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1
Hofsten, Jonas von. "Developmental and reproductive regulation of NR5A genes in teleosts." Doctoral thesis, Umeå : Univ, 2004. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-374.
Texte intégral
2
Giedt, Michelle Suzanne. "JAK/STAT SIGNALING REGULATES GAMETOGENESIS AND AGE-RELATED REPRODUCTIVE MAINTENANCE." UKnowledge, 2018. https://uknowledge.uky.edu/biology_etds/52.
Texte intégralRésumé :
Cell signaling is central to integration of internal and external cues that regulate development and homeostasis. Most development is thought of as pre-adult, but limited developmental processes occur in adults. Gametogenesis incorporates elements of both these facets, with a distinct developmental plan for gamete synthesis which is regulated by integration of homeostatic inputs such as nutrient status, and environmental cues. Signaling pathways integrate and transduce information from these cues to evoke a response. A decline in homeostasis and subsequent cues occurs over time, in the case of reproductive tissues leading to a progressive loss of fertility. The Janus Kinase and Signal Transducer and Activator of Transcription or Jak/Stat signaling pathway is conserved between vertebrates and invertebrates and is necessary for numerous functions needed to maintain organism and reproductive homeostasis, as well as contributing to various developmental events. The pathway in the fruit fly Drosophila melanogaster, is composed of a single receptor, Domeless, one Janus kinase, Hopscotch, one known effector, Stat92E, and the Unpaired family of ligands consisting of Upd, Upd2, and Upd3. Jak/Stat signaling is highly pleiotropic in both sexes with involvement in homeostasis and reproduction, making it an ideal model for studying the role of signaling in reproductive aging. Reduction of pathway activity in females results in a higher proportion of unfertilized eggs, which increases with age, and in males leads to a premature onset of infertility. Central to both is integration through cell signaling to evoke an appropriate response. This dissertation explores two of the requirements for Jak/Stat signaling: the pleiotropic requirement for Jak/Stat activity during oogenesis and male reproductive maintenance.
Jak/Stat functions from the beginning of oogenesis, in the stem cell niche. From there it participates in multiple functions including specification of a subset of somatic cells called the border cells through the polar cells, a pair of cells at either pole of the egg. Pathway stimulation in the border cells drives their migration with the polar cells to the oocyte boundary, where the polar cells each form an extension in a coordinated manner into the micropyle, the means for sperm entrance during fertilization. Loss of Jak/Stat activity in the border cells prevents border cell migration. While border cell migration has been well studied, polar cell involvement after completion of border cell migration is less well known. To investigate the requirements for polar cell activity and Jak/Stat activity after the completion of border cell migration, we reduced Jak/Stat signaling in the polar cells which, while having no effect on border cell migration, results in blocked micropyles due to loss of coordination of extensions during their outgrowth. Reduced function in the polar cells did not significantly affect expression of adhesion molecules. But, the loss of Stat92E is phenocopied by loss of DE-cadherin. Hence, these results indicate a previously unknown autocrine requirement for Jak/Stat activity in the polar cells.
The testes also have a continuous requirement for Jak/Stat activity for stem cell maintenance and differentiation of the germline into mature sperm. Reproductive maintenance not only requires sustained production of gametes, but reproductive tissues are also subject to deterioration of homeostatic functions that contribute to organismal aging. Males from thirty-nine lines of the Drosophila Genetic Reference Panel (DGRP), a panel of inbred, fully sequenced lines, were screened for age at infertility. Data were used to perform a genome-wide association study (GWAS) to identify the genetic architecture of reproductive aging. Candidate variants associated with cell signaling regulators, genes with functions in maintaining cell homeostasis, and organism behavior were uncovered. Notably, several SNPs fell in and near Ptp61F, a negative regulator of Jak/Stat activity. While variants in the primary components of the Jak/Stat pathway were not identified, the general classes of candidate loci functions reflect the requirements for homeostasis, metabolism, and development that have been shown by other studies examining the genetics of aging and fecundity. Thus, we show that Jak/Stat has an amazing amount of pleiotropy that encompasses both the real-time functions of fertility and the time related process of aging.
3
Van, Praet Oliver. "Co-expression and interaction of cubilin and megalin in the adult male rat reproductive system." Thesis, McGill University, 2002. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=29485.
Texte intégralRésumé :
Cubilin is a peripheral membrane protein that co-operates with the endocytic receptor megalin to mediate the endocytosis of ligands in various polarized epithelia. Megalin is expressed in the male reproductive tract where is has been implicated in the process of sperm membrane remodeling. A potential role for cubilin in the male reproductive tract has not been explored. Using RT-PCR, we found that cubilin and megalin mRNAs are expressed in the efferent ducts, corpus and cauda epididymides, and proximal and distal vas deferens. Immunohistological analysis revealed that cubilin was expressed in non-ciliated cells of the efferent ducts, principal cells of the corpus and cauda epididymides and vas deferens. Electron microscopic immunogold labeling showed cubilin in endocytic pits, endocytic vesicles and endosomes of these cells. The expression profile of cubilin in the male reproductive tract was coincident with that of megalin except in principal cells of the caput epididymidis. Double immunogold labeling showed that cubilin and megalin co-localized with one another within the endocytic apparatus and recycling vesicles of efferent duct cells. Neither protein was found in lysosomes. Injection of RAP, an antagonist of megalin interaction with cubilin, reduced the level of intracellular cubilin in cells of the efferent ducts and vas deferens. In conclusion, cubilin and megalin are co-expressed in cells of the epididymis and vas deferens and the endocytosis of cubilin in these tissues is dependent on megalin. Together, these findings highlight the potential for a joint endocytic role for cubilin and megalin in the male reproductive tract.
4
Mercurio, S. "ROLE OF STEROID HORMONES IN ECHINOID REPRODUCTIVE BIOLOGY." Doctoral thesis, Università degli Studi di Milano, 2014. http://hdl.handle.net/2434/230752.
Texte intégralRésumé :
Echinoid reproductive cycle has been extensively studied in several species but the mechanisms regulating gametogenesis processes are still scarcely understood. Apart from environmental factors, different research have suggested a steroid role in gonad maturation and growth. Particularly, in echinoderms steroid involvement in reproduction has been suggested by both studies on seasonal changes of steroid levels during the gonadal cycle and experiments of hormone administration. Nevertheless, the steroid function in echinoid reproductive processes has not been clearly identified, probably due to the low number of studies and the big variability of results reported. Thus, the main aim of this research project was to shed light on echinoid endocrinology and, in particular, to clarify the involvement of sex-steroid hormones in sea urchin reproductive biology. This was achieved employing both in vivo and in vitro approaches.
First of all, considering the lack of studies on the development of effective cell cultures from echinoderm gonads, primary cell cultures from ovaries of the edible sea urchin Paracentrotus lividus were developed. Ovary cell phenotypes, present in culture, were identified and characterized by different microscopic techniques. Although cell cultures could be produced from ovaries at all stages of maturation, the cells appeared healthier and viable, displaying a higher survival rate, when ovaries at early stages of gametogenesis were used. In terms of culture medium, ovarian cells were successfully cultured in modified Leibovitz-15 medium, whereas poor results were obtained in Minimum Essential Medium Eagle and Medium 199. Different substrates were tested but ovarian cells completely adhered only on poly-L-lysine. To improve in vitro conditions and stimulate cell proliferation different serum-supplements were tested. Fetal Calf Serum and an originally developed Pluteus Extract resulted to be detrimental to cell survival, apparently accelerating processes of cell death. In contrast, cells cultured with sea urchin Egg Extract appeared larger and healthier, displaying an increased longevity that allowed to maintain them for up to 1 month. Overall this study provides new experimental bases and procedures for producing successfully long-term primary cell cultures from sea urchin ovaries, providing a simple and versatile experimental tool for research in echinoderm reproductive biology.
Subsequently, in vivo and in vitro experiments, specifically addressed to determine possible 17β-estradiol (E2) and testosterone (T) involvement in echinoid reproduction, were performed. An in vivo long-term experiment of steroid dietary administration was performed in adult specimens of P. lividus. The experimental plan was specifically designed in order to reduce individual variability and synchronize the experimental animals at the same starting maturative condition. We analysed and compared different reproductive parameters (Gonad Index, Maturative Index and maturative stages distribution) in 4 experimental groups: control group (CTL), E2 and T groups fed with pellets containing respectively 17β-estradiol and testosterone, and E2-4 weeks group fed with control pellets for the first 4 weeks and then treated with 17β-estradiol. This latter was chosen in order to verify the existence of a specific E2-sensitive gametogenic stage, as proposed in different asteroid species.
Possible steroid effects on P. lividus female reproduction was also investigated with an in vitro approach. Cells, isolated by ovaries in the same maturative conditions considered in the in vivo experiments, were cultured in presence of E2 and T physiological concentrations for 2 weeks. Effects on ovarian cell morphology and behaviour were investigated. In addition, steroid regulation of the Major Yolk Protein (MYP) expression was analyzed 24 and 48 hours after E2 and T exposure. According to our results, E2 and T do not markedly influence echinoid gonad maturation and, particularly, they do not promote gamete maturation. Hormonal dietary administration did not induce striking variations in the considered reproductive parameters and no effect was observed also when males and females were analyzed separately. In addition, no specific maturative stage sensitive to E2 was found, suggesting the existence of different hormonal mechanisms in asteroids and echinoids. Similar considerations could be reported taking into account the in vitro experiments. E2 and T exposure did not affect ovarian cell size and behaviour nor MYP expression.
The obtained results suggest that these hormones are not directly involved in either gamete maturation, as demonstrated for vertebrates, or in vitellogenesis processes, as reported for several asteroid species. However a possible involvement of steroids in echinoid physiology cannot be completely excluded and their role in the regulation of lipid metabolism and protein synthesis during the different reproductive stages should be strongly considered as suggested by several authors.
Further specific research on steroid hormone mode of action, physiological function and metabolism are therefore needed to completely understand echinoid reproduction and endocrinology.
5
Chen, Hsin-Ying. "T CELL RESPONSE TO INFECTION BY THE PORCINE REPRODUCTIVE AND RESPIRATORY SYNDROME VIRUS." NCSU, 2005. http://www.lib.ncsu.edu/theses/available/etd-01062005-170050/.
Texte intégralRésumé :
The purpose of the research has been to characterize the response of naïve or virus-specific swine T lymphocytes from different lymphoid compartments to Porcine Reproductive and Respiratory Syndrome Virus (PRRSV). Peripheral blood mononuclear cells (PBMCs), tracheobronchial lymph node (TLN) and lateral retropharyngeal lymph node (LLN) cells were labeled with PKH67 green fluorescence dye to measure cell proliferation and surface phenotypes were examined using anti-CD4, anti-CD8 or anti-CD25 monoclonal antibodies. Stimulation with Concanavalin A was also included as a positive control. Our results show that potential virus-specific T lymphocytes were found in the peripheral blood, although no cell proliferation was found in cultures of lymph node cells. We did find that the percentages of CD8+ T cells in cultures of lymph node cells from the virus-infected pig increased after in vitro stimulation with the Powell virus compared to the lymph node cells cultured in media only, suggesting that CD8+ T lymphocytes may play a role in the virus clearance and immune memory to PRRSV.
6
Dias, Tânia Isabel Rodrigues Amaral. "Evaluation of cell death markers and reproductive parameters in models of diabetes mellitus." Master's thesis, Universidade da Beira Interior, 2012. http://hdl.handle.net/10400.6/1121.
Texte intégralRésumé :
Diabetes mellitus (DM) represents one of the greatest threats to modern global health and its incidence is rapidly rising worldwide. It describes a metabolic disorder characterized by hyperglycaemia resulting from defective insulin secretion, resistance to insulin action, or both. There are two types of DM, type-1 DM (T1DM) and type-2 DM (T2DM), both associated with male infertility. T1DM is associated with insulin deprivation and although the overall in vivo effects in the reproductive function is well known, there is a lack of studies concerning about the insulin control over the physiological functions of cells from the reproductive system. Importantly, the in vivo studies are often focused after the disease is fully establish, but it is known that a prediabetic state, which is characterized by insulin resistance, precedes the development of DM, especially T2DM. With our work, we aimed to further investigate the association between DM and male infertility by analyzing several apoptotic markers and reproductive parameters. To do so, we simulated type 1 DM in cultured rat Sertoli cells and analyzed the mRNA and protein expression levels of several cellular markers involved in the mitochondrial apoptotic pathway. We also developed an animal model of prediabetes to evaluate the effect of this pathological state in the reproductive parameters as well as in the mitochondrial apoptotic pathway.
Our results lead us to suggest that insulin interferes with the interaction between pro and anti-apoptotic proteins. As the interaction of these proteins decide the cell fate and exert a strict control over the apoptotic signaling, insulin has a key role in the maintenance of the spermatogenesis. Our rat model shared many of the clinical and metabolic characteristics of the prediabetic state observed in humans such as glucose resistance and progression from normoglycaemia/normoinsulinemia to moderate hyperglycaemia/hyperinsulinemia due to food intake. This prediabetic state induced important alterations in cauda epididymis spermatozoa morphology showing that these animals may develop subfertility or fertility problems. The apoptotic signalling in cauda epididymis spermatozoa of the high-energy (HED) fed animals presented lower Bax mRNA levels and lower cytC protein levels although the apoptotic endpoint, caspase-3 activity, was not altered. This suggests that the apoptotic process may be controlled by other mechanisms rather than the mitochondrial pro-apoptotic proteins, such as the anti-apoptotic cellular systems.
Those two experimental models led us to conclude that the subfertility/infertility problems caused by DM may be mediated by insulin, which has an important effect in the regulation of the interaction between pro and anti-apoptotic proteins and special attention must be taken in the prediabetic state where crucial alterations in rat spermatozoa occurred. Due to the rising incidence and associated complications of DM and male infertility it is crucial to further investigate in these two systems to isolate possible mechanisms and evaluate the overall effects as a strategy to develop possible therapeutics.<br>A diabetes mellitus (DM) representa uma das maiores ameaças à saúde global moderna
e a sua incidência está a aumentar rapidamente em todo o mundo. Esta doença consiste numa
desordem metabólica, caracterizada por hiperglicemia, resultante de uma secreção
defeituosa de insulina, resistência à ação da insulina ou ambas. Existem dois tipos de DM, tipo
1 e tipo 2, ambas as condições relacionadas com a infertilidade masculina. A DM tipo 1 está
associada com a privação de insulina e embora os efeitos globais in vivo na função
reprodutiva sejam bem conhecidos, há uma falta de estudos relativamente ao controlo da
insulina sobre as funções fisiológicas das células do sistema reprodutivo. Sobretudo, os
estudos in vivo são frequentemente feitos depois de a doença estar completamente
estabelecida, mas sabe-se que há um estado pré-diabético, caracterizado por resistência à
insulina, que antecede o desenvolvimento da DM, especialmente da DM tipo 2. Com o nosso
trabalho, pretendemos investigar mais profundamente a ligação entre a DM e a infertilidade
masculina, através da análise de vários marcadores de morte celular e de parâmetros
reprodutivos. Para isso, simulámos o estado de DM tipo 1 humano em células de Sertoli de
rato e analisámos os níveis de expressão de mRNA e proteína de vários marcadores de morte
celular envolvidos na via mitocondrial. Por outro lado, também desenvolvemos um modelo
animal de pré-diabetes de modo a reproduzir esta condição patológica e avaliar alterações
produzidas nos parâmetros reprodutivos, assim como nos níveis de expressão de mRNA e
proteína de marcadores de morte celular envolvidos na via mitocondrial.
