Дисертації з теми "Delayed death"

Autophagy is a physiological process by which the cell eliminates damaged organelles, toxic agents, and long-lived proteins by degradation through lysosomal system. Mitophagy, the specific autophagic elimination of mitochondria, regulates mitochondrial number to match metabolic demand and is a core machinery of quality control to remove damaged mitochondria. A neuroprotective role of physiological autophagy/mitophagy has been discovered. However, recent studies suggested that highly accelerated autophagy/mitophagy might contribute to neuronal death in various pathological situations including cerebral ischemia. In this study, we aimed to investigate the activation of excessive autophagy, particularly, the more specific mitophagy, in neuronal tissues and its contribution to ischemia/hypoxia (I/H)-induced delayed neuronal death. I/H injury was induced by oxygen and glucose deprivation (OGD) followed by reperfusion (RP) on primary cortical neurons in vitro. Cerebral ischemia was induced by unilateral common carotid artery occlusion and hypoxia in neonatal mice in vivo. In order to determine the extent to which autophagy contributes to neuronal death in cerebral ischemia, we performed multiple methods and found that in both primary cortical neurons and SH-SY5Y cells exposed to OGD for 6 h and RP for 24, 48, and 72 h, respectively, an increase of autophagy was observed as determined by the increased ratio of LC3-II to LC3-I and Beclin 1 expression. Using Fluoro-Jade C and monodansylcadaverine double-staining, and electron microscopy we found the increment in autophagy after OGD/RP was accompanied by increased autophagic cell death, and this increased cell death was inhibited by the specific autophagy inhibitor, 3-methyladenine. The presence of large autolysosomes and numerous autophagosomes in cortical neurons were confirmed by electron microscopy. Autophagy activities were increased dramatically in the ischemic brains 3-7 days postinjury from a rat model of neonatal cerebral I/H as shown by increased punctate LC3 staining and Beclin-1 expression. We thus obtained the conclusion that excessive activation of autophagy contributes to neuronal death in cerebral ischemia. BNIP3 (Bcl-2/adenovirus E19 kD interacting protein 3), a member of a unique subfamily of death-inducing mitochondrial proteins, is highly associated with mitochondrial dysfunction and delayed neuronal death in stroke. It is known that BNIP3-induced neuronal death is caspase-independent and characterized by early mitochondrial damage. Recent evidence suggested that the BNIP3 family of proteins might be important regulators of mitophagy. Here, using both stroke models, we found that homodimer (60 kD) of BNIP3/NIX (BNIP3L) were highly expressed in a ‘delayed’ manner. Particularly, significant mitophagic activation was confirmed by electron microscopy. In contrast, both neonatal mitophagy and apoptosis were significantly inhibited in the BNIP3 knockout (KO) mice after I/H, which was also accompanied by a significantly increased autophagic response. In addition, the infarct volume in the BNIP3 KO mice was significantly reduced as compared to wild-type (WT) mice after 7 or 28 days recovery, showing a prominent neuroprotection of BNIP3 gene silencing. A protein-to-protein interaction of mitochondria-localized BNIP3 (60 kD) with the autophagosome marker, LC3, was confirmed by co-ip, immunocytochemistry and further quantified by ELISA, indicating BNIP3 was an effective LC3-binding target on damaged mitochondria. These data demonstrated a novel role of BNIP3 in regulating neuronal mitophagy and cell death during ischemic stroke.

Stroke is a leading cause of death and disability throughout the world; ischemia is the most common form of stroke. Medical procedures such as cardio-pulmonary bypass surgery can cause ischemic stroke can be caused. There are no treatments to limit neural impairment following stroke. The current research investigates neuroprotection offered by treatment with a novel drug combination consisting of Simvastatinâ„¢, Gemfibrozilâ„¢, Troglitazoneâ„¢, and Spironolactoneâ„¢. Animals were treated with the drug cocktail three weeks proceeding and one week subsequent to surgery. Ischemic insult was induced by clamping the carotid arteries for 5 min. Sham subjects underwent similar surgical procedures, but the carotids were not clamped. Twenty-four hrs following the surgical procedure locomotor activity was monitored in an open field for 5 min. Seven to fourteen days following ischemia or the sham procedure animals were sacrificed and sections containing the hippocampal CA1 region were mounted on slides and stained with cresyl violet. The CA1 region was rated on a 4-point scale for level of damage. Rodents generally show increased locomotor activity following transient global ischemia in an open field. In our study, ischemic animals that received vehicle demonstrated increased activity relative to the animals that received the drug treatment on all behavioral measures. Ischemic animals that received vehicle treatment had significantly more neural damage in the hippocampal CA1 region than ischemic animals receiving the drug. The appearance of neurons in the CA1 hippocampal regions of animals in the sham condition was not significantly different from ischemic animals in the drug treatment condition. It is concluded that the drug treatment is effective in offering neuroprotection during transient global ischemia. The next step is to characterize the biochemical mechanisms behind the neuroprotection conferred by the drug treatment. Contrasting the protein expression levels of animals receiving the vehicle treatment with animals receiving the drug treatment following an ischemic insult will assist in elucidating these pathways. Predictions are made regarding the biochemical mechanisms affected by the drug treatment based on previous research on the biochemical pathways affected by each pharmaceutical.

