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1
Hajkova, Petra. « Epigenetic reprogramming in mouse germ cells ». [S.l. : s.n.], 2002. http://deposit.ddb.de/cgi-bin/dokserv?idn=970526938.
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Hajkova, Petra. « Epigenetic reprogramming in mouse germ cells ». Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät I, 2004. http://dx.doi.org/10.18452/15020.
Texte intégralRésumé :
Bei Säugerkeimzellen, Zygoten und Embryos in frühen Stadien kommt der epigenetischen Neuprogammierung eine außergewöhnlich wichtige Rolle in der Regulation der Genomfunktionen in entscheidenden Entwicklungsstadien zu. Die epigenetische Neuprogrammierung in Keimzellen löscht zuerst die Imprinting-Markierungen und Epi-Mutationen und stellt dann geschlechtsspezifische Markierungen (genomische Prägung) wieder her. Die vorliegende Arbeit bezieht sich auf das Löschen epigenetischer Modifikationen in primordialen Mauskeimzellen (primordial germ cells (PGCs)) zwischen dem 10.5 bis 13.5 Tag nach der Befruchtung. Entgegen früheren Annahmen zeigen unsere Ergebnisse, daß primordiale Mauskeimzellen (PGCs) beim Eintritt in die embryonalen Keimdrüsen noch immer DNS Methylierungsmarker besitzen, die ähnlich dem Marker in somatischen Zellen sind. Kurz nach dem Eintritt in die Keimdrüsen werden die DNS Methylierungsmarker, die in Verbindung mit geprägten und nicht geprägten Genen stehen, gelöscht. Für die Mehrzahl der Gene beginnt die Löschung der Marker in männlichen und weiblichen Embryos gleichzeitig und ist innerhalb eines Entwicklungstages abgeschlossen. Diese Kinetik deutet auf einen aktiven Demethylierungsprozess hin, initiiert durch ein somatisches Signal, ausgehend von der embryonalen Keimdrüse. Der Zeitpunkt der Neuprogrammierung in den primordialen Keimzellen ist entscheidend, da er sicherstellt, daß Keimzellen beiden Geschlechts einen epigenetisch äquivalenten Status erhalten, bevor sie geschlechtsspezifisch ausdifferenzieren und anschließend neu elterlich geprägt werden. Vollständiges Verständnis des Prozesses der Neuprogrammierung der Keimzellen ist nicht nur im Hinblick auf genomisches Imprinting wichtig, sondern auch für die Erforschung von Mechanismen für die Wiederherstellung von omnipotenten Zellen bei Klonierung und Stammzellenerhaltung.Epigenetic reprogramming in mammalian germ cells, zygote and early embryos, plays a crucial role in regulating genome functions at critical stages of development. Germ line epigenetic reprogramming assures erasure of all the imprinting marks and epi-mutations and establishment of new sex-specific gametic imprints. The presented work focuses on the erasure of epigenetic modifications that occur in mouse primordial germ cells (PGCs) between day 10.5 to 13.5 post coitum (dpc). Contrary to previous assumptions, our results show that as they enter the genital ridge the PGCs still possess DNA methylation marks comparable to those found in somatic cells. Shortly after the entry of PGCs into the gonadal anlagen the DNA methylation marks associated with imprinted and non-imprinted genes are erased. For most genes the erasure commences simultaneously in PGCs of both male and female embryos and is completed within only one day of development. The kinetics of this process indicates that is an active demethylation process initiated by a somatic signal emanating from the stroma of the genital ridge. The timing of reprogramming in PGCs is crucial since it ensures that germ cells of both sexes acquire an equivalent epigenetic state prior to the differentiation of the definitive male and female germ cells in which, new parental imprints are established subsequently. Complete understanding of the germline reprogramming processes is important not only in the light of genomic imprinting but also for resolving other mechanisms connected with restoring cellular totipotency, such as cloning and stem cell derivation.
3
Ono, Tetsuo. « Novel preservation method of germ cells and somatic cells ». Kyoto University, 2010. http://hdl.handle.net/2433/120542.
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Camacho, Moll Maria Elena. « Germ cell neoplasia in situ (GCNIS) and the pathogenesis of testicular germ cell cancer ». Thesis, University of Edinburgh, 2017. http://hdl.handle.net/1842/28807.
Texte intégralRésumé :
Testicular germ cell cancer (TGCC) has been increasing in incidence over recent decades, and is currently the most common malignancy amongst young men resulting in significant morbidity. These tumours are believed to arise from premalignant germ cell neoplasia in situ (GCNIS) cells, which originate from the aberrant germ cell differentiation from gonocyte to spermatogonia during fetal/early postnatal life. GCNIS cells remain dormant in the testis until puberty when they are activated to become tumours. Therefore, GCNIS cells remain in a pre-invasive stage during early childhood and early adulthood prior to the development of a seminoma or non-seminoma TGCC. GCNIS cells are phenotypically similar to gonocytes with expression of stem cell/early germ cell markers including OCT4, PLAP and LIN28. Furthermore, proteins which are expressed in more mature germ cells (spermatogonia) such as MAGE-A4 have also been shown to be expressed in GCNIS cells and these studies have indicated that GCNIS cells are a heterogeneous population in terms of protein expression profile. The relationship between the protein expression profile of individual GCNIS cells populations and their oncogenic potential has not been fully explored. GCNIS cells are located in the seminiferous tubules supported by somatic Sertoli cells. These cells have been previously reported to exhibit an immature protein expression profile in GCNIS tubules from patients with testis cancer, suggesting that the germ stem cell niche in GCNIS tubules resembles that of a fetal one. Associations between Sertoli cell maturation and GCNIS progression into tumour formation has not been fully investigated. Oncogenes are key players in the regulation of oncogenic potential of cancer cells. Gankyrin is an oncogene that has been shown to down-regulate OCT4, and interact with MAGE-A4 in hepatocellular carcinoma and colorectal cancer, where Gankyrin interaction with MAGE-A4 reduces the oncogenic potential of tumour cells. In this study I aimed to investigate the heterogeneity of GCNIS in relation to disease stage and Sertoli cell development. We also aimed to determine the role of Gankyrin in TGCC cell survival and invasion. The co-expression of early germ cells proteins such as OCT4, LIN28 and PLAP was characterized in GCNIS cells during childhood and adulthood pre-invasive TGCC and in invasive disease characterized by the presence of a testicular tumour. These results show that LIN28 was expressed in 95% of OCT4 GCNIS cells, whereas PLAP expression in GCNIS cells increased as the disease progressed from childhood pre-invasive disease to invasive seminoma (32.3% v 76%; p < 0.05). In contrast there was a reduction in the proportion of MAGE-A4 expressing GCNIS cells with disease progression. The MAGE-A4 expressing population was also less proliferative than the MAGE-A4 negative GCNIS population. The methylation status of GCNIS cells was then investigated. EZH2 a methyltransferase previously reported to be important for TGCC development, was expressed in GCNIS cells at all stages of disease, however the histone 3 modification H3K27me3 (mediated by EZH2) was expressed in a significantly higher percentage of the proliferative OCT4+/MAGE-A4- GCNIS cells compared with the OCT4+/MAGEA4+ population (11.7% v 1.1%; p < 0.01) which could indicate a repressive role for H3K27me3 over MAGE-A4 expression. Next, it was determined whether an association between Sertoli cell maturation status and progression of TGCC could be observed. The maturation status of Sertoli cells was studied using proteins indicative of immature (desmin, cytokeratin, fibronectin and AMH) and mature (vimentin and androgen receptor) Sertoli