Os resultados obtidos levam-nos a sugerir que a insulina interfere com a interação entre
proteínas pró- e anti-apoptóticas. Uma vez que esta interação pode decidir o destino celular
e exercer um controlo rigoroso sobre a sinalização apoptótica, a insulina terá um papel chave
na manutenção da espermatogénese. O modelo de rato utilizado compartilhava muitas das
características clínicas e metabólicas do estado pré-diabético observado em humanos, tais
como resistência à glucose e a progressão de normoglicemia/normoinsulinemia para
hiperglicemia/hiperinsulinemia moderada devido à ingestão de alimentos. Este estado prédiabético induziu alterações significativas na morfologia dos espermatozoides da cauda do
epidídimo, mostrando que esses animais poderão desenvolver problemas de subfertilidade ou
de fertilidade. Na sinalização apoptótica na cauda do epidídimo, os animais sujeitos à dieta
de alta energia (HED) apresentaram níveis mais baixos de mRNA Bax e da proteína citocromo
C, embora a avaliação quantitativa do endpoint apoptótico, a atividade da caspase-3, não
tenha evidenciado quaisquer alterações entre a situação HED e controle. Isto sugere que o
processo apoptótico pode ser controlado por outros mecanismos que não somente os proteicos
pró-apoptóticos mitocondriais, como por exemplo os sistemas anti-apoptóticos celulares. Os resultados obtidos com estes dois modelos experimentais levam-nos a concluir que
os problemas de subfertilidade/infertilidade causados pela DM podem ser mediados pela
insulina, que tem um efeito importante na regulação da interação entre proteínas pró e antiapoptóticas e, por esse motivo, deverá ser dedicada uma atenção especial às disfunções
ocorridas no estado pré-diabético, onde observámos alterações cruciais na morfologia de
espermatozoides epididimais de ratos. Devido à crescente incidência da DM e às complicações
associadas ao nível da infertilidade masculina é fundamental aprofundar o conhecimento
nestes dois sistemas, de modo a isolar possíveis mecanismos envolvidos e a avaliar os efeitos
globais, como uma estratégia para desenvolver possíveis abordagens terapêuticas.
7
Ismail, Rubina Siddiqi. "Expression, hormonal regulation and function of Kit ligand, the ligand for thec-Kit receptor, in the rat reproductive system." Thesis, University of Ottawa (Canada), 1998. http://hdl.handle.net/10393/4364.
Texte intégralRésumé :
Mice having mutations at the Kit and Mast Cell Growth Factor loci suffer from severe development abnormalities in gametogenesis suggesting that the protein products produced at these loci, Kit receptor and kit ligand (KL), serve an essential role in the reproductive system. The purposes of this study were: (1) to identify the expression Kit and KL in the ovary which is the organ in which gametogenesis occurs in the female animal, (2) to determine the expression of Kit and KL in the female reproductive tract, (3) to determine the influence of hormones and growth factors on the expression of Kit and KL, and (4) to identify the functional significance of KL/Kit interactions in the reproductive system. Kit receptor expression was identified in the oocytes of antral follicles and KL expression was seen in granulosa cells in growing and antral follicles. In antral follicles, KL expression was greatest in cumulus granulosa cells and only those mural granulosa cells closest to the oocyte and antral cavity. Low KL expression was seen in the mural granulosa cells closest to the basement membrane. This pattern of KL expression in antral follicles suggested a role for KL in oocyte function. There was an inhibition of oocyte meiotic maturation when oocytes from antral follicles were cultured in the presence of KL compared to untreated oocytes. Luteinizing hormone (LH) is the key stimulator of oocyte maturation within the follicle. In vivo stimulation with LH resulted in a loss of KL expression in cumulus granulosa cells and a shift in the production of membrane-bound KL to more soluble KL in mural granulosa cells. Thus, LH-stimulated oocyte maturation may be partly mediated by the removal of KL in the cumulus granulosa cells and the production of a potentially less active form of KL by mural granulosa cells. The expression of KL in granulosa cells was upregulated at the transcript level by treatment with FSH, LH, or both in vivo and in vitro, as well as with membrane-permeable analogues of cyclic adenosine monophosphate (cAMP), whereas KL mRNA expression decreased with steroid hormone stimulation. KL but not Kit mRNA expression was seen in the ovarian surface epithelium. KL expression in ovarian surface epithelium derived-cells was influenced by both dibutyryl cAMP and transforming growth factor-$\beta$ stimulation. These studies would indicate that the ovarian surface epithelium is a site of KL production and that this expression can be regulated by ovarian factors. Preimplantation embryos have been previously shown to express Kit receptors. In this study KL expression was demonstrated in oviducts and uteri and gonadotropin-stimulated increases in oviductal KL mRNA expression were seen. Furthermore, preimplantation embryo survival and development were enhanced with KL suggesting that KL has stimulatory effects on the mitotic cell cycle of the embryo. (Abstract shortened by UMI.)
8
Uebler, Susanne [Verfasser], Thomas [Akademischer Betreuer] Dresselhaus, and Reinhard [Akademischer Betreuer] Sterner. "Analysis of cell-cell communication during reproductive processes by EA1-like peptides in maize / Susanne Uebler ; Thomas Dresselhaus, Reinhard Sterner." Regensburg : Universitätsbibliothek Regensburg, 2015. http://d-nb.info/1122354932/34.
Texte intégral
9
Uebler, Susanne Verfasser], Thomas [Akademischer Betreuer] [Dresselhaus, and Reinhard [Akademischer Betreuer] Sterner. "Analysis of cell-cell communication during reproductive processes by EA1-like peptides in maize / Susanne Uebler ; Thomas Dresselhaus, Reinhard Sterner." Regensburg : Universitätsbibliothek Regensburg, 2015. http://d-nb.info/1122354932/34.
Texte intégral
10
Perrini, C. "EQUINE AND BOVINE MICROVESICLES DERIVED FROM AMNIOTIC PROGENITOR CELLS IN REGENERATIVE AND REPRODUCTIVE TOPICS." Doctoral thesis, Università degli Studi di Milano, 2017. http://hdl.handle.net/2434/490022.
Texte intégralRésumé :
Durante questo progetto di dottorato sono stati condotti studi per comprendere la capacità delle cellule mesenchimali amniotiche (AMCs) di agire attraverso meccanismi paracrini. La prima fase del disegno sperimentale ha valutato l’efficacia delle AMCs sulla proliferazione delle cellule endometriali (EDCs) in assenza di un contatto cellula-cellula. Le EDCs sono state co-coltivate in un sistema trans-well con le AMCs o sono state coltivate in presenza del loro medium condizionato (CM). In entrambe le condizioni sperimentali è stato osservato un aumento del tasso di proliferazione delle EDCs definendo, così, il ruolo cruciale dei fattori secreti dalle AMCs come mediatori dell'azione delle cellule staminali. Il CM è composto da fattori solubili e non-solubili rappresentati da microvescicole (MVs). Per capire il ruolo delle MVs e del loro contenuto nell’azione paracrina, la seconda fase del disegno sperimentale è stata incentrata sull’individuazione del tipo di MVs secrete dalle AMCs e sulla loro efficacia in processi infiammatori in vitro su due importanti modelli sperimentali nella specie equina quali l’endometrio ed il tendine. La produzione di MVs è stata ottimizzata attraverso un’ultracentrifugazione del CM a 100.000 g per 1 ora. Attraverso lo strumento Nanosight è stato calcolato che la produzione di MVs da AMCs equine è di circa 2550±71 particelle/cellula, con una dimensione media di 258±55 nm. La microscopia elettronica a trasmissione ha confermato la presenza di vescicole cellulari che per dimensioni e modalità di secrezione sono state classificate come shedding vesicles. Per capire se le cellule endometriali (EDCs) e tendinee (TNCs) fossero target delle MVs secrete dalle AMCs, è stata studiata l’incorporazione delle medesime dopo marcatura con un fluorocromo lipofilico quale il PKH-26. Dopo uno studio di una curva dose-risposta è stato osservato che la dose ottimale per l'incorporazione di MVs in EDCs è stata di 40x106 MVs/ml a 72 h e per le TNCs di 40x106 MVs/ml a 24-48h. Per esaminare l'abilità delle MVs nel contrastare un processo infiammatorio, entrambe le linee cellulari sono state stressate in vitro con LPS e trattate con MVs. La vitalità cellulare, l'espressione di alcuni geni pro-infiammatori ed il rilascio delle rispettive citochine sono stati i parametri impiegati per valutare l’efficacia delle MVs. Per entrambe le linee cellulari, il tasso di apoptosi è drammaticamente salito nelle cellule trattate con LPS, rispetto al controllo (CTR). LPS ha indotto un aumento significativo dell'espressione di TNF-α, IL-6 e IL-1β in EDCs e di MMP-1, MMP-9, MMP-13 e TNF-α in TNCs. Le MVs sono state in grado di contrastare l'effetto di LPS, diminuendo il tasso di apoptosi in entrambe le linee cellulari e riducendo il livello di espressione dei geni pro-infiammatori. Una conferma di questi dati è stata ottenuta attraverso l'analisi delle citochine pro-infiammatorie (TNF-β and IL-6) ed anti-infiammatorie (TGF-α) rilasciate dalle EDCs nel terreno di coltura, confermando l'abilità delle MVs di trasportare molecole capaci di contrastare la risposta infiammatoria in cellule stressate con LPS.
Data l'importanza del contenuto in miRNA delle MVs, sia nelle AMCs sia nelle loro MVs è stata studiata la presenza di tre miRNA (miR-335, miR-146a, and miR-26a-2) coinvolti nella regolazione dei processi infiammatori. Questi tre miRNA sono stati identificati sia nelle AMCs sia nelle MVs, di conseguenza, è possibile ipotizzare che l'espressione genica dopo infiammazione con LPS e trattamento con MVs possa essere stato modulato dal trasferimento di miRNAs dalle MVs alle cellule target.
Successivamente a questi studi, si è tentato di capire quanto in un processo rigenerativo possa essere imputato alle MVs e quanto ai fattori solubili presenti nel CM e, a questo proposito, è stata valutata l’abilità delle MVs di inibire la proliferazione delle cellule periferiche mononucleate del sangue (PBMC). La loro azione è stata comparata a quello del CM e del sovranatante (SN) ottenuto dal CM dopo scorporazione delle MVs. Il CM ed il SN hanno manifestato capacità immunosoppressiva ma, anche dopo un trattamento di sonicazione al fine del rilascio del loro contenuto, le MVs non hanno dimostrato quest’abilità. Questi risultati portano a dedurre che le MVs sono in grado di trasportare informazioni alle cellule target attraverso molecole capaci di contrastare l'effetto infiammatorio dovuto all'uso di LPS ma, tenendo in considerazione la mancanza di effetto immunomodulatorio, probabilmente in vivo la loro azione è integrata anche dalla presenza dei fattori solubili presenti nel CM.
I meccanismi paracrini materno-fetali sono di vitale importanza ai fini dell’instaurarsi di una gravidanza, per cui si è ritenuto interessante studiare questi meccanismi durante la produzione in vitro di embrioni bovini. Le differenti componenti del secretoma (CM, SN e MVs) ottenute da AMCs ed EDCs bovine sono state aggiunte al terreno di coltura embrionale a differenti giorni di coltura. I risultati hanno dimostrato che il giorno 5 è il migliore momento per la supplementazione e che le MVs di AMCs portano a migliori risultati qualitativi. Questi dati sono stati confermati dalla valutazione di alcuni geni coinvolti nell'apoptosi e nella protezione contro specie reattive dell'ossigeno. I motivi per i quali le MVs di AMCs si siano dimostrate migliori di quelle secrete dalle EDCs non sono ancora conosciuti ma è probabile che le EDCs coltivate in monostrato in vitro vadano incontro a de-differenziamento che altera la qualità del loro secreto.
In conclusione, è possibile affermare che le AMCs sono una linea cellulare affascinante in quanto derivano da materiale di scarto biologico, e quindi a basso costo, e possono essere impiegate in medicina rigenerativa per la loro capacità di trasportare informazioni alle cellule target. Le MVs possono offrire un nuovo strumento terapeutico privo di cellule nell’ambito della nanomedicina.<br>During this PhD project, studies were carried out to understand the ability of amniotic mesenchymal cells (AMCs) to act by paracrine mechanism. At first, AMCs and their conditioned medium (CM) were investigated in an in vitro model using equine endometrial cells (EDCs) as target. Proliferation of EDCs was studied co-culturing them with AMCs in a trans-well system or in presence of AMC-CM. In both conditions, there was a significant increase in EDC proliferation rate, defining the crucial role of factors secreted by AMCs in stem cells action. CM is composed of soluble factors and no-soluble factors as microvesicles (MVs). In this context, in the second step of this project, the presence and the type of AMC-MVs were identified to understand their role in regenerative medicine. The production of MVs was optimized through a CM ultracentrifugation at 100.000 g for 1 hour. Microvesicle production from equine AMCs was 2550±71 particles/cell, with a mean dimension of 258±55 nm. The transmission electron microscopy confirmed the presence of extra-cellular vesicles that were classified as shedding vesicles for their size and modality of secretion. In order to understand if endometrial cells (EDCs) tendon cells (TNCs) were target of these MVs, an incorporation study was performed labelling MVs with a lipophilic fluorochrome such as PKH-26. By a dose-response curve, the optimal conditions of incorporation were at 72 h at a concentration of 40x106 MVs/ml for EDCs, and at 24-72 h at a concentration of 40x106 MVs/ml for TNCs. In order to study MVs ability to counteract an in vitro inflammation, EDCs and TNCs challenged with LPS and treated with MVs were evaluated by viability cell tests, by expression of some pro-inflammatory genes, and by release of respective cytokines. For both cell types, the apoptosis rate increased dramatically in cells treated with LPS if compared to the control (CTR). LPS significantly upregulated the expression of TNF-α, IL-6 and IL-1β in EDCs and of MMP-1, MMP-9, MMP-13 and TNF-α in TNCs. MVs were able to counteract the action of LPS, decreasing the apoptosis rate and reducing in the expression levels of the pro-inflammatory genes in both cell lines. Coherent results were obtained through the analysis of pro-inflammatory (TNF-β and IL-6) and anti-inflammatory (TGF-α) cytokines released by EDCs in the culture medium, confirming the ability of MVs to transport molecules able to counteract the stress induced by LPS.
