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Expressão de marcadores moleculares em espermatogônias; Expression of molecular markers in spermatogonia

Giassetti, Mariana Ianello
Fonte: Biblioteca Digitais de Teses e Dissertações da USP Publicador: Biblioteca Digitais de Teses e Dissertações da USP
Tipo: Tese de Doutorado Formato: application/pdf
Publicado em 03/06/2015 PT
Relevância na Pesquisa
66.14%
Em mamíferos, a espermatogênese é mantida pela autorrenovação e diferenciação das células-tronco espermatogoniais (SSC). Apesar da grande importância do SSC para a fertilidade masculina, em Bos taurus pouco se sabe sobre a sua identificação e biologia celular. Para roedores, mais de 30 marcadores para células germinativas indiferenciadas já foram descritos. No entanto, ainda não é conhecido um marcador específico apenas para SSC. Quase todos são também expressos por gonócitos, espermatogônias mais diferenciadas ou mesmo células somáticas. Yin Yang 2 (YY2) é um factor de transcrição expresso nas células com a morfologia de gonócitos e SSC, sendo um candidato a marcador de SSC. Assim, a identificação de novos marcadores para SSC e factores que afectam a sua expressão, tais como a idade, são fundamentais para o desenvolvimento da biotecnologia como transgenia e tratamento de infertilidade, nos quais as SSC poderiam ser ferramentas biológicos importantes. Assim, nesta tese temos duas hipóteses principais: 1) a idade do dador afeta a expressão de marcadores moleculares específicos de SSC bovinas assim como potencial de células-tronco dessas células e que as sequências de DNA em que se associa YY2 regulam a expressão génica de SSC em camundongos. Os objetivos específicos...

Testicular germ cells can colonize sexually undifferentiated embryonic gonad and produce functional eggs in fish

Okutsu, Tomoyuki; Suzuki, Kensuke; Takeuchi, Yutaka; Takeuchi, Toshio; Yoshizaki, Goro
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
EN
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46.39%
Understanding the mechanisms that regulate germ-cell development is crucial to reproductive medicine and animal production. Animal gametes originally derive from sexually undifferentiated primordial germ cells (PGCs), which develop into mitotic germ cells (oogonia or spermatogonia) before proceeding to meiosis [Wylie, C. (1999) Cell 96, 165–174]. Spermatogonia are thought to include a population of cells with stem cell activity, which proliferate throughout the lifespan of male animals and produce spermatozoa [Zhao, G. Q. & Garbers, D. L. (2002) Dev. Cell 2, 537–547]. However, the functional differences between PGCs and spermatogonial stem cells are poorly understood. Here we show that transplanted adult testicular germ cells can colonize sexually undifferentiated embryonic gonads and resume gametogenesis. Testicular germ cells containing spermatogonial stem cells isolated from adult male rainbow trout (Oncorhynchus mykiss) were transplanted into the peritoneal cavity of newly hatched embryos of both sexes, and the behavior of the donor cells was observed. The testicular germ cells differentiated into spermatozoa in male recipients and fully functional eggs in female recipients. Furthermore, the donor-derived spermatozoa and eggs obtained from the recipient fish were able to produce normal offspring. These findings indicate that fish testicular germ cells...

Cyp26b1 Expression in Murine Sertoli Cells Is Required to Maintain Male Germ Cells in an Undifferentiated State during Embryogenesis

