TRANSPLANTATION OF TESTIS STEM CELLS IN LARGE ANIMALS

睾丸干细胞在大型动物体内的移植睾丸干细胞在成年男性体内的干细胞中是独一无二的，因为它是唯一可以分裂并将基因贡献给后代的细胞。受者睾丸移植后供者来源的精子发生为精原干细胞的功能提供了唯一明确的生物测定系统，是研究精子发生的有力手段。到目前为止，生殖细胞移植主要在啮齿动物身上进行。这项拟议研究的长期目标是在非啮齿动物物种中建立生殖细胞移植，作为研究生殖系干细胞生物学的独特工具，作为干细胞潜力的生物测定，并允许操纵精子发生的不同方面。猪将被用作非啮齿动物的模式物种，因为它们容易获得，维护成本相对较低，而且成熟迅速。由于它们的大小和睾丸解剖结构，在猪身上获得的结果将适用于其他大型动物物种和人类。该项目将涉及：1)选择生殖系干细胞的供体细胞；2)引入遗传标记以跟踪受者睾丸内供体精子发生的进展并为改良雄性生殖系奠定基础；3)对受者睾丸进行修饰以抑制内源性精子发生并为供体来源的精子生成提供允许的环境；以及4)将供体睾丸组织移植到小鼠背部皮肤下，作为研究精子发生的一种新的体内培养系统，并作为移植分离生殖细胞的替代方法。该系统将为修改雄性生殖系作为产生生殖系转基因动物的替代方法奠定基础，将有助于保存宝贵的遗传物质，并可用于临床应用于接受细胞毒治疗的癌症患者的生育能力恢复。该项目的结果有望为深入研究非啮齿类哺乳动物的生殖系干细胞生物学和雄性生育能力提供一种新的途径。

Transplantation of Testis Stem Cells in Large Animals The testis stem cell is unique among the stem cells in an adult male body in that it is the only cell that divides and can contribute genes to subsequent generations. Donor-derived spermatogenesis in a recipient testis after transplantation provides the only unequivocal bioassay system for functional competence of spermatogonial stem cells and is a powerful approach for the study of spermatogenesis. To date, germ cell transplantation has been performed predominantly in rodents. The long-term objective of the proposed research is to establish germ cell transplantation in a non-rodent mammalian species as a unique tool to study the biology of germ line stem cells, serve as a bioassay for stem cell potential, and allow the manipulation of different aspects of spermatogenesis. Pigs will be used as a non-rodent model species because they are readily obtained, relatively inexpensive to maintain and mature quickly. Due to their size and testicular anatomy, the results obtained in pigs will be applicable to other large animal species and man. The project will address 1) the selection of donor cells for germ line stem cells, 2) the introduction of a genetic marker to follow the progression of donor spermatogenesis in the recipient testis and to lay the foundation for modification of the male germ line, 3) the modification of the recipient testis to suppress endogenous spermatogenesis and to provide a permissive environment for donor-derived spermatogenesis, and 4) grafting of donor testis tissue under the back skin of mice as a novel in vivo culture system for the study of spermatogenesis and as an alternate approach to transplantation of isolated germ cells. This system will lay the foundation for modification of the male germ line as an alternate approach to generate germ line transgenic animals, will serve to preserve valuable genetic material and can be clinically applicable to restore fertility in patients undergoing cytotoxic therapy for cancer. The results of this project are expected to provide a novel approach for in depth investigations into germ line stem cell biology and male fertility in non-rodent mammalian species.