Since MVs contain various active molecules, the presence of three miRNAs (miR-335, miR-146a, and miR-26a-2) was investigated, as they are involved in the regulation of inflammation. The selected miRNAs have been found in both AMCs and their MVs, so the previously observed downregulation of gene expression could be correlated to miRNA transfer from MVs to target cells. Moreover, the ability of MVs to inhibit peripheral blood mononuclear cell (PBMC) was evaluated, but MVs, also after lysis by sonication to release their content, were not able to inhibit PBMC proliferation despite to CM and SN. These results led to hypothesize that MVs brought to the target cells some molecules able to counteract the inflammatory situation due to the LPS but, taking into account the lack of their immunomodulatory action, probably, for an in vivo healing, soluble factor of CM are necessary too.
Paracrine mechanism are essential also in maternal-fetal communication. In this context, we studied these mechanisms during bovine in vitro embryo production. Different components of secretome (CM, SN and MVs) from bovine AMCs and EDCs were supplemented to the embryo culture media at different days of culture. The results demonstrated that the day 5 of culture is the best time point for the supplementation of these components and that AMCs-MVs provided the best environment for the embryo concerning the blastocyst quality. These data were confirmed by the evaluation of genes involved in apoptosis and reactive oxygen species protection. The reasons for which the MVs of AMCs have proved better than those secreted by EDCs are not yet known but it is likely that in vitro culture of EDCs in monolayer may induce a de-differentiation that alters the quality of their secretion.
As conclusion of this project, it is possible to speculate that AMCs are fascinating in view of producing off-the-shelf products, at low cost, and their use in regenerative medicine for their capacity to carry information to the target cells. The MVs may offer a new therapeutic cell-free tool in nanomedicine.
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Lyons, Rachel Ann. "Studies on aspects of cell biology and response to humoral factors of epithelial cells in the human reproductive tract." Thesis, Queen Mary, University of London, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.510918.
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Corrigan, Laura. "Regulation and reproductive functions of membrane-bound vesicles secreted by the Drosophila male accessory gland." Thesis, University of Oxford, 2014. http://ora.ox.ac.uk/objects/uuid:673d46a5-ba88-42d2-9361-51f04d61e01b.
Texte intégralRésumé :
Membrane-bound vesicle secretion provides a novel intercellular communication mechanism, whose roles and regulation remain poorly characterised, particularly in vivo. I have identified two classes of lipid-containing, vesicle-like structures secreted into seminal fluid by epithelial cells of the Drosophila male accessory gland (AG). Exosomes, one class of membrane-bound vesicle formed inside late endosomal multivesicular bodies, are specifically secreted by secondary cells (SCs). The unusual cell biology of SCs allowed me to develop a powerful new high resolution in vivo system to characterise the mechanisms underlying intracellular membrane trafficking events underlying exosome biogenesis using real-time live imaging. I characterise how specific ESCRTs (endosomal sorting complexes required for transport) control SC exosome biogenesis, and identify a novel role for BMP signalling in regulating endolysosomal trafficking events necessary for exosome secretion. I also identify roles for epidermal growth factor receptor (EGFR) and phosphatidylinositide 3-kinase (PI3K) signalling in exosome biogenesis. Importantly, SC exosomes are transferred to females during mating. Here, they fuse with sperm, mirroring in vitro interactions between human prostate exosomes and sperm, and interact with the female reproductive tract epithelium. Blocking SC exosome production specifically suppresses post-mating effects on female receptivity to remating, demonstrating that exosomes have an important reproductive signalling function in vivo, directly or indirectly reprogramming female cells. Finally, I show that main cells, the major epithelial AG cell type, shed lipid-containing microvesicle-like structures from their apical surface. Remarkably, these vesicles carry the seminal peptide, sex peptide, into females during mating and also contribute to the anterior mating plug. In summary, my data reveal previously unsuspected roles for exosomes and microvesicles in Drosophila reproduction that may be evolutionarily conserved. Since these vesicles mediate physiological processes previously thought to involve soluble peptides, my work suggests that current models explaining male reprogramming of female behaviours in flies and higher organisms need substantial revision.
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Jiwakanon, Jatesada. "The porcine endosalpinx at different reproductive stages : morphology, immune cell infiltration and cytokine expression /." Uppsala : Dept. of Clinical Sciences, Swedish University of Agricultural Sciences, 2009. http://epsilon.slu.se/200912.pdf.
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Pellegrino, Giuliana. "Cell neogenesis in the postnatal hypothalamus as a new mechanism of control of the reproductive function." Thesis, Lille 2, 2017. http://www.theses.fr/2017LIL2S042.
Texte intégralRésumé :
Malgré sa complexité, le cerveau intègre en permanence de nouvelles cellules – à la fois neuronales et gliales – au-delà du développement embryonnaire et ce, tout le long de la vie. La période postnatale est caractérisée par une gliogenèse intense. A l’âge adulte, de nouveaux neurones et cellules gliales sont produits dans des régions restreintes à partir de cellules souches/progénitrices (CSP) localisées dans des niches. Les deux niches de CSP adultes les mieux décrites sont la zone sous-ventriculaire des ventricules latéraux, qui produit de nouveaux interneurones olfactifs, et la zone sous-granulaire du gyrus denté de l’hippocampe, où de nouveaux neurones en grain sont produits localement. Des travaux menés ces dernières années ont montré qu’une neuro- et une gliogenèse avaient aussi lieu dans l’hypothalamus postnatal, une petite région du diencéphale ventral qui régule des processus physiologiques vitaux tels que le métabolisme, la reproduction, le sommeil et la thermorégulation. Si l’identité des CSP hypothalamiques reste débattue, de nombreux travaux s’accordent sur l’importance de la neurogenèse hypothalamique postnatale dans le contrôle du métabolisme. Cependant, la possibilité que la genèse postnatale de cellules contribue aussi au contrôle de la fonction de reproduction, une autre fonction clé de l’hypothalamus, restait à explorer. L’objectif premier de mon travail de thèse était de rechercher si la genèse de cellules dans l’hypothalamus postnatal est impliquée dans le contrôle de la reproduction, une fonction physiologique qui requière un haut degré de plasticité. La fonction de reproduction est orchestrée par une petite population de neurones produisant la neurohormone Gonadotrophin-Releasing Hormone (GnRH). Ces neurones, qui naissent en dehors du cerveau, sont en place dans la région préoptique (RPO) de l’hypothalamus à la naissance. Cependant, ils doivent subir une maturation postnatale pour acquérir le profil de sécrétion qui leur permettra d’initier la puberté et d’assurer la fertilité de l’individu. Dans une première étude, grâce à une combinaison d’approches in vitro et in vivo, nous avons mis en évidence une vague d’astrogenèse dans l’environnement des neurones à GnRH au sein de la RPO au cours des deux premières semaines de vie postnatale chez la ratte. Nos résultats suggèrent que les neurones à GnRH utilisent la prostaglandine D2 pour attirer les progéniteurs environnants et que ce recrutement est important pour la maturation sexuelle. Dans une deuxième étude, nous avons recherché si de nouvelles cellules naissent à l’âge adulte dans des régions hypothalamiques qui contrôlent la fonction de reproduction. Nous montrons que des cellules sont produites dans la RPO chez la ratte adulte et que leur taux varie au cours du cycle oestral, suggérant une régulation par les stéroïdes sexuels. De plus, nous montrons que la survenue d’une gestation stimule la néogenèse cellulaire dans une zone de la RPO qui contrôle le comportement maternel. Si la néogenèse hypothalamique adulte a surtout été étudiée chez les rongeurs de laboratoire, il reste à déterminer si ce phénomène existe aussi chez l’homme. Pour aborder cette question, nous avons évalué dans une troisième étude l’expression de marqueurs de CSP dans l’hypothalamus humain adulte, comparativement au rongeur (souris, rat) et à un primate lémurien, le microcèbe. Nous montrons que l’hypothalamus humain adulte contient des populations de cellules au profil antigénique de CSP, dont certaines semblent propres à l’homme. Au total, ces travaux montrent que de nouvelles cellules naissent dans des régions hypothalamiques qui contrôlent la fonction de reproduction au cours de la vie postnatale et à l’âge adulte chez la ratte, et que ce phénomène est important pour la maturation sexuelle. L’observation de CSP putatives dans l’hypothalamus humain adulte suggère que la capacité de l’hypothalamus à produire de nouvelles cellules à l’âge adulte existe aussi dans notre espèce<br>Despite its complexity, the brain keeps adding new cells – both neuronal and glial – beyond embryonic development and throughout life. The postnatal period is characterized by intense and widespread gliogenesis. During adulthood, both glio- and neurogenesis occur in restricted locations from stem/progenitor cells (NPC) residing in niches. The two best-described niches of adult NPC are the subventricular zone of the lateral ventricles, which provides new interneurons to the olfactory bulb, and the subgranular zone of the hippocampal dentate gyrus that locally produces new granule cells. The last decade has seen an accumulation of studies showing that neuro- and gliogenesis also occur in the postnatal hypothalamus, a small portion of the ventral forebrain surrounding the third ventricle that regulates essential physiological processes such as metabolism, reproduction, sleep and thermoregulation. Even though the identity of hypothalamic NPC remains a matter of debate, a growing body of evidence points to postnatal hypothalamic neurogenesis relevance for the control of metabolism. However, a possible contribution of postnatal hypothalamic cell generation to the central control of reproduction, another key function of the hypothalamus, remained to be explored.The main aim of my doctoral researches was to evaluate whether the generation of new cells in the postnatal hypothalamus contributes to the central control of reproduction, a physiological function known to require a high degree of plasticity. The reproductive function is controlled by a small population of neurons producing the neurohormone Gonadotrophin-Releasing Hormone (GnRH). These neurons, which are born in the nasal placodes, are in place at birth in the preoptic area (POA) of the hypothalamus. However, they need a postnatal maturation to reach a mature secretory pattern that will trigger puberty and subsequent fertility.In a first study, using a combination of in vitro and in vivo experiments, we showed that a wave of astrogenesis occurs in the POA from local progenitors in the environment of GnRH neurons during the first weeks of postnatal life in the female rat. We identified prostaglandin D2 as a factor used by GnRH neurons to attract progenitors in their vicinity and showed that impaired progenitor recruitment alters sexual maturation.In a second study, we evaluated whether cell neogenesis still occurs during adulthood in hypothalamic regions relevant for the reproductive function. Our results showed that new cells are born in the POA of adult female rats. The rate of cell neogenesis varies across the estrus cycle, suggesting a regulatory influence of gonadal steroids. Moreover, we showed that gestation impacts the rate of cell neogenesis in a POA region implicated in the control of maternal behavior.While cell neogenesis in the adult hypothalamus has been mainly studied in laboratory rodents, it remains to be known whether this phenomenon also occurs in humans. To start addressing this question, we evaluated in a third study the expression of a panel of NPC markers in the adult human hypothalamus and compared it to that found in rodents (mouse, rat) and a lemur primate, the grey mouse lemur. Our results showed that the adult human hypothalamus contains populations of cells with an antigenic profile of NPC, some of which appear specific to humans.Altogether, this work shows that new cells are born in hypothalamic regions controlling reproduction throughout postnatal and adult life in female rats, and that this process is required for sexual maturation. The identification of NPC marker-expressing cells in the adult human hypothalamus suggests that the capacity for cell neogenesis also exists in the hypothalamus of our species
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Bonetto, Valentina. "From cell to organism: an overview of responses to simulated hypergravity and microgravity." Doctoral thesis, Università del Piemonte Orientale, 2022. http://hdl.handle.net/11579/144698.
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Zabida, Omer Saleh. "The effect of methamphetamine on the blood-testis barrier." University of the Western Cape, 2018. http://hdl.handle.net/11394/6775.
Texte intégralRésumé :
>Magister Scientiae - MSc<br>Introduction
The blood-testis barrier (BTB) is formed by tight junctions between adjacent Sertoli cells. The barrier formed by these tight junction helps to create a specialized environment for spermatogenesis and provide an immunological barrier to protect developing germ cells. Methamphetamine (Meth) is known as neurotoxin however, its effects on the male reproductive system, especially on Sertoli cells and, the BTB are not well established. Therefore, this study aimed to determine the effects of Meth on the TM4 mouse testis Sertoli cell line and on the integrity of the BTB permeability.
Materials and Methods
This study investigated the effect of selected concentrations of Meth (0.1 μM, 1 μM, 10 μM, 20 μM and 100 μM) on TM4 mouse testis Sertoli cell line for 24 until 96 hours, using two treatments: an “acute” study (24 hrs exposure) and a “chronic” study, where treatment occurred on a daily basis over 96 hrs. The following parameters were investigated: viability, cell proliferation, mitochondrial activity, monolayer permeability.
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Le, Nhung Xuan Hong. "ASSOCIATION OF IMMUNE DYSFUNCTION WITH MICROBIAL DYNAMICS AND ABERRANT ESTROGEN METABOLISM IN REPRODUCTIVE DISORDERS." OpenSIUC, 2021. https://opensiuc.lib.siu.edu/dissertations/1918.
