Key Laboratory of Model Animals and Stem Cell Biology in Hunan Province, Hunan Normal University School of Medicine, Changsha 410013, China.
Research Center of Reproduction and Translational Medicine of Hunan Province, Changsha 410008, China.
Cells. 2024 Oct 2;13(19):1642. doi: 10.3390/cells13191642.
Organoids play pivotal roles in uncovering the molecular mechanisms underlying organogenesis, intercellular communication, and high-throughput drug screening. Testicular organoids are essential for exploring the genetic and epigenetic regulation of spermatogenesis in vivo and the treatment of male infertility. However, the formation of testicular organoids with full spermatogenesis has not yet been achieved. In this study, neonatal mouse testicular cells were isolated by two-step enzymatic digestion, and they were combined with Matrigel and transplanted subcutaneously into nude mice. Histological examination (H&E) staining and immunohistochemistry revealed that cell grafts assembled to form seminiferous tubules that contained spermatogonial stem cells (SSCs) and Sertoli cells, as illustrated by the co-expression of PLZF (a hallmark for SSCs) and SOX9 (a marker for Sertoli cells) as well as the co-expression of UCHL1 (a hallmark for SSCs) and SOX9, after 8 weeks of transplantation. At 10 weeks of transplantation, SSCs could proliferate and differentiate into spermatocytes as evidenced by the expression of PCNA, Ki67, c-Kit, SYCP3, γ-HA2X, and MLH1. Notably, testicular organoids were seen, and spermatids were observed within the lumen of testicular organoids after 16 weeks of transplantation, as shown by the presence of TNP1 and ACROSIN (hallmarks for spermatids). Collectively, these results implicate that we successfully established testicular organoids with spermatogenesis in vivo. This study thus provides an excellent platform for unveiling the mechanisms underlying mammalian spermatogenesis, and it might offer valuable male gametes for treating male infertility.
类器官在揭示器官发生、细胞间通讯和高通量药物筛选的分子机制方面发挥着关键作用。睾丸类器官对于研究体内精子发生的遗传和表观遗传调控以及男性不育症的治疗至关重要。然而，具有完整精子发生的睾丸类器官的形成尚未实现。在这项研究中，通过两步酶消化法分离新生小鼠睾丸细胞，将其与 Matrigel 混合，并皮下移植到裸鼠体内。组织学检查（H&E 染色）和免疫组织化学染色显示，细胞移植物组装形成包含精原干细胞（SSCs）和支持细胞的生精小管，如 PLZF（SSCs 的标志）和 SOX9（支持细胞的标志物）的共表达以及 UCHL1（SSCs 的标志）和 SOX9 的共表达所示，移植后 8 周。移植 10 周后，SSCs 可以增殖并分化为精母细胞，这可以通过 PCNA、Ki67、c-Kit、SYCP3、γ-HA2X 和 MLH1 的表达来证明。值得注意的是，在移植 16 周后，可以看到睾丸类器官，并且在睾丸类器官的管腔中观察到精子细胞，这可以通过 TNP1 和 ACROSIN（精子细胞的标志）的存在来证明。总之，这些结果表明我们成功地在体内建立了具有精子发生的睾丸类器官。因此，这项研究为揭示哺乳动物精子发生的机制提供了一个极好的平台，并可能为治疗男性不育症提供有价值的男性配子。
