Made available in DSpace on 2014-06-11T19:22:21Z (GMT). No. of bitstreams: 0 Previous issue date: 2008-08-19Bitstream added on 2014-06-13T19:07:44Z : No. of bitstreams: 1 oliveira_lhb_me_arafcf.pdf: 553720 bytes, checksum: b0f8f04b07802f8bf467e43259c87fba (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Universidade Estadual Paulista (UNESP) / A maior expressão de alvos transcricionais e componentes da via NFkB é uma característica distintiva das células-tronco hematopoéticas (CTH) CD34+ de sangue de cordão umbilical (SCU) comparadas às CTH CD34+ de medula óssea (MO) e pode estar relacionada com o estado mais primitivo das CTH dos neonatos. No entanto, as células CD34+ são um grupo heterogêneo de célulastronco (CT) e progenitoras em diferentes estágios de maturação e diferenças na composição celular entre MO e SCU poderiam contribuir para os resultados mencionados. Estudos recentes têm identificado o marcador de superfície CD133, como um marcador de CT mais primitivas, expresso em uma subpopulação de células CD34bright, com um sugestivo potencial de hemangioblasto. Com o objetivo de caracterizar a composição celular de MO e SCU e identificar mecanismos moleculares envolvidos com a maior primitividade das células CD133+, propusemos avaliar o perfis imunofenotípico (quanto à expressão de CD34 e CD133) por citometria de fluxo e de expressão gênica de células CD34+ e células CD133+ selecionadas imunomagneticamente, de ambas as fontes, pelas técnicas de microarray e PCR em tempo real. Nossos resultados revelaram que enquanto a maioria das células CD133+ são CD34+, independente da fonte, as células CD34+ de SCU possuem uma porcentagem significativamente maior de células CD133+ do que às células CD34+ de MO. A análise de clusterização revelou que as células CD133+ de MO se agrupam com as células de SCU (CD34+ e CD133+), enquanto as CD34+ de MO aparecem como um grupo distinto. A comparação dos perfis de expressão gênica entre as células CD133+ e as células CD34+, revelou a hiper-expressão... / A higher expression of transcription targets and components of the NF-kB pathway is a distinctive feature of umbilical cord blood (UCB) CD34+ hematopoietic stem cells (HSC) when compared to bone marrow (BM) CD34+ HSC and this could be related to the more primitive state of the newborn’s HSC. However, CD34+ cells represent a heterogeneous group of cells composed by stem and progenitors cells in different developmental stages, and differences in cellular composition between both sources could contribute for these finding. The surface marker CD133 has been identified as a very primitive marker, expressed in a subpopulation of CD34bright, with a proposal hemangioblast potential. Thus, in attempt to better characterize the cellular composition of UCB and BM and to identify molecular mechanisms related to the more primitive characteristics of CD133+ cells, we proposed to evaluate the immunophenotypic profile (expression of CD34 and CD133) by flow-cytometry and the gene expression profiles of immunomagnetically selected CD34+ and CD133+ cells, from both sources, by microarray and Real time PCR. Our results highlighted that, while almost all CD133+ cells are CD34+ independently of the evaluated source, the UCB CD34+ cells showed a significantly higher proportion of CD133 expression, compared to BM CD34+ cells. After obtaining the expression profiles from distinct HSC pooled samples generated by microarrays, cluster analysis showed that BM CD133+ cells preferentially grouped with UCB cells (CD34+ and CD133+) instead of BM CD34+ cells, which appeared as a very distinct profile The comparison between CD133+ and CD34+ samples revealed the over-expression of 47 transcriptional factors (TF) in CD133+ cells, many of them well-known and related... (Complete abstract click electronic access below)

INTRODUÇÃO: Exposição a fatores ambientais e estresse durante o período intrauterino estão associados com alterações da trajetória do neurodesenvolvimento de forma sexo-dependente. Mecanismos epigenéticos estão envolvidos a esta associação. OBJETIVOS: Analisar de acordo com a exposição ao estresse na gestação o impacto do sexo e de alterações de metilação do DNA no sangue de cordão umbilical nas medidas antropométricas do neonato. MÉTODOS: Foram recrutadas 94 gestantes e aplicados questionários de medidas exposição ao estresse e fatores de risco durante a gravidez. A coleta de sangue do cordão umbilical seguiu protocolo padronizado. Para analisar o estresse foi utilizada análise de componentes principais (ACP) dos fatores de exposição avaliados: status socioeconômico, educação, ganho de peso, índice de massa corporal pré-gravídico, presença de doença psiquiátrica, estresse psicossocial durante a gravidez. Após o ACP fizemos análise de agrupamento por K-means. As análises de metilação foram realizadas utilizando Illumina Infinium Human Methylation450 (450K) BeadChip. Os dados foram analisados pelos pacotes Minfi e ChAMP (Chip Analysis Methylation Pipeline). A partir das posições diferencialmente