Texte intégralRésumé :
Chronic inflammation is associated with the pathophysiology of obstetrical disorders (e.g. preterm birth [PTB]) and gynecological diseases (e.g. endometriosis); however, the exact mechanism(s) for these conditions are unknown. Numerous immunological conditions and disease states (e.g. inflammatory bowel disease, Crohn’s disease, systemic lupus erythematomus) also disrupt the microbiome homeostasis by inducing a number of changes in the microbial flora when compared to that of healthy individuals. Furthermore, the gastrointestinal (GI) microbiome is one of the principal regulators of circulating estrogens which are known to directly impact the female reproductive disorders endometriosis and PTB. Thus, an alteration of microbial species could indicate a shift in immune balance from homeostatic to pro-inflammatory, and an aberrant estrogen metabolism that precipitates the development of disease stages in endometriosis and/or PTB. The Braundmeier-Fleming lab has developed a systems biology model that investigates the interactions between the immune system, microbial dynamics (in the GI and reproductive system) and estrogen metabolism, in women, as a potential diagnostic tool for endometriosis and PTB. This dissertation, therefore, examined how inflammation triggered by female reproductive disorders (endometriosis or PTB) alter the systemic and localization immune responses, the microbial communities in the urogenital (UG), peritoneal and GI mucosal epithelium, as well as levels of excreted conjugated estrogen. The first specific hypothesis is that inflammation associated with endometriosis alters microbial dynamics and functions that are distinct from those of non-diseased patients. Preliminary data indicated that reproductive tract microbial communities from patients with endometriosis are unique when compared to non-disease patients. Therefore, the central aims of this study are to identify the immune and microbial profiles of patients diagnosed with endometriosis and determine if an alteration of these profiles impact estrogen signaling, thus driving disease pathogenesis. Additionally, I hypothesized that surgery or hormonal therapy will temporarily restore the microbiome and estrogen levels of patients with endometriosis. Differences in systemic (blood) regulatory T cell (Treg) and T-helper 17 (Th17) cell populations (tolerant and inflammatory, respectively) were measured by flow cytometry, and the immune mediators was measured by serum cytokine levels via 10-plex-ELISA kits. Immunohistochemistry was used to identify resident Th17/Treg immune cell distribution within the endometrium and ectopic endometriotic lesions, and RORγt+/FOXP3+ transcripts within these same tissues were analyzed by real-time-qPCR. We implemented high-throughput non genomic sequencing targeting bacterial-V4 16S rRNA and robust bioinformatics analyses to characterize microbial composition/diversity within the GI (fecal swab), vaginal (vaginal swab), and UG (urine) cavities. Alterations in estrogen metabolism, parent estrogens and metabolites, in urine were analyzed via LC-MS/MS. Patients with endometriosis exhibit 1) systemic and localized inflammation within ectopic and endometrial tissues, 2) altered GI/UG microbial dynamics, 3) aberrant levels of endogenous estrogen and estrogen metabolites, 4) dampened inflammation (caused by disease) due to hormonal therapy, 5) altered bacteria populations in the gut and vaginal canal of patients with endometriosis due to hormonal therapy treatment, and 6) increased post-surgical variability in microbial community dynamics. The second specific aims examined the hypothesis that induction of endometriosis in baboons (P. Anubis) results in chronic systemic and tissue specific inflammation through regulation of Th17 and Treg populations. Further, the induction of endometriosis altered GI/UG/peritoneal cavity microbial communities that are distinct from non-diseased animals. Utilizing a non-human primate animal model of induced endometriosis allowed us to characterize factors involved at the early onset of endometriosis and throughout the disease progression. We collected samples from 8 baboons at pre-inoculation (no evidence of disease) and at 3, 6, 9, and 15 months post-induction of the disease. We found that the induction of endometriosis decreased peripheral Tregs cells while Th17 cells increased at all post-induction collections with reduced ratio of total Tregs to Th17 cells indicating systemic inflammation. Microbial community diversities as well as abundances at each sample site (GI, UG [vagina, urine] tracts and peritoneal cavity) were also altered at post-induction. These results therefore suggest that induction of endometriosis in non-human primates caused an inflammatory shift. Disease induction also resulted in altered vaginal, urinary and fecal microbial profiles, which may drive inflammation through the production of inflammatory mediators. The last specific aims studied the hypothesis that patients who deliver preterm have a systemic and placental inflammatory phenotype and abnormal estrogen levels during pregnancy that are distinct from those of patients with term delivery. Biological samples were collected at 8-12 weeks, 20-24 weeks, 32-36 weeks, at delivery and 6 weeks postpartum. Subjects with PTB showed signs of systemic inflammation with an elevation in Th17:Treg ratio, greater Th17 and lower levels of natural Tregs during the 2nd trimester, and lower inducible Tregs during the 3rd trimester and at delivery. Placental tissues from subjects with PTB also had an inflammatory immune phenotype (higher Th17) within the decidua basalis and maternal-fetal interface. Immunological shifts from tolerant to inflammatory were observed in both patient groups, but these shifts occurred early in gestation for subjects with PTB and at a later gestational age for subjects delivering at term. Levels of conjugated parent estrogens and estrogen metabolites were reduced in subjects with PTB, indicative of an abnormal production of estrogen. These analyses gave us a better understanding of the inflammatory cascade with estrogen metabolism associated with pregnancy, and how these effects are correlated with premature labor. The data from this study suggest that the levels of endogenous estrogen and estrogen metabolites of estrogen metabolism were abnormal in PTB and endometriosis disease models of inflammation compared to their respective controls. In the human and non-human primate model of endometriosis studies, we observed that both patients and baboons with endometriosis had systemic and resident inflammatory phenotypes and an alteration in mucosal microbial community dynamics compared to their respective controls. All together, our long-term goal is to identify factors from the microbiome and/or the immune system that would allow us to have early non-invasive diagnostics for endometriosis or to predict which mothers are most at risk to encounter PTB. Furthermore, it would allow us to determine whether the mucosal microbiome may be a good indicator of immune stress, and if alternative therapies can alter microbial community dynamics—thereby eliminating immune stress associated with female reproductive diseases. These findings may have a substantial impact on the obstetrical care and management of patients with endometriosis and women at risk for PTB, as well as provide evidence to support the development of novel therapeutics to treat these diseases.
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Alhabhbeh, Ammar, Purva Sharma, Mohammad Ali Khan, Koyamangalath Krishnan, and Devapiran Jaishanker. "Adenocarcinoma of Prostate with Small Cell Differentiation Presenting As Refractory Hypokalemia." Digital Commons @ East Tennessee State University, 2020. https://dc.etsu.edu/asrf/2020/presentations/29.
Texte intégralRésumé :
Prostate cancer is among the most common malignancies in males in the United States and adenocarcinoma accounts for 95% of all malignancies of prostate. Rarely prostate cancer can also present as small cell carcinoma. Pure small cell carcinoma is rare at time of initial diagnosis (<2%) however neuroendocrine differentiation into small cell carcinoma may emerge in men who have had previous treatment with ADT for prostate adenocarcinoma. These tumors, sometimes called treatment-related neuroendocrine prostate cancers or aggressive-variant prostate cancers, are increasingly recognized in the castration-resistant phases of disease progression. They account for less than 1% of all prostate cancers. A 73-year-old otherwise male had routine health screening in May 2018. Prostate specific antigen (PSA) level was elevated at 9.53 ng/mL. He had not had a screening PSA for at least two prior years but this was a significant change from prior levels. Patient was asymptomatic however the abnormal laboratory evaluation prompted consultation with Urology. Biopsy of prostate gland confirmed prostatic adenocarcinoma with Gleason's score of 5+ 4 = 9 with bilateral gland involvement. Imaging studies including CT scan of abdomen and pelvis, a bone scan and a PET scan showed no clear evidence of metastatic disease. Patient's clinical stage was determined to be IIIC with T2c N0 M0 disease. Patient began treatment with androgen deprivation therapy and received definitive radiation treatment with external bean radiation therapy from July to September 2018. PSA was 0.08 ng/ml at the end of radiation treatment. Patient did well for about 15 months, after which he had multiple hospital admissions for dyspnea, fluid retention and lower extremity edema. He was also found to have refractory hypokalemia. Patient underwent MRI brain which revealed numerous small enhancing calvarial and skull base lesions consistent with bony metastasis in the skull. Patient also underwent PET/CT scan which showed numerous thoracic spine bony lesions, numerous to count bony metastasis throughout the lumbar spine and pelvis, as well as multiple hepatic lesions. Patient underwent biopsy of right hepatic lobe lesion and pathology was consistent with small cell carcinoma with positive neuroendocrine markers including CD56, synaptophysin and TTF-1. Interestingly patient’s PSA was only 0.09ng/dL. Given refractory hypokalemia, paraneoplastic syndrome was suspected and further work-up was initiated. Serum cortisol levels were elevated at 119.6 mcg/dL (3.7-19.4) and ACTH level was 333 pg/mL (7.2 - 63.3). Aldosterone level was <1 ng/dL (0 - 30.0). Patient was diagnosed with paraneoplastic Cushing syndrome. Given aggressive nature of this small cell transformation, patient was started on treatment with systemic chemotherapy with Carboplatin/Etoposide during the hospital stay, with stabilization of potassium levels. Prostate small cell carcinoma poses a challenge for diagnosis and treatment. In contrast to adenocarcinoma of the prostate, serum prostate-specific antigen (PSA) is not predictive of disease severity, nor is it a useful tumor marker for monitoring progression or surveillance. Patients with prostate small cell cancer presents with more diverse symptoms than any other prostate cancer since it tends to metastasize early. Also paraneoplastic syndromes are more common in prostate small cell cancers as well.
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Erasmus, Nicolete. "Investigations on the in vitro effects of aqueous Eurycoma longifolia Jack extract on male reproductive functions." Thesis, University of Western Cape, 2012. http://etd.uwc.ac.za/index.php?module=etd&action=viewtitle&id=gen8Srv25Nme4_2238_1375971626.
Texte intégralRésumé :
<p>Eurycoma longifolia (Tongkat Ali<br>TA) is a Malaysian shrub used to treat various illnesses including male infertility. Considering that TA is also used to improve male fertility and no report <br>regarding its safety has been published, this study investigated the effects of a patented, aqueous TA extract on various sperm and testicular functions. Materials and Methods This study <br>encompasses two parts (part 1: on spermatozoa<br>part 2: on TM3-Leydig and TM4-Sertoli cells). Part 1: Semen samples of 27 patients and 13 fertile donors were divided into two groups, <br>washed and swim-up prepared spermatozoa, and incubated with different concentrations of TA (1, 10, 20, 100, 2000 &mu<br>g/ml) for 1 hour at 37°<br>C. A sample without addition of TA served as control. After incubation with TA, <br>the following parameters were evaluated: viability (Eosin-Nigrosin test), total and progressive motility (CASA), acrosome reaction (triple stain technique), sperm production of reactive oxygen <br>species (ROS<br>dihydroethidium test<br>DHE), sperm DNA fragmentation (TUNEL assay) and mitochondrial membrane potential (&Delta<br>&psi<br>m) (Depsipher kit). Part 2: TM3-Leydig and TM4-Sertoli cells <br>incubated with different concentrations of TA (0.4, 0.8, 1.6, 3.125, 6.25, 12.5, 25, 50 &mu<br>g/ml) and control (without extract) for 48 and 96 hours. After incubation with TA, the following parameters were <br>evaluated: viability (XTT), cell proliferation (protein assay), testosterone (testosterone ELISA test) and pyruvate (pyruvate assay). Results Part 1: For washed spermatozoa, significant <br>dose-dependent trends were found <br>for viability, total motility, acrosome reaction and sperm ROS production. However, these trends were only significant if the highest concentrations were included in the calculation. In the swim-up spermatozoa, ROS production of spermatozoa showed a biphasic relationship with its lowest percentage at 10 &mu<br>g/ml, yet, no significance could be <br>observed (P=0.9505). No influence of TA could be observed for sperm DNA fragmentation nor &Delta<br>&psi<br>m.</p>
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Blake, Crystal. "The Effects of Prenatally Administered Phytoestrogens on the Reproductive and Behavioral Development of Long-Evans Rats." BYU ScholarsArchive, 2008. https://scholarsarchive.byu.edu/etd/2364.
Texte intégralRésumé :
Equol is known to be a selective androgen modulator and has the ability to bind and inhibit 5-alpha dihydrotestosterone (5α-DHT). Equol is also a selective estrogen receptor modulator and is able to bind beta estrogen receptors with high affinity. As estrogen receptors are found in the hypothalamus, pituitary, and gonads, prenatally administered equol could affect the morphological and reproductive development of offspring. To test this hypothesis, during gestational days 14 to 20, forty-two pregnant Long-Evans rats were given one of six treatments: 1) no treatment, 2) injection with dimethyl sulfoxide (DMSO), 3) injection with 10 mg/kg equol, 4) injection with 21.0 mg/kg equol, 5) injection with 63.0 mg/kg equol, or 6) injection with 90.0 mg/kg flutamide. At birth the pups were weighed, anogenital distance measured, and sex was determined. Some of the animals were sacrificed and trunk blood collected from both the mothers and pups. Serum levels of phytoestrogens, estradiol, testosterone, and 5α-DHT levels were determined. Some pups were allowed to grow up to day 29 and were tested on the forced-swim test with the parameters of time mobile, time immobile, swim distance, and average speed measured. The flutamide treated pups had the lowest anogenital distance. The low equol dose animals had the largest anogenital distance. There were no significant differences in 5α-DHT serum levels in the male offspring among the treatments. However in non-injected control female offspring displayed significantly lower 5α-DHT levels than all the other groups. Mothers treated prenatally with equol displayed significantly higher circulating equol levels compared to controls values. Rats injected with 63.0 mg/kg of equol gained the least weight during pregnancy. Their offspring also had the lowest body weights at birth. Male and female offspring displayed similar behaviors in the Porsolt forced-swim test among the treatment groups. The low and high equol groups displayed the least depressive-like behaviors. The offspring from mothers treated with the medium and high equol doses both gained the most weight from birth to day 29. Treating pregnant rats with equol during the last week of gestation does not appear to have any affect on morphological genital development of the offspring.
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Stefansdottir, Agnes. "Development and use of an in vitro technique to investigate the effect of pharmaceutical agents on female germ cell development." Thesis, University of Edinburgh, 2015. http://hdl.handle.net/1842/16209.
Texte intégralRésumé :
With meiosis spanning from embryonic development to the end of reproductive life in females, scientists have faced considerable limitations in studying female meiosis and the effects of toxicants on the developing oocyte. Over the last half century, various culture methods have been developed with the aim of studying the mechanisms of early ovary development, as well as for use in reproductive toxicology. However, very few of the established embryonic ovary culture systems have been used to investigate potential reproductive toxicants on the embryonic ovary, in particular when compared with the vast number of in vitro reproductive toxicity studies on the post-natal ovary. Here, a novel test compound, a topoisomerase II inhibitor: AstraZeneca Test Compound (AZTC), was used to investigate the efficacy and validity of ovarian culture methods when compared with in vivo reprotoxicity studies. AZTC was selected due to preliminary in vivo studies demonstrating its detrimental effects on spermatogenesis in male rats. AZTC targets bacterial type II topoisomerases that might have mammalian homologues involved in meiosis. Topoisomerase-II α was expressed within the female germ cells pre-natally, but became localised to the granulosa and stroma cells post-natally. This occurred both in vivo and in vitro. Ovaries from female rats exposed pre-natally to AZTC in vivo were analysed histologically and a significant increase in the number of primordial follicles was observed within the ovaries, as well as an increase in the number of unhealthy follicles. A novel mouse embryonic ovary culture system was developed by adapting, improving and bridging existing available culture techniques. The culture system supported growth of pre-meiotic mouse germ cells through prophase I of meiosis, the formation of primordial follicles and initiation of follicle growth. Cultured ovaries contained follicles at stages in comparable ratios to those in vivo and appeared morphologically normal and healthy. The culture also supported meiotic progression of oocytes to the pachytene stage, albeit with a slight delay. AZTC was used to validate the novel embryonic ovary culture by comparing the results with those from the in vivo study, where AZTC exposure had also occurred during embryonic development. Similar results were consistently observed between the in vivo and in vitro studies. In vitro effects of AZTC on the post-natal mouse ovary were also investigated, where neonatal mouse ovaries cultured with AZTC had fewer primordial follicles and more unhealthy follicles than did control ovaries. AZTC therefore demonstrated different effects when exposure occurred pre-natally vs. post-natally. The embryonic ovary culture was then used to examine the effects of another topoisomerase II inhibitor, etoposide, on the pre-natal ovary. Etoposide is a chemotherapy agent and has previously been prescribed to pregnant women. A significant reduction in the size of the follicle pool was observed in exposed cultured embryonic ovaries, where primordial and transitional follicles were targeted. Overall, establishment of post-natal culture systems have become a useful addition to in vivo reproductive toxicology studies. The embryonic ovary culture system developed here could become a valuable and powerful tool to screen potential reproductive toxicants, as well as to study the dynamics and regulation of early ovary development.
