cardíaco do tipo B (BNP) em sangue de cordão umbilical no nascimento com os parâmetros da dopplervelocimetria fetal, bem como com o pH no nascimento. Métodos: estudo prospectivo e transversal, com os seguintes critérios de inclusão: mulheres com gestação com feto único e vivo, insuficiência placentária caracterizada pelo aumento do índice de pulsatilidade (IP) da artéria umbilical (AU), membranas íntegras e ausência de anomalias fetais. Foram excluídos da pesquisa os casos com diagnóstico pós-natal de anomalia do recém-nascido e aqueles nos quais não foi realizada a análise do sangue no nascimento. Os seguintes parâmetros da dopplervelocimetria foram analisados: índice de pulsatilidade (IP) da artéria umbilical (AU) e da artéria cerebral média (ACM), relação cerebroplacentária (RCP) e IP para veias (IPV) do ducto venoso (DV). Os parâmetros de Doppler foram transformados em escore zeta. Amostras de sangue do cordão umbilical foram obtidas imediatamente após o parto para a mensuração do pH de artéria umbilical e do BNP. Resultados: Foram incluídas 32 gestações com diagnóstico de insuficiência placentária 21 (65%) com fluxo diastólico positivo e 11 (35%) com diástole zero ou reversa nas AU. A concentração do BNP apresentou correlação significativa com: escore zeta do IP da AU (rho=0,43; P=0,016); escore zeta da RCP (rho=-0,35; P=0,048); escore zeta do IPV do DV (rho=0,61; P<0,001), pH no nascimento (rho=- 0,39; P=0,031) e idade gestacional (rho=-0,51; P=0,003). Na regressão múltipla foram incluídos os parâmetros antenatais e o escore zeta do IPV do DV (P=0,008) demonstrou ser o único fator independente correlacionado com o BNP no nascimento. A correlação entre o BNP e o escore zeta do IPV do DV é representado pela equação de regressão Log[BNP]=2,34+0,13*DV (F=18,8, P<0,001). A correlação entre o BNP e o pH no nascimento é representado pela equação de regressão Log[BNP]=21,36-2,62*pH (F=7,69, P=0,01). Conclusão: os resultados sugerem que a disfunção cardíaca fetal identificada pelas concentrações de BNP no nascimento correlaciona-se de forma independente com as alterações no IPV do DV, além de correlacionarem-se negativamente com os valores do pH no nascimento / This study examined the hypothesis that fetal hemodynamic changes are associated with fetal myocardial dysfunction in pregnancies complicated by placental insufficiency. The objective was to study the correlation between the concentrations of cardiac natriuretic peptide type B (BNP) in umbilical cord blood at birth and fetal Doppler parameters, as well as the pH at birth. Methods: A prospective crosssectional study with the following inclusion criteria: pregnant women with a single fetus, placental insufficiency characterized by increased pulsatility index (PI) of the umbilical artery (UA), intact membranes and absence of fetal abnormalities. There were excluded from the study cases with postnatal diagnosis of abnormality of the newborn and those in which the blood analysis was not performed. The following Doppler parameters were analyzed: UA PI, middle cerebral artery (MCA) PI, cerebroplacental ratio (CPR) and ductus venosus (DV) PI for veins (PIV). The Doppler parameters were converted into zeta score. Blood samples were obtained from the umbilical cord immediately after delivery to measure the pH of the umbilical artery and the BNP. Results: Thirty two pregnancies with placental insufficiency were included, 21 (65%) with positive diastolic flow and 11 (35%) with absent or reversed end diastolic flow in the UA. The concentration of BNP correlated significantly with: UA-PI z-score (rho = 0.43, P = 0.016); CPR z-score (rho = -0.35, P = 0.048); DV-PIV zscore (rho = 0.61, P <0.001), pH at birth (rho = -0.39, P = 0.031) and gestational age (rho = -0.51, P = 0.003). In the multiple regression analysis, antenatal parameters were included and DV-PIV z-score (P = 0.008) was found to be independent parameter correlated with BNP at birth. The correlation between BNP and DV-PIV zscore is represented by the regression equation Log [BNP] = 2.34+0.13*DV (F=18.8, P <0.001). The correlation between BNP and pH at birth is represented by the regression equation Log [BNP] = 21.36-2.62*pH (F=7.69, P = 0.01). Conclusion: The results suggest that fetal cardiac dysfunction identified by BNP concentrations at birth correlated independently with changes in the DV PIV, and correlated negatively with pH values at birth

O objetivo deste estudo foi avaliar as condições de saúde bucal de crianças com desnutrição intra-uterina, acompanhadas no Ambulatório de Nutrição e Puericultura do Departamento de Pediatria da Faculdade de Medicina da Universidade de Santo Amaro (UNISA). Essa instituição adota o padrão ouro da alimentação, crescimento e desenvolvimento, através do incentivo ao aleitamento materno exclusivo até os 4-6 meses de idade e segue as normas aprovadas pelo Departamento de Nutrição da Sociedade Brasileira de Pediatria quanto à suplementação mineral e da vitamina D. A amostra foi constituída de 28 crianças na faixa etária de 6 à 60 meses, nascidas a termo, com mais de 37 semanas, sendo classificadas de acordo com o peso de nascimento como Normais, com peso maior que 3000g, e Pequenas para a Idade Gestacional (PIG), com peso menor que 2500g. Em relação à saúde bucal, foram avaliados os seguintes fatores: visita ao cirurgião dentista, orientação de higiene bucal, limpeza dos dentes, uso de fio dental, placa bacteriana visível na superfície vestibular dos incisivos decíduos superiores, uso de mamadeira, hábitos de sucção (chupeta ou dedo), prevalência de cárie e defeitos de desenvolvimento