cells. These studies demonstrated heterogeneity of Sertoli cells maturation in GCNIS-containing tubules. Desmin, fibronectin, AMH and vimentin expression did not show any association with TGCC progression. Cytokeratin was expressed in Sertoli cells of human fetal testis up to second trimester of fetal life, absent in tubules with active spermatogenesis but heterogeneously present in GCNIS, demonstrating that cytokeratin expression is indicative of the presence of GCNIS. Androgen receptor was weakly present in Sertoli cells from human fetal testis and pre-pubertal pre-invasive TGCC testis whereas in GCNIS of adult pre-invasive testis and invasive samples, androgen receptor was abundantly expressed in Sertoli cells of GCNIS-containing tubules. These combined results for cytokeratin and androgen receptor suggest that Sertoli cells from GCNIS-containing tubules, in pre-invasive and invasive TGCC patients are partially differentiated. Gankyrin expression was characterised in fetal germ cells, GCNIS cells and TGCC tissue. In fetal testis nuclear Gankyrin was absent in OCT4+/MAGE-A4- (gonocyte) population whereas it was present in a subpopulation of OCT4-/MAGE-A4+ (spermatogonia) germ cells. In GCNIS cells from TGCC patients nuclear Gankyrin was expressed in 87%, 63.3%, 91.5% and 79% in childhood pre-invasive, adult pre-invasive, seminoma and non-seminoma GCNIS cells respectively. Finally, in seminoma cells, Gankyrin was expressed in the cytoplasm indicating a change in localisation as the GCNIS cells become invasive. We used siRNA to knockdown Gankyrin in NT2 (a TGCC cell line) cells in-vitro and demonstrated a decrease in cell number, suggesting that Gankyrin might play a role in TGCC progression and invasiveness. Gankyrin down-regulation also resulted in an increase in p53 and p21 mRNA level. Given the role of P53 and p21 in cisplatin cytotoxic effect in TGCC we went on to investigate the role of Gankyrin in cisplatin resistance using NT2 cells. We demonstrate that Gankyrin mediated cisplatin resistance through the p53/p21 pathway, upregulating apoptosis rates through BAX and FAS, whilst there was no effect on cell proliferation, cell cycle or cell migration. In conclusion, we have shown that GCNIS cells are heterogeneous and their phenotype can determine their oncogenic potential. We also show that Sertoli cells from GCNIS-containing tubules undergo partial differentiation displaying markers of immature and mature Sertoli cells, with a heterogeneous association of cytokeratin with GCNIS presence. We also demonstrate that the oncogene Gankyrin has a role in NT2 cells survival and cisplatin resistance indicating that manipulation of Gankyrin may have a role in the treatment of TGCC.
5
Cowan, Gillian. « Fetal germ cell differentiation and the impact of the somatic cells ». Thesis, University of Edinburgh, 2009. http://hdl.handle.net/1842/4164.
Texte intégralRésumé :
Specification of a germ cell lineage and appropriate maturation are essential for the transfer of genetic information from one generation to the next. Germ cells form from pluripotent precursor cells that migrate into the gonadal ridge and undergo commitment to either the female or male lineage. In the fetal ovary, germ cells enter meiotic prophase I, then arrest at the diplotene stage; in the testis germ cells do not begin meiosis until puberty. Abnormal differentiation of germ cells can result in malignant transformation. Somatic cells play a key role in modulating the developmental fate of the germ cells. Research into germ cell development during fetal life has almost exclusively focused on studies in rodents, but we, and others, have reported several fundamental differences in the expression of germ cell specific markers in the human compared with the mouse. The studies described in this thesis have investigated germ cell-specific gene expression and the possible impact of the somatic cells during development. This was achieved by studying human fetal gonads obtained during the 1st and 2nd trimesters of pregnancy and through the use of both wild-type and mutant mouse ES cell lines. Studies on germ cells in the human fetal testis have extended the findings of others, and confirmed that germ cell populations at different stages of maturation co-exist in the human fetal testis, a situation that is in contrast to that in rodents. For example expression of M2A and AP2γ was restricted to the OCT4-positive gonocyte population, while VASA and NANOS1 were localised exclusively to the to the OCT4-negative prespermatogonia. DAZL was expressed in both populations. Analysis also revealed that both the gonocyte and prespermatogonial populations proliferate throughout the 2nd trimester. Recent studies have implicated retinoic acid (RA) in the control of meiotic entry in germ cells of the fetal mouse ovary. In this study we demonstrated for the first time that two genes implicated in the action of RA in mouse gonad, STRA8 and NANOS2, are also expressed in a similar sexspecific- manner in the human fetal gonads, and that the RA receptors are present in both somatic and germ cells suggesting that RA may regulate germ cell function in the human as well as the mouse. However, whilst the mesonephros appears to be the primary site of RA synthesis in the mouse our initial studies indicate that in the human the gonad itself may be a more likely site of RA biosynthesis. In the fetal mouse testis, RA is degraded by the enzyme Cyp26b1 present in the somatic cells and germ cells do not enter meiosis, our novel findings suggest that CYP26B1 is more abundant in the human fetal ovary than the testis, suggesting that meiotic entry may be controlled by an alternative signalling pathway in the human. One of the methods that can aid our understanding of somatic cell gene expression in the gonad is in vitro culture. To date, there have been no published reports of the successful in vitro culture of somatic cells from the human fetal testis. In the current study, populations of human somatic cells were dissociated and maintained in vitro and characterised. Analysis demonstrated that cells expressing mRNAs characteristic of Sertoli cells, Leydig cells and peritubular myoid (PTM) cells were present initially, but long-term culture resulted in downregulation in expression of mRNAs specific for Sertoli cells and Leydig cells, suggesting that these cells either failed to survive or underwent alterations to their phenotype. In contrast PTM/fibroblast cells proliferated in vitro and initially maintained androgen receptor expression. These cultures therefore hold promise for studies into the signalling or cell-cell interactions in testicular somatic cells especially those relevant to the PTM population. Several studies have claimed differentiation of putative germ cells from ES cells. In the current study, analysis of mouse ES cell lines has expanded on results showing that ES cells and early germ cells express a number of genes in common. Kit signalling was shown to be important for ES cell survival as they differentiate although expression of Kit was heterogeneous. We also demonstrated that ES cells that did not express Kit displayed a decreased expression of the early germ cell genes Blimp1, Fragilis and Stella, implicating Kit signalling in the control of germ cell-associated gene expression in ES cells. This may be important to future studies optimising germ cell derivation from ES cells. In conclusion, this study has demonstrated important differences in protein expression patterns in germ cells of the human fetal testis compared to the mouse, and has raised questions about whether the proposed mechanism controlling meiotic entry of germ cells in the mouse can be applied to the human. The establishment of a system for culturing human fetal gonadal somatic cells may lead to further understanding of gene expression and development in the human fetal testis, and data suggest that the Kit/Kitl signalling system may influence germ cell gene expression in mouse ES cells.