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Cheng, Becky. "The Role of the Dosage Compensation Complex as a Pathway for Spiroplasma to Induce Male Lethality in Drosophila melanogaster." Scholarship @ Claremont, 2017. http://scholarship.claremont.edu/cmc_theses/1540.
Texte intégralRésumé :
Drosophila melanogaster and many other insects harbor intracellular bacterial symbionts that are transmitted vertically from infected host mothers to their offspring. Many of these bacteria alter host reproductive developmental processes in order to increase their transmission success. For example, Spiroplasma, a spirochete that naturally infects D. melanogaster, selectively kills males during mid-embryogenesis while sparing females. Previous studies suggested that Spiroplasma interacts genetically with the male-specific dosage compensation pathway, which causes ~2-fold up-regulation of most genes located on the male’s single X chromosome so that their expression matches the levels found in females who have two Xs. To further test this idea, I used confocal microscopy to visualize dosage compensation complex (DCC) localization and activity in infected as well as uninfected embryos. In the presence of Spiroplasma, the DCC became abnormally mis-localized across the nucleus. This pattern was accompanied by abnormal acetylation of histone H4K16, a mark induced by DCC activity and needed for proper X chromatin remodeling. My results imply that Spiroplasma directly targets the DCC by misdirecting it to uncompensated regions of the genome, an effect that leads to abnormal gene mis-regulation and consequent lethality (work from other members in our group). To further investigate this interaction, we transgenically expressed low levels of MSL-2 in both Spiroplasma infected and uninfected embryos in order to cause ectopic formation of the DCC in the female sex. I found that when infected, female embryos expressing the DCC showed significantly reduced viability in comparison to uninfected transgenic females. This result supports the notion that Spiroplasma uses the DCC in a dominant gain-of-function manner to kill embryos.
23
Gunnarsson, David. "Reproductive toxicology of endocrine disruptors : effects of cadmium, phthalates and phytoestrogens on testicular steroidogenesis." Doctoral thesis, Umeå : Department of Molecular Biology, Umeå University, 2008. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-1876.
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Somani, Imtiaz Habib. "Gene knockout mouse models of human Tay-Sachs and Sandhoff diseases and their effects on the male reproductive system." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk2/tape16/PQDD_0006/MQ29790.pdf.
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Cecere, Thomas E. "Porcine circovirus associated disease: Modulation of the host immune response to PCV2 and PRRSV by regulatory T cells." Diss., Virginia Tech, 2012. http://hdl.handle.net/10919/77098.
Texte intégralRésumé :
Porcine circovirus associated disease (PCVAD) is currently one of the most economically important diseases facing the global swine industry. Porcine circovirus type 2 (PCV2) is the primary and essential causative agent of PCVAD, but development of clinical disease typically requires co-infection with other swine pathogens such as porcine reproductive and respiratory syndrome virus (PRRSV). The specific mechanisms of co-infection that lead to clinical disease are not fully understood, but immune modulation by the co-infecting viruses is thought to play a critical role. The ability of dendritic cells (DC) infected with PRRSV, PCV2 or both to induce regulatory T cells (Tregs) was evaluated in vitro. DCs infected with PCV2 significantly increased CD4+CD25+FoxP3+ Tregs (p<0.05) and DCs co-infected with PRRSV and PCV2 induced significantly higher numbers of Tregs than with PCV2 alone (p<0.05). This Treg induction was found to be dependent on TGF-β and not IL-10. Further investigation of the in vivo swine immune response to acute co-infection with PCV2 and PRRSV failed to detect activation of Tregs in peripheral blood mononuclear cells (PBMCs) or bronchoalveolar lavage samples. The Treg response to in vitro and in vivo PRRSV challenge in pigs persistently infected with PCV2 or vaccinated against PCV2 was evaluated. There was no significant difference in Tregs in PBMCs among chronically PCV2-infected, vaccinated PCV2 challenged or negative control pigs. However, following in vitro infection of monocyte-derived dendritic cells with PCV2, PRRSV, or both viruses, co-cultured lymphocytes from chronically infected and PCV2 vaccinated pigs had significantly (p<0.05) decreased Treg expression in the virus infected groups compared to the negative controls. In separate experiments, pigs vaccinated against PCV2 and subsequently challenged with an attenuated PRRSV strain and its pathogenic parental strain developed increased CD4+CD25+FoxP3+ Tregs (p<0.05) in PBMC samples compared to uninfected controls, and this correlated with increased suppressor activity and IL-10 expression. The findings from these studies indicate that the interaction of PCV2 and PRRSV in swine modulates the host immune response mediated in part through the activity of Tregs. However, the extent to which Tregs orchestrate a dysregulated immune response in the pathogenesis of PCVAD in vivo remains to be determined.<br>Ph. D.
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Swanepoel, Magdaleen. "Embryonic stem cell research and cloning a proposed legislative framework in context of legal status and personhood /." Diss., Pretoria : [s.n.], 2006. http://upetd.up.ac.za/thesis/available/etd-07312007-150150/.
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Fabick, Kimberly Michelle. "The Effects of a Resveratrol Derivative on Regulatory Behaviors and Reproductive Health in Male and Female Long-Evans Rats." BYU ScholarsArchive, 2008. https://scholarsarchive.byu.edu/etd/2839.
Texte intégralRésumé :
Phytoestrogens are chemicals produced by plants that act like estrogens and have the ability to bind to the mammalian estrogen receptor system. The purpose of this study is to evaluate a new phytoestrogen analog called 4-acetoxy Resveratrol. Resveratrol is a phytoestrogen that has been found in the skin of grapes. Resveratrol has been shown to be able to bind to the estrogen receptors and has a similar molecular structure as estradiol. Resveratrol has been shown to have many positive health benefits such as improving cardiovascular health, serving as a neuroprotective agent, acting as an anti-inflammatory agent, working as an anti-cancer agent, increasing sperm output, acting as an anti-aging agent, and reducing incidence of prostatic adenocarcinoma. The challenge with using Resveratrol as an oral therapy is that it is quickly metabolized by the liver so for this study we used injections. The injections were 5mg/ Kg, 20 mg/Kg, and 90 mg/Kg of 4-acetoxy Resveratrol. We used intact 160 day old male Long-Evans rats and intact 90 day old female Long-Evans rats. The rats were given injections once a day for 21 days based on their treatment group. The animals were weighed daily and then tested in the Porsolt swim test at day 160 and 90 respectively. At the end of 21 days the rats were sacrificed and white adipose tissue, blood, brains, testis, and prostates were collected. Administration of 4-acetoxy Resveratrol decreased weight gain but not white adipose tissue in the male rats but has no effect in the females. In the male rat administration of 4-acetoxy resveratrol the high group also decreased testosterone, 5α-DHT , and prostate 5α-reductase activity. The high dose of 4-acetoxy Resveratrol also caused a change in prostate histology and decreased prostate weight. 4-acetoxy Resveratrol had no effect on testis weight and only showed a slight increase in depressive-like behaviors. In the females, 4-acetoxy resveratrol had no effect on white adipose tissue deposition, estrous cycle, hypothalamus aromatase activity, or depressive-like behaviors.
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Cao, Qian. "Novel approaches to enhance the protective immune responses of vaccines against Porcine Reproductive and Respiratory Syndrome Virus." Diss., Virginia Tech, 2018. http://hdl.handle.net/10919/92694.
Texte intégralRésumé :
Since late 1980s, porcine reproductive and respiratory syndrome virus (PRRSV) has emerged as the most economically important swine pathogen affecting pig industries worldwide. Vaccination is the principal means that have been used for prevention of PRRSV infection. However, the currently available vaccines for PRRSV are generally considered as not very effective. One of the major obstacles for developing an effective modified live-attenuated vaccine (MLV) with broad protection is the delayed and insufficient immune responses mounted by PRRSV, and the problem is further exacerbated by the antigenic variations of the constantly-evolving field strains of PRRSV.
In order to boost the immune response induced by the MLV vaccine virus, we evaluated the immunogenicity and vaccine efficacy of recombinant PRRSV MLVs expressing porcine IL-15 or IL-18 as adjuvants. The cytokine genes were fused with a GPI modification signal so that they are anchored onto the cell surface upon infection with the recombinant MLV. Both cytokines are successfully expressed on the cell membrane of porcine alveolar macrophage (PAMs) after recombinant MLVs infection in vitro. Subsequently, pigs vaccinated with cytokine-expressing recombinant PRRSV MLVs had an improved antiviral response of cytotoxic lymphocytes including natural killer (NK) cells and T cells, characterized by increased IFN-γ secretion and/or enhanced CD107a expression. The results offer a novel strategy to incorporate cytokine genes into PRRSV genome as potent bio-active adjuvants expressed by the vaccine virus itself.
Since we showed that PRRSV VR2385 down-regulated swine leukocyte antigen class I surface expression, naturally the next logical question is which viral protein is responsible for this down-regulation. To answer the question, we cloned and expressed all known PRRSV structural and non-structural proteins and examined which protein(s) is involved in SLA-I downregulation. Our results identified the newly-discovered nonstructural protein Nsp2TF of PRRSV as the main mediator in down-regulating SLA-I expression. We also demonstrated that the Nsp2TF-knockout mutant virus lost its function of negatively modulating SLA-I presentation compared to the wild-type virus. The results suggest that disruption of the Nsp2TF's ability to down-regulate SLA-I expression may improve the existing PRRSV vaccines towards a better CMI response against the virus.<br>PHD
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Pineyro, Pineiro Pablo Enrique. "Novel approaches towards vaccine developments against porcine circovirus type 2 and porcine reproductive and respiratory syndrome virus." Diss., Virginia Tech, 2015. http://hdl.handle.net/10919/77542.
Texte intégralRésumé :
Porcine circovirus type 2 (PCV2) is the causative agent of porcine circovirus-associated disease (PCVAD). Porcine reproductive and respiratory syndrome (PRRS) is caused by PRRS virus (PRRSV). Both PCV2 and PRRSV have caused devastating diseases in the swine industry worldwide, resulting in immense economic losses. One of the most common co-infections in the swine industry is PCV2 and PRRSV. The aim of this dissertation research is to explore different experimental approaches to develop novel vaccines against the two major pathogens affecting swine production and study the basic mechanisms that may be involved in viral pathogenesis.
Two types of porcine circovirus (PCV), PCV1 and PCV2, have been identified thus far. PCV1, first identified as a contaminant of the PK-15 cell line, is non-pathogenic and has a low prevalence in swine herds. PCV2 is highly prevalent in most swine-producing countries and is associated with clinical PCVAD. The non-pathogenic PCV1 shares similar genomic organization with PCV2. Previously, it has been demonstrated that a genetically modified infectious chimeric PCV1-2a virus can tolerate up to a 27 aa insertion in the C-terminus of the ORF2 without affecting infectivity and produce a dual immune response against PCV2cap and the inserted epitope tag. Therefore, we evaluated the use of the non-pathogenic PCV1 wild-type (wt) virus and chimeric PCV1-2a vaccine virus (vs) to express four known B-cell epitopes of PRRSV. Peptide epitopes of PRRSV-VR2385, including GP2II (aa 40–51, ASPSHVGWWSFA), GP3I (aa 61–72, QAAAEAYEPGRS), GP5I (aa 35–46, SSSNLQLIYNLT), and GP5IV (aa 187–200, TPVTRVSAEQWGRP) were inserted in frame into the C-terminus of the ORF2 of PCV1wt as well as the PCV1-2avs. Four PCV1-PRRSVEPI chimeric viruses and four PCV1-2a-PRRSVEPI chimeric viruses were successfully rescued and shown to be infectious in vitro and co-expressed PCV1cap or PCV2cap with each specific PRRSV epitope. Two independent animal studies were conducted to evaluate whether the non-pathogenic PCV1 can serve as a vaccine delivery vector and whether the PCV1-2a vaccine virus can be used to develop a bivalent vaccine against both PCV2 and PRRSV. We demonstrated that three PCV1-PRRSVEPI chimeric viruses and two PCV1-2a-PRRSVEPI chimeric viruses were infectious in pigs. Importantly, we demonstrated that the PCV1-PRRSVEPI and PCV1-2a-PRRSVEPI chimeric viruses not only induced specific PCV1 or PCV2 IgG antibody but also specific anti-PRRSV epitope antibody responses as well. Regardless of the PCV backbone used, we showed that the PCV-PRRSV chimeric viruses elicited neutralizing antibodies against PRRSV-VR2385. These results provided a proof of concept for the potential use of the non-pathogenic PCV1 as a vaccine delivery system for PRRSV or other swine pathogens and the use of PCV1-2a vaccine virus to generate a bivalent vaccine against both PCV2 and PRRSV.
PRRSV causes a persistent infection and immunosuppression. Immunomodulation of the host immune system is caused by modulation of numerous interleukins, such as type I interferons, tumor necrosis factor alpha (TNF-α), interleukin-1 (IL-1), interleukin-6 (IL-6), and interleukin-12 (IL-12) in infected pigs. Antigen-presenting cells (APCs) are the first line of defense, and their infection plays an important role in innate-mediated immune regulation during early immune responses. Among the APCs, pulmonary alveolar macrophages (PAMs), pulmonary interstitial macrophages (PIMs), and dendritic cells (DCs) are the main targets for PRRSV replication. The role of PRRSV-DCs interaction is not fully understood, and current research focuses on the production and regulation of interferons through DC-SIGN receptors. In this study, we evaluated the immunomodulation of MoDCs by PRRSV through interactions with the pDC-SIGN receptor, by blocking pDC-SIGN with recombinant hICAM-3-Fc or anti-pDC-SIGN mAb. Our results indicate that recombinant hICAM-3-Fc enhances mRNA expression of proinflammatory cytokines and that anti-pDC-SIGN mAb inhibits mRNA expression of TNF-α and IL-1α and enhances the expression of IL-12 induced by PRRSV in MoDCs. The results will help understand the molecular mechanisms of PRRSV pathogenesis.<br>Ph. D.
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Crozier, Tamara May. "The effects of reproductive experience on prefrontal cortex dependent learning and memory and pyramidal cell morphology in the rat dam." Thesis, University of British Columbia, 2010. http://hdl.handle.net/2429/29544.