Le vieillissement des populations est corrélé à l’augmentation des pathologies neurodégénératives liées à l’âge, plus particulièrement la maladie d’Alzheimer. La recherche de marqueurs précoces de la maladie ainsi que l’élaboration de nouvelles stratégies thérapeutiques constituent un enjeu de taille. Parmi les mécanismes moléculaires de la formation des plaques amyloïdes actuellement explorés, les formes oligomériques tronquées de peptide amyloïde (Aß), notamment le peptide Aß3(?pE)??42? retrouvé à des stades précoces de la maladie, joueraient un rôle déterminant. Ces travaux de thèse ont permis de montrer, dans un premier temps, que l’injection intracérébrale de ce peptide chez la souris entraîne des altérations de la mémoire de travail et des capacités d’apprentissage, associées à une accumulation d’espèces réactives dérivées de l’oxygène dans des régions cérébrales spécifiques (hippocampe et bulbes olfactifs) de ces animaux. Des essais menés in vitro sur des cultures primaires de neurones de souris montrent leur implication dans les voies apoptotiques impliquant l’activation des caspases et la cascade métabolique de l’acide arachidonique. La seconde étape de ces travaux a constitué en l’étude des effets protecteurs d’un peptide antiapoptotique d’origine endogène, l’humanine (HN) et son variant S14G (HNG). In vitro, un effet protecteur de ces peptides a été mesuré après traitement de neurones en culture par le peptide A[bêta]3?(pE)42.??? Les résultats les plus marquants résident dans les observations faites in vivo : en effet, ces peptides inhibent l’effet délétère de l’injection intracérébroventriculaire du peptide Aß3?(pE??)42?? en restaurant les performances mnésiques des animaux dans les tests comportementaux. A la lumière de ces résultats, les peptides HN pourraient constituer de nouveaux outils thérapeutiques dans le traitement ou la prévention des dommages cellulaires précoces liés à la présence des oligomères solubles du peptide Aß
Aging of population is correlated to the increase of neurodegenerative disease, more particularly Alzheimer disease. Defining early diagnostic markers and new therapeutic strategies are highly relevant. Among the molecular pathways which are currently developed, N-terminal-truncated forms of amyloid-ß (Aß) peptide have been recently suggested to play a pivotal role in the disease. Among them, Aß3(?pE)42 ?peptide is the dominant Aß species in amyloid plaques. We first investigated the effects of soluble oligomeric Aß3(pE) 42 after intracerebroventricular injection on mice learning capacities and the molecular mechanisms of in vitro neurotoxicity. Mice injected with soluble Aß3(pE) 42 displayed impaired spatial working memory and delayed memory acquisition. These cognitive alterations were associated with free radical overproduction in hippocampus and olfactory bulbs. In vitro, Aß3(pE) 42 oligomers induced a redox-sensitive neuronal apoptosis involving caspase activation and an arachidonic acid-dependent pathway. The second goal of this work was to investigate the protective effects of the apoptosis rescue endogenous peptide humanin (HN) and its S14G mutant (HNG). In vitro, we measured their inhibitory effect on neuronal death and apoptotic events resulting from soluble Ab oligomer treatment. What’s of particular interest is the in vivo restoration of soluble Aß3(pE) 42 oligomer-induced mnesic impairment. Thus, HN peptides might serve as new drug candidates for treatment or prevention of early cellular damages linked to soluble A[bêta] oligomers

This thesis attempts to identify and explain what influences the length of time an inmate spends on Florida's death row. A systematic random sample of 33 Florida death row inmates was drawn from the Florida Department of Corrections death row roster and the Florida Commission on Capital Cases inmate roster. Documented for each death row inmate was how long he spent on Florida's death row navigating the various stages and steps in Florida's post-conviction capital punishment process. The data show that petitions to the state trial courts and appeals to the Florida Supreme Court take the longest time in Florida's post-conviction capital punishment process. It also shows a considerable amount of "dead time," which refers to any additional time that an inmate spends on death row with no legal actions pending. A theory of "benign neglect" is proposed as the most likely explanation for the excessive delays.
M.S.
Department of Criminal Justice and Legal Studies
Health and Public Affairs
Criminal Justice MS

Master of Public Health - MPH
Access to appropriate health care service during childbirth is a great challenge to many women in Africa and Namibia is no exception. More than 70% of women in Otjozondjupa region experienced some form of delay during childbirth, and while maternal mortality continued to rise over the years in Namibia it is currently at about 265/100 000, which is too high for a middle-income country. Hence, this study aimed to get a deeper understanding of the perceptions of women who experienced any of the three delays in accessing appropriate health care during childbirth in Otjiwarongo hospital.