6
Al-Thani, Rawda. « Primordial germ cells of the chick embryo ». Thesis, University of Reading, 1992. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.315524.
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Li, Ying. « Transgenic birds from transformed primordial germ cells ». Thesis, University of Reading, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.385118.
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Yoon, Christina Migyung 1970. « Idenficiation of the zebrafish primordial germ cells ». Thesis, Massachusetts Institute of Technology, 1997. http://hdl.handle.net/1721.1/43551.
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Somers, Christopher Michael Quinn James S. « Germline mutations at expanded simple tandem repeat DNA loci in sentinel mice / ». *McMaster only, 2004.
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10
Leitch, Harry Gordon. « Pluripotency and the germline ». Thesis, University of Cambridge, 2012. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.610336.
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Grabole, Nils. « Functional analysis of Prdm14 in primordial germ cells and stem cells ». Thesis, University of Cambridge, 2013. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.608092.
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Mruk, Dolores Dorothy. « A study on the dynamics of sertoli-germ cell interactions : new perspectives on male fertility control / ». Thesis, Hong Kong : University of Hong Kong, 1999. http://sunzi.lib.hku.hk/hkuto/record.jsp?B22079002.
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Okumura, Leah M. « Germ cell nuclear factor is not required for the down-regulation of pluripotency markers in fetal ovarian germ cells ». Thesis, Massachusetts Institute of Technology, 2012. http://hdl.handle.net/1721.1/77781.
Texte intégralRésumé :
Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Biology, 2012.Cataloged from PDF version of thesis.
Includes bibliographical references.
In mouse, germ cells retain expression of the pluripotency markers Oct4 and Nanog longer than any other cells in the body. While somatic cells repress these markers during gastrulation, female germ cells continue to express them until around the time of meiotic initiation. It is not yet clear why pluripotency markers are downregulated with this particular timing, nor is it understood what factors are involved in their repression. I have examined in fetal ovarian germ cells the expression and function of Gcnf (germ cell nuclear factor), an orphan nuclear receptor known to regulate both Oct4 and Nanog in gastrulating embryos. I have found that Gcnf is expressed in a female germ-cell-specific manner at the time when Oct4 and Nanog are down-regulated there. Gcnf mutants in which the ligand binding domain is disrupted display defects after gastrulation comparable to those observed in Gcnf-null mutants and those lacking the DNA binding domain. In contrast, the germ cells Gcnfligand binding domain mutants show no failure in repression of pluripotency markers, and other aspects of female germ cell development appear normal as well. Thus, it appears that the ligand binding domain of GCNF is not required for fetal ovarian germ cell development.
by Leah M. Okumura.
Ph.D.
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Wang, Qiufan Claire, et 王秋帆. « Mechanisms of junctional restructuring at the sertoli-sertoli and sertoli-germ cell interfaces during spermatogenesis ». Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2008. http://hub.hku.hk/bib/B40887686.
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Wang, Qiufan Claire. « Mechanisms of junctional restructuring at the sertoli-sertoli and sertoli-germ cell interfaces during spermatogenesis ». Click to view the E-thesis via HKUTO, 2008. http://sunzi.lib.hku.hk/hkuto/record/B40887686.
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Wigmore, Kip. « The primordial germ cells of the goat fetus ». Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1999. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp01/MQ43237.pdf.
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Seisenberger, Stefanie. « Reprogramming the epigenome of mouse primordial germ cells ». Thesis, University of Cambridge, 2012. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.610019.
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Vick, Lorraine Mary. « Genetic manipulation of fowl via primordial germ cells ». Thesis, University of Reading, 1993. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.317960.
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Matias, Neuza. « Regulation of Abscission in Female Drosophila Germ Cells ». Thesis, Paris 11, 2015. http://www.theses.fr/2015PA112211/document.
Texte intégralRésumé :
En fin de cytocinèse, le fin pont cytoplasmique qui relie les deux cellules filles est clivé au niveau d’une structure dense en microtubules, le midbody, et permet ainsi la séparation physique de leurs deux cytoplasmes. Les mécanismes cellulaires et moléculaires de ce processus, appelé abscission, sont très étudiés dans des modèles de cellules en culture. Cependant, ils restent encore mal connus dans le contexte d’un organisme en développement. L’ovogenèse de drosophile est un modèle de choix pour l’étude de la régulation développementale de l’abscission. En effet, des cellules à abscission complète (cellules souches germinales) et incomplète (cystes germinaux) sont situées côte à côte au sein de la même unité développementale, le germarium. Les cellules souches se divisent asymétriquement, pour donner une autre cellule souche et un cystoblaste individualisé. Celui-ci entre en alors en différenciation, un programme au cours duquel il réalise quatre cycles cellulaires synchrones au cours desquels la cytocinèse est incomplète. Un cyste germinal de seize cellules interconnectées est ainsi formé. La durée de l’abscission est régulée précisément et dépend du contexte développemental. Notre laboratoire a récemment montré que les kinases Aurora B et Cdk1/ Cyclin B sont des régulateurs de la durée d’abscission dans les cellules germinales de drosophile et en cellules en culture de vertébrés. Mon travail a consisté à explorer la fonction de la protéine Shrub, un membre du complexe ESCRT-III, au cours de l’abscission dans la lignée germinale femelle de drosophile. Nous avons montré que Shrb est localisé au midbody des cellules souches en fin de cytocinèse, et promeut l’abscission. En effet, nous avons montré qu’une réduction du niveau de Shrub dans la lignée germinale provoque un fort délai de l’abscission des cellules souches, supérieur à la durée de leur cycle cellulaire. La cellule souche et son cystoblaste restent donc connecté jusqu’à la mitose suivante, formant ainsi des structures de plusieurs cellules connectées, appelées stem-cyst . L’abscission tardive au sein du stem cyst libère un progéniteur binucléé qui entre en différenciation. En conséquence, des chambres ovariennes à 32 cellules, au lieu de 16, sont formées. De plus, la fonction de Shrub dans l’abscission semble être contrecarrée par Aurora B, puisqu’une réduction des niveaux d’Aurora B dans des hétérozygotes Shrub réduit le nombre de stem-cysts et de chambres à 32 cellules observés. Enfin, nous avons identifié un nouveau facteur impliqué lors de l’abscission, la protéine Lethal giant discs (lgd), dont la perte de fonction induit, comme celle de Shrub, la formation de stem-cysts. En accord avec son rôle dans l’abscission, nous avons montré que Lgd est localisé au midbody. Lgd est requis pour la fonction de Shrub dans la voie endosomale, mais son implication lors de la cytocinèse était inconnue. Nous avons montré qu’un niveau réduit de Lgd augmente le nombre de stem-cysts des hétérozygotes Shrub, indiquant que Lgd et Shrub fonctionnent ensemble pour l’abscission des cellules souches. De façon surprenante, un nombre réduit de chambres à 32 cellules est observé dans ces ovaires, suggérant une fonction antagoniste de Lgd sur Shrub dans les cystes germinaux. Dans ces cystes, une abscission tardive se produit, qui divise en deux cystes de 16 cellules les cystes de 32 cellules, et expliquant ainsi le paradoxe observé (plus de stem-cysts, mais moins de chambres à 32 cellules)At the end of cytokinesis, a thin cytoplasmic intercellular bridge is cleaved to allow physical separation of the two daughter cells. This process is called abscission, and its cellular and molecular events have been extensively explored in yeast and isolated mammalian cells. However, how abscission is regulated in different cell types or in a developing organism remains poorly understood.Drosophila oogenesis is a great model to study how abscission is regulated