Texte intégralRésumé :
Pregnancy, parturition, and motherhood, collectively known as reproductive experience, bring about profound and enduring changes in the hormonal, neural, and behavioral profile of the female rat. Much of the research to date investigating the effects of reproductive experience on learning and memory and cellular morphology in the rat dam has focused on the hippocampus. These studies revealed enhancements in spatial working and reference memory as well as alterations in pyramidal cell morphology following reproductive experience. Interestingly, it has long been established that other brain regions undergo persistent changes in response to reproductive experience including the prefrontal cortex, yet there remains a paucity of research investigating this area. Thus, the objective of the following experiments was to determine the effects of reproductive experience on prefrontal cortex-dependent learning and memory as well as pyramidal cell morphology in the prelimbic region in nulliparous, primiparous, and multiparous rats. For Experiment 1, age-matched nulli-, primi- and multiparous rats were tested for seventeen consecutive days using the delayed spatial win-shift task. This experiment revealed that multiparous rats committed fewer within-phase and omission errors than nulli- or primiparous rats on Blocks 2, 3, and 4 as well as committing fewer across-phase errors in Blocks 2 and 4 than either the nulli- or primiparous groups. Furthermore, the total number of within-phase errors significantly and negatively correlated with an increase in the total time engaged in nursing behaviors. Using Golgi impregnation, pyramidal cell morphology in Laminae 2/3 and 5 of the prelimbic region of the prefrontal cortex was examined in Experiment 2. The results of Experiment 2 revealed that multiparous rats have more total branch points in the apical region of Lamina 2/3. In addition, arched-back nursing was found to significantly positively correlate with the number of branch points in apical and basal regions of Lamina 5. Passive nursing significantly correlated with the number of basal branch points in Lamina 5 and apical length in Lamina 2/3. The findings from these studies suggest that multiparity may be necessary in realizing the effects of enhanced learning and memory and morphological changes associated with the prefrontal cortex in female rats.
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NOTARSTEFANO, VALENTINA. "Development of new molecular tools for the characterization of human Granulosa cells: new implications for the research on human infertility." Doctoral thesis, Università Politecnica delle Marche, 2018. http://hdl.handle.net/11566/252927.
Texte intégralRésumé :
Nella routine della riproduzione assistita, la selezione degli ovociti si basa sulle loro caratteristiche
morfologiche, nonostante la scarsa correlazione tra tali parametri e la competenza dell’ovocita. Pertanto,
la ricerca si sta concentrando sull’identificazione di marker che possano supportare l’attuale metodica di
valutazione, in particolare studiando le cellule della Granulosa (GCs), che svolgono compiti fondamentali
all’interno del microambiente del follicolo: produzione di estradiolo e progesterone, regolazione
dell’avanzamento della meiosi e dell’attività trascrizionale dell’ovocita, produzione di nutrienti essenziali
per l’ovocita, e accumulo di metaboliti secreti. Pertanto, la progressione degli step della follicologenesi
dipende strettamente dalla comunicazione bidirezionale tra ovocita e le cellule somatiche che lo
circondano.
Lo studio delle GCs ha messo in luce molti aspetti circa le funzionalità ovariche, la follicologenesi, e i
meccanismi con cui fattori endogeni ed esogeni possono alterare questi delicati processi.
In questo senso, lo scopo principale del presente PhD è stato quello di caratterizzare le GCs, applicando
per la prima volta la tecnica Fourier Transform Infrared Microspectroscopy (FTIRM), in modo da
sviluppare un nuovo metodo per valutare la qualità degli ovociti umani. Questo potrebbe rappresentare
uno strumento nuovo, affidabile e oggettivo per la valutazione della qualità degli ovociti nella routine
della riproduzione assistita.
Oltre al tema principale, sono state applicate le tecniche FTIRM, Raman Microspectroscopy e qPCR per
studiare i meccanismi di intaccamento del metabolismo, della composizione biochimica e dell’attività
cellulare causati da fattori endogeni ed esogeni, in particolare l’invecchiamento riproduttivo,
l’endometriosi ovarica e inquinanti plastici.
Le informazioni ottenute su queste cellule contribuiscono alla comprensione dei meccanismi di
danneggiamento della follicologenesi, tramite un approccio multidisciplinare che ha visto l’accoppiamento
di analisi spettroscopiche e qPCR. Grazie ai risultati ottenuti, è stato proposto un approccio innovativo
per l’analisi della qualità degli ovociti tramite caratteristiche spettrali delle GCs, suggerendo la possibilità
di un’applicazione dell’FTIRM come strumento diagnostico di facile utilizzo nella routine di riproduzione
assistita.<br>In assisted reproductive routine, oocyte selection is based on its morphological features, which seem not
to be related to its intrinsic competence. Hence, several efforts have been made to identify markers to be
added to the actual evaluation, in particular focusing on the crucial roles of Granulosa cells (GCs) in the
follicular microenvironment: production of estradiol and progesterone, regulation of the meiosis steps and
the transcriptional activity in the oocyte, production of essential nutrients for the oocyte, and
accumulation of secreted metabolites. Hence, the progression through the steps of folliculogenesis heavily
relies upon bi-directional interactions between germ cells and the surrounding somatic cells.
The study of GCs has shown to be determining to highlight particular features of ovarian mechanisms
and folliculogenesis, and also to identify the endogenous and exogenous factors that can impair these
delicate processes.
In this sense, the main aim of the PhD project was to characterize GCs, applying for the first time Fourier
Transform Infrared Microspectroscopy (FTIRM) to develop a method for evaluating the quality of human
oocytes. This could represent a new, reliable and objective tool for oocyte quality assessment in assisted
reproduction routine.
Besides this main topic, the impairment induced by endogenous and exogenous factors on the biochemical
composition, metabolism and cellular activity of GCs was also investigated by FTIRM, Raman
Microspectroscopy and qPCR, shedding new light on the mechanisms governing folliculogenesis. In
particular, the research was focused on the impairment determined by reproductive aging, ovarian
endometriosis, and plastic pollutants.
The information obtained on GCs contribute to the understanding of the mechanisms of impairment of
folliculogenesis, by a multidisciplinary approach made of spectroscopic analysis and qPCR. Thanks to the
results, an innovative approach to evaluate oocyte quality by spectral features of GCs was proposed,
suggesting the possibility to apply FTIRM as a clinical feasible diagnostic tool in assisted reproduction
routine.
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Shaan, Lakshmanappa Yashavanth. "Development and Evaluation of Efficacy of Novel Porcine Reproductive and Respiratory Syndrome (PRRS) Virus Vaccine Candidates in Pigs." The Ohio State University, 2018. http://rave.ohiolink.edu/etdc/view?acc_num=osu1532064253191032.
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Madouasse, Aurélien. "An evaluation of milk recording, somatic cell counts and reproductive performance in a large cohort of dairy herds in England and Wales." Thesis, University of Nottingham, 2010. http://eprints.nottingham.ac.uk/11203/.
Texte intégralRésumé :
Milk recording consists in the regular, usually monthly, collection of a milk sample from all the lactating cows of a dairy herd. A large sample of milk recording data collected in England and Wales between 2004 and 2006 was used in this thesis. A sample of 8,211,988 recordings in 2,128 herds, representing 16 % of the dairy herds in activity in December 2006, were described and analysed. Calvings followed a seasonal pattern with 80 % more calving in September than in May. Milk production was highest in May (26.5 kg) and lowest in October (24.1 kg). Butterfat was stable, close to 4 % from October to March and reached a minimum at 3.7 % in June and July. Protein stayed between 3.2 and 3.3 % all the year. Geometric mean somatic cell count was between 177 and 180 between October and March and reached 205,000 cells/mL in July and August. At the individual cow level, the mean milk yield, percentage of butterfat, percentage of protein, fat to protein ratio and somatic cell count (geometric mean) were 26.4 kg, 3.96 %, 3.29 %, 1.21 and 90,000 cells/mL, between 5 and 305 days in milk. Changes in individual cow somatic cell counts around a threshold of 200,000 cells/mL between consecutive recording dates were used to predict bulk milk somatic cell count at both the herd-year and test-day levels. The main contributors to bulk milk somatic cell counts were cows staying above the threshold for 2 consecutive test-days. Milk yields and composition at the start of lactation were used to predict the calving to conception interval. Higher milk yield, lower percentage of protein, lower percentage of lactose, higher somatic cell count and higher percentage of butterfat were associated with lower probabilities of conception before 145 days in milk.
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Bowers, Hannah Elizabeth, and Jennifer Hall. "THE EFFECTS OF ESTROGEN-INDUCED STROMAL CELL EFFECTORS, OSTEOPONTIN AND VIMENTIN, ON CHLAMYDIA INFECTIONS IN A NON-POLARIZED CELL CULTURE MODEL." Digital Commons @ East Tennessee State University, 2018. https://dc.etsu.edu/asrf/2018/schedule/98.
Texte intégralRésumé :
Chlamydia is the most reported sexually transmitted infection in the US and is caused by the obligate intracellular bacterium Chlamydia trachomatis. Typically, this presents as a lower genital tract infection (cervicitis or urethritis), but can ascend to the upper genital tract, causing pelvic inflammatory disease, tubal infertility, epididymitis, or ectopic pregnancy. While chlamydia infections can be cured with a single-dose oral antibiotic, repeat infections are common and having multiple chlamydial infections increases a woman’s risk of developing serious chronic conditions. Previous research has shown that estrogen has a positive effect on C. trachomatis infections—an important finding, connecting fluctuating estrogen levels in females to variance in pathogenesis.The mechanism behind this hormonal influence remains unknown; however, previous work in our laboratory indicates that estrogen-stimulated stromal cell effectors play a role in enhancing C. trachomatis infections in a polarized endometrial epithelial Ishikawa (IK)/stromal (SHT-290) cell co-culture model. Specifically, our data indicate that estrogen exposure stimulates osteopontin and vimentin release from stromal cells in co-culture with endometrial epithelial cells. Furthermore, we noted that Chlamydia-infected, polarized Ishikawa cells exposed to a combination of recombinant osteopontin and estrogen released significantly more infectious chlamydia than cultures exposed to estrogen alone. Most tissue culture models being used today employee non-polarized cells. Given the fact that epithelial cell polarization is known to impact C. trachomatis serovar E development, in the current study we sought to determine if the estrogen-induced stromal cell effectors, osteopontin and vimentin, affect C. trachomatis viability and infectivity in non-polarized Ishikawa cells. Non-polarized Ishikawa cells were exposed to osteopontin or vimentin in the presence or absence of estrogen, infected with C. trachomatis serovar E, and collected for examination of chlamydial infectivity and progeny production. Our initial data show that osteopontin and vimentin impact chlamydial progeny production in a concentration dependent fashion, with higher concentrations of recombinant effectors +/- estrogen significantly decreasing progeny production. These data suggest that polarization of host cells influences the way hormone-stimulated effectors interact with the cell to impact on chlamydial infection. Future research goals are to explore other stromal effectors such as fibronectin with estrogen and to study the cell signaling mechanism osteopontin and vimentin use to affect chlamydial infections in polarized epithelial cell cultures.
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Lu, Zhigang [Verfasser]. "Towards an in-depth view of the reproductive biology of Schistosoma mansoni : from gonad isolation to sub-transcriptomics and cell culture / Zhigang Lu." Gießen : Universitätsbibliothek, 2016. http://d-nb.info/1082503878/34.
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Mokhtar, Helen. "Characterisation of the T cell response to the porcine reproductive and respiratory syndrome virus and its application towards the development of improved vaccines." Thesis, University of Surrey, 2015. http://epubs.surrey.ac.uk/807548/.
Texte intégralRésumé :
Porcine reproductive and respiratory syndrome (PRRS) is one of the most important pig diseases worldwide. The causative PRRS virus (PRRSV) is rapidly evolving and there is an urgent need for the development of safer and more efficacious vaccines to improve PRRS control. Immunity to PRRSV is not well understood but there are data suggesting that virus-specific T cell IFN-γ responses play an important role. Therefore, this project focussed on characterising the T cell response PRRSV and applied this knowledge to develop a novel vaccine strategy. The first part of the project aimed to identify the antigenic targets of the T cell response to PRRSV by utilising a proteome-wide synthetic peptide library and a cohort of PRRSV immune animals. The T cell IFN-γ response was directed at a range of viral proteins but the M and NSP5 proteins stood out as major antigens. Further experiments confirmed M and NSP5 as well conserved targets of in many instances dominant T cell responses. Characterisation of the responding T cell populations showed NSP5-specific responders to be CD8 T cells with a predominant CD44highCD62LlowCD27lowCD25- phenotype. The majority of cells were polyfunctional as assessed by co-expression of TNF-α and mobilisation of the cytotoxic degranulation marker CD107a. Both CD8 and CD4 T cells responded to M with a comparable phenotype to that observed for NSP-specific T cells. In addition, conserved antigenic regions of each protein were identified and specificity shown to associate with major histocompatibility complex haplotype, rather than PRRSV strain. Finally, a vaccine study was conducted using M and NSP5 proteins as T cell antigens formulated as a particulate vaccine with a molecular adjuvant. Vaccination primed antigen-specific CD4 but not CD8 T cell responses and did not confer significant protection of animals from viraemia upon challenge infection. Analysis of the lungs during the resolution of infection showed high levels of virus and M/NSP5 specific CD8 T cell IFN-γ responses, suggesting that vaccine priming of a CD8 T cell response is required for protection from PRRSV infection. It is hoped that this work will inform future PRRSV vaccine design, as well as contributing to the wider field of T cell vaccinology.
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Jent, Karen Ingeborg. "Making stem cell niches : an ethnography of regenerative medicine in Scotland and the United States." Thesis, University of Cambridge, 2018. https://www.repository.cam.ac.uk/handle/1810/279088.
Texte intégralRésumé :
This thesis presents the findings from an ethnography of stem cell science based on fieldwork with researchers in two connected laboratories in Scotland and the United States. It explores stem cell scientists' complicated interactions with live stem cell cultures within national projects of translational regenerative medicine. This analysis both draws upon and contributes to the social studies of biomedicine, reproductive studies and science and technology studies. I examine how stem cell scientists, involved in an international research initiative, navigate the challenging landscapes of translational regenerative medicine and attempt to transform fragile live cell cultures into successful biotechnical, medical and economic products. By considering translational regenerative medicine as an effort to reformulate the relationship between biology and technology in terms of applicability and utility, I illuminate tensions between the specific practices of care that enable stem cell growth in vitro and the elusive goals of national projects of biotechnological innovation. A major focus of this study is the means by which scientists in the two laboratories manage the inherent uncertainties of both cell culture and translational science. By exploring how researchers react to unstable and unpredictable cellular behaviour in the laboratory, while also managing the expectations of government and external funding bodies, I provide a portrait of the complex sociality of contemporary bioscience. In addition to the international collaboration between the two laboratories, I explore scientists' interdisciplinary work with medical specialists and public engagement with stakeholders in regenerative medicine. In doing so, I pay attention to the ways in which scientists themselves deal with and reflect on the relational and interdependent nature of their endeavours. Drawing on twenty-two months of ethnographic fieldwork and fifty qualitative interviews, I show how stem cell scientists' new engagement practices also inform scientific work and the care of stem cells in the laboratory. In short, I argue that translation of science across different sites at once creates and depends on new social relations between stem cells, people and communities. After providing an overview of the literature, central questions and methodology that frame this thesis, I introduce the opportunities and challenges that translational regenerative medicine goals create for the care of stem cells in vitro. From there, I zoom out beyond the tissue culture flask to demonstrate how the necessity for science applicability creates new responsibilities for scientists to connect with stakeholders in regenerative medicine outside of the laboratory. I conclude that a consideration of scientists' ties and societal links is significant for an understanding of the connection between the biological and the technological.
