Department of Anatomical Sciences, Medical Sciences Faculty, Tarbiat Modares University, Tehran, Iran.
Department of Molecular Genetics, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran.
Urol J. 2020 Nov 23;18(2):214-224. doi: 10.22037/uj.v16i7.6092.
Generating functional gametes for patients with male infertility is of great interest. We investigated different cultural systems for proliferation of SSCs derived from obstructive azoospermic patients.
Testicular cells were obtained from men with obstructive azoospermia. After enzymatic digestion process, cells were assigned to various groups: culture of SSCs in the dish without cover (control group), co-culture of SSCs with infertile Sertoli cells (I), co-culture of SSCs with fertile Sertoli cells (II), culture of SSCs on nanofiber (covered with laminin) (III), culture of testicular cell suspension (IV). Then cells were cultured and colony formation, gene-specific methylation (by MSP), quantitative genes expression of pluripotency (Nanog, C-Myc, Oct-4) and specific germ cell (Integrin α6, Integrin β1, PLZF) genes were evaluated in five different culture systems.
Our findings indicate a significant increase in the number and diameter of colonies in IV group in compare to control group and other groups. Expression of germ specific genes in IV group were significantly increased (P ≤ 0.05) and levels of expression of pluripotency genes were significantly decreased in this group (P ≤ 0.05) compared with other groups. Gene-specific pattern of methylation of examined genes showed no changes in culture systems during the culture era.
A microenvironment capable of controlling the proliferation of cell colonies can be restored by testicular cell suspension.
为男性不育患者生成功能性配子具有重要意义。我们研究了不同的培养体系，以增殖来源于梗阻性无精子症患者的精原干细胞。
从梗阻性无精子症患者中获取睾丸细胞。在酶消化过程后，将细胞分为以下几组：在无盖培养皿中培养精原干细胞（对照组）、将精原干细胞与不育的支持细胞共培养（I 组）、将精原干细胞与可育的支持细胞共培养（II 组）、在纳米纤维上培养精原干细胞（覆盖层粘连蛋白）（III 组）、培养睾丸细胞悬液（IV 组）。然后在五个不同的培养体系中培养细胞并评估集落形成、基因特异性甲基化（MSP）、多能性基因（Nanog、C-Myc、Oct-4）和特定生殖细胞（Integrin α6、Integrin β1、PLZF）的定量基因表达。
与对照组和其他组相比，IV 组的集落数量和直径明显增加。IV 组的生殖特异性基因表达显著增加（P≤0.05），而多能性基因的表达水平在该组显著降低（P≤0.05），与其他组相比。在培养过程中，检查基因的基因特异性甲基化模式在培养体系中没有变化。
睾丸细胞悬液可以恢复控制细胞集落增殖的微环境。
