Institute of Clinical Laboratory Medicine, Nanjing Jinling Hospital, Nanjing University Clinical School of Medical College, Nanjing, China.
Urology. 2013 Mar;81(3):696.e1-8. doi: 10.1016/j.urology.2012.11.005. Epub 2013 Jan 3.
To investigate the expression pattern of Ras-related protein 1 (Rap1) during testicular development and to clarify whether its expression is developmentally regulated and whether this expression is involved in the process of germ cell apoptosis.
The expression pattern of Rap1 in the adult rat testicle was compared with that in the developing rat testicle using immunoblotting and immunohistochemical analyses. After the adult rats were treated with methoxyacetic acid (MAA), which selectively depletes primary spermatocytes, we correlated Rap1 expression with apoptotic dynamics using terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL), double immunofluorescent staining, and coimmunoprecipitation assays.
During testicular development, Rap1 was expressed in the nucleus of gonocytes and in the Golgi apparatus of spermatocytes. The expression pattern of Rap1 during spermatogenesis was also shown to be stage specific. After 12 hours of MAA treatment, we found that Rap1 was translocated into the nucleus of some spermatocytes and the overlapping rate of Rap1 and NF-κB was much greater than that of Rap1 and TUNEL staining. In addition, Rap1 protein could be co-immunoprecipitated with NF-κB protein.
In cooperation with NF-κB, Rap1 might be involved in the early stage of the apoptotic process occurring in the MAA-treated rat testicle. Additional characterization of this small guanosine triphosphatase in the apoptotic dynamics should provide information on the early diagnosis of MAA-induced male infertility.
研究 Ras 相关蛋白 1（Rap1）在睾丸发育过程中的表达模式，阐明其表达是否受发育调控，以及这种表达是否参与生精细胞凋亡过程。
采用免疫印迹和免疫组织化学分析比较 Rap1 在成年大鼠睾丸和发育中大鼠睾丸中的表达模式。在成年大鼠用甲氧基乙酸（MAA）处理后，该药物选择性消耗精原细胞，我们用末端脱氧核苷酸转移酶介导的脱氧尿苷三磷酸 - 生物素缺口末端标记（TUNEL）、双重免疫荧光染色和共免疫沉淀分析将 Rap1 表达与凋亡动力学相关联。
在睾丸发育过程中，Rap1 表达在性母细胞的核中和精母细胞的高尔基器中。Rap1 在精子发生过程中的表达模式也表现出阶段特异性。在 MAA 处理 12 小时后，我们发现 Rap1 易位到一些精母细胞的核中，并且 Rap1 和 NF-κB 的重叠率远高于 Rap1 和 TUNEL 染色的重叠率。此外，Rap1 蛋白可以与 NF-κB 蛋白共免疫沉淀。
Rap1 与 NF-κB 合作，可能参与 MAA 处理大鼠睾丸中发生的凋亡过程的早期阶段。对这种小 GTP 酶在凋亡动力学中的进一步表征应提供有关 MAA 诱导男性不育症早期诊断的信息。