developmentally, as within the same developmental unit, the germarium, we find cells undergoing abscission next to others where this process is blocked. Indeed, the germline stem cell (GSC) divides asymmetrically to give rise to another GSC and to an individualized cystoblast. This cell then enters a well-studied process of differentiation, where through four rounds of mitosis with incomplete cytokinesis, gives rives to a cyst of 16 interconnected cells. The duration of abscission, seems to be tightly regulated and dependent on the developmental context. Our lab has recently discovered that AurB and CycB/Cdk1 function as abscission timers in Drosophila GSC and isolated mammalian cells. Thus, my work consisted in exploring how this process is regulated in the Drosophila female germline.We showed that the ESCRT-III protein Shrb localizes to the midbody of the dividing GSC, functioning to promote abscission. Indeed, we found that reduced levels of Shrb resulted in the blockage, or strong delay, of abscission in the GSC and formation of a structure similar to a cyst. In these so called stem-cysts, the GSC keeps dividing while interconnected to its daughter cells. As a consequence, we saw the appearance of egg chambers formed of 32 cells, instead of 16. Furthermore, Shrb function in abscission seems to be counteracted by AurB, as reducing AurB levels in Shrb heterozygous resulted in decreased stem-cysts and 32-cell cysts. Finally, Lethal giant discs (lgd), required for Shrb function in the endosomal pathway, was also seen localizing at the midbody and regulating abscission in GSCs. Removing one copy of Lgd from Shrb heterozygous increased the number of stem-cysts, but surprisingly the number of 32-cell cysts was reduced. This paradoxical result was explained with the observation of late abscission events in mitotic cysts, which divided the 32-cell cysts in the middle, leading to the formation of two cysts of 16 cells
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Pogool, Satian. « Factors controlling migration of avian primordial germ cells ». Thesis, University of Manchester, 2002. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.531435.
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Avian embryos have played an important role in the study of vertebrate development but further understanding of their development may also benefit the commercial poultry industry. Manipulation and use of primordial germ cells (PGCs) as a vehicle for constructed genotypes will be increasingly important for future improvement of commercial poultry including the turkey. However, the manipulation and use of PGCs will depend on a basic understanding of PGC migration. PGCs are known to migrate considerable distances before colonising the gonad. Previous work in the chick and quail have suggested that avian PGCs arise from the ventral surface of the area pellucida before the blastoderm undergoes gastrulation, translocate to the germinal crescent during primitive streak stages and penetrate the extraembryonic blood vessels (stage 10) after they have formed. PGCs are subsequently carried by the blood stream into the embryo proper during stages 12-14 and then leave the blood vessels and migrate to the developing gonads. However, the factors controlling PGC migration are poorly understood. The developmental stages of older turkey embryos are not well documented. Therefore, this study started by classifying them. It was found that turkey embryos at stage 4 and younger were slightly different from chick embryos, but from stages 5 onwards, turkey embryos could be staged using criteria described for the chick (Hamburger and Hamilton, 1951). Avian PGCs can be recognized by their distinctive morphology being large cells with large and eccentrically placed nuclei, and a cytoplasm containing refractile granules. These characteristics were confirmed for turkey PGCs. In order to manipulate PGCs, cells need to be identified using appropriate markers. A panel of markers previously used for chick, quail, mouse, rat and rabbit were tested with turkey PGCs at different stages. It was found that turkey PGCs could be detected by the histochemical stains, periodic acid-Schiff (PAS) and alkaline phosphatase (AP) or using antibodies to stage-specific embryonic antigen-1 (SSEA-1), fucosylated polylactosamine carbohydrate groups (EMA-1) and ovomucin-like protein (OLP). At primitive streak stages, turkey PGCs were located in the area pellucida. During stages 11 - 15, they were found in the blood reaching maximal numbers at stage 14. After stage 15, most PGCs were located outside the circulatory system. From stage 18, PGCs began to settle in gonadal ridges and after stage 28, PGCs were found only in the gonads. Turkey PGCs were isolated from the blood with a micropipette, during visual selection under the microscope, and their Identity confirmed using the above markers. These PGCs could be cultured on a gonadal stromal cell layer or on coverslips coated with rat-tail collagen. Isolated PGCs adhered to coated coverslips were prepared and examined with scanning electron microscope (SEM). Such cells had typical features of PGCs seen in situ and some had the morphology of motile cells. PGC movement in vitro, in response to a variety of potential chemoattractants, was then studied. Conventional chemotaxis assays could not be used to study such small numbers of cells. Therefore, the assay was carried out using a Dunn chemotaxis chamber which allows direct observation of cells and detailed analysis of their movement from a timed series of images. A positive chemotactic response was observed with cells exposed to a gradient of medium conditioned by stromal cells or 10 ng/ml transforming growth factor beta-1 (TGFbeta1). Cells exposed to 100 ng/ml stem cell factor (SCF) or control medium showed no such response. These observations indicate that TGF?l can play a role in directed migration of PGCs in vitro, but further experiments are required to determine whether this factor is similarly involved in vivo.
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Mifsud, William. « Studies on the ontogeny of the mammalian germ line ». Thesis, University of Cambridge, 2011. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.609789.
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Hamaguchi, Satoshi. « Sex differentiation of germ cells and their supporting cells in Oryzias latipes ». Laboratory of Freshwater Fish Stocks, Nagoya University, 1992. http://hdl.handle.net/2237/13766.
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Sneddon, Sharon F. « Oestrogen regulation of gene expression in male germ cells and Sertoli cells ». Thesis, University of Edinburgh, 2005. http://hdl.handle.net/1842/29373.
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The aims of this study were to investigate the role of steroid hormones, in particular oestrogens, in murine spermatogenesis. A major focus of these investigations was the role played by ERβ in the modulation of germ cell and somatic cell function. Studies were conducted both using a transformed murine Sertoli cell line (SK11), which has maintained a differentiated Sertoli cell phenotype and spermatogonial stem cells, which were successfully isolated and characterised. ERβ mRNA and protein were shown to be expressed in the SK11 cells both in the undifferentiated and differentiated states. Transient transfections using ERE or ARE-luciferase reporter constructs and stimulation with steroid ligands revealed that the cells contained functional steroid hormone receptors. Knockdown of ERβ mRNA and protein was achieved in the cells after targeted deletion using a short hairpin RNAi containing vector; this blunted the ability of the cells to respond to oestrogen. Isolation of spermatogonial stem cells was carried out using immunomagnetic beads. The stem cell population were shown to express Oct-4 and GFRα-1 mRNAs, both of which are stem cell markers, but not c-kit, which is a marker of differentiated germ cells. Taqman Q-RT-PCR demonstrated that the stem cell population expressed ERβ. Oct-4 mRNA expression was shown to be reduced by RNAi; this induced the cells to undergo differentiation in vitro characterised by increased expression of c-kit. In conclusion, the current studies have extended our understanding of the impact of steroid hormones on testicular function and have revealed for the first time that spermatogonial stem cells are ERβ positive. The SK11 cell line has been found to provide a suitable model system for the study of steroid regulation of Sertoli cell function. In addition, the use of RNAi has provided and exciting new avenue by which to manipulate gene expression levels in testicular germ and somatic cells.