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Richmond, Owen Benjamin. "Immune modulation mechanisms of porcine circovirus type 2." Diss., Virginia Tech, 2015. http://hdl.handle.net/10919/73766.
Texte intégralRésumé :
Porcine circovirus associated disease (PCVAD) is an umbrella term for a multitude of diseases and syndromes that have a negative impact on the health and economics of pig production operations throughout the world. Porcine circovirus type 2 is the causative agent of PCVAD; however the presence of PCV2 alone is rarely enough to cause clinical disease. In order for the full development of PCVAD the presence of a co-infecting pathogen is required. The mechanisms by which co-infection leads to disease remain ongoing areas of research, but it is thought that host immune modulations by PCV2 or a co-infecting pathogen are critical in the pathogenesis of PCVAD. In the first study of this dissertation the ability of PCV2 to induce regulatory T-cells (Tregs) and alter cytokine production was evaluated in vivo. The addition of PCV2 to a multiple viral challenge resulted in a significant increase in Tregs. Levels of IL-10 and IFN-γ were also found to be altered when PCV2 was added to a multiple viral challenge. In further experiments, monocyte derived dendritic cells (MoDC) were infected with different combinations and strains of PCV2 and PRRSV in vitro and evaluated for expression levels of programmed death ligand-1 (PD-L1), IL-10, CD86, swine leukocyte antigen-1 (SLA-1), and swine leukocyte antigen-2 (SLA-2). Expression levels of PD-L1 were significantly increased in PCV2 and PRRSV co-infected MoDCs. SLA-1, SLA-2, and CD86 expression levels were significantly decreased in the MoDC treatment groups containing both PCV2 and virulent stains of PRRSV. MoDC IL-10 expression was significantly increased by PCV2 and virulent strains of PRRSV co-infection. Finally, we investigated the role of the PD-L1/programmed death ligand-1 (PD-1) axis in porcine lymphocyte anergy, apoptosis, and the induction of Tregs. Lymphocyte populations with normal PD-1 expression had significantly higher percentages of anergic and apoptotic lymphocytes, and CD4+CD25HighFoxP3+ Tregs when compared to a PD-1 deficient lymphocyte population. The findings from these studies indicate host immune modulation by PCV2 in vivo and the development of a regulatory phenotype of dendritic cell following PCV2/PRRSV co-infections in vitro that may contribute to a dysfunctional adaptive immune response and the overall pathogenesis of PCVAD.<br>Ph. D.
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Gaynor, Louise Michelle. "Immunogenetic regulation of Natural Killer cell function in pregnancy." Thesis, University of Cambridge, 2017. https://www.repository.cam.ac.uk/handle/1810/270329.
Texte intégralRésumé :
Uterine NK (uNK) cells are a distinct subset of NK cells in the decidua of humans and rodents during pregnancy, which are essential for remodelling of the spiral arteries supplying the feto-placental unit. Similarly to peripheral NK cells, uNK cells express Natural Killer receptors (NKRs) that engage MHC class I molecules. Evidence from human genetic association studies suggests that, in the presence of allogeneic cognate paternal MHC class I ligands, inhibitory uterine NKRs are associated with disorders of pregnancy arising from impaired decidual vascular remodelling. Conversely, enhancement of human uNK cell activity through activating NKRs is associated with high birth weight. Evidence from mouse models corroborates that uNK cell activity is modulated by interactions between NKRs and MHC class I, but has largely focussed on the effect of paternal MHC. In this study, the contribution of maternal immunogenetic regulation of NK cell function to reproductive outcome was assessed independently of parental MHC disparity in mice. To evaluate the role of NKR genes in isolation, I used congenic B6.BALB-TC1 (TC1) mice that differ from C57BL/6 (B6) mice only within the region of chromosome six encoding NKRs that recognise MHC class I. Absence of a major inhibitory NKR for self-MHC, Ly49I, in TC1 mice causes a compensatory shift in the NKR repertoire expressed and preserves a majority subpopulation of educated NK cells. B6 and TC1 splenic and uterine NK cells are similarly functionally reactive and mature, and no significant differences could be detected in spiral arterial remodelling or fetal growth between these strains in MHC-syngeneic matings. This supports data from human immunogenetic studies showing that maternal uterine NKRs are not associated with differences in pregnancy outcome in the absence of novel paternal MHC class I ligands, and highlights the importance of maternal and paternal co-regulation of uNK cell activity during pregnancy. No mouse models of uNK cell activation are currently available with which to corroborate human immunogenetic associations between activating uterine NKRs and high birth weight. Male m157-transgenic (m157-Tg) mice, which ubiquitously express viral m157 glycoprotein ligands for the activating NKR Ly49H, were mated with B6 females. Exclusive expression of m157 glycoprotein by trophoblast improved placental efficiency, but did not enhance fetal growth. Some fertility clinics surmise that uNK cell activation initiates the pathogenesis of spontaneous abortion. It has been suggested that this may occur due to reduced expression by human uNK cells of miR-483-3p, which stimulates endogenous insulin-like growth factor (IGF)-1 production and uNK cell cytotoxicity in vitro. It is demonstrated here that neither miR-483-3p nor IGF-1 regulate murine NK cell development, maturation or function. No discernible reproductive phenotype is evident in miR-483 deficient females. It can be inferred that post-transcriptional control by miR-483 is not biologically relevant to murine NK cell function. Although m157-Tg mice may provide an interesting model to further study uNK cell-mediated placental adaptations, it remains important to identify a murine model of enhanced uNK cell function to corroborate human immunogenetic associations with high birth weight and to challenge the supposition that uNK cell activation is harmful to pregnancy.
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Dean, Afshan. "Androgens and the masculinisation programming window." Thesis, University of Edinburgh, 2012. http://hdl.handle.net/1842/6516.
Texte intégralRésumé :
The commonest reproductive disorders of young men (namely low sperm counts, testicular germ cell cancer) may originate in fetal life similar to established disorders (cryptorchidism, hypospadias) that manifest at birth. These disorders are interlinked and may comprise a testicular dysgenesis syndrome (TDS), a concept supported by animal model studies. The latter have identified the likely time-frame within which TDS disorders may be induced, namely within the so-called masculinisation programming window (MPW). During this critical period, sufficient testosterone (androgen) must be produced by the fetal testis to program the male reproductive tract so that it will differentiate and grow normally after the MPW. Impaired androgen production or action within the MPW can result in smaller reproductive organs and their abnormal formation and function (e.g. cryptorchidism, hypospadias). The MPW is thus of fundamental importance in determining normal, or abnormal, male reproductive development and function for later life. There are two big unanswered questions about the MPW. First, what determines its timing? Second, what mechanisms are controlled by androgens specifically within this time-window and not at later time points? Three approaches were undertaken to address the first question experimentally in rats. First, investigation of whether the availability of androgens and or androgen receptors (AR) plays a role in determining the onset or ‘opening’ of the MPW. Second, investigation of whether the expression of AR co-regulators was a factor in determining androgen sensitivity during the MPW. Third, investigation of whether prostaglandins played a role in mediating androgen action in the MPW, as studies in the 1980s had suggested this possibility. To address what mechanisms are controlled by androgens specifically within the MPW, the expression of selected genes in the genital tubercle was investigated before, during and after the MPW in fetuses that had been exposed to treatments that modulated androgen action. Selection of genes was based on microarray studies and data reported in the literature (ie candidate genes). The studies reported in this thesis show that neither availability of androgens nor the AR are important in determining onset of the MPW, and providing exogenous androgens either prior to or during the MPW does not advance or enhance masculinisation. These studies also showed that females may have a slightly different window of susceptibility to androgen action than do males. Key AR co-regulators have been characterized in the male reproductive tract for the first time, two of which (BRG1, CBP) show changes in expression through development of the testis consistent with a role in Sertoli cells. Another AR co-regulator, RWDD1, was found to switch off in the absence of androgen action in the genital tubercle, pointing to a potential role during and/or after the MPW. Studies involving gestational exposure to indomethacin (a compound which inhibits prostaglandin synthesis) during the MPW showed no detectable effect on masculinisation. Finally, evaluation of candidate genes for mediating androgen action in the genital tubercle during the MPW, failed to identify their key involvement, thus they are unlikely to be involved in penis development and disorders such as hypospadias.
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Liu, Yanhe. "Human oocytes and embryos viewed by time-lapse videography, and the development of an embryo deselection model." Thesis, Edith Cowan University, Research Online, Perth, Western Australia, 2016. https://ro.ecu.edu.au/theses/1787.
Texte intégralRésumé :
Despite its wide application today, in vitro fertilization (IVF) treatment continues to have relatively low efficacy, largely due to inaccuracy in selecting the best quality embryo(s) from the cohort for transfer. Novel methodologies for improved selection are being developed, and time-lapse observation of human embryos is gaining increasing popularity due to the more detailed morphokinetic information obtained plus uninterrupted culture conditions. The morphokinetic information enables the use of quantitative timings in developmental milestones of embryos and qualitative measures of abnormal biological events, to assist embryo selection/deselection. This project aimed to identify current limitations in the use of such measures and to develop recommendations for improvement in clinical application.
In the current study, most data were collected retrospectively from infertile couples seeking IVF treatment at a fertility clinic, with consent to use time-lapse incubation (Embryoscope) for embryo culture. Comparisons of time-lapse measures were made between embryos with confirmed implantation and non-implantation outcomes following uterine transfers. Thereafter, an embryo deselection model was proposed based on the retrospective findings, followed by prospective validation.
It was found in the current study that the reference starting time point (t0) in certain existing time-lapse systems was inaccurate due to (i) the early biological variations between sibling oocytes, (ii) technical limitations in current equipment and protocols, and (iii) different insemination methods used (Papers 1&2). The above variations may be minimized by using pronuclear fading (PNF, a biological time point) as t0 rather than insemination (a procedural time point) (Paper 2). An example of such application was the comparison of embryo development between patients with high and low serum progesterone levels on the trigger-day (Paper 3). Furthermore, the growth rate of embryos reported in the literature is subject to multiple clinical or laboratory factors, and this was in agreement with the present study where a published time-lapse algorithm emphasizing quantitative timing parameters was shown to lose its discriminatory power in implantation prediction when applied in two different laboratories (Paper 4). Interestingly, the qualitative measures seemed to have better inter-laboratory transferability due to the embryo growth patterns appearing independent of clinical and technical factors (Paper 4). Two novel qualitative measures were reported in the present study, namely reverse cleavage and less than 6 intercellular contact points at the end of the 4-cell stage, showing negative correlations with embryo implantation outcomes (Papers 5&6). A qualitative embryo deselection model was therefore proposed, including several qualitative measures with implantation rates being potentially increased from 22.4% to 33.6% (Paper 6). Finally, an embryo deselection model combining both qualitative and quantitative measures was reported with the use of PNF as t0, showing significant prediction of implantation outcomes in embryos regardless of insemination method (Paper 7).
In conclusion, this thesis demonstrates the usefulness of time-lapse embryo selection during IVF treatment in one specific laboratory. However, any new time-lapse parameter or model for embryo selection requires external validation by properly designed large-scale studies. Future clinical research and the development of integrated engineering and computer technology may further improve the efficacy of time-lapse selection of human embryos.
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Taylor, Jacqueline Susan. "The Effect of Pyrethroid Compounds on the Expression of Estrogen Receptors in Mouse Sertoli Cells and Implications for Male Infertility." Thesis, University of Canterbury. Biological Sciences, 2006. http://hdl.handle.net/10092/1482.
Texte intégralRésumé :
Male fertility is largely controlled by the hypothalamic-pituitary axis, a careful balance between stimulating and suppressing gene expression and the secretion of hormones. The critical factors for male fertility have in the past been thought to be limited to testosterone and the gonadotropins. Estrogen has only recently been demonstrated to be both a crucial requirement for fertility and a cause of infertility. Reports in the early 1990s demonstrated a decrease in mean sperm counts over the last 50 years. A hypothesis for this observation is the increase of xenoestrogens in the environment that are able to mimic and potential disrupt the natural estrogens involvement in fertility. Although the mechanisms of estrogens involvement are not yet defined, the Sertoli cells are a potential sites of action as they possess receptors for the hormone and are able to locally produce it. Sertoli cells both act to protect and provide for the male germ cells and the developing spermatozoa. Pyrethroids are common synthetic insecticides of which some have previously shown estrogenic activity. Therefore this investigation examined the effects of pyrethoids, whose estrogenicity was confirmed via the yeast assay, on the estrogen receptor expression in mouse Sertoli cells as a model for general effects of estrogenic chemicals on male fertility. The results first confirmed the estrogenicity of some pyrethroids and these pyrethroids when exposed to mouse Sertoli cells effected estrogen receptor mRNA expression however in a different way to the natural ligand 17β-estradiol.
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PERRINO, STEFANIA PIA. "La natura giuridica dell'embrione." Doctoral thesis, Università degli Studi di Milano-Bicocca, 2021. http://hdl.handle.net/10281/314067.
Texte intégralRésumé :
Il problema qualificatorio dell’embrione costituisce il banco di prova per il civilista dinanzi all’irrompere delle innovazioni in ambito bio-tech.
Se lo status del concepito ha rappresentato il terreno di contesa tra le tesi dottrinali più disparate e della giurisprudenza più recente, lo statuto giuridico dell’embrione extra-corporeo costituisce un tema nuovo ma ugualmente controverso.
L’elaborato muove da una considerazione generale: non esiste una disciplina unitaria a tutela dell’embrione, fatta eccezione per una serie di laconiche disposizioni normative e sanzioni, penali ed amministrative, contenute nella l. 19 febbraio 2004, n. 40, dedicata prevalentemente alle tecniche di fecondazione assistita. Il legislatore nel 2004 ha colto l’occasione per regolare, da un lato e con poche norme, il diritto sulla scienza del nuovo millennio e, dall’altro, la procreatica asessuata per il superamento di condizioni di infertilità e la soddisfazione del progetto parentale di molti.
Ne deriva che il successo raggiunto dalla medicina riproduttiva degli ultimi quindici anni non si è accompagnato ad un utilizzo dello strumento legislativo capace di cogliere la cifra dell’avanzamento scientifico e ne costituisce prova palmare la «sfibrante dialettica» di cui è destinataria la normativa in materia. La “legge 40” non ha mai ottenuto un risolutorio intervento legislativo volto a disciplinare la fecondazione eterologa, la diagnosi genetica preimpianto e la condizione degli embrioni crioconservati e prodotti in soprannumero. Istituti, questi, transitati nell’ordinamento italiano solo per effetto dei numerosi interventi demolitori della Corte costituzionale.
