Gene expression profiling of supporting, interstitial/stromal, and germ cells during the formation of the testis cords
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ABSTRACT: The aim of this study was to analyze the gene expression profile for three main cell lines (supporting, interstitial/stromal, and germ cells) isolated from developing gonads during the process of the testis cord and ovarian cyst formation (between TS13 and TS34).
Project description:The aim of this study was to analyze the gene expression profile for three main cell lines (supporting, interstitial/stromal, and germ cells) isolated from developing gonads at the critical period of sexual differentiation (between 11.5, and 13.5 dpc). Three cell lines (supporting, interstitial/stromal, and germ cells) were isolated from murine fetal XX and XY gonads at three time points (11.5, 12.5, and 13.5 dpc). Transgenic mouse strains with the expression of cell type specific fluorescent markers were used to isolate the cell lines. Cells were sorted using FACS method and then the RNA was extracted.
Project description:The aim of this study was to analyze the gene expression profile for three main cell lines (supporting, interstitial/stromal, and germ cells) isolated from developing gonads at the critical period of sexual differentiation (between 11.5, and 13.5 dpc).
Project description:The goal of this study is to determine the complete gene expression profile for each cell type of the developing gonad during the critical window in which it adopts the testis or ovarian fate. Transgenic mice with cell type specific fluorescent markers were used to isolate germ cells, supporting cells, interstitial cells (including steroidogenic precursors), and endothelial cells in the developing testis and ovary. The gonads were dissociated in trypsin, and the fluorescent cells were isolated by FACS. The RNA was collected from the isolated cells and their gene expression profiles were determined by microarray analysis.
Project description:One of the earliest morphological changes during testicular differentiation is the establishment of an XY specific vasculature. The testis vascular system is derived from mesonephric endothelial cells that migrate into the gonad. In the XX gonad, mesonephric cell migration and testis vascular development is inhibited by WNT4 signaling. In Wnt4 mutant XX gonads, endothelial cells migrate from the mesonephros and form a male-like coelomic vessel. Consequently, the XX Wnt4 mutant mice presented an opportunity to focus a gene expression screen on the processes of mesonephric cell migration and testicular vascular development. We compared differences in gene expression between XY Wnt4+/+ and XX Wnt4+/+ gonads and between XX Wnt4-/- and XX Wnt4+/+ gonads to identify genes similarly upregulated in wildtype XY gonads and XX mutant gonads that might play important roles in testis-specific vascular development. Keywords: genetic sex and phenotypic comparisons
Project description:The goal of this study is to determine the complete gene expression profile for each cell type of the developing gonad during the critical window in which it adopts the testis or ovarian fate.
Project description:The project aimed at identifying the cofactors regulating Polycomb complex PRC2 enzymatic activity in gonads. We used a knock-in mouse model where the enzymatic subunit of PRC2 (either EZH2 or EZH1) is tagged (Flag-tag) to purify PRC2 from mouse adult testis. This experiment revealed the existence of a new cofactor for PRC2 that we called GPIF (AU022751).
Project description:Purpose: Genome-wide DNA-binding analysis for Stat92E in Drosophila testis cyst cells by DNA adenine methyltransferase identification(DamID).Methods: DNA adenine methyltransferase identification (DamID) on Stat92E driven by c587Gal4ts;hopTum-l
Project description:Testicular germ cell tumours (TGCTs) of young adult men, seminoma (SEM) and nonseminoma (NSEM), develop from a precursor cell, carcinoma in situ (CIS), which resembles foetal gonocytes and retains embryonic pluripotency. We used microarrays to analyse microRNA (miRNA) expression in 12 human testis samples with CIS cells and compared it to the miRNA expression profiles of normal adult testis, testis with Sertoli-cell-only (SCO) that lack germ cells, testis tumours (SEM and embryonal carcinoma, EC; an undifferentiated component of NSEM) and foetal male and female gonads. Principal component analysis revealed distinct miRNA expression profiles characteristic for each of the different tissue types. We identified several miRNAs that were unique to testis with CIS cells, foetal gonads, and testis tumours. These included miRNAs from the hsa-miR-371~373 and -302~367 clusters that have previously been reported in germ cell tumours and three miRNAs (hsa-miR-96, -141 and -200c) that were also expressed in human epididymis. We found several miRNAs that were upregulated in testis tumours: hsa-miR-9, -105 and the hsa-miR-182~183~96 cluster were highly expressed in SEM, while the hsa-miR-515~526 cluster was high in EC. We conclude that miRNA expression profile changes during testis development and that the miRNA profile of adult testis with CIS cells shares characteristic similarities with the expression in foetal gonocytes.
Project description:NSAIDs and ACE that affect prostaglandin synthesis are widely used by pregnant women. Epidemiological studies have hypothesized a potential relation of testis dysgenesis syndromes such as cryptorchidism and hypospadias to exposure to these molecules during both the first and the second trimesters of gestation. To decipher whether the embryonic gonads themselves are targets for these molecules, we analysed the impact of precocious in utero exposure to NSAIDs and ACE alone or in combination on the early development of the testis during sex determination, using therapeutic doses similar to those administrated in human medications. We found that in utero exposure to ACE, aspirin or ibuprofen affects the germ cell proliferation in embryonic testis. The whole transcriptome of 13.5 dpc (days post coïtum) treated testis suggests different mechanisms of action of these drugs and a functional interaction between both molecules used in combination, in accelerating the germ cell differentiation. We identified that ACE and ibuprofen exposure through the up-regulation of Dnmt3L expression induces advanced epigenetic reprograming of the germline and enhanced glycogen storage within the testis cords through the activation of extracellular matrix genes expression. In addition, we identified for the first time the prostaglandin production pattern in the embryonic gonad and showed that PGD2, PGE2 and PGI2 were the targets of ACE and NSAIDs drugs. These features might affect the formation and maturation of postnatal testis and secondary reproductive organs leading to male infertility in adult age.
Project description:From fish to human, FOXL2 is considered one of the most conserved markers of ovarian granulosa cell identity. To determine if the sole expression of FOXL2 can determine ovarian differentiation, we created a mouse model that allows the conditional expression of FOXL2. Rosa26-CAG-LSL-Foxl2 mice were crossed to Sf1-Cre mice to induce the expression of FOXL2 in the SF1+ somatic cells of the fetal gonads.When FOXL2 was induced in the somatic cells of the undifferentiated testis, the Sertoli cells and consequently the other cell lineages composing the fetal gonads were feminized, resulting in a partial testis-to-ovary sex reversal We created a mouse genetic model that conditionaly express FOXL2 in the somatic cells of the fetal gonads. All embryos used in this study resulted from the crossing between Rosa26-CAG-LSL-Foxl2+/f and Sf1-cre+/Tg mice. XX and XY fetal gonads were collected at embryonic day E14.5. This microarray analysis led to the identification of the genes misregulated upon ectopic induction of FOXL2 in the fetal testis, and showed that FOXL2 expression resulted in feminization of both somatic and germ cells of the fetal gonad.