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Tee, Wee Wei. « Functional analysis of PRMT5 in mouse pluripotent stem cells and germ cells ». Thesis, University of Cambridge, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.608638.
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STALLOCK, JAMES PATRICK. « THE ROLE OF BAX IN APOPTOSIS OF ECTOPIC PRIMORDIAL GERM CELLS IN THE MOUSE ». University of Cincinnati / OhioLINK, 2003. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1046891667.
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Xia, Weiliang, et 夏偉梁. « Role of cytokines in junction restructuring and germ cell migration inmammalian testes ». Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2006. http://hub.hku.hk/bib/B37101134.
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Petyim, Somsim. « Gene regulation during development of human primordial germ cells ». Thesis, University of Nottingham, 2013. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.665476.
Texte intégralRésumé :
A critical event during embryonic development is the segregation of the germ line from the soma. The germ line founders, or primordial germ cells (PO C), are the precursors of the gametes, which enable the transmission of the genetic material to future generations. In rodents, the PGC precursors are segregated from somatic lineages before the start of gastrulation. In several model organisms segregation of the germ line is accomplished by maternally inherited transcriptional repressors contained in the egg's germ plasm. In mammals, PGC specification is regulated by epigenesis, a mechanism by which pluripotent cells ill the epiblast respond to specification signals according to a temporal and spatial program of differentiation. In the mouse embryo, the transcriptional repressor Blimp] plays a fundamental role in restricting nascent POC to somatic differentiation signals. The aim of this project was to determine whether the mechanism of lineage specification is conserved in humans.
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Adhikari, Deepak. « Signaling pathways in the development of female germ cells ». Doctoral thesis, Umeå universitet, Institutionen för medicinsk kemi och biofysik, 2014. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-88309.
Texte intégralRésumé :
Primordial follicles are the first small follicles to appear in the mammalian ovary. Women are born with a fixed number of primordial follicles in the ovaries. Once formed, the pool of primordial follicles serves as a source of developing follicles and oocytes. The first aim of this thesis was to investigate the functional role of the intra-oocyte signaling pathways, especially the phosphatidylinositol-3 kinase (PI3K) and mammalian target of rapamycin complex 1 (mTORC1) pathways in the regulation of primordial follicle activation and survival. We found that a primordial follicle remains dormant when the PI3K and mTORC1 signaling in its oocyte is activated to an appropriate level, which is just sufficient to maintain its survival, but not sufficient for its growth initiation. Hyperactivation of either of these signaling pathways causes global activation of the entire pool of primordial follicles leading to the exhaustion of all the follicles in young adulthood in mice. Mammalian oocytes, while growing within the follicles, remain arrested at prophase I of meiosis. Oocytes within the fully-grown antral follicles resume meiosis upon a preovulatory surge of leutinizing hormone (LH), which indicates that LH mediates the resumption of meiosis. The prophase I arrest in the follicle-enclosed oocyte is the result of low maturation promoting factor (MPF) activity, and resumption of meiosis upon the arrival of hormonal signals is mediated by activation of MPF. MPF is a complex of cyclin dependent kinase 1 (Cdk1) and cyclin B1, which is essential and sufficient for entry into mitosis. Although much of the mitotic cell cycle machinery is shared during meiosis, lack of Cdk2 in mice leads to a postnatal loss of all oocytes, indicating that Cdk2 is important for oocyte survival, and probably oocyte meiosis also. There have been conflicting results earlier about the role of Cdk2 in metaphase II arrest of Xenopus oocytes. Thus the second aim of the thesis was to identify the specific Cdk that is essential for mouse oocyte meiotic maturation. We generated mouse models with oocytespecific deletion of Cdk1 or Cdk2 and studied the specific requirements of Cdk1 and Cdk2 during resumption of oocyte meiosis. We found that only Cdk1 is essential and sufficient for the oocyte meiotic maturation. Cdk1 does not only phosphorylate the meiotic phosphoproteins during meiosis resumption but also phosphorylates and suppresses the downstream protein phosphatase 1, which is essential for protecting the Cdk1 substrates from dephosphorylation.
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Intarapat, S. « Isolation and characterisation of chick embryonic primordial germ cells ». Thesis, University College London (University of London), 2012. http://discovery.ucl.ac.uk/1344049/.
Texte intégralRésumé :
Embryonic stem cells (cESCs) can be isolated from chick embryos, with the ability to contribute to all somatic lineages in chimaeras, but not to the germ line. However, lines of chicken embryonic germ cells (cEGCs), which are able to contribute to the germ line, can be established from chicken primordial germ cells (cPGCs). However very little is known about these cells, or about the changes that accompany the establishment of gonadal cells as self-renewing cell lines. This thesis presents a detailed study of the properties of cPGCs and the parent tissue from which they are derived. Gene expression profiles for 30 genes related to pluripotency and/or differentiation were are compared between gonads at the indifferent stage (stage 26-28HH), in primary gonocytes, established PGCs and cESCs. The results reveal great heterogeneity in the expression of various markers in culture. Several genes associated with pluripotency change dramatically upon culture. The most salient of these changes is that while cSox3 (but not Sox2) is expressed in the gonads, whereas their expression becomes reversed upon culture (becoming more similar to mammalian stem cells). This suggests that these two SoxB1 class genes have swapped functions in chick. In the process of studying the expression pluripotency markers in later (stage HH35) gonads, we made an unexpected discovery: both male and female embryos show left-right asymmetric patterns of expression of some, but not all, of these markers. Expression of pluripotency (cPouV, cNanog, cSox2 and ERNI) in the left gonad is much higher than those in the right gonad of both sexes. The expression of pluripotency markers is irrespective of its colonisation by primordial germ cells, and it appears that this left-right decision is made independently of whether the gonad will regress or be retained. These findings offer a new model system for investigating the roles of pluripotency-related markers during normal development as well as in stem cell lines.
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Xia, Weiliang. « Role of cytokines in junction restructuring and germ cell migration in mammalian testes ». Click to view the E-thesis via HKUTO, 2006. http://sunzi.lib.hku.hk/hkuto/record/B37101134.
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Jiang, Shan. « Establishing genomic imprinting by cell differentiation a model system using embryonic germ cells / ». Available to US Hopkins community, 2000. http://wwwlib.umi.com/dissertations/dlnow/3099377.
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Lokman, Muhammad. « Development of post-meiotic germ cells from human embryonic stem cells in vitro ». Thesis, University of Sheffield, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.531176.