The purpose of the current study is to evaluate the relationship between discounting immediate and former life statuses prior to death related to quality of life across the lifespan. A discounting survey was completed by 83 participants in which they were asked to make hypothetical choices regarding returning back to an age they preferred or remaining at their current age prior to dying. In addition, participants completed surveys measuring quality of life, death depression, and death anxiety. Results indicated that participants who reported low quality of life and experienced high depression and anxiety towards death responded more impulsively to the discounting survey, engaging in steeper discounting. Additionally, results indicated that a positive correlation between participants aged 30 to 50 years old and AUC exists, indicating that as age increases, impulsivity decreases. However, this same trend was not evident in participants over the age of 65, yielding no correlation. Strengths and limitations, implications of the current study’s findings, and opportunities for future research are discussed.

This thesis studies the design and implementation of an ultra-sonic water depth sounder. The depth sounder is implemented in a hand-held smart console used by divers. Since the idea of echo sounding is to measure the flight time between transmitting the signal and receiving the echo, the main challenge of this task is to find a time-of-flight (ToF) estimation for a signal in noise. It should be suitable for this specific application and robust when implemented in the device. The thesis contains an investigation of suitable ToF methods. More detailed evaluations of the matched filter, also known as the correlation method, and the linear phase approach are done. Aspects like pulse frequency and duration, speed of sound in water and underwater noise are taken into account. The ToF-methods are evaluated through simulation and experiments. The matched filter approach is found suitable based on these simulations and tests with signals recorded by the console. This verification leads to the implementation of the algorithm on the device. The algorithm is tested in real time, the results are evaluated and improvements suggested.
Denna rapport behandlar skattning av vattendjup med hjälp av ultraljudspulser och implementation av detta. Djupmätaren implementeras i en handhållen dykarkonsoll. Eftersom grundidén i ekolodning är att mäta tiden mellan att pulsen skickas iväg och att ekot tas emot är en stor del av utmaningen att hitta en lämplig metod för att skatta flykttiden för en signal i brus. Metoden ska passa för detta användingsområde och vara robust. Rapporten tar upp tidigare forskning gjord inom flykttidsestimering. De metoder som utvärderas för implementation är det matchade filtret, också kallad korrelationsmetoden, och linjär fas-metoden. Andra aspekter som avvägs och utreds är pulsfrekvens och pulsvaraktighet, ljudets hastighet och brus under vattnet. Metoderna för att skatta flykttid utvärderas genom simuleringar. Det matchade filtret bedöms vara lämpligt baserat på dessa simuleringar och experiment med data inspelad med konsollen. Denna verifikation leder till att algoritmen implementeras på konsollen. Den implementerade algoritmen testas i realtid, resultaten utvärderas och förbättringar föreslås.

Peripheral neuropathy refers to diseases of or injuries to the peripheral nerves in the human body. The damage can interfere with the vital connection between the central nervous system and other parts of the body, and can significantly reduce the quality of life of those affected. In the US, approximately between 15 and 20 million people over the age of 40 have some forms of peripheral neuropathy. The diagnosis of peripheral neuropathy often requires an invasive operation such as a biopsy because different forms of peripheral neuropathy can affect different types of nerve fibers. There are non-invasive methods available to diagnose peripheral neuropathy such as the nerve conduction velocity test (NCV). Although the NCV is useful to test the viability of an entire nerve trunk, it does not provide adequate information about the individual functioning nerve fibers in the nerve trunk to differentiate between the different forms of peripheral neuropathy. A novel technique was proposed to estimate the individual nerve fiber diameters using group delay and simulated annealing optimization. However, this technique assumed that the fiber depth is always constant at 1 mm and the fiber activation due to a stimulus is depth independent. This study aims to incorporate the effect of fiber depth into the fiber diameter estimation technique and to make the simulation more realistic, as well as to move a step closer to making this technique a viable diagnostic tool. From the simulation data, this study found that changing the assumption of the fiber depth significantly impacts the accuracy of the fiber diameter estimation. The results suggest that the accuracy of the fiber diameter estimation is dependent on whether the type of activation function is depth dependent or not, and whether the template fiber diameter distribution contains mostly large fibers or both small and large fibers, but not dependent on whether the fiber depth is constant or variable.