Li, Hui; MacLean, Glenn; Cameron, Don; Clagett-Dame, Margaret; Petkovich, Martin
Fonte: Public Library of Science Publicador: Public Library of Science
Tipo: Artigo de Revista Científica
Publicado em 19/10/2009 EN
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46.35%
In mammals, germ cells within the developing gonad follow a sexually dimorphic pathway. Germ cells in the murine ovary enter meiotic prophase during embryogenesis, whereas germ cells in the embryonic testis arrest in G0 of mitotic cell cycle and do not enter meiosis until after birth. In mice, retinoic acid (RA) signaling has been implicated in controlling entry into meiosis in germ cells, as meiosis in male embryonic germ cells is blocked by the activity of a RA-catabolizing enzyme, CYP26B1. However, the mechanisms regulating mitotic arrest in male germ cells are not well understood. Cyp26b1 expression in the testes begins in somatic cells at embryonic day (E) 11.5, prior to mitotic arrest, and persists throughout fetal development. Here, we show that Sertoli cell-specific loss of CYP26B1 activity between E15.5 and E16.5, several days after germ cell sex determination, causes male germ cells to exit from G0, re-enter the mitotic cell cycle and initiate meiotic prophase. These results suggest that male germ cells retain the developmental potential to differentiate in meiosis until at least at E15.5. CYP26B1 in Sertoli cells acts as a masculinizing factor to arrest male germ cells in the G0 phase of the cell cycle and prevents them from entering meiosis...

Vaccinia-Related Kinase 1 Is Required for the Maintenance of Undifferentiated Spermatogonia in Mouse Male Germ Cells

Choi, Yoon Ha; Park, Choon-Ho; Kim, Wanil; Ling, Hua; Kang, Aram; Chang, Matthew Wook; Im, Sun-Kyoung; Jeong, Hyun-Woo; Kong, Young-Yun; Kim, Kyong-Tai
Fonte: Public Library of Science Publicador: Public Library of Science
Tipo: Artigo de Revista Científica
Publicado em 13/12/2010 EN
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46.31%
Vaccinia-related kinase 1 (VRK1) is a crucial protein kinase for mitotic regulation. VRK1 is known to play a role in germ cell development, and its deficiency results in sterility. Here we describe that VRK1 is essential for the maintenance of spermatogonial stem cells. To determine whether VRK1 plays a role in these cells, we assessed the population size of undifferentiated spermatogonia. Flow cytometry analyses showed that the number of undifferentiated spermatogonia was markedly reduced in VRK1-deficient testes. VRK1 was highly expressed in spermatogonial populations, and approximately 66% of undifferentiated spermatogonia that were sorted as an Ep-CAM+/c-kit−/alpha-6-integrin+ population showed a positive signal for VRK1. Undifferentiated stem cells expressing Plzf and Oct4 but not c-kit also expressed VRK1, suggesting that VRK1 is an intrinsic factor for the maintenance of spermatogonial stem cells. Microarray analyses of the global testicular transcriptome and quantitative RT-PCR of VRK1-deficient testes revealed significantly reduced expression levels of undifferentiated spermatogonial marker genes in early postnatal mice. Together, these results suggest that VRK1 is required for the proliferation and differentiation of undifferentiated spermatogonia...

Licensing of gametogenesis, dependent on RNA binding protein DAZL, as a gateway to sexual differentiation of fetal germ cells

Gill, Mark E.; Hu, Yueh-Chiang; Lin, Yanfeng; Page, David C.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
46.16%
Mammalian oocytes and spermatozoa derive from fetal cells shared by the sexes. These primordial germ cells (PGCs) migrate to the developing somatic gonad, giving rise to oocytes or spermatozoa. These opposing sexual fates are determined not by the PGCs’ own sex chromosome constitution (XX or XY), but by the sexual identity of the fetal gonad that they enter. We asked whether PGCs undergo a developmental transition that enables them to respond to feminizing or masculinizing cues from fetal ovary or testis. We conducted in vivo genetic studies of DAZL, an RNA-binding protein expressed in both ovarian and testicular germ cells. We found that germ cells in C57BL/6 Dazl-deficient fetuses—whether XX or XY—migrate to the gonad but do not develop either male or female features. Instead, they remain in a sexually undifferentiated state similar to that of migrating PGCs. Thus, germ cells in C57BL/6 Dazl-deficient fetuses do not respond to sexual cues from ovary or testis, whereas the earlier processes of germ cell specification and migration are unaffected. We propose that PGCs of both XX and XY fetuses undergo licensing, an active developmental transition that enables the resultant gametogenesis-competent cells to respond to feminizing or masculinizing cues produced by the fetal ovary or testis and hence to embark on oogenesis or spermatogenesis. In C57BL/6 mice...

