Spermatogonial stem cells (SSCs) self-renew and produce a large number of differentiated germ cells to maintain normal spermatogenesis. However, the growth factors crucial for SSC self-renewal and the mechanism underlying this process remain unclear. In the present study, a serum-free culture media was used to evaluate the effect of several growth factors on the expression of some SSC markers and self-renewal related genes. The putative SSCs were cultured on buffalo Sertoli cell feeder layer in KO-DMEM +10% KOSR. The colony formation was observed between 7 and 10 days. The putative SSC colonies also expressed markers specific for undifferentiated type A spermatogonia and pluripotency markers. After 15 days, relative mRNA expression study revealed that 20 ng/mL concentration of Glial cell line-derived neurotrophic factor (GDNF) upregulated the expression of PLZF, TAF4B, and THY1. Furthermore, supplementation of a combination of 20 ng/mL GDNF, 10 ng/mL basic fibroblast growth factor (bFGF), 1000 IU/mL leukemia inhibitory factor (LIF), and 1 ng/mL colony stimulating factor 1 (CSF1) upregulated the expression of PLZF, TAF4B, BCL6B, and ID4 genes. These results demonstrated that our defined culture media in combination with GDNF, bFGF, LIF, and CSF1 well supported SSC self-renewal.

中文翻译：

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优化无血清培养条件，促进假定的水牛（Bubalus bubalis）精原干细胞的繁殖。

精原干细胞（SSCs）自我更新并产生大量分化的生殖细胞，以维持正常的精子发生。但是，对于SSC自我更新至关重要的生长因子以及该过程的潜在机制仍不清楚。在本研究中，使用无血清培养基评估几种生长因子对一些SSC标记和自我更新相关基因表达的影响。假定的SSC在KO-DMEM + 10％KOSR中的水牛Sertoli细胞饲养层上培养。在7至10天之间观察到菌落形成。假定的SSC菌落还表达了未分化的A型精原细胞和多能性标记物的特异性标记物。15天后，相对mRNA表达研究表明，浓度为20 ng / mL的神经胶质细胞源性神经营养因子（GDNF）可以上调PLZF，TAF4B和THY1的表达。此外，补充20 ng / mL GDNF，10 ng / mL碱性成纤维细胞生长因子（bFGF），1000 IU / mL白血病抑制因子（LIF）和1 ng / mL菌落刺激因子1（CSF1）的组合上调了PLF，TAF4B，BCL6B和ID4基因的表达。这些结果表明，我们定义的培养基与GDNF，bFGF，LIF和CSF1的结合很好地支持了SSC的自我更新。BCL6B和ID4基因。这些结果表明，我们定义的培养基与GDNF，bFGF，LIF和CSF1的结合很好地支持了SSC的自我更新。BCL6B和ID4基因。这些结果表明，我们定义的培养基与GDNF，bFGF，LIF和CSF1的结合很好地支持了SSC的自我更新。