Characterization of male germline stem cells in a rhesus model of infertility

恒河猴不育模型中雄性生殖干细胞的表征

• 批准号: 8090012
• 负责人: Kyle Edwin Orwig
• 金额: $ 6万
• 依托单位: MAGEE-WOMEN'S RES INST AND FOUNDATION
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-08-01 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8090012
• 关键词: Adolescent; Adult; Animal Model; Animals; Autologous; Biological; Biological Assay; Biology; Busulfan; Cell Transplantation; Cell physiology; Cells; Cellular biology; Characteristics; Clinic; Collaborations; Colony-Forming Units Assay; Consensus; Cryopreservation; Data; Development; Endocrine; Equilibrium; Evaluation; Event; Fertility; Foundations; Future; Generations; Germ Cells; Goat; Growth Factor; Health Sciences; Homologous Transplantation; Human; Human Characteristics; Immune; Infertility; Institutes; Investigation; Kinetics; Knowledge; Macaca mulatta; Male Infertility; Mammals; Modeling; Molecular; Molecular Biology; Morphology; Mus; Natural regeneration; Nuclear; Nude Mice; Phenotype; Physiology; Plants; Population; Primates; Principal Investigator; Process; Proliferating; Rattus; Reagent; Regenerative Medicine; Reproduction; Research Ethics Committees; Rodent; Seminiferous tubule structure; Site; Spermatogenesis; Spermatogonia; Stem Cell Research; Stem cell transplant; Stem cells; System; Techniques; Technology; Testing; Testis; Therapeutic; Tissue Banking; Tissue Banks; Tissues; Translating; Transplantation; Treatment Protocols; Undifferentiated; Universities; Xenograft procedure; ZNF145 gene; abstracting; cell type; experience; fluorescence activated cell sorter device; in vivo; male; meetings; nonhuman primate; novel strategies; progenitor; regenerative; research study; self-renewal; sperm cell; stem; stem cell niche; stem cell population; therapeutic evaluation; tool

Abstract Spermatogonial stem cells (SSCs) are at the foundation of mammalian spermatogenesis. In rodents, the SSC pool is comprised of type Asingle (As) spermatogonia that balance self-renewing and differentiating divisions to exactly meet the biological demand of the spermatogenic system. Spermatogenesis is highly conserved in mammals and it is tempting to extrapolate rodent SSC data to higher species, including humans. However, in humans and nonhuman primates (NHPs), spermatogenesis arises from Adark (Ad) and Apale (Ap) spermatogonia, and the identity of the stem cell is subject to debate. The fundamental differences between species highlight the need to investigate the biology of NHP SSCs, which have greater relevance to human physiology. To facilitate the investigation of spermatogonial stem cells in primate testes, an in vivo functional assay is needed. In preliminary studies, we developed primate- to-nude mouse xenotransplantation as a biological assay to evaluate SSC function. In addition, we identified a busulfan treatment regimen that causes long- term infertility in rhesus macaques to facilitate autologous and homologous testis cell transplantation. This model will be used to test the full regenerative potential of rhesus SSCs and stem cell/niche interactions. We will exploit the rhesus-to-nude mouse xenotransplantation assay in the current application to characterize SSC activity in juvenile and adult (Aims 1) and identify phenotypic characteristics of rhesus SSCs using a fluorescence-activated cell sorter (Aim 2). Progress in Aim 2 will enable the isolation, enrichment and molecular characterization of rhesus SSCs. Because of evolutionary distance between rhesus and mouse, rhesus SSCs will engraft and persist in mouse seminiferous tubules, but will not produce complete spermatogenesis. Therefore, autologous and homologous rhesus-to-rhesus transplantations will be performed in Aim 3 to test the full regenerative potential of putative SSC-enriched rhesus testis cell populations identified in Aim 2. This aim will also provide information about the quality of the stem cell niche in busulfan-treated testes, the immunological tolerance of autologous vs. homologous donor cells, and the effect of cryopreservation on SSC regenerative potential. Finally, our progress characterizing NHP SSCs (from Aims 1-3) will guide our initial studies in Aim 4 to characterize human spermatogonial stem cells. These studies will provide fundamental information about stem cells and the stem cell niche in nonhuman primate testes and lay the groundwork to translate these discoveries for future investigations of the biology and therapeutic potential of human SSCs. Project Narrative The proposed studies will provide the experimental framework to study the biology and molecular characteristics of rhesus spermatogonial stem cells (SSCs) and translate the results to facilitate the investigation of human SSCs. The studies will also establish a nonhuman primate model of infertility and examine the regenerative potential of SSCs to establish donor spermatogenesis and/or restore fertility. Responsible development of these ideas in the rhesus model may have implications for treating some cases of male infertility.

抽象的 精子干细胞（SSC）是哺乳动物精子发生的基础。在啮齿动物中 SSC池由asingle类型（AS）组成，平衡自我更新和 区分分裂以准确地满足精子系统的生物学需求。 精子发生在哺乳动物中是高度保守的，很容易推断啮齿动物SSC数据 到包括人类在内的较高物种。但是，在人类和非人类灵长类动物（NHP）中， 精子发生来自Adark（AD）和Apale（AP）精子，以及干细胞的身份 可能会辩论。物种之间的基本差异突出了调查的必要性 NHP SSC的生物学，与人类生理学有更大相关。促进 在灵长类动物睾丸中研究精子干细胞，需要进行体内功能测定。在 初步研究，我们开发了灵长类动物 - 裸露的小鼠异种移植作为生物学测定 评估SSC功能。此外，我们确定了一种升压方案，该治疗方案导致长期 恒河猕猴中的术语不育症，以促进自体和同源睾丸细胞移植。 该模型将用于测试恒河猴和干细胞/小众的全部再生潜力 互动。我们将利用电流中的恒河与裸露的鼠标异种移植测定法 在少年和成人中表征SSC活动的应用（目标1）并识别表型 使用荧光激活的细胞分散者（AIM 2）的恒河类SSC的特征。目标2 将使恒河类SSC的分离，富集和分子表征。由于 恒河主和小鼠之间的进化距离，恒河所SSC会植入并持续在小鼠中 精神小管，但不会产生完全的精子发生。因此，自体和 在AIM 3中将进行同源的恒河 - 逆转移植，以测试完整的再生 在AIM 2中确定的假定SSC富含恒晶的细胞种群的潜力。此目标也将 提供有关干细胞生态位质量的信息 自体和同源供体细胞的免疫耐受性以及冷冻保存的影响 关于SSC再生潜力。最后，我们表征NHP SSC的进度（来自AIMS 1-3）将 指导我们在AIM 4中的最初研究以表征人类精子干细胞。 这些研究将提供有关干细胞和干细胞生态位的基本信息 非人类灵长类动物睾丸并为将这些发现转化为未来的基础 人类SSC的生物学和治疗潜力的研究。项目叙述 拟议的研究将为研究生物学和 恒河生精子干细胞（SSC）的分子特征，并将结果转化为 促进对人类SSC的研究。研究还将建立一个非人类的灵长类动物模型 不孕症并检查SSC建立供体精子发生的再生潜力 和/或恢复生育能力。在恒河校模型中负责发展这些想法 治疗某些男性不育症的含义。