MicroRNA-mediated posttranscriptional regulation is required for maintaining undifferentiated properties of blastoderm and primordial germ cells in chickens

Lee, Sang In; Lee, Bo Ram; Hwang, Young Sun; Lee, Hyung Chul; Rengaraj, Deivendran; Song, Gwonhwa; Park, Tae Sub; Han, Jae Yong
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
46.29%
MicroRNAs (miRNAs) play a critical role in determining the differentiation fate of pluripotent stem cells and germ cells in mammals. However, the mechanism(s) of miRNA-mediated posttranscriptional regulation with regard to lineage specification and differentiation in chick development require further investigation. Therefore, we conducted miRNA expression profiling to explore specific miRNA signatures in undifferentiated blastoderm and primordial germ cells (PGCs). We identified seven miRNAs that are highly expressed in blastoderm and 10 that are highly expressed in PGCs. In this study, miR-302a and miR-456 for blastoderm and miR-181a* for PGCs were analyzed further for their target transcripts and regulatory pathways. Both miR-302a and miR-456 bound directly to the sex-determining region Y box 11 transcript and could act as posttranscriptional coregulators to maintain the undifferentiated state of the chicken blastoderm through the suppression of somatic gene expression and differentiation. Moreover, miR-181a* showed a bifunctional role in PGCs by binding to two different transcripts. miR-181a* inhibited the somatic differentiation of PGCs by silencing homeobox A1 expression. Additionally, miR-181a* prevented PGCs from entering meiosis through the repression of the nuclear receptor subfamily 6...

Mutation in the type IB bone morphogenetic protein receptor alk6b impairs germ-cell differentiation and causes germ-cell tumors in zebrafish

Neumann, Joanie C.; Chandler, Garvin L.; Damoulis, Vanessa A.; Fustino, Nicholas J.; Lillard, Katherine; Looijenga, Leendert; Margraf, Linda; Rakheja, Dinesh; Amatruda, James F.
Fonte: National Academy of Sciences Publicador: National Academy of Sciences
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
46.25%
Germ-cell tumors (GCTs), which arise from pluripotent embryonic germ cells, exhibit a wide range of histologic differentiation states with varying clinical behaviors. Although testicular GCT is the most common cancer of young men, the genes controlling the development and differentiation of GCTs remain largely unknown. Through a forward genetic screen, we previously identified a zebrafish mutant line, tgct, which develops spontaneous GCTs consisting of undifferentiated germ cells [Neumann JC, et al. (2009) Zebrafish 6:319–327]. Using positional cloning we have identified an inactivating mutation in alk6b, a type IB bone morphogenetic protein (BMP) receptor, as the cause of the zebrafish GCT phenotype. Alk6b is expressed in spermatogonia and early oocytes, and alk6b mutant gonads display impaired BMP signal transduction, altered expression of BMP target genes, and abnormal germ-cell differentiation. We find a similar absence of BMP signaling in undifferentiated human GCTs, such as seminomas and embryonal carcinoma, but not in normal testis or in differentiated GCTs. These results indicate a germ-cell–autonomous role for BMP signal transduction in germ-cell differentiation, and highlight the importance of the BMP pathway in human GCTs.