Rapid and deep transport of solutes in soils can potentially pollute groundwater resources. Field estimates of solute leaching depth based on randomized sampling provide extremely variable field average estimates that confound the treatment effects of the leaching study with the high spatial variation of soil hydraulic properties. The purpose of this study was to investigate the spatial scale of variation of solute (Bromide) leaching depth, and apply this scale of variation to study the leaching depth of Bromide as a function of a sinusoidal application of transport causing factors, i.e., rainfall amount, intensity and application time delay. Solute leaching depth varied over different spatial scales. The deepest leaching was observed on plots where the Br center of mass ranged from 19-30 cm depth. Deep leaching occurred with large quantities of low intensity precipitations (5.5 to 6 cm/day) and short time delays (≤ 17 hours), respectively. The hydraulic gradient presented cyclic variation at 8 m wavelength across the 10-30cm depth compartment. Spectral analysis indicated that spatial variation of the leaching depth was mainly affected by precipitation amount and intensity and only a small portion of the leaching depth variation was caused by time delay. Cross-spectral analysis identified common cyclic variation between the Br leaching depth and precipitation amount, intensity and time delay over 32, 32 and 8 m wavelengths, respectively. Simulated Br concentration over depth and horizontal distance and soil water matric potential ψm were in good agreement with experimental observations, the latter revealing a satisfactory Br and water mass balance.

Status epilepticus (SE) is a serious neurological disorder, characterised by prolonged and/or frequent seizure activity. Following SE, a selective and delayed neuronal death (DND) occurs in limbic regions of the brain, particularly in the hippocampus. The objective of this thesis was to investigate the molecular basis of SE-induced DND in the Wistar rat hippocampus. Following the induction of SE, moribund (i.e. dead/dying) neurons were identified by histological staining, DNA fragmentation and an increase in activated microglia. Clusterin, a glycoprotein implicated in apoptotic cell death was also observed to accumulate in the soma and axons of moribund neurons 72-144 hr following SE. Morphological evidence suggested that dying neurons exhibited many of the classical features of apoptosis (i.e. apoptotic body formation, oligo-nucleosomal DNA fragmentation and rapid phagocytosis of debris) and therefore raised the possibility that SE-induced DND might be programmed (i.e. requiring de novo protein synthesis). To investigate this hypothesis I have examined the temporal and anatomical expression of a number of proteins which may have a critical role in SE-induced DND. The expression of the inducible transcription factors (ITFs) was examined as they couple extracellular stimulation to the transcription of late effector gene(s), resulting in long-term phenotypical changes in the neuron and therefore, they may couple SE-inducing stimulation with DND. A high correlation was shown between neurons which exhibited a delayed and prolonged ITFP expression and those which were selectively vulnerable to SE-induced DND (e.g. CA1 and CA3 pyramidal cells and dentate hilar neurons). However, administration of the protein synthesis inhibitor anisomycin following the induction of SE reduced the ITFPs expression, but resulted in an increase in SE-induced DND after 48 hr. However, the levels of brain-derived neurotrophic factor (BDNF)-like immunoreactivity were also shown to attenuate at this time after this procedure. Thus, protein synthesis inhibitors administered following SE may attenuate the level of trophic support and promote cell death. To further investigate the role of the ITFPs in nerve cell death, etoposide, a DNA topoisomerase II inhibitor, which is known to facilitate apoptosis was infused into the hippocampus. The results suggested that a complex ITFP expression occurred which preceded nerve cell death. Moreover, this nerve cell death occurred earlier (12-24 hr) and was not anatomically selective. Furthermore, following the etoposide infusion, clusterin was expressed in the hippocampal pyramidal cells, in the dentate hilar neurons and in the dentate granule cells, however the latter exhibited the strongest BDNF-like immunoreactivity. In summary, circumstantial evidence suggests that the ITFPs may form a critical component in the cascade of events which couple toxic stimulation to nerve cell death. However, this thesis demonstrates that the ITFPs have a complex role in DND, as although the ITFPs may be sufficient to induce DND, they may not always be necessary (e.g. in the absence of sufficient trophic support).

