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Murine Spermatogonial Stem Cells: Rapamycin- vs Vehicle-exposed in vivo (2-wk treatment)

ABSTRACT: Male FVB strain mice aged 12-days-old through 26-days-old were administered daily intraperitoneal injections of rapamycin (4mg/kg body weight) or control vehicle (5% Tween-80, 5% PEG-400), beginning at postnatal day (P)12. Mice were euthanized at P26 and their testes were isolated for germ cell enrichment. Single cell suspensions of germ cells were prepared from isolated testes and subjected to magnetic-activated cell sorting (MACS). This procedure enriches the undifferentiated spermatogonia fraction, which represents the spermatogonial stem cell (SSC) population. Total RNA from cells double-positive for the SSC surface markers thymus cell antigen 1, theta (THY1) and glial cell line-derived neurotrophic factor family receptor alpha 1 (GFRA1) was isolated for gene expression microarray analysis. Magnetic-activated cell sorting (MACS) was used to enrich undifferentiated spermatogonia from a pool of primary testicular cells isolated from 5 littermate rapamycin (RM)-treated male mice and from 5 littermate control vehicle (VEH)-treated male mice. With MACS-enriched cells from pool generating RNA, no technical replicates performed.

ORGANISM(S): Mus musculus

SUBMITTER: Christopher Payne

PROVIDER: E-GEOD-37062 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Murine Spermatogonial Stem Cells: Rapamycin- vs Vehicle-exposed in vivo (2-wk treatment)

Project description:Male FVB strain mice aged 12-days-old through 26-days-old were administered daily intraperitoneal injections of rapamycin (4mg/kg body weight) or control vehicle (5% Tween-80, 5% PEG-400), beginning at postnatal day (P)12. Mice were euthanized at P26 and their testes were isolated for germ cell enrichment. Single cell suspensions of germ cells were prepared from isolated testes and subjected to magnetic-activated cell sorting (MACS). This procedure enriches the undifferentiated spermatogonia fraction, which represents the spermatogonial stem cell (SSC) population. Total RNA from cells double-positive for the SSC surface markers thymus cell antigen 1, theta (THY1) and glial cell line-derived neurotrophic factor family receptor alpha 1 (GFRA1) was isolated for gene expression microarray analysis.

2012-05-01 | GSE37062 | GEO

RNA-seq reveals an abnormal transcriptome as a result of DDX20 loss in spermatogonia.

Project description:To investigate the role of DDX20 in spermatogenesis, we generated Ddx20 flox/flox Ddx4-Cre mice to make a germ cell-specific Ddx20 knockout, and isolated spermatogonia from four-day-old mouse testes by THY1 magnetic beads.We then performed gene expression profiling analysis using data obtained from RNA-seq of THY1 + spermatogonia.

2023-10-01 | GSE217685 | GEO

Proteomic analysis of the effects of DDX20 deletion in spermatogonia

Project description:To investigate the role of DDX20 in spermatogenesis, we generated Ddx20 flox/flox Ddx4-Cre mice to make a germ cell-specific Ddx20 knockout, and isolated spermatogonia from four-day-old mouse testes by THY1 magnetic beads. We then performed proteomic analysis using protein lysates obtained from THY1 + spermatogonia.

2024-04-24 | PXD038170 | iProX

A regulatory role for CHD4 in maintenance of the spermatogonial stem cell pool

Project description:Maintenance and self-renewal of the spermatogonial stem cell (SSC) population is the cornerstone of male fertility. In this manuscript we have identified a key role for the nucleosome remodelling protein Chromodomain Helicase DNA binding protein 4 (CHD4) in regulating SSC function. Gene expression analyses revealed that CHD4 expression is largely restricted to spermatogonia in the mouse testis, and is particularly enriched in SSCs. Using spermatogonial transplantation techniques and RNAi mediated knockdown it was established that loss of Chd4 expression significantly impairs SSC regenerative capacity, resulting in a ~50% reduction in colonisation of recipient testes. A single cell RNA-seq comparison depicted reduced expression of ‘self-renewal’ genes such as Gfra1 and Pten following Chd4 knockdown, along with increased expression of signature progenitor genes, Neurog3 and Dazl. Co-immunoprecipitation analyses demonstrated that CHD4 regulates gene expression in spermatogonia not only though its traditional association with the remodelling complex NuRD, but also via interaction with the GDNF-responsive transcription factor SALL4. Cumulatively, the results of this study depict a previously unappreciated fundamental role for CHD4 in controlling fate decisions in the spermatogonial pool.