Sin3a Is Required by Sertoli Cells to Establish a Niche for Undifferentiated Spermatogonia, Germ Cell Tumors, and Spermatid Elongation

Payne, Christopher J.; Gallagher, Shannon J.; Foreman, Oded; Dannenberg, Jan-Hermen; DePinho, Ronald A.; Braun, Robert E.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
Publicado em /08/2010 EN
Relevância na Pesquisa
36.28%
Microenvironments support the maintenance of stem cells and the growth of tumors through largely unknown mechanisms. While cell-autonomous chromatin modifications have emerged as important determinants for self-renewal and differentiation of stem cells, a role for non-cell autonomous epigenetic contributions is not well established. Here, we genetically ablated the chromatin modifier Sin3a in fetal Sertoli cells, which partly comprise the niche for male germline stem cells, and investigated its impact on spermatogenic cell fate and teratoma formation in vivo. Sertoli cell-specific Sin3a deletion resulted in the formation of few undifferentiated spermatogonia after birth while initially maintaining spermatogenic differentiation. Stem cell-associated markers Plzf, Gfra1, and Oct4 were downregulated in the mutant fetal gonad, while Sertoli cell markers Steel and Gdnf, which support germ cells, were not diminished. Following birth, markers of differentiating spermatogonia, Kit and Sohlh2, exhibited normal levels, but chemokine signaling molecules CXCL12/SDF1 and CXCR4, expressed in Sertoli cells and germ cells, respectively, were not detected. In the juvenile, mutant testes exhibited a progressive loss of differentiating spermatogonia and a block in spermatid elongation...

Effects of multiple doses of cyclophosphamide on mouse testes: Accessing the germ cells lost, and the functional damage of stem cells

Drumond, Ana Luiza; Weng, Connie C.; Wang, Gensheng; Chiarini-Garcia, Helio; Eras-Garcia, Leticia; Meistrich, Marvin L.
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
46%
Spermatogenesis is sensitive to the chemotherapeutic drug cyclophosphamide, which decreases the patients’ sperm count. Since the recovery of fertility is dependent on regeneration from stem cells, in the present study we evaluated the ability of cyclophosphamide-exposed stem spermatogonia from mice to regenerate spermatogenesis in situ and after transplantation. When seven doses of cyclophosphamide were given at 4-day intervals, the differentiating germ cells were largely eliminated but ~50% of the undifferentiated type A spermatogonia remained. We monitored the recovery and found that sperm production recovered to 64% of control within the time expected. When the cyclophosphamide-surviving spermatogonia were transplanted into recipient mice, recovery of spermatogenesis from the cyclophosphamide-exposed donor cells was observed, but was reduced when compared to cells from cryptorchid donors. Thus, multidose regimens of cyclophosphamide did not eliminate the stem spermatogonia, but resulted in cell loss and residual damage.

Temporal Transcriptional Profiling of Somatic and Germ Cells Reveals Biased Lineage Priming of Sexual Fate in the Fetal Mouse Gonad

Jameson, Samantha A.; Natarajan, Anirudh; Cool, Jonah; DeFalco, Tony; Maatouk, Danielle M.; Mork, Lindsey; Munger, Steven C.; Capel, Blanche
Fonte: Public Library of Science Publicador: Public Library of Science
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
46.24%
The divergence of distinct cell populations from multipotent progenitors is poorly understood, particularly in vivo. The gonad is an ideal place to study this process, because it originates as a bipotential primordium where multiple distinct lineages acquire sex-specific fates as the organ differentiates as a testis or an ovary. To gain a more detailed understanding of the process of gonadal differentiation at the level of the individual cell populations, we conducted microarrays on sorted cells from XX and XY mouse gonads at three time points spanning the period when the gonadal cells transition from sexually undifferentiated progenitors to their respective sex-specific fates. We analyzed supporting cells, interstitial/stromal cells, germ cells, and endothelial cells. This work identified genes specifically depleted and enriched in each lineage as it underwent sex-specific differentiation. We determined that the sexually undifferentiated germ cell and supporting cell progenitors showed lineage priming. We found that germ cell progenitors were primed with a bias toward the male fate. In contrast, supporting cells were primed with a female bias, indicative of the robust repression program involved in the commitment to XY supporting cell fate. This study provides a molecular explanation reconciling the female default and balanced models of sex determination and represents a rich resource for the field. More importantly...