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Wong, Ching-hang. « Cell-cell interactions and cell junction dynamics in the mammalian testis ». Click to view the E-thesis via HKUTO, 2005. http://sunzi.lib.hku.hk/hkuto/record/B31993084.
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Kasai, Shinya. « Haploinsufficiency of Bcl-x leads to male specific defects in fetal germ cells : differential regulation of germ cell apoptosis between the sexes ». Kyoto University, 2004. http://hdl.handle.net/2433/148262.
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Ewen-Campen, Benjamin Scott. « An evolutionary perspective on germ cell specification genes in insects ». Thesis, Harvard University, 2014. http://dissertations.umi.com/gsas.harvard:11549.
Texte intégralRésumé :
This dissertation investigates the embryonic specification of a specific group of cells: the germ cells. Germ cells, which give rise to sperm and egg, are the only cells in sexually-reproducing animals that directly contribute hereditary information to the next generation. Germ cells are therefore a universal cell type across animals, and represent a profound novelty that likely arose near the base of the animal phylogeny. Yet despite their conserved, essential function in all animals, there is surprising diversity in the mechanisms that specify these cells during embryonic development. In this dissertation, I address the diversity of germ cell specification mechanisms in insects. I focus on two species, the milkweed bug Oncopeltus fasciatus (Hemiptera) and the cricket Gryllus bimaculatus (Orthoptera), which both branch basally to the Holometabola (those insects which undergo metamorphosis, including the well-studied fruit fly Drosophila melanogaster), and thus provide important phylogenetic breadth to our understanding of germ cell specification across insects. Using functional genetic approaches, I show that germ cell specification in both Oncopeltus and Gryllus differs fundamentally from germ cell specification in Drosophila. Specifically, I provide evidence that germ cells arise via inductive cell signaling during mid-embryogenesis, rather than via maternally-supplied cytoplasmic determinants localized in the oocyte, as is the case for Drosophila. These data suggest that Drosophila employs an evolutionarily derived mode of germ cell specification. In further support of this hypothesis, I show that several of the genes required for Drosophila germ cell specification perform other functions in both Oncopeltus and Gryllus. I demonstrate that one of these genes, oskar, which is the only gene both necessary and sufficient for germ cell specification in Drosophila, instead functions in nervous system of the cricket, both during embryonic development and in the adult brain. I suggest that the evolution of the derived mode of germ cell specification seen in Drosophila may have involved co-opting oskar into the germ cell specification pathway from an ancestral role in the nervous system.
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McCormick, Rachel Jacqueline. « Investigations into Xpat, a novel gene expressed in the germ plasm and primordial germ cells of Xenopus laevis ». Thesis, University of Warwick, 2001. http://wrap.warwick.ac.uk/3093/.
Texte intégralRésumé :
To determine the expression pattern of XPAT (Xenopus primordial germ cell associated transcript) protein in Xenopus oocytes, XPAT-GFP fusion proteins were generated. When XPAT was amino-terminally tagged with GFP it became localised to the nuclei of stage VI Xenopus oocytes. However, when carboxy-terminally tagged with GFP, XPAT also translocated to the vegetal pole of stage VI oocytes. XPAT-GFP formed particles (1 to 2.5mm in diameter) which aggregated into large (10 to 50mm) granular structures at the vegetal pole. These particles looked exactly like those seen after in situ hybridisation to germ plasm RNAs. The granules of XPAT-GFP were larger than endogenous germ plasm granules seen in stage VI oocytes; they were more consistent with those observed in 2-cell embryos during germ plasm aggregation. Studies involving the use of the anticytoskeletal drugs colcemid, nocodazole and cytochalasin D and the microtubule stabilising agent taxol indicated that microtubular transport was important in the location of XPAT-GFP. Several attempts were made to raise antibodies to XPAT peptides, but at present the endogenous expression pattern of XPAT protein is unresolved. To investigate possible domain structure of XPAT, one carboxy-terminal and three amino-terminal deletion variants of XPAT-GFP were constructed. An N-terminal deletion protein lacking the first 61 amino acids of XPAT was able to form small particles, but none of the deletion proteins exhibited vegetal localisation or formed large aggregates in Xenopus oocytes. The N-terminal deletion proteins all became predominantly localised to the nucleus; protein motif analysis revealed that XPAT contains a putative bipartite NLS in its carboxy-terminal region. The C-terminal deletion protein, which lacked the putative NLS, was evenly distributed throughout the nucleus and cytoplasm of Xenopus oocytes. XPAT was shown to be able to bind to homopolymeric RNAs in vitro. When Xpat mRNA was depleted from stage VI Xenopus oocytes (by injection of an antisense oligo) levels of DEADSouth and XVLG1 mRNAs decreased substantially.
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Angeles, Vanessa Therese. « Characterization of NANOS expression and function in human germ cells ». Diss., Search in ProQuest Dissertations & ; Theses. UC Only, 2009. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:3390030.
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Severino, Jacqueline 1990. « X chromosome status : a gatekeeper of germ cells meiotic entry ». Doctoral thesis, TDX (Tesis Doctorals en Xarxa), 2021. http://hdl.handle.net/10803/671536.
Texte intégralRésumé :
X chromosome reactivation in female mouse germ cells is essential for the transmission of one active
X chromosome to the progeny. However, despite its key role in development, the mechanistic details
and kinetics still remain elusive, as previous studies were restricted by a scarcity of cells in vivo and a
lack of adequate in vitro systems. Here, I present the characterization of X-chromosome dynamics
during germ cell formation, which was possible thanks to the development of a tailor-made in vitro
system which allows the accurate profiling of X-chromosome activity. We recapitulate X-inactivation
starting in epiblast-like cells (EpiLCs), progressing in primordial germ cell-like cells (PGCLCs),
followed by X-reactivation in germ cells upon meiotic entry. We show that PGCLCs undergoing
X-inactivation can enter meiosis more efficiently, while PGCLCs bypassing X-inactivation, with the
consequent lack of X-reactivation, show a reduced meiotic potential. We conclude that tracing the X
chromosome status during germ cell formation facilitates the dissection of the relationship between X
chromosome dynamics and proper germline fate acquisitionEn las hembras de ratón, la reactivación del cromosoma X en las células germinales es esencial para la transmisión de un cromosoma X activo a la descendencia. Sin embargo, a pesar del papel crucial del cromosoma X durante el desarrollo, el mecanismo y la dinámica de su reactivación siguen siendo difíciles de alcanzar, ya que los estudios realizados anteriormente estaban restringidos por la escasez de células in vivo y la falta de sistemas in vitro adecuados. En este estudio he desarrollado un sistema que me permite seguir y caracterizar de manera detallada la actividad del cromosoma X durante la formación de las células germinales femeninas. Empezando por células similares a las células del epiblasto y diferenciandolas hacia células parecidas a células germinales primordiales (PGCLCs), hemos conseguido recapitular la inactivación del cromosoma X. A continuación hemos observado la reactivación del cromosoma X a medida que las células germinales primordiales entran en meiosis. Mostramos que las PGCLCs que se someten a inactivación del cromosoma X pueden entrar en meiosis forma más eficiente, mientras que las PGCLCs que no logran inactivar el cromosoma X, con la consiguiente falta de su posterior reactivación, muestran un menor potencial para entrar en meiosis. Concluimos que el seguimiento del estado del cromosoma X durante la formación de las células germinales nos permitió analizar la relación entre la dinámica del cromosoma X y la correcta especificación y desarrollo de la línea germinal.