Sicché la legge tutela, per il tramite di divieti, talvolta dichiarati incostituzionali, una entità, quale è l’embrione extrauterino, di cui il legislatore non ha chiari i contorni, né tantomeno la natura giuridica. E dunque, pure in difetto di un chiaro inquadramento, se in passato uno statuto speciale poteva essere evinto dall’interprete in capo all’embrione attraverso le lenti di tali divieti, occorre chiedersi quale sia il suo statuto all’esito di una interpretazione attenta, sistematica ed attuale.
L’elaborato, allora, propone un inquadramento giuridico dell’embrione ed una ricostruzione della disciplina applicabile, all’esito dell’esame puntuale della stratificata trama normativa dedicata alla procreazione medicalmente assistita e, più in generale, alla cessione e all’impiego di tessuti ed alle cellule umane nel Biodiritto italiano e sovranazionale.
Per quanto concerne l’embrione, oggetto della presente disamina, sono state analizzate diverse impostazioni interpretative, capaci di orientare il giurista nella selezione delle regole applicabili. Queste teorie vengono suddivise a seconda del metodo: le prime nel solco del metodo dell’adattamento, l’ultima, propugnata nel presente elaborato, secondo il metodo della innovazione.
Tra le prime si evidenziano le teorie dell’embrione come concepito, come soggetto di diritto, come fanciullo, nonché le teorie dell’embrione come cosa in senso giuridico e come parte del corpo.
Tuttavia, l’adeguamento delle citate categorie dogmatiche risalenti non ha fornito risultati appaganti, poiché costituisce un espediente incapace di fronteggiare le questioni più complesse emerse nel trattamento della entità in discorso. Allora, seguendo gli insegnamenti di autorevoli interpreti del diritto civile, si intraprende il percorso più arduo, ossia l’applicazione del metodo dell’innovazione. Per questo si procede all’elaborazione della teoria processuale, in virtù della quale va contestato un inquadramento unitario e va, invece, preso in considerazione il processo evolutivo che contraddistingue la “vita nascente”. Sicché per ciascuna fase e contesto il giurista individua un trattamento differenziato a seconda del segmento temporale di riferimento.<br>The qualifying problem of the embryo constitutes the test bed for the civilist in the face of the bursting of innovations in the bio-tech field.
While the status of the conceived has been the battleground between the most disparate doctrinal theses and the most recent jurisprudence, the legal status of the extracorporeal embryo is a new but equally controversial issue.
The paper moves from a general consideration: there is no unitary discipline to protect the embryo, except for a series of laconic regulatory provisions and penalties, criminal and administrative, contained in the Law February 19, 2004, n. 40, dedicated mainly to assisted fertilization techniques. The legislator in 2004 took the opportunity to regulate, on the one hand and with few rules, the science law of the new millennium and, on the other hand, the asexual procreatic for the overcoming of infertility conditions and the satisfaction of the parental project of many.
It follows that the success achieved by reproductive medicine in the last fifteen years has not been accompanied by the use of the legislative instrument capable of grasping the figure of scientific advancement and constitutes handheld proof of it the "exhausting dialectic" of which the legislation on the subject is addressed. The "law 40" has never obtained a resolute legislative intervention to regulate heterologous fertilization, pre-implantation genetic diagnosis and the condition of cryopreserved and supernumerary produced embryos. Institutes, these, passed through the Italian legal system only as a result of the numerous demolition interventions of the Constitutional Court.
So the law protects, by means of prohibitions, sometimes declared unconstitutional, an entity, such as the extrauterine embryo, of which the legislator has no clear contours, nor the legal nature. And therefore, even in the absence of a clear framework, if in the past a special statute could have been avoided by the interpreter at the embryo through the lens of these prohibitions, it is necessary to ask what is its status as a result of a careful, systematic and current interpretation.
The paper, then, proposes a legal framework of the embryo and a reconstruction of the applicable discipline, the result of a detailed examination of the stratified regulatory framework dedicated to medically assisted procreation and, more generally, the transfer and use of human tissues and cells in the Italian and supranational Biolaw.
With regard to the embryo, the subject of this examination, different interpretative approaches have been analyzed, able to guide the jurist in the selection of the applicable rules. These theories are divided according to the method: the first in the groove of the method of adaptation, the last, advocated in this paper, according to the method of innovation.
Among the first are the theories of the embryo as conceived, as a subject of law, as a child, and the theories of the embryo as a thing in the legal sense and as part of the body.
However, the adaptation of the above mentioned dogmatic categories has not provided satisfactory results, since it is a gimmick incapable of dealing with the most complex issues that have emerged in the treatment of the entity in question. Then, following the teachings of authoritative interpreters of civil law, the most difficult path is taken, namely the application of the method of innovation. For this we proceed to the elaboration of the procedural theory, by virtue of which a unitary framework must be challenged and, instead, the evolutionary process that characterizes the "nascent life" must be taken into consideration. So for each phase and context the jurist identifies a differentiated treatment according to the time segment of reference.
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Gawriluk, Thomas R. "Targeted Knockout of Beclin-1 Reveals an Essential Function in Ovary and Testis." UKnowledge, 2014. http://uknowledge.uky.edu/biology_etds/19.
Texte intégralRésumé :
An estimated 12% of couples worldwide are infertile. The contributing factor is approximately equal between men and women with nearly 25% diagnosed as idiopathic. Despite the increasing numbers of couples seeking assistance from infertility clinics, few molecular mechanisms have been identified for treatment. Autophagy is an evolutionarily conserved cellular process for bulk degradation and recycling of cytosolic components through the lysosome to maintain homeostasis. Several studies have observed increased levels of autophagy during ovarian folliculogenesis and gonadal steroidogenesis; however, no genetic studies to determine the significance of autophagy exist.
To investigate the function of autophagy in the ovary and testis, a directed genetic knockout approach was used to independently knockout two key autophagy genes, Becn1 and Atg7. Chapter 2 reports that deficiency of Becn1 results in 56% fewer primordial follicles at postnatal day 1. In addition, Atg7 knockout mice do not have identifiable primordial follicles, suggesting that autophagy is necessary for survival of female germ cells during embryogenesis. Chapter 3 presents that Becn1 is necessary to sustain pregnancy and the deficiency of Becn1 in granulosa cells is a novel genetic model to study preterm labor due to impaired corpora lutea function. The results indicate that Becn1 is necessary for lipid droplet formation and subsequent progesterone production in luteal cells. In contrast, Atg7 is not necessary and deficiency results in overproduction of progesterone throughout pregnancy, suggesting that the defect in Becn1 conditional knockout mice is additional to autophagy. Chapter 4 presents that Sertoli cell expression of Becn1 is required for spermatogenesis after 8 weeks of age. Beyond 9-weeks-old, Becn1 conditional knockout mice are unable to sire a litter due to a failure of spermatogenesis and a Sertoli-cell-only phenotype in a majority of the seminiferous tubules. Atg7 was also identified as a necessary factor for spermatogenesis beyond 26-weeks-old. Together the data presented in Chapter 4 suggests that autophagy is necessary for adult Sertoli cell function. Primarily, this dissertation presents data from the first functional studies on autophagy in the reproductive tract. The results demonstrate an understanding of the functional significance for Becn1 and Atg7 in both the ovary and testis.
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Hadjisavas, Michael. "Induction of mitogenesis and cell-cell adhesion by porcine seminal plasma." Title page, contents and abstract only, 1992. http://web4.library.adelaide.edu.au/theses/09PH/09phh1293.pdf.
Texte intégralRésumé :
Includes list of publications by the author. Includes bibliographical references (leaves 103-123) Evaluates the nature of the interactions occurring between semen and cells of the uterus that occur following mating in pigs. Describes a novel ability of porcine seminal plasma to induce dose dependent cell-cell adhesion and mitogenesis amongst peripheral blood lymphocytes in vitro.
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Fraga, Cecília Archer de Menezes Castro. "Medicina da produção leiteira : estudos de relação entre o desempenho reprodutivo e as mastites." Master's thesis, Universidade de Lisboa, Faculdade de Medicina Veterinária, 2018. http://hdl.handle.net/10400.5/15046.
Texte intégralRésumé :
Dissertação de Mestrado Integrado em Medicina Veterinária<br>O impacto das mastites na eficiência reprodutiva dos bovinos leiteiros, tem sido estudado ao longo dos últimos anos, por diversos autores. O objetivo do presente estudo, foi analisar a possível associação causal entre a ocorrência de mastites subclínicas e a alteração do desempenho reprodutivo.
Recorrendo a uma base de dados de contraste leiteiro, na qual existiam registos provenientes de 9 explorações nacionais, introduzidos entre 1996 e 2016, procedeu-se à análise da relação da contagem de células somáticas, com o intervalo parto-conceção. Os resultados obtidos demonstraram que, à medida que aumenta a contagem de células somáticas, aumenta o intervalo parto-conceção. Tais resultados foram estatisticamente significativos e mais pronunciados para os dados relativos à segunda lactação. Observou-se ainda que, para cada aumento do número de eventos, cuja contagem de células somáticas se encontra acima do limiar de distinção entre infetado e saudável, o intervalo parto-conceção aumenta 28,7 dias para a primeira lactação e 27,9 dias para a segunda. Tais resultados, sugerem a hipótese de haver uma relação da cronicidade da infeção, com a fertilidade.
Assim, à semelhança do reportado por autores anteriores, os resultados obtidos apontam a existência de uma relação entre as mastites subclínicas e o desempenho reprodutivo dos bovinos leiteiros.<br>ABSTRACT - Dairy Production Medicine: studies of the relationship between reproductive performance and mastitis - The impact of mastitis on reproductive performance of dairy cattle has been studied throughout the last years, by several authors. The objective of this study was to evaluate the possible causal association between the occurrence of subclinical mastitis and altered reproductive performance.
Through the analysis of a database, in which there was access to milk recording data from 9 national dairy farms, introduced from 1996 to 2016, the relationship between the somatic cell count and the calving-to-conception interval was analyzed. The obtained results showed that, as the somatic cell count increases, the calving-to-conception interval increases. These results were statistically significant and more pronounced on the second lactation. Furthermore, it was observed that, with increases in the number of mastitis episodes, in which somatic cell counts were above the considered threshold between infected and healthy, the calving-to-conception interval progressively increased 28,7 days for the first lactation and 27,9 days for the second. These results support the existence of a relationship between the chronicity of the episodes and fertility.
As such, similarly to results presented by previous authors, this study supports the existence of a direct relationship between mastitis and the reproductive performance of dairy cattle.<br>N/A
47
Kieckbusch, Jens. "How do natural killer cells contribute to reproductive success?" Thesis, University of Cambridge, 2015. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.708449.
Texte intégral
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Sazzad, TM Shahriar. "An automated approach to identify nongrowing follicles using digitized images of type P63 histopathology ovarian slides." Thesis, Edith Cowan University, Research Online, Perth, Western Australia, 2017. https://ro.ecu.edu.au/theses/2032.
Texte intégralRésumé :
Many developing countries are still facing challenges with limited access to fertility health services. Women face problems in conceiving due to many factors such as increasing age. In vitro fertilization (IVF) treatments can assist these women but are considered too expensive. Medical pathology laboratories are searching for novel technologies that can improve microscopic slide testing of female ovarian reproductive tissues. Current electronic methods for the assessment of human ovaries are not suitable for analysis of ovarian reproductive tissues. Ultrasound method cannot be used to identify small ovarian Non-Growing Follicles (NGFs) that are responsible for reproduction. A computer assisted approach to overcome the problems associated with manual microscopic analysis of ovarian reproductive tissues could be beneficial in increasing the accuracy and speed of the analysis. Few studies have reported on the use of images and other artificial intelligence techniques for ovarian tissue samples and have mostly concentrated on the analysis of cancer cells or ovarian animal tissues which are different from human ovarian reproductive tissues. Other studies using human ovarian reproductive tissues have been limited in terms of accuracy. This research examines the possibility of developing an automated computer approach which will improve the practices of these pathology laboratories to analyse female ovarian reproductive tissues and assist medical practitioners to provide necessary fertility treatment. The objective of the research was to study existing computerized methods used in various tissue assessments; identify the gaps and limitations; and to propose a novel method on digitized colour images acquired from ovarian reproductive tissue slides. The following major research question has been addressed by the research study: “How to develop an automated approach to assist pathology experts to identify ovarian reproductive NGFs (Non-growing follicles) or simply ovarian reproductive tissues using digital images acquired from type P63 (counter and non-counter stained) histopathology ovarian biopsy slides?” In order to answer this question, the research was carried out in a number of phases to examine existing computerized techniques for impact assessment of the ovarian reproductive tissue analysis. The research used a mixed method approach based on a case study using experimental and engineering methodologies. The study also employed quantitative and statistical data analysis methods. The research was carried out as a series of research activities including data collection, image processing, development of proposed approach, assessment factors (different magnification and different stains), validation of results with manual microscopic analysis results and development of the framework. A series of 7 different approaches were examined which started with basic image analysis technique. Modification and further medications were carried out to find the best possible approach which maintains the “gold standard” criteria in comparison to manual microscopic analysis results. A novel proposed approach was developed which used two phases: (1) phase one include pre-processing (intensity correction, filter operation, colour image segmentation, intensity clustering, feature extraction approach to find out the most suitable features); and (2) phase two for identifications of potential ovarian NGFs using shape, size and colour features that were extracted in phase one. It was found that the accuracy rate was above 90% for all magnifications and stains used in this research study which maintains the “gold standard” criteria in comparison to manual microscopic analysis results. To increase the accuracy rate and to diminish the false error rate classification approach was incorporated. The proposed approach established the most effect techniques in comparison to existing available approaches. A novel intensity correction was proposed and incorporated at the beginning of pre-processing, fast reliable novel filter operation was developed and incorporated for filter operation, colour image segmentation was considered to use the colour features for identification of region of interest (ROIs) from other tissues, extraction of features to capture NGFs’ characteristics, and incorporation of domain knowledge to identify NGFs. Validation was carried out with experts’ manual microscopic analysis results and similar regions were analysed to minimize the experts’ observation variability issues to improve the accuracy rate. A prototype software tool was developed in MATLAB platform, which enables a non-expert to easily use and analyse the ovarian reproductive tissues without changing any processing parameter automatically by giving the image magnification and image type as input parameter. The proposed approach was found to reduce the time and effort required for the analysis without any human intervention. The novelty of the research is that the approach was fully automated; non-experts will be able to use this approach for analysis; and no change of processing parameter is essential for new image batch/batches. The approach was also accurate, reliable and provided repeatable results in comparison to manual microscopic analysis results. Further work could explore the modification of tissue parameters that could be used for other tissue analysis.
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Pangestu, Mulyoto 1963. "Drying biological material for use in assisted reproductive technology." Monash University, Institute of Reproduction and Development, 2002. http://arrow.monash.edu.au/hdl/1959.1/7879.
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Shores, Ellen Marie. "Theca cell function in the pig." Thesis, University of Nottingham, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.297990.
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