The receptor tyrosine phosphatase Lar regulates adhesion between Drosophila male germline stem cells and the niche

Srinivasan, Shrividhya; Mahowald, Anthony P.; Fuller, Margaret T.
Fonte: Company of Biologists Publicador: Company of Biologists
Tipo: Artigo de Revista Científica
Publicado em 15/04/2012 EN
Relevância na Pesquisa
45.91%
The stem cell niche provides a supportive microenvironment to maintain adult stem cells in their undifferentiated state. Adhesion between adult stem cells and niche cells or the local basement membrane ensures retention of stem cells in the niche environment. Drosophila male germline stem cells (GSCs) attach to somatic hub cells, a component of their niche, through E-cadherin-mediated adherens junctions, and orient their centrosomes toward these localized junctional complexes to carry out asymmetric divisions. Here we show that the transmembrane receptor tyrosine phosphatase Leukocyte-antigen-related-like (Lar), which is best known for its function in axonal migration and synapse morphogenesis in the nervous system, helps maintain GSCs at the hub by promoting E-cadherin-based adhesion between hub cells and GSCs. Lar is expressed in GSCs and early spermatogonial cells and localizes to the hub-GSC interface. Loss of Lar function resulted in a reduced number of GSCs at the hub. Lar function was required cell-autonomously in germ cells for proper localization of Adenomatous polyposis coli 2 and E-cadherin at the hub-GSC interface and for the proper orientation of centrosomes in GSCs. Ultrastructural analysis revealed that in Lar mutants the adherens junctions between hub cells and GSCs lack the characteristic dense staining seen in wild-type controls. Thus...

Mei-P26 regulates the maintenance of ovarian germline stem cells by promoting BMP signaling

Li, Yun; Maines, Jean Z.; Tastan, Ömür Y.; McKearin, Dennis M.; Buszczak, Michael
Fonte: Company of Biologists Publicador: Company of Biologists
Tipo: Artigo de Revista Científica
Publicado em 01/05/2012 EN
Relevância na Pesquisa
56.21%
In the Drosophila ovary, bone morphogenetic protein (BMP) ligands maintain germline stem cells (GSCs) in an undifferentiated state. The activation of the BMP pathway within GSCs results in the transcriptional repression of the differentiation factor bag of marbles (bam). The Nanos-Pumilio translational repressor complex and the miRNA pathway also help to promote GSC self-renewal. How the activities of different transcriptional and translational regulators are coordinated to keep the GSC in an undifferentiated state remains uncertain. Data presented here show that Mei-P26 cell-autonomously regulates GSC maintenance in addition to its previously described role of promoting germline cyst development. Within undifferentiated germ cells, Mei-P26 associates with miRNA pathway components and represses the translation of a shared target mRNA, suggesting that Mei-P26 can enhance miRNA-mediated silencing in specific contexts. In addition, disruption of mei-P26 compromises BMP signaling, resulting in the inappropriate expression of bam in germ cells immediately adjacent to the cap cell niche. Loss of mei-P26 results in premature translation of the BMP antagonist Brat in germline stem cells. These data suggest that Mei-P26 has distinct functions in the ovary and participates in regulating the fates of both GSCs and their differentiating daughters.

MicroRNAs 221 and 222 regulate the undifferentiated state in mammalian male germ cells