metiladas (PDMs) foi feito análise de enriquecimento de processos biológicos com a ferramenta WebGestalt. Para avaliar impacto do sexo e alterações de metilação no desfecho antropométrico do neonato usamos modelos de análise linear de regressão múltipla. RESULTADOS: A coorte final para a avaliação do estresse foi composta por 89 pares mãe/recém-nascidos, sendo 50 meninas e 39 meninos. A ACP mostrou que os primeiros 3 componentes explicaram 60% da variabilidade da amostra. Sendo o primeiro componente (CP1) estresse psíquico, o segundo CP estresse social e o CP3 exposição a tóxicos. O biplot dos primeiros dois componentes sugeriu a separação das mães em dois grupos, confirmados pela análise de agrupamentos. Usando o ponto de corte de p-valor < 0,01 e deltabeta-valor>5%, encontramos 110 posições PDMs entre os grupos e restringindo este valor para p-valor < 0,01 e delta beta valor > 10% encontramos 13 PDMs. Usando apenas as crianças adequadas para idade gestacional fizemos análise de metilação diferencial entre os sexos. Foram encontradas 426 PDMs. Nenhuma das 13 PDMs encontradas entre os dois grupos pertenciam ao conjunto das PDMs entre sexos. No modelo de regressão linear multivariada controlando para sexo da criança e idade da mãe não encontramos nenhuma PDM associada aos desfechos antropométricos do neonato. Na análise estratificada por grupos os sítios cg24702040 (MAP3K21), cg21550016 (PAX8) foram estatisticamente significantes para perímetro abdominal e cg18706028 (CCKBR) e cg21550016 (PAX8) foram estatisticamente significantes para índice do perímetro cefálico para a idade. Este estudo sugere que o estresse materno independente do sexo pode afetar o crescimento fetal, mediado por respostas epigenéticas em genes relacionados à resposta ao estresse, regulação negativa da via de sinalização do receptor do fator de crescimento epidérmico, biogênese da sinapse e processo apoptótico / BACKGROUND: Exposure to environmental factors and stress during the intrauterine period are associated with changes in the neurodevelopmental trajectory in a sex-dependent manner. Epigenetic mechanisms are involved in this association. OBJECTIVES: Analyze according to exposure to stress during pregnancy the impact of sex and DNA methylation alterations on umbilical cord blood in the anthropometric measurements of the neonate METHODS: A total of 94 pregnant women were recruited and questionnaires were used to measure stress exposure and risk factors during pregnancy. Umbilical cord blood collection followed a standardized protocol. In order to analyze the stress, the principal components analysis (PCA) of the exposure factors evaluated were: socioeconomic status, education, weight gain, pre-gravid body mass index, presence of psychiatric illness, and psychosocial stress during pregnancy. After the PCA we did group analysis by k-means. Methylation analyzes were performed using Illumina Infinium Human Methylation 450 (450K) BeadChip. The data were analyzed by the Minfi and ChAMP (Chip Analysis Methylation Pipeline) packages. From the differentially methylated positions (DMPs) was made analysis of enrichment of biological processes with the tool WebGestalt. To evaluate gender impact and methylation alterations in the neonatal anthropometric outcome we used multiple regression linear analysis models. RESULTS: The final cohort for the evaluation of stress was composed of 89 mother/newborn pairs, being 50 girls and 39 boys. The PCA showed that the first 3 components accounted for 60% of the variability of the sample. Being the first component (PC1) psychic stress, the second PC social stress and PC3 exposure to toxic. The biplot of the first two components suggested the separation of the mothers into two groups, confirmed by cluster analysis. Using the cutoff point of p-value < 0.01 and delta beta-value > 5%, we found 110 DMPs between the groups and restricting this value to p-value < 0.01 and delta beta-value > 10 % we found 13 DMPs. Using only children suitable for gestational age we did differential methylation analysis between genders. There were 426 DMPs found. None of the 13 DMPs found between the two groups belonged to the pool of DMPs between the sexes. In the multivariate linear regression model controlling for child sex and age of the mother we did not find any DMP associated with the anthropometric outcomes of the neonate. In group-stratified analysis the cg24702040 (MAP3K21), cg21550016 (PAX8) sites were statistically significant for abdominal perimeter and cg18706028 (CCKBR) and cg21550016 (PAX8) were statistically significant for head cephalic circumference for age. This study suggests that maternal stress independent of sex can affect fetal growth, mediated by epigenetic responses in genes related to stress response, negative regulation of the epidermal growth factor receptor signaling pathway, biogenesis of the synapse and apoptotic process

