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NANOS3 function in human germ cell development.

ABSTRACT: Human infertility is common and frequently linked to poor germ cell development. Yet, human germ cell development is poorly understood, at least in part due to the inaccessibility of germ cells to study especially during fetal development. Here, we explored the function of a highly conserved family of genes, the NANOS genes, in the differentiation of human germ cells from human embryonic stem cells. We observed that NANOS-1, -2 and -3 mRNAs and proteins were expressed in human gonads. We also noted that NANOS3 was expressed in germ cells throughout spermatogenesis and oogenesis and thus, focused further efforts on this family member. NANOS3 expression was highest in human germ cell nuclei where the protein co-localized with chromosomal DNA during mitosis/meiosis. Reduced expression of NANOS3 (via morpholinos or short hairpin RNA) resulted in a reduction in germ cell numbers and decreased expression of germ cell-intrinsic genes required for the maintenance of pluripotency and meiotic initiation and progression. These data provide the first direct experimental evidence that NANOS3 functions in human germ cell development; indeed, NANOS3 is now one of just two genes that has been directly shown to function in germ cell development across diverse species from flies, worms, frogs and mice to humans [the other is BOULE, a member of the Deleted in Azoospermia (DAZ) gene family]. Findings may contribute to our understanding of the basic biology of human germ cell development and may provide clinical insights regarding infertility.

SUBMITTER: Julaton VT

PROVIDER: S-EPMC3090199 | biostudies-literature | 2011 Jun

REPOSITORIES: biostudies-literature

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NANOS3 function in human germ cell development.

Julaton Vanessa T Angeles VT Reijo Pera Renee A RA

Human molecular genetics 20110319 11

Human infertility is common and frequently linked to poor germ cell development. Yet, human germ cell development is poorly understood, at least in part due to the inaccessibility of germ cells to study especially during fetal development. Here, we explored the function of a highly conserved family of genes, the NANOS genes, in the differentiation of human germ cells from human embryonic stem cells. We observed that NANOS-1, -2 and -3 mRNAs and proteins were expressed in human gonads. We also no ...[more]

PMID: 21421998

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Project description:BackgroundUsing an organotypic culture system termed human Fetal Testis Assay (hFeTA) we previously showed that 0.01 μM BPA decreases basal, but not LH-stimulated, testosterone secreted by the first trimester human fetal testis. The present study was conducted to determine the potential for a long-term antiandrogenic effect of BPA using a xenograft model, and also to study the effect of BPA on germ cell development using both the hFETA and xenograft models.MethodsUsing the hFeTA system, first trimester testes were cultured for 3 days with 0.01 to 10 μM BPA. For xenografts, adult castrate male nude mice were injected with hCG and grafted with first trimester testes. Host mice received 10 μM BPA (~ 500 μg/kg/day) in their drinking water for 5 weeks. Plasma levels of total and unconjugated BPA were 0.10 μM and 0.038 μM respectively. Mice grafted with second trimester testes received 0.5 and 50 μg/kg/day BPA by oral gavage for 5 weeks.ResultsWith first trimester human testes, using the hFeTA model, 10 μM BPA increased germ cell apoptosis. In xenografts, germ cell density was also reduced by BPA exposure. Importantly, BPA exposure significantly decreased the percentage of germ cells expressing the pluripotency marker AP-2γ, whilst the percentage of those expressing the pre-spermatogonial marker MAGE-A4 significantly increased. BPA exposure did not affect hCG-stimulated androgen production in first and second trimester xenografts as evaluated by both plasma testosterone level and seminal vesicle weight in host mice.ConclusionsExposure to BPA at environmentally relevant concentrations impairs germ cell development in first trimester human fetal testis, whilst gonadotrophin-stimulated testosterone production was unaffected in both first and second trimester testis. Studies using first trimester human fetal testis demonstrate the complementarity of the FeTA and xenograft models for determining the respective short-term and long term effects of environmental exposures.

Project description:BackgroundRadiotherapy, chemotherapy, and surgery have made crucial strides in glioblastoma treatment, yet they often fail; thus, new treatment and new detection methods are needed. Aberrant expression of Nanos3 has been functionally associated with various cancers. Here, we sought to identify the clinical significance and potential mechanisms of Nanos3 in human glioblastoma.MethodsNanos3 expression was studied in nude mouse glioblastoma tissues and glioblastoma cell lines by immunohistochemistry, Western blot, and RT-PCR. Clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 gene editing assay was performed to generate the Nanos3 knockdown glioblastoma cell lines. The effects of Nanos3 on glioblastoma cells proliferation, migration, invasion, chemoresistance, germ cell characteristics, and tumor formation were analyzed by CCK8, transwell, cell survival experiments and alkaline phosphatase staining in vitro and in nude mouse models in vivo. Correlation between the expression of stemness proteins and the expression of Nanos3 was evaluated by Western blot.ResultsWe found that Nanos3 was strongly expressed in both glioblastoma cell lines and tissues. Western blot and sequencing assays showed that the Nanos3 knockdown glioblastoma cell lines were established successfully, and we discovered that Nanos3 deletion reduced the proliferation, migration, and invasion of glioblastoma cells in vitro (P < 0.05). Nanos3 knockdown enhanced the sensitivity of glioblastoma cells to doxorubicin (DOX) and temozolomide (TMZ) (P < 0.05), and Nanos3+/- glioblastoma cell lines did not show the characteristics of the germline cells. In addition, Nanos3 deletion inhibited subcutaneous xenograft tumor growth in vivo (P < 0.001). Moreover, the oncogenesis germline protein levels of CD133, Oct4, Ki67, and Dazl decreased significantly in glioblastoma cells following Nanos3 knockdown.ConclusionsBoth in vitro and in vivo assays suggest that Nanos3, which is a cancer-germline gene, initiates the tumorigenesis of glioblastoma via acquiring the oncogenesis germline traits. These data demonstrate that ectopic germline traits are necessary for glioblastoma growth.

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NANOS3 function in human germ cell development.

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NANOS3 function in human germ cell development.

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