Yang, Qi-En; Racicot, Karen E.; Kaucher, Amy V.; Oatley, Melissa J.; Oatley, Jon M.
Fonte: Company of Biologists Publicador: Company of Biologists
Tipo: Artigo de Revista Científica
Publicado em 15/01/2013 EN
Relevância na Pesquisa
46.18%
Continuity of cycling cell lineages relies on the activities of undifferentiated stem cell-containing subpopulations. Transition to a differentiating state must occur periodically in a fraction of the population to supply mature cells, coincident with maintenance of the undifferentiated state in others to sustain a foundational stem cell pool. At present, molecular mechanisms regulating these activities are poorly defined for most cell lineages. Spermatogenesis is a model process that is supported by an undifferentiated spermatogonial population and transition to a differentiating state involves attained expression of the KIT receptor. We found that impaired function of the X chromosome-clustered microRNAs 221 and 222 (miR-221/222) in mouse undifferentiated spermatogonia induces transition from a KIT– to a KIT+ state and loss of stem cell capacity to regenerate spermatogenesis. Both Kit mRNA and KIT protein abundance are influenced by miR-221/222 function in spermatogonia. Growth factors that promote maintenance of undifferentiated spermatogonia upregulate miR-221/222 expression; whereas exposure to retinoic acid, an inducer of spermatogonial differentiation, downregulates miR-221/222 abundance. Furthermore, undifferentiated spermatogonia overexpressing miR-221/222 are resistant to retinoic acid-induced transition to a KIT+ state and are incapable of differentiation in vivo. These findings indicate that miR-221/222 plays a crucial role in maintaining the undifferentiated state of mammalian spermatogonia through repression of KIT expression.

Differentiation of murine male germ cells to spermatozoa in a soft agar culture system

Abu Elhija, Mahmoud; Lunenfeld, Eitan; Schlatt, Stefan; Huleihel, Mahmoud
Fonte: Nature Publishing Group Publicador: Nature Publishing Group
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
46.13%
Establishment of an in vitro system that allows the development of testicular germ cells to sperm will be valuable for studies of spermatogenesis and future treatments for male infertility. In the present study, we developed in vitro culture conditions using three-dimensional agar culture system (SACS), which has the capacity to induce testicular germ cells to reach the final stages of spermatogenesis, including spermatozoa generation. Seminiferous tubules from testes of 7-day-old mice were enzymatically dissociated, and intratubular cells were cultured in the upper layer of the SACS in RPMI medium supplemented with fetal calf serum (FCS). The lower layer of the SACS contained only RPMI medium supplemented with FCS. Colonies in the upper layer were isolated after 14 and 28 days of culture and were classified according to their size. Immunofluorescence and real-time PCR were used to analyse specific markers expressed in undifferentiated and differentiated spermatogonia (Vasa, Dazl, OCT-4, C-Kit, GFR-α-1, CD9 and α-6-integrin), meiotic cells (LDH, Crem-1 and Boule) and post-meiotic cells (Protamine-1, Acrosin and SP-10). Our results reveal that it is possible to induce mouse testicular pre-meiotic germ cell expansion and induce their differentiation to spermatozoa in SACS. The spermatozoa showed normal morphology and contained acrosomes. Thus...

Fate of induced pluripotent stem cells following transplantation to murine seminiferous tubules

Durruthy Durruthy, Jens; Ramathal, Cyril; Sukhwani, Meena; Fang, Fang; Cui, Jun; Orwig, Kyle E.; Reijo Pera, Renee A.
Fonte: Oxford University Press Publicador: Oxford University Press
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
36.34%
Studies of human germ cell development are limited in large part by inaccessibility of germ cells during development. Moreover, although several studies have reported differentiation of mouse and human germ cells from pluripotent stem cells (PSCs) in vitro, differentiation of human germ cells from PSCs in vivo has not been reported. Here, we tested whether mRNA reprogramming in combination with xeno-transplantation may provide a viable system to probe the genetics of human germ cell development via use of induced pluripotent stem cells (iPSCs). For this purpose, we derived integration-free iPSCs via mRNA-based reprogramming with OCT3/4, SOX2, KLF4 and cMYC alone (OSKM) or in combination with the germ cell-specific mRNA, VASA (OSKMV). All iPSC lines met classic criteria of pluripotency. Moreover, global gene expression profiling did not distinguish large differences between undifferentiated OSKM and OSKMV iPSCs; however, some differences were observed in expression of pluripotency factors and germ cell-specific genes, and in epigenetic profiles and in vitro differentiation studies. In contrast, transplantation of undifferentiated iPSCs directly into the seminiferous tubules of germ cell-depleted immunodeficient mice revealed divergent fates of iPSCs produced with different factors. Transplantation resulted in morphologically and immunohistochemically recognizable germ cells in vivo...