do esmalte (DDE). Também foi avaliada a condição nutricional (NCHS) na data do exame odontológico. Não houve diferença estatística significante quanto ao tempo médio de aleitamento, que foi em torno de nove meses para ambos os grupos (teste de Mann-Whitney, Z=0,02 / Z crítico=1,96). As crianças Normais e as PIG não apresentaram diferenças estatisticamente significantes quando se consideram as possíveis alterações dos fatores avaliados, com exceção do uso do fio dental, o qual foi significante nas crianças PIG (teste Exato de Fisher p=0,0126). Através do coeficiente de correlação de Spearman verificou-se que tanto para o grupo PIG quanto para o Normal, os três primeiros fatores favoráveis à saúde bucal foram: limpeza dos dentes, ausência de placa visível e orientação sobre higiene bucal. Concluiu-se, que apesar da condição nutricional de nascimento, as crianças PIG não mostraram diferença quanto à saúde bucal, o que pode ser explicado devido às ações educativas voltadas para prevenção e promoção de saúde global que são ministradas nos Ambulatórios Materno-Infantil e de Puericultura da UNISA, com a orientação da Disciplina de Odontopediatria da Faculdade de Odontologia da mesma Entidade. / This study’s objective was value the oral conditions of children with intra-uterine under nutrition, accompanied at the Nutrition an childcare outpatient department of the Pediatric department of Medicine college of Santo Amaro ‘ s University (UNISA). This institution adopt the old Standard of feeding, growth and development, by the breast feeding incentive, exclusive until 4-6 mouths of age and follows the norms approved by the Nutrition Department of the Brazilian ‘ s Pediatric Society that refers to the mineral supplement and supplement of D vitamin. The sample was constituted by 28 children aged between 6 and 60 months, born at the right moment, with more than 37 weeks, being classified according with the weigh of birth as Normal, weighting over 3000g, and small to their gestational age (SGA), weighting less than 2500g. About the oral health, the following factors were valued: visit to a dentist surgeon, oral hygiene orientation, teeth cleanliness, usage of dental floss, visible bacterial plaque at the vestibular surface of the superior incisors, usage of feeding bottle, suck habits (dummy or finger), prevalence of caries and development defects of the enamel (DDE). The nutrition condition was valued (NCHS) at the dental exam. There was no significant difference at the breast feeding time that was about 9 mouths for both groups(Mann-Whitney test, Z+0,02 / critics Z =1,96). The normal children and the ones SGA did not present significant differences when took into account the possible alteration of the valued factors, except of the usage of dental floss, that was significant at children that was small for the gestational age (Exact Fisher test p=0,0126). By the Spearman ‘ s correlation’s coefficient was observed that to both groups, the three more important factors to the oral health were: teeth cleanliness, no visible plaque and oral hygiene orientation. The conclusion was that in spite of the nutritional condition of birth, the children that were small for the gestational age, did not show differences in the oral health, what can be explained by the educational actions done to prevention and promotion of global health that are learned at the Motherly-Infant and child care outpatient department of UNISA, with orientation of Pediatric dentistry of Dental College discipline. Key-words: intra-uterine growth retardation – fetal undernutrition – breast feeding – oral health children

幹細胞療法是近年的研究熱點之一，然而幹細胞在組織修復中的實際應用受到移植後幹細胞存活率低的制約，約80% 的幹細胞在移植至組織後不能存活。 人類臍帶血管周皮 (HUCPV) 幹細胞為多功能間充質幹細胞移植提供豐富的細胞來源。 在合適的誘導環境下，它們具有向多種間充質細胞系分化的能力。 與從骨髓或臍帶血中提取的間充質幹細胞比較，人類臍帶血管周皮幹細胞的體外增殖更為容易。 在本研究中，我們從人類臍帶血管周圍組織中分離人類臍帶血管周皮幹細胞，並採用流式細胞技術分選細胞表面標記物CD34、CD45呈陰性同時CD44 、CD90、 CD105、 CD146呈陽性的HUCPV細胞。HUCPV細胞在體外培養以及三維支架的環境下具有分化為骨和軟骨的能力。 / 在本研究中，我們主要研究腦和生殖器官表達基因（BRE）在HUCPV細胞中的功能。 BRE蛋白與其他已知蛋白的同源性均不高，目前尚未鑑定出任何功能性的結構域。 至今為止，BRE基因的已知功能大多數是通過對腫瘤模型的研究發現的。 據報導，BRE能夠提高DNA損傷的腫瘤細胞的存活率，但BRE在幹細胞中的作用仍不清楚。 我們發現，當HUCPV細胞分化後，其BRE的表達水平降低。 此外，利用BRE-siRNA降低HUCPV細胞中BRE基因的表達，能夠促進HUCPV細胞向骨和軟骨分化的進程。 因此，我們假設BRE對維持HUCPV細胞的幹細胞功能具有重要的作用。 由於經過BRE基因沉默處理的HUCPV細胞與對照組相比並無顯著的表型差別，我們採用微陣列（microarray）以及比較蛋白組學的方法研究兩者間的區別，從而找出BRE基因的功能以及可能涉及BRE的信號通路。 / 通過微陣列技術，我們深入地分析了BRE基因表達沉默後HUCPV細胞的轉錄組。 在經過BRE基因沉默處理的HUCPV細胞中，我們發現與維持幹細胞多向分化潛能有關的OCT4、 FGF5和FOXO1A等基因的表達顯著下調。 另外，BRE基因的沉默能夠影響表觀遺傳調控基因以及TGF-β 信號通路組成部件的表達，而TGF -β 信號通路是維持幹細胞自我更新的重要通路。 這些結果提示，BRE作為一個重要的調控因子，在維持HUCPV細胞的多向分化潛能的同時能夠防止細胞分化。 / 在比較蛋白組學的研究中，我們發現BRE基因的沉默能夠降低細胞骨架結合蛋白的表達，例如actin, annexin II 及 tropomyosin。 此外，我們利用免疫共沉澱的方法證明了BRE蛋白與actin及 annexin II蛋白直接結合。 細胞骨架的改變可能為HUCPV細胞的分化提供了一個有利的環境，因而BRE基因的沉默能夠促進HUCPV細胞向骨和軟骨分化。 支持這一推論的其中一個依據是Lim et al., 2000; Solursh, 1989; Zhang et al., 2006，文獻報導肌動蛋白多聚化抑製劑能夠促進軟骨形成的過程。 綜上所述，本研究為進一步研究BRE基因在HUCPV細胞中的功能以及與BRE直接作用的蛋白打下了基礎。 / Stem cells therapy has gained considerable attention in recent years. However, the practical use of stem cells for tissue repair has been hindered due to their low survival rate after grafting into tissues, for approximately 80% of the stem cells died after implantation. Human umbilical cord perivascular (HUCPV) stem cells offer a new and rich resource of multipotent mesenchymal stem cells. These cells possess the ability to differentiate into various mesenchymal cell lineages when induced. HUCPV cells can be more easily amplified in culture than mesenchymal stem cells extracted from bone marrow or umbilical cord blood. In this study, HUCPV cells were isolated from the perivascular regions of human umbilical cords. The HUCPV cells were sorted using flow cytometer for CD34⁻, CD44⁺, CD45⁻, CD90⁺, CD105⁺ and CD146⁺ surface markers. These HUCPV cells were found to be capable of differentiating into osteogenic lineage in monolayer culture and chondrogenic lineage in pellet culture. These cells were also found to be capable of differentiating into osteogenic and chondrogenic lineage in silk fibroin which acted as three-dimensional scaffolds for the cells to grow on. / The function of the Brain and Reproductive Organ-Expressed (BRE) gene in the context of HUCPV cells was investigated. The BRE protein shares no homology with any other known gene products and contains no known functional domain. To date, most of what we know about the function of this gene has been conducted in the tumor model. It has been reported that BRE can enhance the cellular survival of cancer cells following DNA damage. The role of BRE in stem cells has never been examined. We have established that BRE expression was down-regulated when HUCPV cells started to differentiate. In addition, silencing BRE expression, using BRE-siRNA, in HUCPV cells could accelerate osteogenic and chondrogenic differentiation. Hence, we hypothesized that BRE played an important role in maintaining the stemness of HUCPV cells. Because there was a lack of phenotypic difference between the BRE-silenced HUCPV cells and cells transfected with the control-siRNA, we decided to profile these cells using microarray and proteomic analyses. The aim was to elucidate the function of the BRE gene and establish whether BRE was involved in any signaling pathways. / In the microarray analysis, we examined the transcriptome of HUCPV cells in response to BRE-silencing in depth. Amongst the genes that we identified were significantly down-regulated by BRE-silencing and involved in the maintenance of pluripotency in ES cells were OCT4, FGF5 and FOXO1A. BRE-silencing also altered the expression of epigenetic genes and also components of the TGF-β signaling pathway. This pathway is crucially involved in maintaining stem cell self-renewal. Therefore, we propose that BRE acts like a modulator that promotes stemness and at the same time inhibits the differentiation of HUCPV cells. / In the comparative proteomic study, BRE-silencing resulted in decreased expression patterns of cytoskeletal binding proteins such as actin, annexin II and tropomyosin. In addition, co-immunoprecipitation experiments revealed that the BRE protein can bind directly with actin and annexin II. It is possible that altering the cytoskeleton may provide a favorable environment for HUCPV cells to differentiate. This may explain why we were able to accelerate osteogenic and chondrogenic differentiation following BRE-silencing. In support of the view, it has been reported that chondrogenesis could be enhanced after cells have been treated with actin polymerization inhibitors (Lim et al., 2000; Solursh, 1989; Zhang et al., 2006). In sum, our studies provide an insight into the function of the BRE gene in HUCPV cells and the proteins that BRE can directly act on. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Chen, Elve. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 135-159). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese. / Thesis/Assessment Committee --- p.i / Abstract --- p.ii / 摘要 --- p.v / Acknowledgements --- p.viii / List of Figures --- p.ix / List of Tables --- p.xiii / Table of Abbreviations --- p.xiv / Contents --- p.xviii / Chapter 1 --- p.1 / Literature Review --- p.1 / Chapter 1.1 --- Stem cells --- p.1 / Chapter 1.2 --- Embryonic stem cells (ESCs) --- p.2 / Chapter 1.3 --- Epiblast-derived stem (EpiS) cells --- p.2 / Chapter 1.4 --- Somatic stem cells (SSCs) --- p.3 / Chapter 1.5 --- Induced pluripotent stem (iPS) cells --- p.5 / Chapter 1.6 --- Human umbilical cord perivascular (HUCPV) cells --- p.7 / Chapter 1.7 --- CD146 --- p.8 / Chapter 1.8 --- Stem cell senescence --- p.9 / Chapter 1.9 --- Brain and reproductive organ-expressed (BRE) protein --- p.12 / Chapter 1.10 --- Stem cell self-renewal --- p.14 / Chapter 1.11 --- Apoptosis --- p.16 / Chapter 1.12 --- Stem cell niche --- p.21 / Chapter 1.13 --- Stem cell homing --- p.22 / Chapter 1.14 --- Objective --- p.22 / Chapter 2 --- p.24 / Accelerated osteogenic and chondrogenic differentiation of HUCPV cells by modulating the expression of BRE --- p.24 / Chapter 2.1 --- Introduction --- p.24 / Chapter 2.2 --- Rationale --- p.27 / Chapter 2.3 --- Materials and Methods --- p.27 / Chapter 2.3.1 --- Extraction of HUCPV cells from umbilical cord --- p.27 / Chapter 2.3.2 --- Cell culture condition --- p.28 / Chapter 2.3.3 --- Flow cytometry analysis and cell sorting --- p.28 / Chapter 2.3.4 --- In vitro osteogenic differentiation --- p.29 / Chapter 2.3.5 --- In vitro chondrogenic differentiation --- p.29 / Chapter 2.3.6 --- Alcian blue staining --- p.29 / Chapter 2.3.7 --- Alizarin red S staining --- p.30 / Chapter 2.3.8 --- Immunofluorescence analysis --- p.30 / Chapter 2.3.9 --- Quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR) --- p.31 / Chapter 2.3.10 --- Transfection with siRNA --- p.35 / Chapter 2.3.11 --- Microarray --- p.35 / Chapter 2.3.12 --- Cell lysis and immunoprecipitation --- p.36 / Chapter 2.3.13 --- SDS-PAGE and Western blot --- p.36 / Chapter 2.3.14 --- Isoelectric focusing and 2-dimensional gel electrophoresis --- p.37 / Chapter 2.3.15 --- Migration (wound healing) assay --- p.38 / Chapter 2.4 --- Results --- p.38 / Chapter 2.4.1 --- HUCPV cells were capable to differentiate into osteoblasts and chondrocytes --- p.38 / Chapter 2.4.2 --- BRE expression is down-regulated when HUCPV cells begins to differentiate --- p.40 / Chapter 2.4.3 --- Silencing of BRE expression accelerates induction of osteogenesis and chondrogenesis --- p.40 / Chapter 2.4.4 --- Microarray analysis of BRE-silenced HUCPV cells --- p.42 / Chapter 2.4.4.1 --- Stemness factors --- p.43 / Chapter 2.4.4.2 --- Epigenetic regulation --- p.43 / Chapter 2.4.4.3 --- Signaling pathways crucial for stemness maintenance --- p.44 / Chapter 2.4.4.4 --- TGF-β signaling --- p.44 / Chapter 2.4.4.5 --- FGF signaling --- p.44 / Chapter 2.4.4.6 --- NOTCH signaling --- p.45 / Chapter 2.4.4.7 --- WNT signaling --- p.46 / Chapter 2.4.4.8 --- Homeobox transcription factors (HOX) --- p.46 / Chapter 2.4.4.9 --- Cell cycle regulation --- p.47 / Chapter 2.4.4.10 --- Chemokines and cytokines regulation --- p.48 / Chapter 2.4.4.11 --- Apoptosis --- p.49 / Chapter 2.4.5 --- BRE-silencing alters the cellular proteome of HUCPV cells --- p.50 / Chapter 2.4.5.1 --- BRE-silencing alters the cytoskeletal binding proteins of HUCPV cells --- p.51 / Chapter 2.4.5.2 --- BRE-silencing alters the expressions of stemness-related proteins in HUCPV cells --- p.52 / Chapter 2.4.5.3 --- BRE-silencing alters the expressions of apoptosis-related proteins in HUCPV cells --- p.53 / Chapter 2.5 --- Discussion --- p.86 / Chapter 2.5.1 --- Microarray study discussion --- p.87 / Chapter 2.5.2 --- Proteomic study discussion --- p.89 / Chapter 3 --- p.93 / Replicative senescence alters the transcriptome and proteome of HUCPV cells --- p.93 / Chapter 3.1 --- Introduction --- p.93 / Chapter 3.2 --- Materials and methods --- p.93 / Chapter 3.3 --- Results --- p.93 / Chapter 3.3.1 --- Microarray analysis of aged HUCPV cells --- p.94 / Chapter 3.3.1.1 --- Stemness factors --- p.95 / Chapter 3.3.1.2 --- Epigenetic regulation --- p.96 / Chapter 3.3.1.3 --- Senescence associated markers --- p.96 / Chapter 3.3.1.4 --- Chemokines and cytokines regulation --- p.97 / Chapter 3.3.1.5 --- Matrix metalloproteinases regulation --- p.97 / Chapter 3.3.1.6 --- WNT signaling --- p.98 / Chapter 3.3.1.7 --- Toll-like receptor signaling pathway --- p.98 / Chapter 3.3.2 --- Proteomic profiling of aged HUCPV cells --- p.98 / Chapter 3.4 --- Discussion --- p.117 / Chapter 3.4.1 --- Aging alters the transcriptome of HUCPV cells --- p.117 / Chapter 3.4.2 --- Aging alters the proteome of HUCPV cells --- p.118 / Chapter 4 --- p.121 / Osteogenic and chondrogenic differentiation capacities of HUCPV cells in silk fibroin scaffold --- p.121 / Chapter 4.1 --- Introduction --- p.121 / Chapter 4.2 --- Materials and methods --- p.121 / Chapter 4.2.1 --- Extraction of silk fibroin --- p.121 / Chapter 4.2.2 --- Fabrication of porous silk fibroin scaffold --- p.122 / Chapter 4.2.3 --- Scanning electron microscopy --- p.123 / Chapter 4.2.4 --- Cell culture --- p.123 / Chapter 4.3 --- Results --- p.124 / Chapter 4.4 --- Discussion --- p.132 / Chapter 5 --- p.133 / Conclusions --- p.133 / References --- p.135

<p>Intrauterusna restrikcija rasta (IUGR) se odnosi na stanje u kojem fetus nije u mogućnosti da ostvari svoj genetski potencijal za rast. IUGR je ozbiljan klinički problem i nedavno je povezan sa bolestima odraslog doba kao &scaron;to su hipertenzija, insulin nezavistan diabetes melitus, dislipidemije i ishemijske bolesti srca. Eritropoetin je glavni regulator proliferacije i diferencijacije eritroidnih progenitorskih ćelija zahvaljujući svojoj antiapoptotičkoj aktivnosti. Cilj istraživanja je bio da se ispita uticaj IUGR na bubrege, kao i uticaj eritropotina na bubrege sa IUGR. Eksperimentalna studija je sprovedena u gajili&scaron;tu Pasterovog zavoda u Novom Sadu na 60 mi&scaron;eva rase NMRI. IUGR je izazivana aplikacijom deksametazona gravidnim ženkama. Po rođenju su mladunci bili podeljeni u sedam grupa. Mladuncima je u 1. i 7. danu života davan darbepoetin alfa (DA) u dozama od 1, 4 i 10&mu;g/kg. Dve grupe su predstavljale potomke majki koje su tokom trudnoće dobile DA. Nakon 4 nedelje su uzimani uzorci bubrega i vr&scaron;ena je morfolo&scaron;ka i stereolo&scaron;ka analiza glomerula. Aplikacija deksametazona (100 &mu;g/kg) trudnim mi&scaron;icama dovodi do potomstva sa IUGR. Primena DA kod novorođenih mi&scaron;eva sa IUGR dovodi do bržeg porasta telesne mase u prvih 7 dana života (&bdquo;catch-up― rasta). Mi&scaron;evi rođeni sa IUGR imaju manju povr&scaron;inu glomerula bubrega. Primena DA nakon rođenja i u 7. danu života (4 i 10 &mu;g/kg) kod novorođenih mi&scaron;eva sa IUGR dovodi do hipertrofije glomerula bubrega. IUGR nema uticaja na broj glomerula bubrega mi&scaron;eva. Primena DA nema uticaja na broj glomerula bubrega mi&scaron;eva. Mi&scaron;evi rođeni sa IUGR imaju manju debljinu korteksa bubrega. Primena DA (4 i 10 &mu;g/kg) kod mi&scaron;eva rođenih sa IUGR dovodi do povećanja debljine korteksa bubrega. Davanje DA kod IUGR značajno povećava povr&scaron;inu glomerula i debljinu korteksa bubrega.