Identification of transcripts involved in meiosis and follicle formation during ovine ovary development

<p>Abstract</p> <p>Background</p> <p>The key steps in germ cell survival during ovarian development are the entry into meiosis of oogonies and the formation of primordial follicles, which then determine the reproductive lifespan of the ovary. In sheep, these steps occur during fetal life, between 55 and 80 days of gestation, respectively. The aim of this study was to identify differentially expressed ovarian genes during prophase I meiosis and early folliculogenesis in sheep.</p> <p>Results</p> <p>In order to elucidate the molecular events associated with early ovarian differentiation, we generated two ovary stage-specific subtracted cDNA libraries using SSH. Large-scale sequencing of these SSH libraries identified 6,080 ESTs representing 2,535 contigs. Clustering and assembly of these ESTs resulted in a total of 2,101 unique sequences depicted in 1,305 singleton (62.11%) and 796 contigs (37.9%) ESTs (clusters). BLASTX evaluation indicated that 99% of the ESTs were homologous to various known genes/proteins in a broad range of organisms, especially ovine, bovine and human species. The remaining 1% which exhibited any homology to known gene sequences was considered as novel. Detailed study of the expression patterns of some of these genes using RT-PCR revealed new promising candidates for ovary differentiation genes in sheep.</p> <p>Conclusion</p> <p>We showed that the SSH approach was relevant to determining new mammalian genes which might be involved in oogenesis and early follicle development, and enabled the discovery of new potential oocyte and granulosa cell markers for future studies. These genes may have significant implications regarding our understanding of ovarian function in molecular terms, and for the development of innovative strategies to both promote and control fertility.</p

Toxicity Assays in Nanodrops Combining Bioassay and Morphometric Endpoints

BACKGROUND: Improved chemical hazard management such as REACH policy objective as well as drug ADMETOX prediction, while limiting the extent of animal testing, requires the development of increasingly high throughput as well as highly pertinent in vitro toxicity assays. METHODOLOGY: This report describes a new in vitro method for toxicity testing, combining cell-based assays in nanodrop Cell-on-Chip format with the use of a genetically engineered stress sensitive hepatic cell line. We tested the behavior of a stress inducible fluorescent HepG2 model in which Heat Shock Protein promoters controlled Enhanced-Green Fluorescent Protein expression upon exposure to Cadmium Chloride (CdCl(2)), Sodium Arsenate (NaAsO(2)) and Paraquat. In agreement with previous studies based on a micro-well format, we could observe a chemical-specific response, identified through differences in dynamics and amplitude. We especially determined IC50 values for CdCl(2) and NaAsO(2), in agreement with published data. Individual cell identification via image-based screening allowed us to perform multiparametric analyses. CONCLUSIONS: Using pre/sub lethal cell stress instead of cell mortality, we highlighted the high significance and the superior sensitivity of both stress promoter activation reporting and cell morphology parameters in measuring the cell response to a toxicant. These results demonstrate the first generation of high-throughput and high-content assays, capable of assessing chemical hazards in vitro within the REACH policy framework

TOPAZ1, a Novel Germ Cell-Specific Expressed Gene Conserved during Evolution across Vertebrates

BACKGROUND: We had previously reported that the Suppression Subtractive Hybridization (SSH) approach was relevant for the isolation of new mammalian genes involved in oogenesis and early follicle development. Some of these transcripts might be potential new oocyte and granulosa cell markers. We have now characterized one of them, named TOPAZ1 for the Testis and Ovary-specific PAZ domain gene. PRINCIPAL FINDINGS: Sheep and mouse TOPAZ1 mRNA have 4,803 bp and 4,962 bp open reading frames (20 exons), respectively, and encode putative TOPAZ1 proteins containing 1,600 and 1653 amino acids. They possess PAZ and CCCH domains. In sheep, TOPAZ1 mRNA is preferentially expressed in females during fetal life with a peak during prophase I of meiosis, and in males during adulthood. In the mouse, Topaz1 is a germ cell-specific gene. TOPAZ1 protein is highly conserved in vertebrates and specifically expressed in mouse and sheep gonads. It is localized in the cytoplasm of germ cells from the sheep fetal ovary and mouse adult testis. CONCLUSIONS: We have identified a novel PAZ-domain protein that is abundantly expressed in the gonads during germ cell meiosis. The expression pattern of TOPAZ1, and its high degree of conservation, suggests that it may play an important role in germ cell development. Further characterization of TOPAZ1 may elucidate the mechanisms involved in gametogenesis, and particularly in the RNA silencing process in the germ lin

Des souris et des femmes : une ovogenèse fœtale similaire ?

Des souris et des femmes : une ovogenèse fœtale similaire

Mammalian ovary differentiation - A focus on female meiosis

Over the past 50 years, the ovary development has been subject of fewer studies as compare to the male pathway. Nevertheless due to the advancement of genetics, mouse ES cells and the development of genetic models, studies of ovarian differentiation was boosted. This review emphasizes some of new progresses in the research field of the mammalian ovary differentiation that have occurred in recent years with focuses of the period around prophase I of meiosis and of recent roles of small non-RNAs in the ovarian gene expression

Mammalian gonodal differentiation : the pig model

SupplémentInternational audienc

Etude d'un nouveau gène exprimé dans les gonades de mammifères: TOPAZ1

National audienc

Male Infertility and Genetic screening: Guidelines in 2021

International audienceFor many years, genetic screening for male infertility was limited to a few analyses: karyotyping, screening for Y microdeletions, and tests for the most frequent cystic fibrosis transmembrane conductance regulator (CFTR) gene variants. The development of newtechnologies, such as chromosome microarray or new genome sequencing, has broadened access to wholegenome analyses. Over the last decade, many genetic defects have been described, and new strategies seem to emerge. Hence, by focusing on peripheral (rather than central) failures of spermatogenesis, the objectives of the present study were to review the latest data on clinical practice (rather than the physiopathology of these genetic abnormalities) and suggest new guidelines for the genetic screening of male infertility

L'atrésie folliculaire, un gaspillage programmé

National audienc

Genetic defects in human azoospermia

Résumé Comme pour beaucoup de maladies humaines, les analyses génétiques en cas d’azoospermie étaient initialement limitées à la réalisation d’un caryotype, conduisant au diagnostic de réarrangements chromosomiques comme pour le syndrome de Klinefelter ou autres syndromes. L’avènement de la biologie moléculaire, dans les années 1980, a permis l’élargissement du dépistage génétique à la recherche des microdélétions du chromosome Y et aux anomalies du gène CFTR (cystic fibrosis transmembrane conductance regulator). Il a fallu attendre plusieurs décennies et l’apparition des techniques d’analyses du génome entier pour que soit réalisée l’identification d’autres anomalies génétiques associés à l’azoospermie humaine. Si les anomalies des gènes TEX11 et ADGRG2 sont fréquemment décrites dans la littérature pour les hommes présentant une azoospermie, la plupart des altérations génétiques découvertes à ce jour sont privées, identifiées dans un petit nombre de familles souvent consanguines. L’objectif dans cette revue est de fournir un aperçu actualisé de toutes les anomalies génétiques décrites dans la littérature et associées à l’azoospermie humaine tout en essayant de fournir des guides de conduite diagnostique en fonction du phénotype de l’azoospermie. En plus des mutations homozygotes et délétères, les polymorphismes et les défauts épigénétiques sont également brièvement abordés. Néanmoins, comme ces variations ne sont que de potentiels facteurs de prédisposition à l’azoospermie, une étude spécifique sera nécessaire pour compiler l’ensemble des données de la littérature pour chaque variant génétique