Malaria is caused by mosquito-borne parasites of the genus Plasmodium which were responsible for ~435,000 of deaths annually, with >90% caused by the deadliest species, P. falciparum. Over the last two decades, global implementation of vector control and artemisinin combination therapies have resulted in significant reductions in the global burden of malaria. Of current concern is the spread of multi-drug resistant parasites that have severely limited the efficacy of antimalarials, including front-line artemisinins, highlighting the urgent need to identify new antimalarials for use as treatments. The aim of this thesis was to investigate novel antimalarial development avenues and identify new chemotypes that could be used in the near future as treatments. The macrolide antibiotic azithromycin is known to target the malaria parasites remnant plastid organelle (the apicoplast’s) bacterial-like ribosome and causes slow-killing ‘delayed death’, where the parasite dies in the second replication cycle (4 days). Azithromycin has also been shown to inhibit invading merozoites and kill blood stages within the first replication cycle (2 days) via an unidentified mechanism, proposed to be independent of delayed death. Thus, we hypothesised that azithromycin could be redeveloped into an antimalarial with two different mechanisms of action against parasites: delayed death and quick-killing. Over 100 azithromycin analogues that featured a high proportion of different structural profiles were obtained, leading to improved quick-killing activities over azithromycin. Quick-killing was also confirmed to be completely unrelated to delayed death, as blood stage parasites lacking the apicoplast were equally susceptible to quick-killing of azithromycin and analogues. Two different avenues were also confirmed for azithromycin’s antimalarial re-development: delayed death and quick-killing or quick-killing only, which could be modulated depending on the location of added functional groups. Azithromycin and analogues were found to be active across blood stage development, with only short treatments required to kill parasites. The metabolomics signatures of parasites treated with azithromycin and analogues suggested that quick-killing acts multi-factorially, with the parasite’s food vacuole and mitochondria being likely targets. Finally, in vitro activities of two subtypes of tri-peptide proteasome-like inhibitors, vinyl sulfone and aldehydes, were addressed against P. falciparum and the zoonotic malaria parasite P. knowlesi. All compounds exhibited low-nanomolar activities against both Plasmodium spp. and showed excellent selectivity for parasites over human cells, suggesting these inhibitors provide viable chemical scaffolds for optimisation. There was no evidence of increased protein ubiquitination upon treating parasites with these compounds, suggesting they do not target the proteasome. We also investigated whether hypoxia inducible pro-drug proteasome-like inhibitors could be used to reduce host toxicity of antimalarials. However, these pro-drugs could be not activated in in vitro culture conditions and there was limited evidence suggesting this strategy would be applicable in malaria. These studies build on previous findings on the drug-killing efficacy, mechanism of action and possible application of redeveloping azithromycin analogues as new and improved antimalarials. I also identified new proteasome inhibitor-like scaffolds as starting points for further development. This body of work provides thorough biological characterisation of a panel of compounds that could lead to new avenues for antimalarial development.
Thesis (Ph.D.) -- University of Adelaide, School of Biololgical Sciences, 2020

碩士
中國文化大學
心理輔導研究所
95
This research was about the resilience process of one Christian mother facing developmental delay child’s death. The purposes were to understand the resilience process and the connotation of the resilience. The participant was a member from the Angel Heart’s Family Social Welfare Foundation. Her child had passed away about a year and a half ago. This research took the approach of narrative analysis; using in-depth interview collected data and validated the results through triangulation. The results were displayed with the form of story that expressed its narrative style and special context. Five turning points were shown: keep off distress feeling, accept the new job descending from God speaking to her, in-depth awareness emerge through interview, the meaning of life resurrection through her child’s death, discover that she can still connection with her child closely. There were six themes emerged in the resilience process: resources intervention, coping emotion, facing challenge, reconstruct meaning, hope for the future. These themes could also perform in phases. In the resilience process, the essential of dynamic come out in evidence, the strength of environment took an important role in guided the grief process to resilience. The strength of personality emerged the most, and the strength of spirituality pushed forward individual result in transformation and developed extended driving force firmly. In the connotation of the resilience, this research dividing resilience into the strength of personality, environment and spirituality. Five categories emerge in the strength of personality: ability, characteristic, cognition, behavior, experience. Three categories emerge in the strength of environment: intervention of support network, provide the resources, change of the context. Three categories emerge in the strength of spirituality: identification of religion, intervention from God, spiritual religion behavior. 46 contexts of resilience were emerged from 11categories above. All the results were being discussed. On the side, the study raised several feasible suggestions for the practice as well as research of the counseling according the findings of this research. Finally, researcher provides own opinions to the Christian bereavement mothers and the society. At last, researcher reviewed the process of studying this research in envoi.