臍帶血幹細胞移植自1988年首次被應用在異體的臨床治療至今，在各國衍生出許多以「利他」為目的的公捐臍帶血庫，以及鼓吹「自存自用」的私人臍帶血銀行。類似的情況也在台灣發生，第一家私人公益臍帶血庫在1997年底成立，第一家私人臍帶血銀行在2000年8月成立。然而，有關臍帶血儲存在台灣的發展歷史，卻甚少被討論。本論文嘗試以「創新的擴散」的四大因素—創新、溝通管道、時間及社會體系，詮釋自費儲存臍帶血在台灣的擴散過程和實質的內涵。 藉由中國時報報系和聯合報系有關臍帶血儲存的相關報導，估算出台灣自費儲存臍帶血的接受率在時間軸上的變化，發現台灣自費儲存臍帶血的接受率從2000年到2004年快速地成長，2004年以後成長趨緩。另外，從資料蒐集、文獻整理，以及台灣最大的親子社群網站”Babyhome”(寶貝家庭親子網)中有關臍帶血儲存的相關議題討論，觀察到台灣社會體系中的主管機關、私人臍帶血銀行、醫界和學界、公捐血庫、媒體等各成員之間的互動，直接或間接地影響潛在接受者對自費儲存臍帶血的意願和認知。在創新和溝通管道的探討中，則利用初探性質的網路問卷調查，探索台灣的接受者對於自費儲存臍帶血的創新認知屬性和資訊來源的溝通管道，並歸納出台灣的接受者對於自費儲存臍帶血的主要認知為：相容於「保險」的概念，以及代表「心安」的相對優勢。 是否應該自費儲存臍帶血實為一爭議性的話題，本研究也透過以上的分析，探討自費儲存臍帶血在台灣擴散所造成的後果並提出相關建議，期望台灣社會體系能夠從自費儲存臍帶血的擴散中，獲得創新的最大的利益又不至於造成社會體系的瓦解。 / Since the first umbilical cord blood transplant was performed in 1988 for allogeneic transplantation, many cord blood banks came out in countries, including public cord blood banks for an altruistic purpose and private blood banks which claim to bank umbilical cord blood for autologous use. Similar cases happened in Taiwan, the first public cord blood bank run by a private organization was founded in the end of 1997, then the first private cord blood bank was founded in August 2000. The history about banking umbilical cord blood in Taiwan, however, is rarely discussed. Based on the four main elements of “Diffusion of Innovations”, innovation, communication channels, time and social system, this study attempts to interpret the diffusion and substantial content of banking umbilical cord blood for autologous use in Taiwan. I estimated the adoption rate of banking umbilical cord blood for autologous use in Taiwan in the course of time by searching news reports from the databases of China Times Inc. and United Daily News Group. The adoption rate grown rapidly from 2000 to 2004 and the growth became slowly after 2004. By data collection, the survey of literatures and searching the relevant topics about storing umbilical cord blood from BabyHome, the biggest social networking website for parent-child relationship in Taiwan, I also found that the interaction among authority body, private cord blood bank, medical professional bodies, public cord blood bank and media, which are the members of Taiwan social system would affect the cognition and willingness of potential adopters directly or indirectly. In addition, I used a pilot online- questionnaire to explore the characteristics of innovation and communication channels of banking umbilical cord blood in Taiwan, which concludes that, the main characteristics of innovation sensed by the adopter who banking umbilical cord blood in Taiwan are compatibility with the concept of insurance and the relative advantage of feeling at ease. To store or not to store umbilical cord blood in private cord blood bank is actually a controversial issue. Through the analysis described above, the consequence of banking umbilical cord blood diffused in Taiwan is studied. I also make some suggestions and hope that Taiwan social system could gain the maximum social benefit from the diffusion of banking umbilical cord blood, and would not collapse by it.

The aim of this thesis was to study maternal and infant effects of delayed cord clamping (≥180 seconds, DCC) compared to early (≤10 seconds, ECC) in a randomised controlled trial. Practice and guidelines regarding when to clamp the cord vary globally, and different meta-analyses have shown contradictory conclusions on benefits and disadvantages of DCC and ECC. The study population consisted of 382 term infants born after normal pregnancies and randomised to DCC or ECC after birth. The primary objective was iron stores and iron deficiency at 4 months of age, but the thesis was designed to investigate a wide range of suggested effects associated with cord clamping. Paper I showed that DCC was associated with improved iron stores at 4 months (45% higher ferritin) and that the incidence of iron deficiency was reduced from 5.7% to 0.6%. Neonatal anaemia at 2-3 days was less frequent in the DCC group, 1.2% vs. 6.3%. There were no differences between the groups in respiratory symptoms, polycythaemia, or hyperbilirubinaemia. In paper II we demonstrated that DCC versus ECC was not associated with higher risk for maternal post partum haemorrhage and rendered a comparable ratio of valid umbilical artery blood gas samples. In paper III, the Ages and Stages Questionnaire was used to assess neurodevelopment at 4 months. The total scores did not differ, but the DCC group had a higher score in the problem-solving domain and a lower score in the personal-social domain. Immunoglobulin G level was 0.7 g/L higher in the DCC group at 2–3 days, but did not differ at 4 months. Symptoms of infection up to 4 months were comparable between groups. Finally, in paper IV, iron stores and neurodevelopment were similar between groups at 12 months. Gender specific outcome on neurodevelopment at 12 months was discovered, implying positive effects from DCC on boys and negative on girls. We conclude that delaying umbilical cord clamping for 180 seconds is safe and associated with a significantly reduced risk for iron deficiency at 4 months, which may have neurodevelopmental effects at a later age.

