精子发生相关基因SPATA48在人和小鼠睾丸组织中的表达研究

Expression of spermatogenesis related gene SPATA48 in human and mouse testis

目的通过分析精子发生相关基因SPATA48在人和小鼠睾丸组织中的表达,探讨SPATA48在精子发生中的作用。方法收集2010年1月至2017年12月在深圳市第二人民医院就诊的非梗阻性无精子症患者和正常生育者睾丸活检样本各60例;腹腔注射白消安方法建立小鼠无精子症模型,小鼠在不同时期处死后解剖留取各个脏器;提取小鼠各脏器组织、两组睾丸组织以及人精母细胞、精原细胞和支持细胞中mRNA;并购买人肺、肾、肝脏、脾脏等组织mRNA样本。利用RT-PCR方法检测SPATA48基因在各种细胞和组织器官、两组人睾丸组织中的表达水平,并利用免疫组化方法检测SPATA48蛋白在各种细胞和两组人睾丸组织中的表达水平,对各检测指标进行相关统计分析。结果 RT-PCR结果表明SPATA48基因特异表达在睾丸组织中,肺、肾脏、肝脏、脾脏等其他组织中均不表达;非梗阻性无精子症患者睾丸组织中无SPATA48基因表达,三种睾丸细胞仅精母细胞存在阳性表达。无精子症模型小鼠,随白消安处理时间的延长,SPATA48基因表达显著减弱(P<0.05)。免疫组化结果显示,在正常生育者睾丸中SPATA48蛋白主要表达在精母细胞和圆形精子细胞中,在非梗阻性无精子症患者睾丸中无SPATA48蛋白表达。结论 SPATA48特异表达在人和小鼠睾丸组织中,无精子发生的睾丸组织无表达,提示SPATA48基因在精子发生中具有重要作用。

Objective： To investigate the role of spermatogenesis related gene SPATA48 in spermatogenesis by analyzing the expression of SPATA48 in human and mouse testis tissue. Methods： The testicular biopsy samples from the patients with non- obstructive azoospermia and the patients with normal fertility, 60 samples for each group were collected in Shenzhen Second People＇s Hospital from January 2010 to December 2017. The mouse azoospermia model was established by intraperitoneal injection of busulfan. The mice were executed and collect various organs at different times. The mRNA was extracted from various organs , the testicular tissues of two patient groups, as well as human spermatocytes, spermatogonia and Sertoli cells. The mRNA samples of human lung, kidney, liver, spleen and other tissue were purchased. The expression of the gene in human and mouse cells,testis tissues and other organs was analyzed by RT PCR. The expression of SPATA48 in the cells,testis tissues was also determined by immunohistochemistry. The test indicator was statistical analyzed. Results： The RT PCR results showed that SPATA48 gene was not expressed in lung, kidney, liver, spleen and other tissues and was highly expressed in the testis. There was no expression of SPATA48 in the testis with non obstructive azoospermia. SPATA48 gene was not expressed in human spermatogonia and Sertoli ceils except spermatocytes. In the azoospermia model mice, SPATA48 gene expression was significantly decreased with the prolonged treatment time of busulfan（P〈0.05）. Immunohistochemistry and RT PCR analysis showed that SPATA48 protein was mainly expressed in spermatocytes and round spermatids in the testis of patients with normal fertility. There is no SPATA48 protein expression in the testis of patients with non obstructive azoospermia. Conclusions： SPATA48 was highly expressed in human and mouse testis tissues; there is no expression of SPATA48 in the testis of patients with azoospermia; which suggests that SPATA48 gene plays an important role in the regulation of spermatogenesis.