Dissertação de Mestrado apresentada no Instituto Superior de Psicologia Aplicada para obtenção de grau de Mestre na especialidade de Psicologia Clínica
Esta revisão de literatura visa ilustrar o fenómeno da parentalidade tardia na perspectiva da psicologia, assim como dos aspectos psicologizantes presentes nestas famílias e nas suas relações, numa tentativa de completar o vazio da literatura recente, que não acompanha a crescente incidência desta tendência nos países industrializados. Explorámos os factores psicológicos e ambientais que possam explicar a tomada de decisão, as motivações, as dificuldades e necessidades associadas à criação e educação de um filho com 40 anos a menos que os pais. Uma revisão de literatura sobre este fenómeno deverá abranger uma multiplicidade de variáveis, que enquadrem a parentalidade tardia no seu contexto, aprofundando as questões de base relacionadas com a gravidez e a adaptação à parentalidade, reflectindo sobre o comportamento dos pais tardios na transição para a parentalidade e insistindo na apreciação do impacto da idade parental no desenvolvimento dos filhos. Apesar da escassez de literatura sobre o impacto psicológico deste fenómeno nos membros das famílias, a grande maioria dos estudos parece relacionar o adiamento da parentalidade com alterações e mudanças do papel da mulher na sociedade, nomeadamente com o investimento na carreira, o que, concomitantemente, poderá trazer algumas incompatibilidades na conjugação de papéis diferentes, não parecendo ser, contudo, esse factor determinante para as relações intergeracionais. Há uma tendência para relacionar a idade parental com comportamentos sobre-protectores, o que, conjugado com a possibilidade de uma angústia de morte mais evidente dos filhos tardios, poderá resultar em maiores dificuldades de autonomia, separação e independência.
The present research’s aim is to illustrate the psychological perspective on the delayed parenthood phenomena, as well on the psychologically predisposing aspects of the families and its relationships, in an attempt to fulfill the void left by the modern literature that does not keep up with this ever more often tendency in the industrialized nations. We have explored the psychological and environmental factors that can provide some explanation on the decision making, the motivations, the difficulties and necessities related with the nurture and education of a son 40 years younger than his parents. So it can take account of the multiple variables that provide the social layout of the delayed parenthood, a thorough literary review on this phenomenon must examine the basic issues related with pregnancy and parenthood adaptation by reflecting on the parental behavior during the transition to parenthood and stressing out the impact of the parents age on the child’s development. Even though the literature fails to clarify the psychological impact of this phenomenon on the family members, the majority of studies seem to relate the parenthood postponement with the changes on the woman’s role in society, namely with her career investment, which, therefore can originate incompatibilities during the conjugation of the different roles, although there are no evidences that this factor should be determinant to the intergenerational relationships. The tendency to relate parental age with over-protective behaviors, could, if associated with the possibility of death angst present in late children, aggravate the difficulties in separation, autonomy and independence.
Nesta investigação, estudou-se a parentalidade tardia na tentativa de completar o vazio da literatura recente, que não acompanha a crescente tendência do aumento deste fenómeno nos países industrializados. Explorámos os factores psicológicos e ambientais que possam explicar a tomada de decisão, as motivações, as dificuldades e necessidades associadas à criação e educação de um filho com 40 anos a menos que os pais. Entrevistámos quatro mães, quatro filhos e uma prima e encontrámos características comuns nestas famílias, nomeadamente, que a parentalidade tardia ocorre essencialmente por duas vias: como um “acidente”/“surpresa”, ou de forma planeada, em que houve decisão de adiar (casamento tardio). Os pais tardios tendem a exibir comportamentos sobre-protectores, promovendo a dependência parental e o isolamento social, contribuindo, a longo-prazo, para sentimentos de “não-pertença” face a contextos extra-familiares e a situações sociais e relacionais. Os filhos de pais tardios tendem a introjectar a angústia de morte dos pais, tornando-se mais vulneráveis às ameaças de abandono, e, para se defenderem da angústia de separação, vão construindo um superego severo, auto-punitivo, que se manifesta sob o medo de desiludir, de chatear e de evitar sobrecarregar os pais, expresso num discurso hiper-moralizador e disciplinado. A ansiedade filial pronuncia-se desde muito cedo – mesmo quando os pais ainda não precisam de ajuda – como manifestação do receio de perder os pais precocemente, obrigando os sujeitos a inverter os papéis e a assumir como suas responsabilidades que outrora foram dos seus pais.
In the current investigation, we approach the delayed parenthood phenomenon in an attempt to fulfill the void left by the modern literature that does not keep up with this ever more often tendency in the industrialized nations. We have explored the psychological and environmental factors that can provide some explanation on the decision making, the motivations, the difficulties and necessities related with the nurture and education of a son 40 years younger than his parents. For that matter we interviewed four mothers, four sons and one cousin and we came across similar characteristics between these families, namely, the fact that the delayed parenthood derives from one of two circumstances: from an “accident”/ “surprise”, or in the form of a planned way, in which there was a decision to postpone (delayed marriage). These parents tend to show over-protective behaviors, promoting the parental dependence and the social isolation and, on a long-term, contributing to the rise of a sense of “not belonging” in social and relational context and in the extra-familiar milieu. The sons of these late parents tend to introject the parents’ death angst, becoming more vulnerable to the threats of abandonment, which, in order to defend themselves from the separation anxiety, results in the rise of a sever and punitive super-ego that manifests itself in the fear of upsetting and letting down the parents, in an attempt to avoid overloading the parents and through a hyper-moralizing and disciplined speech. The filial anxiety expresses itself since early childhood – even when the parents don’t need help – as a manifestation of the fear of losing the parents before time, forcing the individuals to reverse the roles and to assume as their own the responsibilities that once belonged to the parents.