Der ischämische Schlaganfall ist global eine der bedeutendsten Volkskrankheiten. Die derzeit verfügbaren kurativen Therapieoptionen werden vorrangig durch ein enges therapeutisches Zeitfenster limitiert. Ziel der aktuellen Schlaganfallforschung ist die Entwicklung von über dieses Zeitfenster hinaus wirksamen Therapien. Ein vielversprechender neuer Ansatz ist die experimentelle Behandlung mit humanen Nabelschnurblutzellen. Diese Arbeit erforscht das therapeutische Zeitfenster für die systemische Therapie des ischämischen Schlaganfalls mittels mononukleärer Nabelschnurblutzellen (hUCB MNC) in spontanhypertensiven Ratten nach permanentem Verschluss der Arteria cerebri media (pMCAO). Hierzu wurden die Therapiezeitpunkte 4, 24, 72, 120 Stunden und 14 Tage nach experimentellem Schlaganfall in einem komplexen Studiendesign inklusive neurofunktioneller Tests, magnetresonanztomographischer und immunhistochemischer Verfahren untersucht. In vitro wurde der Einfluss kokultivierter hUCB MNC auf Nekrose und Apoptose in neuralem Gewebe unter Sauerstoff-Glukose-Deprivation betrachtet. Die Studie ergab eine verbesserte funktionelle Rekonvaleszenz und eine geringere Ausprägung von Atrophie und Astroglianarbe bei Therapie innerhalb eines 72- Stunden-Zeitfensters. In vitro wurde eine signifikante Reduktion von Nekrose und Apoptose durch kokultivierte hUCB MNC beobachtet. Eine histologische Relokalisierung der intravenös applizierten Zellen war in keiner Therapiegruppe möglich. Die Integration der hUCB MNC ins Hirnparenchym stellt somit keine conditio sine qua non für die funktionelle Erholung nach Schlaganfall dar. Trotz des beobachteten erweiterten Zeitfensters ist die Translation dieses Therapieansatzes in die klinische Realität kritisch zu diskutieren, da weiterführende Studien unserer Arbeitsgruppe eine limitierte Wirksamkeit unter sehr praxisnahen Bedingungen (z.B. Einsatz kryokonservierter hUCB MNC) gezeigt haben. / Experimental treatment strategies using human umbilical cord blood mononuclear cells (hUCB MNCs) represent a promising option for alternative stroke therapies. An important point for clinical translation of such treatment approaches is knowledge on the therapeutic time window. Although expected to be wider than for thrombolysis, the exact time window for hUCB MNC therapy is not known. Our study aimed to determine the time window of intravenous hUCB MNC administration after middle cerebral artery occlusion (MCAO). Male spontaneously hypertensive rats underwent MCAO and were randomly assigned to hUCB MNC administration at 4h, 24h, 72h, 120h or 14d. Influence of cell treatment was observed by magnetic resonance imaging on days 1, 8 and 29 following MCAO and by assessment of functional neurological recovery. On day 30, brains were screened for glial scar development and presence of hUCB MNCs. Further, influence of hUCB MNCs on necrosis and apoptosis in post-ischemic neural tissue was investigated in hippocampal slices cultures. Transplantation within a 72h time window resulted in an early improvement of functional recovery, paralleled by a reduction of brain atrophy and diminished glial scarring. Cell transplantation 120h post MCAO only induced minor functional recovery without changes in the brain atrophy rate and glial reactivity. Later transplantation (14d) did not show any benefit. No evidence for intracerebrally localized hUCB MNCs was found in any treatment group. In vitro hUCB MNCs were able to significantly reduce post-ischemic neural necrosis and apoptosis. Our results for the first time indicate a time window of therapeutic hUCB MNC application of at least 72 hours. The time window is limited, but wider than compared to conventional pharmacological approaches. The data furthermore confirms that differentiation and integration of administered cells is not a prerequisite for poststroke functional improvement and lesion size reduction.