</p> / <p>Intrauterine growth restriction (IUGR) refers to a condition in which a foetus is not able to achieve its genetic potential for growth. IUGR is a serious clinical problem, and has recently been linked with diseases of adulthood, such as hypertension, insulin-independent diabetes mellitus, dyslipidemia, and ischemic heart disease. Erythropoietin is the major regulator of proliferation and differentiation of erythroid progenitor cells, thanks to its anti-apoptotic activities. The aim of this study was to investigate the effect of IUGR on the kidneys, and the impact of erythropoietin on the kidneys with IUGR. The experimental study was conducted in Pasteur Institute of Novi Sad on 60 mice of NMRI race. IUGR has been imposed with the application of dexamethasone to pregnant females. After birth, the pups were divided into seven groups. DA was administered to the pups on the 1st and 7th day of life (dose 1, 4 and 10 &mu;g/kg). Two groups represented the offspring of the mothers who during pregnancy received DA. After 4 weeks, kidney samples were taken and morphological and stereological analysis of the glomeruli was performed. The application of dexamethasone (100 &mu;g/kg) to pregnant mice leads to their offspring with IUGR. Application of DA to newborn mice with IUGR leads to faster weight gain in the first 7 days of life (&quot;catch-up&quot; growth). Mice born with IUGR have a reduced glomerular surface. Application of DA after birth and on the 7th day of life (4 and 10 &mu;g/kg) in mice with IUGR leads to hypertrophy of the kidney glomeruli. IUGR has no effect on the number of kidney glomeruli. Application of DA has no effect on the number of kidney glomeruli. Mice born with IUGR have a reduced cortical thickness. Application of DA (4 and 10 &mu;g/kg) in mice born with IUGR leads to increased thickness of the kidney cortex. Application of DA to mice with IUGR significantly increases the surface area of the kidney glomeruli and cortical thickness.</p>

Submitted by Neusa Fagundes (neusa.fagundes@unioeste.br) on 2019-03-29T16:56:55Z No. of bitstreams: 2 Camila_Kuhn2018.pdf: 2657327 bytes, checksum: 9eb518e6dcb5d576b52f1606ea47d2fa (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2019-03-29T16:57:16Z (GMT). No. of bitstreams: 2 Camila_Kuhn2018.pdf: 2657327 bytes, checksum: 9eb518e6dcb5d576b52f1606ea47d2fa (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2018-04-03 / Studies show that maternal obesity can affect fetal development, resulting in diseases in adult life, such as diabetes mellitus type 2, cardiovascular disease, obesity itslef. To reduce the effects of obesity and its comorbidities, bariatric surgeries stand out among the most effective interventions, with Roux-en-Y gastric bypass (RYGB) being the most frequently performed type of bariatric procedure. However, there are still few studies in the literature that evaluate the effects of obesity and bariatric surgeries on the morphology of skeletal muscle tissue in adult offspring. Thus, the objective of this study was to evaluate microscopic parameters of muscle fibers and neuromuscular junctions (NMJs) of the extensor digitorum longus muscle (EDL) in obese rats’ offspring submitted or not to RYGB surgery. Three-week Wistar rats were randomly divided into three groups: Control Group (CTL) which received a standard diet; 2) Cafeteria False Operated (CAF FO) and 3) Cafeteria RYGB (CAF RYGB), the latter two received a cafeteria diet before and after the surgical procedure until the weaning of the offspring. In the 18th of life, the surgical procedure and false operation were performed in the CAF RYGB and CAF FO groups, respectively. The mating of the animals occurred five weeks after the surgical procedure. The birth of the offspring was postnatal day 0, and weaning occurred at three weeks of age, and only the male offspring were separated for the experiment. The offspring of the first generation (F1) were named CTL-F1, OB-F1, CAF FO-F1 and CAF RYGB and received standard diet. At 17 weeks the animals were euthanized and the EDL muscle collected for analysis of fiber muscles and NMJs. When the CTL-F1 and OB-F1 groups were analyzed, the latter had an increase in body weight, retroperitoneal and periepididymal fats, and capillary/fiber ratio. Reduction in the number of nuclei, conjunctive and morphological changes in the parameters evaluated in the ultrastructure. The area and larger diameter of NMJs also showed reduction. The analysis between CAF RYGB-F1 and CAF FO-F1 groups showed reduction of body weight, ELD muscle weight, retroperitoneal and periepididimal fat, nasoanal length, fiber area and nuclei/fiber ratio in the CAF RYGB-F1 group. This group also presented increase in the number of fibers of type I and IIa and number of capillaries, as well as reduction in the area of the NMJs and morphological alterations in the ultrastructure. These results demonstrate that both obesity and bariatric surgery expose the offspring, through metabolic programming, to effects on the morphology of skeletal muscle tissue, being found greater aggravations in the muscular fiber of the offspring of mothers submitted to RYGB. / Estudos apontam que a obesidade materna pode afetar o desenvolvimento fetal, resultando em doenças na vida adulta, tais como diabetes mellitus tipo 2, doenças cardiovasculares e a própria obesidade. Para reduzir os efeitos da obesidade e as suas comorbidades, as cirurgias bariátricas destacam-se entre as intervenções mais eficazes, sendo a derivação gástrica em Y de Roux (DGYR) o tipo de procedimento bariátrico mais frequentemente realizado. No entanto, ainda são escassos na literatura estudos que avaliem os efeitos da obesidade e das cirurgias bariátricas na morfologia do tecido muscular esquelético da prole adulta. Diante disso, o objetivo deste estudo foi avaliar a morfologia e a morfometria das fibras musculares e as junções neuromusculares (JNMs) do músculo extensor longo dos dedos (ELD) da prole de ratas obesas submetidas ou não à cirurgia de DGYR. Para tanto, ratas Wistar de três semanas de vida foram separadas aleatoriamente em três grupos: 1) Controle (CTL), que recebeu dieta padrão; 2) Cafeteria Falso operado (CAF FO) e 3) Cafeteria DGYR (CAF DGYR); esses dois últimos