2021-06-17 | PXD022066 | Pride

Expression profiling of rainbow trout testicular development and spermatogenesis

Project description:Trout spermatogenesis proceeds seasonally in a way that all morphological and cellular events tend to be synchronized. We thus used gonads at key stages of the male reproductive cycle together with isolated germ cell populations to study the changes in gene expression underlying testis development. Statistical analysis followed by clustering of expression profiles allowed the discrimination of sequential events of gene activation or repression during testis development and/or throughout the germline. 38 samples covering 6 developmental stages (as defined in Gomez et al. 1998 Biol. Reprod. 58, 483-491) and 3 isolated germ cell populations were analysed. This included: - Testes at early stages containing slowly-dividing type A spermatogonia (Stage_I, n=5) or growing numbers of actively-dividing type B spermatogonia (Stages_IIa, n= 4; Stage_IIb, n= 4) - Maturing testes containing in addition large numbers of meiotic spermatocytes (Stage_IIIb, n=3) and post-meiotic spermatids (Stage_V, n=4) - Spawning testes containing essentially mature spermatozoa (Stage_VIII, n=3) - Fractions of isolated germ cells enriched in spermatogonia (Spermatogonia, N=6), spermatocytes (Spermatocyte, N=6) and spermatids (Spermatid, N=3).

2010-05-06 | E-GEOD-16029 | biostudies-arrayexpress

Murine Testes: Germ Cell-Specific Sin3a Knockout vs. Hemizygous vs. Wild Type at Postnatal Day 0

Project description:Chromatin modifier Swi-independent 3a (SIN3A), together with associated histone deacetylases, influences gene expression during development and differentiation through multiple transcription factors in a cell-specific manner. Sin3a is essential for the maintenance of inner cell mass cells of mouse blastocysts, embryonic fibroblasts, and myoblasts, but is not required for the survival of trophectoderm or Sertoli cells. To better understand how this transcriptional regulator modulates cells at different developmental stages within a single lineage, we used conditional gene targeting in mice to ablate Sin3a from perinatal quiescent male gonocytes and from postnatal differentiating spermatogonia. Gene expression in the whole testes of age-matched Ddx4(Vasa/Mvh)-cre mediated conditional Sin3a gene-targeted(VSKO) mice, germ cell-specific Sin3a hemizygous (HEMI) mice, and wild type (WT) mice was measured at postnatal day 0. Two biological replicate animals were used for each condition.

2012-10-01 | E-GEOD-38096 | biostudies-arrayexpress

The miRNA profiling of goat CD49f-positive and negative testicular cells

Project description:Purpose: miRNAs, a member of the small RNA, play critical roles in the mammalian spermatogenesis. Spermatogonia was the foundation of spermatogenesis and valuable for the study of spermatogenesis. However, it is still not clear that the expression profiling of the miRNAs in spermatogonia of dairy goat. Methods: The CD49f was one of the surface markers for spermatogonia enrichment by MACS. Therefore, we used CD49f microbeads antibody to purify CD49f-positive and negative cells of dairy goat testicular cells by MACS (Magnetic Activated Cell Sorting), and then in-depth analyzed the miRNA expression in these cells using Illumina sequencing technology. Results: The results of miRNAs expression profiling in purified CD49f-positive and negative testicular cells showed that 933 were miRNAs upregulated in CD49f-positive cells and 916 were miRNAs upregulated in CD49f-negative cells with a 2-fold increase, respectively; some spermatogonial stem cells(SSCs) specific miRNAs and marker genes in testis had a higher level expression in CD49f-positive testicular cells, such as miR-221, miR-23a, miR-29b, miR-24, miR-29a, miR-199b, miR-199a, miR-27a, miR-21. Conclusions: our comparative miRNAome data provided some useful miRNAs profiling data of dairy goat spermatogonia cells and suggested CD49f could be used to enrich dairy goat spermatogonia-like cells, including SSCs.

2014-04-25 | GSE57041 | GEO

The miRNA profiling of goat CD49f-positive and negative testicular cells

2014-04-25 | E-GEOD-57041 | biostudies-arrayexpress

In Vitro Derivation and Propagation of Spermatogonial Stem Cell Activity from Mouse Pluripotent Stem Cells

Project description:The in vitro derivation and propagation of spermatogonial stem cells (SSCs) from pluripotent stem cells (PSCs) is a key goal in reproductive science. We show here that when aggregated with embryonic testicular somatic cells (reconstituted testes), primordial germ cell-like cells (PGCLCs) induced from mouse embryonic stem cells differentiate into spermatogonia-like cells in vitro and are expandable as cells that resemble germline stem cells (GSCs), a primary cell line with SSC activity. Remarkably, GSC-like cells (GSCLCs), but not PGCLCs, colonize adult testes and, albeit less effectively than GSCs, contribute to spermatogenesis and fertile offspring. Whole-genome analyses reveal that GSCLCs exhibit aberrant methylation at vulnerable regulatory elements, including those critical for spermatogenesis, which may restrain their spermatogenic potential. Our study establishes a strategy for the in vitro derivation of SSC activity from PSCs, which, we propose, relies on faithful epigenomic regulation.

2016-12-06 | GSE76245 | GEO

In Vitro Derivation and Propagation of Spermatogonial Stem Cell Activity from Mouse Pluripotent Stem Cells

2016-12-06 | GSE87341 | GEO

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