La transplantation de sang de cordon ombilical (TSCO) est utilisée pour traiter les enfants atteints de maladies hématologiques en l’absence de donneurs apparentés compatibles. Elle est associée avec des risques plus élevés d’échec de greffe et d’infections opportunistes dans les premiers mois qui suivent la transplantation en comparaison avec une greffe de moelle osseuse. Par contre, la TSCO comporte un risque plus faible de maladie du greffon contre l’hôte et une incidence comparable de rechute de leucémie. Ces quatre complications impliquent directement les lymphocytes T. Dans le but de mieux comprendre le schéma particulier des évènements qui suivent la TSCO et d’améliorer le pronostic des patients, nous avons étudié le potentiel fonctionnel, la persistance et la reconstitution antivirale des lymphocytes T au sein d’un groupe d’enfants transplantés de sang de cordon ombilical (SCO). Étant donné que le SCO contient une majorité de lymphocytes T naïfs, nous avons étudié les lymphocytes T spécifiques au HLA-A2:Melan-A26-35 A27L; seul répertoire naïf et abondant caractérisé chez l’homme. Nous avons observé que les lymphocytes T du SCO se différencient en populations effectrices, s’oligoclonalisent, produisent de l’IFN-γ et lysent spécifiquement leur cible suite à la stimulation. Néanmoins, ces cellules produisent moins d’IFN-γ et sont moins bifonctionnelles que leurs homologues issus du sang périphérique d’adultes. Chez les patients, les lymphocytes T du SCO s’épuisent après la TSCO : ils s’oligoclonalisent dramatiquement, sont principalement en différenciation terminale, et une importante fréquence exprime PD-1 (« programmed death-1 ») dans les 3 à 6 premiers mois post-greffe. Très peu de patients sont capables de développer des réponses antivirales durant cette période et la fréquence de lymphocytes T qui expriment PD-1 semble aussi avoir un impact sur le risque subséquent de faire une rechute de leucémie. La deuxième vague de lymphocytes T émergeant à 6 mois post-TSCO mène à une population fonctionnelle et diversifiée. En conclusion, la fonctionnalité des lymphocytes T présents dans les 3 à 6 premiers mois post-TSCO doit être rétablie pour améliorer les risques d’infections opportunistes et de rechute de leucémie. / Umbilical cord blood (UCB) is increasingly used as a source of hematopoietic progenitor cells to treat a variety of disorders in children. UCB transplantation (UCBT) is associated with a reduced incidence of graft-versus-host disease, a robust graft-versus-leukemia effect, more frequent graft failures and a higher incidence of opportunistic infections (OI) compared to bone marrow transplantation; four processes in which donor-derived T lymphocytes are known to be predominantly involved. UCB T cells are mostly naïve. To examine the differential functionality of UCB T cells, CD8+ T cells specific for the melanoma-associated HLA-A2-restricted Melan-A26-35 A27L peptide were isolated from UCB and UCBT recipients, as it represents an abundant preimmune repertoire in human. Following in vitro stimulation, UCB T cells proliferated, oligoclonalized, acquired cell surface markers reflective of effector/memory differentiation, expressed cytolytic activity and produced IFN-γ. While functional properties of UCB T cells resembled their counterparts in adult peripheral blood, they were more likely to reach terminal differentiation following stimulation, produced less IFN-γ and were less frequently bifunctional (IFN-γ and cytolysis). Following UCBT, T cells became exhausted: they oligoclonalized dramatically, exhibited a terminal differentiation phenotype and a high frequency also expressed PD-1 (“ programmed death 1 ”) in the first 3-6 months post-UCBT. Moreover, very few patients developed an antiviral response during this period. Finally, the frequency of PD-1+ CD8+ T cells was significantly higher in subjects who subsequently experienced leukemic relapse. A second wave of T cells emerging at 6 months post-UCBT was observed and characterized by an increase in the repertoire diversity till 1 year, the development of a naïve population with polyfunctional potential and the progressive reconstitution of antiviral responses. This study reports to the biological properties of UCB-derived CD8+ T cells and provides a rationale for the characteristics of UCBT in terms of immune reconstitution and OI. These results also suggest that the first wave of CD8+ T cells in the first 3-6 months post-UCBT should be targeted in priority to improve both OI and leukemic relapse risks.

Past and present uses of DDT and pyrethroids have led to their incorporation into humans, mainly through the food chain and sometimes by direct exposure. The present work focuses on establishing the levels of DDT, its analogous compounds (DDE and DDD), and pyrethroids in humans and the human environment in Manhiça, a rural area where they have been used as insecticides for indoor residual spraying (IRS) and insecticide treated nets (ITN) in malaria control programs. Thatch samples from human dwellings, breast milk from pregnant women and cord blood from newborns were analyzed for assessment of the concentration levels of these compounds. The results showed that DDT and its analogues were already present in humans and dwellings before reintroduction of this insecticide for IRS. As consequence of these applications DDT concentrations increased significantly. The higher proportion of 4,4’-DDT than 4,4’-DDE evidenced that the observed amounts were due to recent applications of this insecticide. Concerning pyrethroids, their presence has been identified in both breast milk and human dwellings showing that both agricultural applications and use for ITN may be responsible for their occurrence in humans and human environments of Manhiça. / L’ús en el passat i en temps actuals del DDT i piretroides ha donat lloc a la seva incorporació en els humans, principalment mitjançant la cadena tròfica i a vegades per exposició directa. Aquest treball té com objectiu establir els nivells de DDT i els seus compostos anàlegs (DDE i DDD), i piretroides en humans i l’ambient humà a Manhiça, una àrea rural on aquests s’han utilitzat com insecticides per aplicació interna (indoor residual spraying, IRS) i tractament de xarxes de protección (insecticide treated nets, ITN) en programes de control de la malària. Per a esbrinar els nivells de concentració d’aquests compostos s’analitzaren mostres de palla de cabanes, de llet materna i de sang de cordó de nou nats. Els resultats mostraren que el DDT i els seus compostos anàlegs ja eren presents en humans i cabanes abans de la reintroducció d’aquest insecticida per IRS. L’ús del DDT en aquest programa féu augmentar considerablement les concentracions d’aquest insecticida. La major proporció de 4,4’-DDT que 4,4’-DDE posà de manifest que les quantitats observades corresponien a aplicacions recents d’aquest insecticida. Respecte als piretroides, s’han trobat en mostres de llet materna i cabanes tot mostrant que tant les aplicacions agrícoles com el seu ús en ITN poder esser la causa de la seva presència en els humans i els ambients humans de Manhiça.