UTF1, a Putative Marker for Spermatogonial Stem Cells in Stallions

Jung, Heejun; Roser, Janet F.; Yoon, Minjung
Fonte: Public Library of Science Publicador: Public Library of Science
Tipo: Artigo de Revista Científica
Publicado em 01/10/2014 EN
Relevância na Pesquisa
36.31%
Spermatogonial stem cells (SSCs) continuously undergo self-renewal and differentiation to sustain spermatogenesis throughout adulthood in males. In stallions, SSCs may be used for the production of progeny from geldings after cryopreservation and therapy for infertile and subfertile stallions. Undifferentiated cell transcription factor 1 (UTF1) is a putative marker for undifferentiated spermatogonia in humans and rats. The main purposes of this study are to determine the following: 1) changes in the expression pattern of UTF1 at various reproductive stages of stallions, 2) subpopulations of spermatogonia that express UTF1. Testicular samples were collected and categorized based on the age of the horses as follows: pre-pubertal (<1 yr), pubertal (1–1.5 yr), post-pubertal (2–3 yr), and adult (4–8 yr). Western blot analysis was utilized to determine the cross-activity of the UTF1 antibody to horse testes tissues. Immunohistochemistry was conducted to investigate the UTF1 expression pattern in germ cells at different reproductive stages. Whole mount staining was applied to determine the subpopulation of UTF1-positive spermatogonia. Immunohistological analysis showed that most germ cells in the pre-pubertal and pubertal stages were immunolabeled with UTF1...

Interactions of diploid embryonal carcinoma cells and normal cell mass cells with 2.5 day mouse embryos following aggregation

Waters, Barbara Kathleen.
Fonte: Brock University Publicador: Brock University
Tipo: Electronic Thesis or Dissertation
ENG
Relevância na Pesquisa
45.92%
Spontaneous teratocarcinomas are ovarian or testicular tumors which have their origins in germ cells. The tumors contain a disorganized array of benign differentiated cells as well as an undifferentiated population of malignant stem cells, the embryonal carcinoma or EC cells. These pluripotent stem cells in tissue culture share many properties with the transient pluripotent cells of the early embryo, and might therefore serve as models for the investigation of developmental events ill vitro. The property of EC cells of prime interest in this study is an in vivo phenomenon. Certain EC cell lines are known to be regulated ill vivo and to differentiate normally in association with normal embryonic cells, resulting in chimeric mice. These mice have two genetically distinct cell populations, one of which is derived from the originally malignant EC cells. This has usually been accomplished by injection of the EC cells into the Day 3 blastocyst. In this study, the interactions between earlier stage embryos and EC cells have been tested by aggregating clumps of EC cells with Day 2 embryos. The few previous aggregation studies produced a high degree of abnormality in chimeric embryos, but the EC cells employed had known chromosomal abnormalities. In this study...

MicroRNA expression profile during adipogenic differentiation in mouse embryonic stem cells

Knelangen, J.; Van der Hoek, M.; Kong, W.C.; Owens, J.; Fischer, B.; Santos, A.
Fonte: Amer Physiological Soc Publicador: Amer Physiological Soc
Tipo: Artigo de Revista Científica
Publicado em //2011 EN
Relevância na Pesquisa
45.89%
Pluripotent embryonic stem cells (ESC) have the potential to differentiate into any cell type of the three germ layers. Differentiation processes depend on genetic and epigenetic factors. The guidance of cell fate determination by microRNAs (miRs) seems important for embryonic development and cell lineage decisions. MiRs are short, single-stranded, noncoding RNA molecules that regulate through posttranscriptional modulation, a subset of target genes involved in cell differentiation and specific cell function. We have used microarray profiling of miRs in the mouse embryonic stem cell line CGR8. Comparison of the miR profiles of undifferentiated stem cells with mesodermal progenitors cells (day 5), preadipocytes (day 10), and adipocytes (day 21) showed that the expression level of 129 miRs changed (twofold) during adipogenic differentiation. We identified 10 clusters of differentially expressed miRs, which contain putative markers and regulators of mesodermal differentiation and cell fate determination into adipocytes. Notably, the adipocyte-specific miRs 143 and 103 were upregulated from day 10 onward. We have therefore demonstrated and characterized the dynamic profile of miR expression during murine adipogenic differentiation in vitro...