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Molaro, Antoine. « Inheritance and evolution of epigenetic reprogramming in Mammalian germ cells ». Phd thesis, Université Pierre et Marie Curie - Paris VI, 2012. http://tel.archives-ouvertes.fr/tel-00833274.
Texte intégralRésumé :
During mammalian post-implantation development, germ cells are induced from the somatic tissues of the embryo. Following their induction, primordial germ cells undergo a genome-wide erasure and de novo re-establishment of DNA methylation marks. This epigenetic reprogramming re-instates pluripotency and allows parental imprints to be deposited. In the male germ line, a unique RNAi pathway involving PIWI proteins and their associated small RNAs (piRNAs) is necessary for proper de novo methylation. PIWI mutant mice are infertile and display methylation defects over transposon sequences. Using a transgenic approach, we investigated the signals necessary for piRNA production. We show that artificial piRNAs can be produced from reprogrammed loci outside of their native context. We then studied the genome-wide impact of piRNA loss on germ cell methylation. Whereas most of the genome is properly methylated, only a small group of transposons transiently reactivated in primordial germ cells is affected. Also we identified important structural differences in de novo methylation profiles between human sperm and ES cells. Finally, we compared sperm methylation profiles between human and chimpanzee and showed that the genome and the epigenome can evolve independently. Taken together, our results highlight the surprising plasticity of genome and epigenome interactions during development and evolution
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Hurtado, Gonzalez Pablo Ignacio. « The consequences of fetal exposure to analgesics for germ cells ». Thesis, University of Edinburgh, 2018. http://hdl.handle.net/1842/29631.
Texte intégralRésumé :
Despite the general advice of avoiding medication during pregnancy, the majority of pregnant woman use one or more ‘over the counter’ analgesics. During the last few years there has been growing evidence that analgesic exposure, such as paracetamol, ibuprofen or indomethacin, during pregnancy can have detrimental effects on rodent and human fetal gonads. The majority of previous studies have focused in alterations in testosterone production and male reproductive disorders. However, few studies have analysed the effect of these analgesics on fetal germ cells and possible consequences on fertility. During my thesis, I first focused on the effect of paracetamol and indomethacin exposure during pregnancy on rat fetal gonads. These showed that both paracetamol and indomethacin are able to alter the expression of genes important for fetal gonad and germ cell development. Previous studies on germ cells and analgesics have focused on rat models, but there is a lack of similar studies performed in human models. Therefore, I investigated the consequences of exposure of therapeutically relevant doses of paracetamol and ibuprofen on human gonads, with a special attention to the germ cells. Fetal gonads from the 1st and 2nd trimester were used in two different models: hanging drop cultures for 1st trimester testes and ovaries and a xenograft system for 2nd trimester fetal testes. Fetal gonad culture in the presence of paracetamol or ibuprofen reduced AP2γ+ (gonocyte) GC number in both 1st trimester fetal testes (22-28% reduction) and ovaries (43-49% reduction). 2nd trimester fetal testes were exposed to three different regimes, 1 or 7 days paracetamol and 7 days ibuprofen, which led to reductions of 17% and 30%, respectively in AP2γ+ GC number for paracetamol and a 53% reduction in total germ cell number for ibuprofen.
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LOLICATO, FRANCESCA. « Molecular mechanisms involved in spermatogonial germ cells proliferation and differentiation ». Doctoral thesis, Università degli Studi di Roma "Tor Vergata", 2008. http://hdl.handle.net/2108/514.
Texte intégralRésumé :
Lo scopo del seguente lavoro, è comprendere alcuni dei meccanismi molecolari coinvolti nella regolazione della proliferazione e del differenziamento degli spermatogoni nel testicolo prepuberale di topo. Lo studio ha riguardato la caratterizzazione della RNA-binding-protein Nanos 3 e il suo coinvolgimento nella progressione del ciclo cellulare degli spermatogoni e l’analisi dell’espressione genica negli spermatogoni in seguito a trattamento con i fattori Kit ligand e acido retinico, coinvolti rispettivamente nella proliferazione e nel differenziamento in questo tipo cellulare.
Nel topo sono stati identificati tre omologhi di Nanos , Nanos1, Nanos2 and Nanos3. L’ortologo Nanos3 è espresso nella gonade embrionale maschile e femminile e dopo la nascita è presente esclusivamente nel testicolo. E’ noto che l’eliminazione di questo gene comporta la perdita completa della cellule germinali in entrambi i sessi, ma la funzione di Nanos3 nel testicolo dopo la nascita, non è stata ancora analizzata. Nel nostro studio dimostriamo che Nanos3 è espresso negli spermatogoni indifferenziati del testicolo prepuberale di topo e che l’over espressione di questo gene comporta un aumento della percentuale di cellule nella fase G1, suggerendone il coinvolgimento nel rallentamento della progressione del ciclo cellulare degli spermatogoni. Dimostriamo inoltre, che il meccanismo di azione di Nanos 3, quale potenziale repressore della traduzione, è conservato nel topo e prevede l’interazione con una seconda RNA-binding protein murina, Pumilio2. In accordo con il possibile coinvolgimento di Nanos 3 nell’inibizione del differenziamento degli spermatogoni, il trattamento con il fattore differenziativo acido retinoico all-trans (ATRA), induce una notevole down-regolazione della sua espressione. Questi risultati permettono di concludere che nel testicolo di topo dopo la nascita, Nanos3 è un fattore importante nel mantenimento dello stato indifferenziato degli spermatogoni attraverso la regolazione del loro ciclo. Nella seconda parte del lavoro, è stata realizzata un’ analisi, mediante macroarray, dei geni regolati dal trattamento con KL o ATRA negli spermatogoni ottenuti da topi di 7 giorni di età. L’analisi ha evidenziato che il pattern di espressione dei geni che risultano indotti da KL è in accordo con i cambiamenti che intervengono nel ciclo cellulare durante le divisioni cellulari successive degli spermatogoni di tipo A e B. KL induce, infatti, l’espressione dei geni correlati alle fasi iniziali della meiosi negli spermatogoni in differenziamento e regola negativamente l’espressione dei geni tipicamente presenti negli spermatogoni durante la fase mitotica. Inoltre, l’analisi dei dati ottenuti da spermatogoni trattati con acido retinoico, ha confermato che l’ ATRA induce un pattern di espressione genica compatibile con la progressione verso il programma differenziativo della meiosi, suggerendo che l’ATRA e KL, indipendentemente, sono in grado di promuovere l’entrata nella meiosi negli spermatogoni.The aim of this work is to understand some of the molecular mechanisms which regulate spermatogonial germ cells proliferation and differentiation in the prepuberal mouse testis. In this study we characterized the RNA-binding protein Nanos3 and its role in the regulation of spermatogonial cell-cycle progression. Furthermore we performed a transcriptome analysis of spermatogonia stimulated with KL and retinoic acid, which have been previously shown to regulate their proliferaton and differentiaton, respectively. In the mouse, three Nanos homologs have been identified, Nanos1, Nanos2 and Nanos3. The Nanos3 ortholog is expressed in both male and female gonads of early embryo and, after birth, it is found only in the testis. Nanos3 targeted disruption results in the complete loss of germ cells in both sexes during embryonic development, however the role of Nanos3 in the testis during the postnatal period has not been explored yet. We found that Nanos3 is expressed in undifferentiated spermatogonia of the prepuberal testis and that its up-regulation causes accumulation of cells in the G1 phase, suggesting that this protein is able to delay the cell cycle progression of spermatogonial cells. We also demonstrate a conserved mechanism of action of Nanos3 as potential translational repressor, involving the interaction with the murine RNA-binding protein Pumilio2. According to the possible role of Nanos3 in inhibiting spermatogonia cell differentiation, treatment with the differentiating factor all-trans retinoic acid (ATRA) induces a dramatic down-regulation of its expression. These results allowed us to conclude that, in the prepuberal testis, Nanos3 is important to maintain undifferentiated spermatogonia via the regulation of their cell cycle. In the second part of the work we performed a wide genome analysis of gene expression regulated by treatment with KL or ATRA of spermatogonia from 7-day-old mice. The analysis revealed that the pattern of RNA expression induced by KL is compatible with the qualitative changes of the cell cycle that occur during the subsequent cell divisions in type A and B spermatogonia. Moreover, KL up-regulates in differentiating spermatogonia the expression of early meiotic genes whereas it down-regulates typical spermatogonial markers. Since KL modifies the expression of several genes known to be up-regulated or down-regulated in spermatogonia during the transition from the mitotic to the meiotic cell cycle, these results are consistent with a role of the KL/Kit interaction in the induction of their meiotic differentiation. Microarray analysis on stimulated spermatogonia showed that ATRA induces a pattern of gene expression which is compatible with their ongoing differentiating program toward meiosis, suggesting that ATRA and KL, independently, are able to promote meiotic entry of postnatal spermatogonia.
