上海交通大学学报（医学版）

上海交通大学学报（医学版）

• 论著（基础研究） • 上一篇    下一篇

一组miRNAs在睾丸发育中的表达及miR-125a对精原干细胞发育的调节作用

梅星星1，李小勇1，吴际1,2   

  1. 1.上海交通大学Bio-X研究院遗传发育与精神神经疾病教育部重点实验室, 上海 200240; 2.宁夏医科大学生育力保持教育部重点实验室, 银川 750004
  • 出版日期:2015-05-28 发布日期:2015-06-04
  • 通讯作者: 吴际, 电子信箱: jiwu@sjtu.edu.cn。
  • 作者简介:梅星星（1990—）， 男， 硕士生; 电子信箱: xingxing_0220@126.com。
  • 基金资助:

    国家重大科学研究计划 (2013CB967401)； 国家自然科学基金 (81370675, 81200472)

Expressions of a group of miRNAs during testis development and regulation effect of miR-125a on development of spermatogonial stem cells

MEI Xing-xing1, LI Xiao-yong1, WU Ji1,2   

  1. 1.Key Laboratory for the Genetics of Developmental and Neuropsychiatric Disorders (Ministry of Education), Bio-X Institutes, Shanghai Jiao Tong University, Shanghai 200240, China; 2.Key Laboratory of Fertility Preservation and Maintenance of Ministry of Education, Ningxia Medical University, Yinchuan 750004, China
  • Online:2015-05-28 Published:2015-06-04
  • Supported by:

    National Basic Research Program of China, 2013CB967401; National Natural Science Foundation of China, 81370675, 81200472

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摘要：

目的 探究一组精原干细胞（SSCs）中高表达的miRNAs在睾丸不同发育阶段的表达变化及miR-125a对体外培养SSCs增殖和凋亡的调节。方法 采用实时荧光定量RT-PCR研究34个miRNAs在睾丸不同发育阶段的表达变化。通过miR-125a过表达和抑制慢病毒颗粒感染体外培养SSCs，利用CCK8实验和EdU标记及Hoachst 33342、碘化丙啶（PI）双染研究miR-125a对体外培养SSCs增殖和凋亡的影响。最后通过实时荧光定量RT-PCR和Western blotting验证促凋亡基因Bak1是否为miR-125a的靶基因。结果 不同miRNAs在睾丸发育过程中具有不同的表达模式。miR-125a能促进体外培养SSCs数目的增加，减少其凋亡的发生。miR-125a过表达能降低体外培养SSCs中Bak1 mRNA和蛋白表达量，而miR-125a的抑制则增加体外培养SSCs中Bak1 mRNA和蛋白的表达量。结论 不同miRNAs在睾丸发育中有不同的表达模式；miR-125a能调控SSCs增殖和凋亡，Bak1在SSCs中是miR-125a的靶基因。

关键词: 精子发生, 精原干细胞, miRNAs, miR-125a, Bak1

Abstract:

Objective To explore variations of expressions of a group of miRNAs that highly express in spermatogonial stem cells (SSCs) during different stages of testis development and the regulation of proliferation and apoptosis of SSCs cultured in vitro by miR-125a. Methods The variations of expressions of 34 miRNAs during different stages of testis development were explored by the real-time RT-PCR. SSCs cultured in vitro were infected with lentivirus particles with over-expression and expression suppression of miR-125. The effects of miR-125a on the proliferation and apoptosis of SSCs cultured in vitro were explored by CCK8 assay, EdU incorporation, and double staining of Hoachst 33342 and propidium iodide (PI). Finally, real-time RT-PCR and Western blotting were adopted to verify whether the proapoptotic gene Bak1 was the target gene of miR-125a. Results Different miRNAs had different expression patterns during the testis development. MiR-125a increased the number of SSCs cultured in vitro and decreased the apoptosis. Over-expression of miR-125a decreased mRNA and protein expressions of Bak1 in SSCs cultured in vitro, while expression suppression of miR-125a increased mRNA and protein expressions of Bak1 in SSCs cultured in vitro. Conclusion Different miRNAs have different expression patterns during the testis development. MiR-125a can regulate the apoptosis and proliferation of SSCs and Bak1 is a target gene of miR-125a in SSCs.

Key words: spermatogenesis, spermatogonial stem cells, miRNAs, miR-125a, Bak1