Establishment of Etv5 gene knockout mice as a recipient model for spermatogonial stem cell transplantation.
Establishment of Etv5 gene knockout mice as a recipient model for spermatogonial stem cell transplantation.
复制标题
Etv5基因敲除小鼠精原干细胞移植受体模型的建立。
DOI：
10.1242/bio.056804
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发表时间：
2021-01-06
期刊：
影响因子：
2.4
通讯作者：
Wu Z
中科院分区：
文献类型：
--
作者：
Zhang X;Zhao X;Li G;Zhang M;Xing P;Li Z;Chen B;Yang H;Wu Z
精原干细胞（SSC）移植是畜牧生产中实现优良动物保护和生产的一种替代生殖方法。建立无内源性生殖细胞的受体动物是有效的SSC移植的重要条件。然而，缺乏ssc的天然突变体很难获得，内源性生殖细胞的药物消融也很难用于实际应用。在本研究中，我们使用小鼠模型研究先天性生殖细胞消融受体的制备。我们使用CRISPR/Cas9系统在小鼠中敲除（KO） Ets-variant基因5 （Etv5）。Etv5−/−小鼠的睾丸重量明显低于野生型（WT）小鼠。Etv5−/−小鼠精小管生殖细胞层随着年龄的增长逐渐消退。在12周龄时，Etv5−/−小鼠的小管中几乎缺乏所有具有支持细胞表型的生精细胞，并且附睾中完全没有精子。随后，我们将具有增强绿色荧光蛋白（EGFP）的同种异体ssc移植到3-（未成熟）或7周龄（成熟）的Etv5−/−小鼠中。移植后2个月，所有未成熟睾丸均能部分恢复精管生殖细胞层和精子形成，而在成熟成年睾丸中未见恢复。经PCR检测，受体睾丸组织和附睾精子中存在外源基因Etv5和EGFP，表明精子来源于移植供体细胞。我们的研究表明，尽管Etv5−/−小鼠能够适应和支持外源生殖细胞移植，但这一过程的效率很低，无法支持移植的SSCs完整的精子发生。然而，使用Etv5−/−小鼠作为SSC移植的受体模型是可行的，但仍需进一步研究以建立优化的移植过程。摘要：通过对特定基因进行基因修饰，实现生殖系消融术小鼠精原干细胞移植。
Spermatogonial stem cell (SSC) transplantation is an alternative reproductive method to achieve conservation and production of elite animals in livestock production. Creating a recipient animal without endogenous germ cells is important for effective SSC transplantation. However, natural mutants with depletion of SSCs are difficult to obtain, and drug ablation of endogenous germ cells is arduous to perform for practical use. In this study, we used mouse models to study the preparation of recipients with congenital germ cell ablation. We knocked out (KO) Ets-variant gene 5 (Etv5) in mice using the CRISPR/Cas9 system. The testicular weight of Etv5−/− mice was significantly lower than that of wild-type (WT) mice. The germ cell layer of the seminiferous tubules gradually receded with age in Etv5−/− mice. At 12 weeks of age, the tubules of Etv5−/− mice lacked almost all spermatogenic cells with a Sertoli cell-only phenotype, and sperm were completely absent in the epididymis. We subsequently transplanted allogeneic SSCs with enhanced green fluorescent protein (EGFP) into 3- (immature) or 7-week-old (mature) Etv5−/− mice. Partially restoration of germ cell layers in the seminiferous tubules and spermatogenesis was observed in all immature testes but not in mature adult testes at 2 months post-transplantation. The presence of heterologous genes Etv5 and EGFP in recipient testicular tissue and epididymal sperm by PCR indicated that sperm originated from the transplanted donor cells. Our study demonstrates that, although Etv5−/− mice could accommodate and support foreign germ cell transplantation, this process occurs in a quite low efficiency to support a full spermatogenesis of transplanted SSCs. However, using Etv5−/− mice as a recipient model for SSC transplantation is feasible, and still needs further investigation to establish an optimized transplantation process. Summary: Spermatogonial stem cell transplantation in mice with germline ablation by genetic modification of a specific gene.