L’hémorragie sous-arachnoïdienne (HSA) est une pathologie redoutable résultant fréquemment de la rupture d’un anévrisme intracrânien. Elle est associée à une mortalité élevée et d’importants déficits neurologiques. Le saignement entraîne l’augmentation de la pression intracrânienne, la diminution du flux cérébral sanguin, l’apparition de l’inflammation cérébrale et la mort neuronale. Ces évènements de la phase d’insulte cérébrale précoce (72 premières heures) conditionnent le devenir du patient. Le vasospasme était initialement considéré comme la cause principale des ischémies cérébrales retardées (DCI), mais sa diminution pharmacologique n’a montré aucun bénéfice pour les patients. Cependant, l’infiltration rapide des leucocytes dans le SNC suivant le saignement semble impliquée dans le développement des DCI. Notre hypothèse est que l’activation précoce du système immunitaire à la suite de l’HSA est responsable de la mort neuronale retardée et de la survenue des déficits constatés chez les patients souffrant d’HSA. Nos buts étaient de caractériser la contribution des leucocytes dans l’inflammation cérébrale et la mort neuronale dans un modèle murin d’HSA, de moduler cette inflammation par l’utilisation de la protéine MFG-E8, une protéine anti-inflammatoire favorisant la clairance apoptotique, et de confirmer la présence d’une signature immunologique comparable chez les patients HSA. Notre modèle murin nous a permis d’induire chirurgicalement l’HSA et d’injecter par voie intrapéritonéale la protéine MFG-E8. La composition cellulaire du sang (humain et murin) et du cerveau des souris a été analysée par cytométrie en flux. Le plasma (humain et murin) et le liquide céphalo-rachidien (LCR) des patients ont été analysés par dosage cytokinique. Certains cerveaux de souris étaient inclus en paraffine pour l’imagerie par microscopie confocale. La lignée cellulaire de microglie nous a permis d’étudier la modulation de la capacité de phagocytose et de production de ROS par l’exposition au sérum ou au LCR de patients HSA. Dans une première étude, nous avons démontré le rôle de l’inflammation cérébrale précoce dans le développement de la mort neuronale et des symptômes chez les souris HSA. Nous avons également caractérisé la présence de marqueurs inflammatoires systémiques chez les patients HSA. Dans une deuxième étude, nous avons montré que le traitement par la protéine MFG-E8 chez les souris HSA entraînait la diminution de l’inflammation périphérique ainsi que de la présence des marqueurs M1, de l’activation des astrocytes et de la mort neuronale dans le cerveau aboutissant à la diminution de la sévérité des symptômes. L’étude de l’activation immunitaire chez les patients HSA, nous a permis d’observer une signature immunologique similaire à notre modèle murin. Nous avons montré que les patients HSA présentaient une augmentation des cellules immunitaires innées et une immunodépression lymphocytaire en comparaison avec des donneurs sains. Nous avons également décrit l’importance du grade et du genre des patients par la caractérisation d’un profil inflammatoire plus sévère chez les patients hauts gradés et chez les hommes. Finalement, nos résultats confirment l’existence d’une signature immunologique similaire entre les patients HSA et notre modèle murin aboutissant, dans les deux cas, à l’augmentation de l’activation de l’inflammation systémique et cérébrale. Cette signature immunologique est dépendante du sexe et du grade des patients. La diminution de la gravité des symptômes par le traitement avec la protéine MFG-E8 dans notre modèle souris confirme l’implication incontestable de l’inflammation dans l’apparition des déficits moteurs secondaires à la mort neuronale, et le potentiel thérapeutique de cette protéine MFG-E8 dans le développement de nouvelles thérapies.
Subarachnoid hemorrhage (SAH) is a redoubtable pathology resulting frequently from the rupture of an intracranial aneurysm. It is associated to an important mortality and severe neurologic deficits. The bleeding leads to an increase in the intracranial pressure, to a decrease in cerebral blood flow, to the development of cerebral inflammation and to neuronal death. These events of early brain injury (first 72 hours) determine the patient’s prognosis. The vasospasm was first thought to be the main cause of delayed cerebral ischemia (DCI), but its pharmacological decrease was not being associated to any benefits for the patients. However, rapid leucocytic infiltration in the CNS secondary to the bleeding seems implicated in the development of DCI. Our hypothesis is that the early immune system activation in SAH is responsible for delayed neuronal death and for the onset of symptoms in SAH patients. Our goals were to characterize the contribution of leucocytes in cerebral inflammation and neuronal death in our SAH mice model, to modulate the inflammation by using MFG-E8 protein, an anti-inflammatory protein promoting the apoptotic clearance, and to confirm this similar immunologic signature in SAH patients. Our mouse model allows us to surgically induce SAH and to inject the MFG-E8 protein by intraperitoneal injection. The cellular composition of blood (human and mouse) and of mouse brains were analyzed by flow cytometry. The plasma (human and mouse) and the cerebrospinal fluid (CSF) were analyzed by cytokine assay. Some mice brains were paraffin-embedded for confocal microscopy imaging. Microglia cell lines allowed us to evaluate the modulation of phagocytosis and reactive oxygen species (ROS) production secondary to the exposition to SAH patients’ serum and CSF. In the first study, we have demonstrated the impact of early cerebral inflammation on neuronal death and the occurrence of symptoms in SAH mice. We also have characterized the presence of systemic inflammatory markers in SAH patients. In the second study, we have shown that MFG-E8 protein treatment in our SAH mice model is linked to a decrease in peripheric inflammation as well as to a decrease of M1 markers, astrocytic activation and neuronal death in the brain leading to a decrease of symptoms severity. iv The study of immune activation in SAH patients allowed us to observe an immune signature like in our mouse model. We have revealed that SAH patients have an increase in innate immune cells and in lymphocytic immunosuppression in comparison to healthy donors. We have also described the importance of gender and SAH grade by the characterization of a more severe inflammatory profile in high-grade and in male patients. To conclude, our results confirm the existence of a similar immune signature between SAH patients and our mouse model leading in both cases to an increase in systemic and cerebral inflammation. This immunologic signature depends on the patients’ gender and on the grade of SAH. The decrease of symptom severity with MFG-E8 protein treatment in our mice model confirms the unquestionable implication of inflammation in the occurrence of motor deficits secondary to neuronal death and the therapeutic potential of MFG-E8 for the development of new therapies.