receberam dieta de cafeteria antes e após o procedimento cirúrgico, até o desmame da prole. Na 18ª semana de vida, foi realizado o procedimento cirúrgico e a falsa operação nos grupos CAF DGYR e CAF FO, respectivamente. O cruzamento dos animais ocorreu cinco semanas após o procedimento cirúrgico. O nascimento dos animais foi considerado o dia zero pós-natal e o desmame se deu na terceira semana vida, quando somente os machos foram separados para o experimento. A prole da primeira geração (F1) foi nomeada em CTL-F1, OB-F1, CAF FO-F1 e CAF DGYR-F1 e todos os animais receberam dieta padrão. Na 17ª semana, os animais foram eutanasiados e o músculo ELD coletado para análise das fibras musculares e JNMs. Quando analisado os grupos CTL-F1 e OB-F1, esse último apresentou aumento do peso corpóreo, das gorduras retroperitoneal e periepididimal, e relação capilar/fibra. Além disso, houve a redução do número de núcleos, conjuntivo e alterações morfológica nos parâmetros avaliados na ultraestrutura. A área e diâmetro maior das JNMs também apresentaram redução. A análise entre os grupos CAF DGYR-F1 e CAF FO-F1 evidenciou redução do peso corporal, do peso do músculo ELD, da gordura retroperitoneal e periepididimal, don comprimento nasoanal, da área das fibras e relação núcleo/fibra no grupo CAF DGYR-F1. Esse grupo também apresentou aumento no número de fibras do tipo I e IIa e no número de capilares, assim como redução na área das JNMs e alterações morfológicas na ultraestrutura. Esses resultados demonstram que tanto a obesidade como a cirurgia bariátrica expõem a prole, por meio da programação metabólica, com efeitos na morfologia do tecido muscular esquelético, sendo encontrado maiores agravos na fibra muscular da prole de mães submetidas à DGYR.

O Streptococcus do Grupo B (EGB) é um dos principais agentes de infecção no período neonatal, sendo responsável por altos índices de morbimortalidade materno-fetal. Este estudo tem por objetivo avaliar a passagem transplacentária de anticorpos anti-Streptococcus B e imunoglobulina G em recém-nascidos de termo e pré-termo, bem como comparar seus níveis séricos. Foi realizado estudo transversal incluindo 44 recém-nascidos (18 pré-termo e 26 de termo) do Berçário Anexo à Maternidade do Hospital das Clínicas da FMUSP no período de dezembro de 2006 a julho de 2007. Após consentimento esclarecido, foram obtidas amostras de sangue das mães e do cordão umbilical de seus respectivos recém-nascidos, realizadas dosagens de IgG total por nefelometria e de anticorpos anti-EGB através do ensaio imunoenzimático (ELISA). Observou-se que nos dois grupos de mães da casuística, compostos por 16 mães de RN pré-termo e 26 mães de RN de termo, não houve diferença significativa em relação aos níveis séricos de anticorpos anti-EGB. O nível sérico médio de anticorpos maternos anti-EGB foi de 1697,98, com variação de 456 a 5200 (em títulos). O nível sérico médio de anticorpos anti-EGB das mães de RNPT foi de 1570,72, com variação de 588 a 3829 (em títulos), enquanto nas mães de RNT o nível médio foi de 1786,08, com variação de 456 a 5200. Os níveis séricos de anticorpos anti-Streptococcus do grupo B dos recém-nascidos foram significantemente mais baixos nos RN pré-termo em relação aos RN de termo. O nível sérico médio de anticorpos anti-EGB dos RNPT foi de 1059,22, com variação de 416 a 3924 (em títulos), enquanto nos RNT foi 2025,50, com variação de 542 a 5476. Houve correlação positiva entre os níveis de imunoglobulina G e de anticorpos anti-Streptococcus B com a idade gestacional, demonstrando correlação entre prematuridade e baixos níveis séricos de anticorpos anti-EGB. A associação entre idade gestacional inferior a 37 semanas e diminuição dos níveis de anticorpos anti-EGB concorda com a maior vulnerabilidade dos neonatos pré-termo à infecção por esta bactéria. Os autores concluem que houve passagem transplacentária de imunoglobulina G e anticorpos anti-Streptococcus B nos RN de termo e pré-termo, sendo, porém, os níveis séricos significantemente mais baixos nos RNPT, tanto em relação às suas mães quanto em relação aos níveis observados nos RN de termo. Os recém-nascidos de termo apresentaram níveis séricos de anticorpos anti-Streptococcus B semelhantes aos maternos, devido ao incremento da transferência transplacentária no final da gestação. Houve correlação positiva entre os níveis de imunoglobulina G e de anticorpos anti-Streptococcus B com a idade gestacional, enfatizando a importância da prematuridade como fator determinante das baixas concentrações séricas destes componentes imunológicos. Não houve diferença significante entre as mães dos recém-nascidos de termo e pré-termo em relação aos níveis séricos de anticorpos anti-Streptococcus B. / Group B Streptococcus (GBS) is one of the leading causes of infections in mothers and newborn babies, and it is responsible for high mortality rates. The purpose of this study was to evaluate the transplacental transfer of anti-Streptococcus B antibodies and immunoglobulin G in term and preterm newborns and compare the serum levels between these two groups. A transversal study was conducted with 44 newborns (18 preterm and 26 term infants) admitted to the Nursery next to FMUSP Hospital das Clínicas Maternity in the period of December 2006 to July 2007. After they gave their informed consent, blood and umbilical cord samples were collected from the mothers and from their respective newborn babies. Total IgG measurements were performed using nephelometry and the presence of antibodies anti-GBS was evaluated by the immunoenzimatic test (ELISA). In both groups of