Expression of intermediate filaments and other special markers by testicular germ cell tumors. With reference to embryogenesis

Hanai, J.; Lin, M.; Wada, A.; Ishiguro, S.; Miki, T.; Sakaguchi, H.; Kanda, H.
Fonte: Murcia : F. Hernández Publicador: Murcia : F. Hernández
Tipo: Artigo de Revista Científica Formato: application/pdf
ENG
Relevância na Pesquisa
56.12%
Distribution of intermediate filament proteins (IFs) and severa1 special markers was studied in 39 testicular germ cell tumors and 8 embryos and foetuses. The similarity and difference between development of germ cell tumor and embryogenesis were immunohistochemically investigated. Seminoma and embryonal carcinoma, as tumoral counterparts of undifferentiated germ cells, were characterized by little IF expression. This study revealed that the maturing and differentiating process in germ cell tumor is different from normal embryonal development and the tumor cells showed leaping maturing steps in tumorigenesis. Immunostaining for IFs helped to discover the further differentiation occurring in embryonal carcinoma and to demonstrate heterogeneous elements in non-seminoma germ cell tumors, which sometimes might not be apparent by light microscopical observation of H&E staining section. According to the findings, two patterns in mixed germ cell tumors are suggested; Le., combined and diffuse types. The mechanism of tumorigenesis of the two types is supposed to be different. Clinically, the prognosis of most patients with testicular germ cell tumor is fairly good because of the improved chemotherapies that are dependent on histological diagnosis.

Distinct Transcriptional Mechanisms Direct Expression of the Rat Dmrt1 Promoter in Sertoli Cells and Germ Cells of Transgenic Mice1

Lei, Ning; Karpova, Tatiana; Hornbaker, Kaori I.; Rice, Daren A.; Heckert, Leslie L.
Fonte: Society for the Study of Reproduction, Inc. Publicador: Society for the Study of Reproduction, Inc.
Tipo: Artigo de Revista Científica
EN
Relevância na Pesquisa
46.18%
DMRT1 is a transcription factor expressed only in Sertoli cells and undifferentiated spermatogonia of the postnatal testis, where it is required for proper cellular differentiation and fertility. To elucidate the transcriptional regulatory regions that provide DMRT1's cell-specific expression, transgenic mice containing a LacZ reporter gene driven by variable amounts of rat Dmrt1 5′ flanking sequence, 9 kb and smaller, were evaluated. Examination of transgene expression by RT-PCR indicated that multiple promoter regions direct Dmrt1 to the testis and that sequences upstream of 2.8 kb are needed for both Sertoli cell expression and limiting transcriptional influence imposed by surrounding chromatin. Thus, whereas many of the transgenes were expressed in the testis, the ones with smaller promoters were significantly more prone to expression at ectopic sites or to complete silencing. Transgene expression in Sertoli cells and germ cells was assessed by immunohistochemistry and RT-PCR following busulfan treatment to remove germ cells. Both evaluations indicated expression of the 9- and 3.2-kb promoters in Sertoli cells and germ cells, whereas activity of smaller promoters was largely restricted to germ cells. In all, the present study provides in vivo evidence that distinct promoter sequences participate in Dmrt1 regulation in somatic cells and germ cells...