Σκοπός της παρούσας μελέτης είναι η σύγκριση των συγκεντρώσεων 6 ορμονών, Ε3, β-hCG, hPL, FSH, LH και κορτιζόλη στο αίμα του ομφαλίου λώρου των νεογέννητων των καπνιστριών γυναικών σε σχέση με τις μη καπνίστριες μητέρες. Μέθοδοι: Οι παραπάνω ορμόνες μετρήθηκαν στο αίμα ομφαλίου λώρου σε 100 νεογνά των οποίων οι μητέρες κάπνιζαν (ομάδα μελέτης) και 100 παιδιά των οποίων οι μητέρες δεν κάπνιζαν (ομάδα ελέγχου). Αποτελέσματα: H μέση τιμή των συγκεντρώσεων E3, hPL, β-hCG, FSH, LH και κορτιζόλης στα νεογνά μη καπνιζόντων μητέρων ήταν 212 ng/mL, 2.00 microg/mL, 57.5 mIU/mL, 0.10 mIU/mL, 0.20 mIU/mL, and 14.3 microg/mL, αντιστοίχως· στα νεογνά των καπνιστριών μητέρων ήταν 163, 1.39, 45.4, 0.10, 0.20, and 25.1, αντιστοίχως (p=0.008, 0.004, 0.037, 0.498, 0.286, 0.004, respectively). Διαπιστώθηκε σημαντική αλλά αρνητική συσχέτιση μεταξύ του αριθμού των τσιγάρων ανά ημέρα και των E3 (r=-0.163, P=0.021), hPL (r=-0.191, P=0.007) και β-hCG (r=-0.143, P=0.044), ενώ η συσχέτιση με την κορτιζόλη ήταν θετική (r=0.259, P<0.0001). Πολλαπλή γραμμική εξαρτημένη ανάλυση έδειξε ότι το μητρικό κάπνισμα ήταν καθοριστικός παράγοντας για τις συγκεντρώσεις των ορμονών E3, hPL, β-hCG, FSH, και κορτιζόλη του αίματος του ομφαλίου λώρου. Συμπέρασμα: Το κάπνισμα συσχετίζεται με μείωση των συγκεντρώσεων των ορμονών E3, hPL, β-hCG και FSH του αίματος του ομφαλίου λώρου. Ενώ, συσχετίζεται με αυξημένη συγκέντρωση κορτιζόλης. Η διαταραγμένη ενδοκρινική ισορροπία του εμβρύου από το κάπνισμα του καπνού μπορεί να έχει αρνητικές επιδράσεις στο έμβρυο και το παιδί εφόσον ο εμβρυϊκός εγκέφαλος είναι στόχος ορμονικών δράσεων. / To determine the effect of maternal cigarette smoking on cord blood concentrations of E3, hPL, beta-hCG, FSH, LH, and cortisol. Hormone concentrations were measured in term neonates of 100 smoking and 100 non-smoking mothers. The median E3, hPL, beta-hCG, FSH, LH and cortisol cord blood concentrations in the non-smoking mothers' offspring were 212 ng/mL, 2.00 microg/mL, 57.5 mIU/mL, 0.10 mIU/mL, 0.20 mIU/mL, and 14.3 microg/mL, respectively; in the smoking they were 163, 1.39, 45.4, 0.10, 0.20, and 25.1, respectively (P=0.008, 0.004, 0.037, 0.498, 0.286, 0.004, respectively). There was a significant but poor negative correlation between number of cigarettes/day and E3 (r=-0.163, P=0.021), hPL (r=-0.191, P=0.007), and beta-hCG (r=-0.143, P=0.044), whereas the correlation with cortisol was positive (r=0.259, P<0.0001). Multiple linear regression analyses showed that maternal smoking is a determinant of cord blood E3, hPL, beta-hCG, FSH, and cortisol concentrations. Tobacco smoking is associated with a reduction in cord blood E3, hPL, and beta-hCG concentrations, whereas it is associated with increased cortisol concentrations. The disturbed endocrine equilibrium of the fetus induced by tobacco smoking could have adverse consequences on the fetus and child since fetal brain is a target organ for hormonal actions.