mothers (16 preterm`s mothers and 26 term`s mothers) the serum levels of anti-Streptococcus B antibodies were similar and there was no statistical significance between them. The mean serum levels of anti-GBS antibodies in mothers was 1697,98, ranging from 456 to 5200 (in titles). The mean serum levels of anti-GBS antibodies in mothers of preterm babies was 1570,72, ranging from 588 to 3829 (in titles), while in term`s mothers the mean level was 1786,08, ranging from 456 to 5200. Anti-Streptococcus B antibodies in the newborns demonstrated significantly lower levels in preterm newborn compared to the levels of the term newborns. The mean serum levels of anti-GBS antibodies in preterm newborns was 1059,22, ranging from 416 to 3924 (in titles), while in term newborns the mean level was 2025,50, ranging from 542 to 5476. There was a positive correlation between the levels of immunoglobulin G and anti-Streptococcus B antibodies and the gestational age which shows the correlation between prematurity and low levels of anti-Streptococcus B antibodies. The association between gestational age less than 37 weeks and reduction of anti-GBS antibody levels corroborates with the fact that preterm neonates are more vulnerable to the infection by this bacteria. The authors have come to the conclusion that transplacental transfer of immunoglobulin G and anti-Streptococcus B antibodies have been proved in term and preterm newborns. The transfer of immunoglobulin and antibodies was less effective in preterm newborns, whose serum levels were significantly lower compared to the levels of their mothers and the term newborns. Term newborns showed levels of anti-Streptococcus B antibodies similar to their mothers due to the increase of transplacental transfer of antibodies during the end of gestation. The positive correlation between the levels of immunoglobulin G and anti-Streptococcus B antibodies with gestational age, proves the importance of prematurity as a determining factor of the low serum concentrations in the components of this newborn\'s immunological repertoire. There was no significant difference between the levels of anti-Streptococcus B antibodies in mothers of term and preterm newborns.

A fim de investigar a associação entre alterações do tônus uterino e a ocorrência de anormalidades da freqüência cardíaca fetal (FCF) após analgesia obstétrica, em especial após o bloqueio combinado raqui-peridural, foi conduzido estudo prospectivo randomizado e encoberto, com setenta e sete parturientes que solicitaram analgesia regional durante o trabalho de parto. As pacientes do grupo estudo (41 casos) receberam duplo bloqueio com sufentanil e bupivacaína e as do grupo controle (36 casos) foram submetidas à peridural com as mesmas drogas. Monitorizou-se o tônus uterino por meio de cateter intra-amniótico de aferição da pressão intra-uterina e a freqüência cardíaca fetal durante 15 minutos antes e 30 minutos após a indução da analgesia. Pesquisaram-se os seguintes desfechos após a administração da analgesia: ocorrência de aumento do tônus uterino superior ou igual a 10mmHg em relação aos valores pré-analgesia e presença de desacelerações prolongadas da FCF ou bradicardia. Foram ainda quantificados os escores maternos de dor, a pressão arterial materna e o uso de ocitocina, antes e após o bloqueio. Observou-se associação significativa tanto do aumento de tônus uterino quanto das alterações da FCF com a analgesia combinada, nos primeiros 15 minutos após sua administração. O aumento de tônus foi encontrado em 17 de 41 casos no grupo estudo e em 6 de 36 pacientes nos controles (p=0,02). Alterações da FCF foram vistas em 11 dos 17 casos de aumento de tônus nas parturientes que receberam analgesia combinada, versus 1 de 6 no grupo peridural (p<0,001) A análise de regressão logística apontou o tipo de analgesia como fator independentemente associado ao aumento de tônus uterino, mesmo inserindo-se o uso de ocitocina como covariável. Também revelou o aumento de tônus como único fator independentemente associado ao desenvolvimento de anormalidades da FCF, mesmo com a hipotensão materna como covariável. Demonstrou-se ainda correlação entre o rápido declínio da dor com a técnica combinada e a probabilidade estimada de ocorrência simultânea de aumento do tônus e alterações da FCF. Os presentes achados reforçam a teoria de que o duplo bloqueio pode levar a aumento transitório de tônus uterino e culminar em anormalidades da FCF / In order to investigate the association between uterine tone elevation and fetal heart rate (FHR) abnormalities following labor analgesia - mainly with combined spinal-epidural (CSE) technique - a prospective double-blinded randomized study was conducted with seventy seven parturients who requested labor analgesia. Study group (41 cases) received CSE with sufentanil and bupivacaine and control group (36 cases) received epidural analgesia with the same drugs. Intra-uterine pressure was monitored with intra-amniotic pressure device and FHR with external transducer, both for at least 15 minutes before and 30 minutes after analgesia induction. The primary outcomes were the occurrence of an elevation of 10mmHg or more on uterine tone compared to the values before analgesia and the presence of prolonged fetal heart rate decelerations or fetal bradycardia. Maternal pain scores, blood pressure and use of oxytocin were also computed. A significant association was noticed between elevation of uterine tone and fetal heart rate abnormalities with combined spinal-epidural analgesia, at the first 15 minutes of administration. Uterine tone