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El, Sharnouby Sherif Maher. « Methodology for genome-wide epigenetic profiling of the Drosophila male germline ». Thesis, University of Cambridge, 2011. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.609941.
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Cassidy, Liam David. « The evolution of cancer in germline BRCA2 mutation carriers ». Thesis, University of Cambridge, 2014. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.648683.
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Chung, Shui-wah. « Cell-cell interactions in the rat testis : biology and future perspectives / ». Hong Kong : University of Hong Kong, 1999. http://sunzi.lib.hku.hk/hkuto/record.jsp?B2056692X.
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Roewer, Jesse F. « CONSEQUENCE OF PREMATURE AND CHRONIC LUTEINIZING HORMONE RECEPTOR ACTIVATION ON TESTICULAR SPERMATOGENIC CELL DEVELOPMENT ». OpenSIUC, 2010. https://opensiuc.lib.siu.edu/theses/253.
Texte intégralRésumé :
Luteinizing hormone (LH), one of the two gonadotropin hormones released from the anterior pituitary gland, binds to its receptor (LHR) in the gonads to stimulate steroid hormone production, in addition to ovulation and gametogenesis. Mutations of the receptors amino acid sequence have the ability to either constitutively activate or inactivate it. All activating mutations result in male-limited precocious puberty. Males with this condition undergo puberty around 4 years of age, and have a premature elevation in testosterone levels and premature skeletal development. In order to understand how chronic ligand-mediated activation of the LHR affects gonadal development and function, a mouse model expressing a yoked hormone-receptor (YHR) complex, engineered by covalently linking the hormone human chorionic gonadotropin to the rat LHR, has been studied. YHR+ males have prepubertally elevated testosterone and decreased gonadotropin levels, smaller testis, and smaller average seminiferous tubule diameters when compared to wild type (WT) animals. In a preliminary breeding study it was shown that YHR+ males were sub-fertile. Based the phenotype exhibited by the YHR+ mice, it was hypothesized that increased levels of testosterone in addition to decreased gonadotropin hormone levels in neonatal and prepubertal mice that results from premature activation of the luteinizing hormone receptor causes spermatogenesis to be impaired. The first objective of this study was to determine if there was a difference in the testicular germ cell and Sertoli cell populations in the WT and YHR+ animals using flow cytometry and systematic Sertoli cell counting, respectively. There was no difference in the Sertoli cell population between YHR+ animals and WT controls, but there were significantly fewer total germ cells in YHR+ animals at 10 days and from 4 weeks through adulthood. The second objective was to calculate the daily sperm production in the testis and epididymis of WT and YHR+ animals in order to determine if there is a further decrease in the total sperm count due to an epididymal dysfunction. Interestingly, there were significantly fewer sperm calculated in the caput/corpus region of the epididymis in YHR+ males at 12 weeks of age, but not in the testis and cauda epididymis. Furthermore, the daily sperm production in WT and YHR+ mice at 16 weeks of age were not significantly different. The final objective was to determine if the decrease in germ cells observed in YHR+ animals is the result of decreased proliferation or an increase in either germ cell or Sertoli cell apoptosis. Quantification of germ cell and Sertoli cell proliferation revealed no significant difference between the WT and YHR+ animals. Similar findings were found after quantification of apoptotic germ cell and Sertoli cells. Taken together, these data suggest that premature elevation in testosterone and persistently lower levels of circulating follicle stimulating hormone (FSH) are affecting Sertoli cell function, which is causing a reduced germ cell to Sertoli cell ratio in the YHR+ mice. These data suggest that the decrease in testis weight and seminiferous tubule diameter in YHR+ mine is due to a decrease in germ cell rather than Sertoli cell number.
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Wong, Ching-hang, et 黃政珩. « Cell-cell interactions and cell junction dynamics in the mammalian testis ». Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2005. http://hub.hku.hk/bib/B31993084.
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Yamaguchi, Shinpei. « Conditional Knockdown of Nanog Induces Apoptotic Cell Death in Mouse Migrating Primordial Germ Cells ». Kyoto University, 2010. http://hdl.handle.net/2433/97931.
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Murase, Yusuke. « Long-term expansion with germline potential of human primordial germ cell-like cells in vitro ». Doctoral thesis, Kyoto University, 2021. http://hdl.handle.net/2433/261607.
Texte intégralRésumé :
京都大学0048
新制・課程博士
博士(医学)
甲第22880号
医博第4674号
新制||医||1047(附属図書館)
京都大学大学院医学研究科医学専攻
(主査)教授 篠原 隆司, 教授 近藤 玄, 教授 万代 昌紀
学位規則第4条第1項該当
Doctor of Medical Science
Kyoto University
DFAM
49
Nakaki, Fumio. « Induction of mouse germ-cell fate by transcription factors in vitro ». Kyoto University, 2014. http://hdl.handle.net/2433/188684.
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Yamada, Yukiko. « Deciphering molecular mechanisms that regulate programmed cell death of primordial germ cells in Drosophila melanogaster ». [Ames, Iowa : Iowa State University], 2007. http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:3337369.
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