Le système immunitaire néonatal est caractérisé par une susceptibilité accrue aux infections par des pathogènes intracellulaires et des réponses vaccinales altérées. Ces caractéristiques soulignent la capacité limitée du nouveau-né à développer des réponses immunes à médiation cellulaire.<p>Les LTγδ représentent le prototype des lymphocytes T non-conventionnels :ils sont capables de réagir très rapidement après activation et sont caractérisés par une activité qui n’est pas restreinte aux molécules du CMH. Les LTγδ qui expriment le TCR Vγ9Vδ2 constituent la population majoritaire (60-90%) de LTγδ dans le sang périphérique humain adulte. Ces cellules reconnaissent spécifiquement les phosphoantigènes, des molécules non-peptidiques de faible poids moléculaire tel que l’HMB-PP. Cet activateur naturel est un intermédiaire de la voie non-mévalonate de biosynthèse des isoprénoïdes, essentielle à de nombreux agents pathogènes. Ces cellules sont également capables de réagir aux aminobisphosphonates, drogues couramment utilisées en thérapie du cancer. Le zolédronate (Zometa) est le plus puissant des aminobisphosphonates. Grâce à leur capacité à sécréter rapidement des cytokines comme l’IFN-γ et le TNF-α ainsi qu’à leur puissante activité cytotoxique, les LT Vγ9Vδ2 jouent un rôle important dans le contrôle des infections et des cancers.<p><p>Au contraire des LTγδ de l’adulte, la fonction des LTγδ du nouveau-né est peu connue. Cependant, des évidences suggèrent que ces lymphocytes pourraient jouer un rôle particulièrement important en début de vie. Dans ce contexte, nous avons caractérisé en détail la réponse des LTγδ du sang de cordon ombilical humain après stimulation avec l’HMB-PP et le zolédronate. Contrairement à l’HMB-PP, le zolédronate induit la prolifération et la production d’IFN-γ par les LT Vγ9 du nouveau-né. L’IL-23 étant une cytokine produite de façon optimale par les DCs en début de vie, nous avons choisi d’étudier son influence sur le profil d’activation de ces cellules. L’ajout d’IL-23 au zolédronate augmente l’expression d’IFN-γ et génère une population distincte de LT Vγ9 négatifs pour l’IFN-γ et produisant de l’IL-17. De plus, l’IL-23 favorise la synthèse de plusieurs médiateurs cytotoxiques (perforine, granzymes A et B, granulysine) induits par le zolédronate. Alors que l’effet co-stimulateur de l’IL-23 sur la production d’IFN-γ et de molécules cytotoxiques est également observé au sein des LT Vγ9 de l’adulte, l’induction d’une sous-population IL-17+ IFN-γ- est unique aux LT Vγ9 du nouveau-né.<p>Afin de mieux définir les conditions requises pour induire la polarisation des LT Vγ9 du nouveau-né en cellules productrices d’IL-17, nous avons évalué l’effet de diverses cytokines ainsi que celui de dérivés microbiens sur l’activation de ces cellules au zolédronate. Le zolédronate en présence d’IL-1β ou de LPS génère une population distincte de LT Vγ9 IL-17+ IFN-γ-. De plus, l’ajout d’IL-1β au zolédronate n’augmente pas le pourcentage de LT Vγ9 producteurs d’IFN-γ. Nous avons ensuite comparé le phénotype des LT Vγ9 IL-17+ à celui des LT Vγ9 IFN-γ+. Nos résultats indiquent qu’une fraction des LT Vγ9 sécrétant l’IL-17 exprime sélectivement CCR6 contrairement aux LT Vγ9 producteurs d’IFN-γ.<p><p>En conclusion, ce travail a permis de révéler la capacité des LT Vγ9 du nouveau-né humain à se différencier en cellules productrices d’IFN-γ, d’IL-17 et de molécules cytotoxiques. Ces cellules pourraient donc constituer une première ligne de défense capable d’orchestrer une réponse immune efficace à l’encontre de dérivés microbiens ou de signaux endogènes de danger. / Doctorat en Sciences biomédicales et pharmaceutiques / info:eu-repo/semantics/nonPublished