elevation was observed in 17 out of 41 CSE subjects and only 6 out of 36 controls (p=0.02). Fetal heart rate abnormalities were seen in 11 out of 17 cases that had hypertonus with combined analgesia and in only one of the 6 epidural patients (p<0.001). Logistic regression analysis revealed the mode of analgesia as the independent factor for the elevation of uterine tone, even with oxytocin use as a covariate. It also pointed out the uterine tone elevation as the only independent factor related to the development of fetal heart rate abnormalities, even with maternal hipotension as a covariate. A correlation was found between the fast onset pain relief provided by CSE analgesia and the estimated probability of uterine tone elevation and simultaneous fetal heart rate abnormalities. The present results strengthen the hypothesis that CSE analgesia can lead to a transient increase in uterine tone, leading to fetal bradycardia

O presente estudo analisou a hipótese de que as alterações hemodinâmicas fetais estão associadas à disfunção miocárdica fetal em gestações complicadas pela insuficiência placentária. Os objetivos do estudo foram correlacionar os valores do peptídeo natriurético cardíaco do tipo B (BNP) em sangue de cordão umbilical no nascimento com os parâmetros da dopplervelocimetria fetal, bem como com o pH no nascimento. Métodos: estudo prospectivo e transversal, com os seguintes critérios de inclusão: mulheres com gestação com feto único e vivo, insuficiência placentária caracterizada pelo aumento do índice de pulsatilidade (IP) da artéria umbilical (AU), membranas íntegras e ausência de anomalias fetais. Foram excluídos da pesquisa os casos com diagnóstico pós-natal de anomalia do recém-nascido e aqueles nos quais não foi realizada a análise do sangue no nascimento. Os seguintes parâmetros da dopplervelocimetria foram analisados: índice de pulsatilidade (IP) da artéria umbilical (AU) e da artéria cerebral média (ACM), relação cerebroplacentária (RCP) e IP para veias (IPV) do ducto venoso (DV). Os parâmetros de Doppler foram transformados em escore zeta. Amostras de sangue do cordão umbilical foram obtidas imediatamente após o parto para a mensuração do pH de artéria umbilical e do BNP. Resultados: Foram incluídas 32 gestações com diagnóstico de insuficiência placentária 21 (65%) com fluxo diastólico positivo e 11 (35%) com diástole zero ou reversa nas AU. A concentração do BNP apresentou correlação significativa com: escore zeta do IP da AU (rho=0,43; P=0,016); escore zeta da RCP (rho=-0,35; P=0,048); escore zeta do IPV do DV (rho=0,61; P<0,001), pH no nascimento (rho=- 0,39; P=0,031) e idade gestacional (rho=-0,51; P=0,003). Na regressão múltipla foram incluídos os parâmetros antenatais e o escore zeta do IPV do DV (P=0,008) demonstrou ser o único fator independente correlacionado com o BNP no nascimento. A correlação entre o BNP e o escore zeta do IPV do DV é representado pela equação de regressão Log[BNP]=2,34+0,13*DV (F=18,8, P<0,001). A correlação entre o BNP e o pH no nascimento é representado pela equação de regressão Log[BNP]=21,36-2,62*pH (F=7,69, P=0,01). Conclusão: os resultados sugerem que a disfunção cardíaca fetal identificada pelas concentrações de BNP no nascimento correlaciona-se de forma independente com as alterações no IPV do DV, além de correlacionarem-se negativamente com os valores do pH no nascimento / This study examined the hypothesis that fetal hemodynamic changes are associated with fetal myocardial dysfunction in pregnancies complicated by placental insufficiency. The objective was to study the correlation between the concentrations of cardiac natriuretic peptide type B (BNP) in umbilical cord blood at birth and fetal Doppler parameters, as well as the pH at birth. Methods: A prospective crosssectional study with the following inclusion criteria: pregnant women with a single fetus, placental insufficiency characterized by increased pulsatility index (PI) of the umbilical artery (UA), intact membranes and absence of fetal abnormalities. There were excluded from the study cases with postnatal diagnosis of abnormality of the newborn and those in which the blood analysis was not performed. The following Doppler parameters were analyzed: UA PI, middle cerebral artery (MCA) PI, cerebroplacental ratio (CPR) and ductus venosus (DV) PI for veins (PIV). The Doppler parameters were converted into zeta score. Blood samples were obtained from the umbilical cord immediately after delivery to measure the pH of the umbilical artery and the BNP. Results: Thirty two pregnancies with placental insufficiency were included, 21 (65%) with positive diastolic flow and 11 (35%) with absent or reversed end diastolic flow in the UA. The concentration of BNP correlated significantly with: UA-PI z-score (rho = 0.43, P = 0.016); CPR z-score (rho = -0.35, P = 0.048); DV-PIV zscore (rho = 0.61, P <0.001), pH at birth (rho = -0.39, P = 0.031) and gestational age (rho = -0.51, P = 0.003). In the multiple regression analysis, antenatal parameters were included and DV-PIV z-score (P = 0.008) was found to be independent parameter correlated with BNP at birth. The correlation between BNP and DV-PIV zscore is represented by the regression equation Log [BNP] = 2.34+0.13*DV (F=18.8, P <0.001). The correlation between BNP and pH at birth is represented by the regression equation Log [BNP] = 21.36-2.62*pH (F=7.69, P = 0.01). Conclusion: The results suggest that fetal cardiac dysfunction identified by BNP concentrations at birth correlated independently with changes in the DV PIV, and correlated negatively with pH values at birth