L’accouchement et ses conséquences demeurent une des principales causes d’incapacité et de décès pour les femmes dans les pays en développement et comprendre l’utilisation tardive des soins obstétricaux d’urgence au moment d’une complication obstétricale constitue un véritable défi en santé publique. La présente étude qualitative relate l’expérience d’accouchement difficile au Mali, en milieu rural. Dans un contexte de pluralité de systèmes de soins, l’objet de cette étude consiste à déterminer les raisons de l’arrivée tardive des femmes au centre de santé de 1ier ligne, à saisir la compréhension qu’elles ont de l’utilisation des services sanitaires, enfin à reconstruire les processus de prise de décisions de recourir aux soins modernes. Cette étude s’inscrit dans une démarche de type ethnographique. Des entretiens semi dirigés et l’observation des interactions entre les femmes et les professionnels de santé ont constitué le corpus de données. Nous retenons qu’une série de facteurs entrent en jeu pour comprendre le problème de l’utilisation tardive des soins obstétricaux d’urgence. Des contraintes exogènes liées à la distance et l’immédiateté de l’urgence obstétricale c'est-à-dire la mobilisation des ressources et des moyens de transport, accompagnées de contraintes endogènes telles que la subordination des femmes au consentement familial, la perception de la qualité des soins et de la compétence des soignants, influencent systématiquement le choix de recourir aux soins modernes. Le phénomène de gestion collective de la complication obstétricale s’inscrit dans un contexte d’intéractions complexes où l’opinion des femmes est totalement minimisée.
Childbirth and its consequences remain a leading cause of disability and death for women in developing countries and the late use of emergency obstetric care during obstetric complications is a challenge in public health. This qualitative study describes the experience of difficult childbirth in Mali, in rural areas. In a context of multiple systems of care, the purpose of this study is first to determine the reasons for the late arrival of women in the health center, to grasp their understanding of the use of health care services, and finally to reconstruct the decision making process to use modern health care. This study is part of an ethnographic approach. Semi-structured interviews and observation of interactions between women and health professionals were conducted. Numerous factors come into play to understand the problem of late use of emergency obstetric care. Exogenous constraints related to the distance and immediacy of the obstetric emergency that is to say the mobilization of resources and means of transport, influence the choice of women to use modern obstetric cares. In the same way, endogenous constraints such as the subordination of women in the family consent, perception of quality of care and skill of caregivers, systematically influence the choice to use modern health care. Collective management of obstetric complications we have observed, occur in a context where the opinion of women is completely minimized.

The utilization of in vitro tests with a tiered testing strategy for detection of mild ocular irritants can reduce the use of animals for testing, provide mechanistic data on toxic effects, and reduce the uncertainty associated with dose selection for clinical trials. The first section of this thesis describes how in vitro methods can be used to improve the prediction of the toxicity of chemicals and ophthalmic products. The proper utilization of in vitro methods can accurately predict toxic threshold levels and reduce animal use in product development. Sections two, three and four describe the development of new sensitive in vitro methods for predicting ocular toxicity. Maintaining the barrier function of the cornea is critical for the prevention of the penetration of infections microorganisms and irritating chemicals into the eye. Chapter 2 describes the development of a method for assessing the effects of chemicals on tight junctions using a human corneal epithelial and canine kidney epithelial cell line. In Chapter 3 a method that uses a primary organ culture for assessing single instillation and multiple instillation toxic effects is described. The ScanTox system was shown to be an ideal system to monitor the toxic effects over time as multiple readings can be taken of treated bovine lenses using the nondestructive method of assessing for the lens optical quality. Confirmations of toxic effects were made with the utilization of the viability dye alamarBlue. Chapter 4 describes the development of sensitive in vitro assays for detecting ocular toxicity by measuring the effects of chemicals on the mitochondrial integrity of bovine cornea, bovine lens epithelium and corneal epithelial cells, using fluorescent dyes. The goal of this research was to develop an in vitro test battery that can be used to accurately predict the ocular toxicity of new chemicals and ophthalmic formulations. By comparing the toxicity seen in vivo animals and humans with the toxicity response in these new in vitro methods, it was demonstrated that these in vitro methods can be utilized in a tiered testing strategy in the development of new chemicals and ophthalmic formulations.