p. 1-204 / Submitted by Santiago Fabio (fabio.ssantiago@hotmail.com) on 2013-05-02T19:30:43Z No. of bitstreams: 1 999999999999999999.pdf: 821652 bytes, checksum: a675e81264e0276fcd87b509e674c613 (MD5) / Approved for entry into archive by Maria Creuza Silva(mariakreuza@yahoo.com.br) on 2013-05-04T17:40:30Z (GMT) No. of bitstreams: 1 999999999999999999.pdf: 821652 bytes, checksum: a675e81264e0276fcd87b509e674c613 (MD5) / Made available in DSpace on 2013-05-04T17:40:30Z (GMT). No. of bitstreams: 1 999999999999999999.pdf: 821652 bytes, checksum: a675e81264e0276fcd87b509e674c613 (MD5) Previous issue date: 2007 / Fatores associados ao retardo no crescimento intra-uterino ainda não foram totalmente esclarecidos. Recém-nascidos (RN) pequenos para a idade gestacional (PIG) apresentam alterações nos valores hematológicos comparados aos RN adequados para a idade gestacional (AIG) e pré-termos. Persistem também limitações metodológicas inerentes a acurácia dos métodos de determinação da idade gestacional. Objetivos: Determinar os fatores associados ao RCIU, as características hematológicas dos recém nascidos portadores do RCIU e avaliar a acurácia dos métodos da avaliação da idade gestacional utilizados no diagnóstico antropométrico desses recém-nascidos. Metodologia: Estudo envolvendo puérperas e 564 RN em 2 maternidades públicas da cidade de Salvador. Foram classificados de AIG, RN com o peso ≥10th e <90th, de PIG aqueles com peso <10th, avaliados pela curva de Williams; e pré-termo, aqueles com <37 semanas de gestação. Os dados foram coletados utilizando-se de questionário padronizado. Foram tomadas as medidas antropométricas da criança e da puérpera e coletou-se o sangue do cordão umbilical. A idade gestacional foi calculada pelos métodos da data da última menstruação, ultrassonografia e físico de Capurro. Estudo 1 de desenho caso-controle. Estudos 2 e 3 de desenhos de corte transversal. Análise estatística: No estudo 1, utilizou-se a regressão logística não condicional para testar a associação entre a variável dependente e as preditoras e Odds Ratio foi adotado como medida de associação. No estudo 2, a média e seu respectivo DP, valor máximo e mínimo foram usados para descrever os valores hematológicos do cordão umbilical. Para o estudo 3, realizou-se a diferença entre as medianas da idade gestacional segundo os métodos, utilizando-se do teste não paramétrico “Wilcoxon Signed Rank Test”. A correlação entre as idades gestacional estimadas pelo DUM, USG e Capurro e entre estes métodos e o peso ao nascer foi realizada utilizando-se do coeficiente de correlação de Pearson. Utilizou-se o Coeficiente Kappa para avaliar a concordância entre os métodos na estimação do estado antropométrico do recém-nascido. A validade dos métodos na estimação da idade gestacional foi avaliada por meio do cálculo da sensibilidade, especificidade, VPP, VPN e pela curva ROC. Foram utilizados os pacotes estatístico SPSS.11 e o Stata 8, aceitando-se a significância de 5% nas estimativas de interesse. Resultados: Com o estudo 1, identificou-se que a primeira gestação (OR:2,85; 1,73-4,71), o hábito de fumar (OR: 2,65; 1,35-5,19) e a gestação anterior desfavorável (OR:2.10; 1,21-3,64) se comportaram como fatores de risco para retardo no crescimento intra-uterino. Os resultados do estudo 2 indicaram que valores mais altos de hemácias (4,21×1012/l±0,48), hemoglobina (14,50 g/dl ±1,42), hematócrito (43,50%±4,36), ferritina (162,61 µg/l ±100,10); RDW (13,34±0,85%) e leucócitos (12,82 109/l ±3,39) foram identificados nos RN PIG, quando comparados com aqueles dos AIG a termo e pré-termos. Os RN AIG pré-termos apresentaram valores mais altos de VGM e HGM. Os valores médios das plaquetas foram mais altos nos RN AIG a termo. A partir dos resultados do estudo 3, observou-se que o método de Capurro incrementou a idade gestacional em intervalos menores de 39 semanas e a USG a aumentou a partir deste patamar, em relação ao método DUM. Os métodos DUM e USG apresentaram maior correlação entre a estimativa da idade gestacional e o estado antropométrico para o conjunto dos RN (r=0,668) e para a identificação do RN AIG (r=0,685). Considerando a idade gestacional estimada pelo DUM e a relação com o peso ao nascer, o coeficiente de correlação foi mais elevado (r=0,609) na identificação dos RN PIG’s. Na predição de RN PIG’s, maior sensibilidade foi observada para a USG (96,6%) e maior especificidade para o Capurro (75,5%). Acurácia mais elevada na estimativa da idade gestacional foi observada para a USG na 41ª, tomando como referencia o DUM (ROC=77%). Conclusão: O RCIU associou-se à primeira gestação, à gestação anterior desfavorável e ao hábito de fumar. RN portadores de RCIU apresentaram valores ematológicos mais elevados em relação os RN AIG e pré-termos e o método físico de Capurro apresentou pior desempenho na identificação de RCIU. / Salvador