Family Planning Research Institute/Center of Reproductive Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
Hum Reprod. 2012 Apr;27(4):991-7. doi: 10.1093/humrep/der481. Epub 2012 Jan 26.
Cell-free seminal mRNA (cfs-mRNA) exists in human ejaculate at high concentrations and with high stability, and contains many tissue-specific transcripts secreted from the male reproductive system. Owing to the sensitivity of RNA technology, cfs-mRNAs are ideal candidates for non-invasive biomarkers of physiopathological conditions. This study applied cfs-mRNA in identifying the presence of either germ cells or complete obstruction in men with azoospermia.
RT-PCR was performed to amplify the germ cell-specific (DDX4), seminal vesicle-specific (SEMG1) and prostate-specific (TGM4) mRNAs from cfs-mRNAs, which were isolated from the seminal plasma of men with non-obstructive azoospermia (NOA) or obstructive azoospermia (OA). The 39 patients with NOA, diagnosed by testicular biopsy, included 8 men with maturation arrest (MA), 3 men with incomplete sertoli cell only (iSCO) syndrome and 28 men with complete SCO (cSCO). The 29 patients with OA, confirmed by the presence of sperm in the testis or epididymis, included 8 men with congenital bilateral absence of the vas deferens (CBAVD) and 21 men with non-CBAVD. Healthy individuals and vasectomized men were enrolled as controls.
TGM4 was detected in all participants. Consistent with their diagnosis, DDX4 was detected in all patients with MA or iSCO but was absent in most cases of cSCO (n = 21, 75.0%) or non-CBAVD (n = 18, 85.7%), and in all men with vasectomy or CBAVD. The presence of DDX4 in the other seven men with cSCO and three patients with non-CBAVD suggests the presence of germ cells in the testis, and incomplete obstruction, respectively. SEMG1 was undetectable in three patients with CBAVD with bilateral absence of the seminal vesicles, and in two non-CBAVD cases with low ejaculate volume.
These results suggest that, with high sensitivity and representativity, cfs-mRNA could be non-invasive biomarkers for identifying the presence of germ cells or complete obstruction in azoospermia.
游离于精液中的 mRNA（cfs-mRNA）在人体精液中浓度高、稳定性强，其中包含了许多来自男性生殖系统的组织特异性转录本。由于 RNA 技术的敏感性，cfs-mRNA 是生理病理条件下非侵入性生物标志物的理想候选物。本研究应用 cfs-mRNA 鉴定非梗阻性无精子症（NOA）或梗阻性无精子症（OA）患者中是否存在生精细胞或完全梗阻。
从非梗阻性无精子症（NOA）或梗阻性无精子症（OA）患者的精液中分离出 cfs-mRNA，然后使用逆转录 PCR 扩增其生精细胞特异性（DDX4）、精囊特异性（SEMG1）和前列腺特异性（TGM4）mRNA。39 名经睾丸活检诊断为 NOA 的患者，包括 8 名未成熟阻滞（MA）患者、3 名不完全唯支持细胞综合征（iSCO）患者和 28 名完全唯支持细胞综合征（cSCO）患者。29 名经睾丸或附睾中存在精子证实为 OA 的患者，包括 8 名先天性双侧输精管缺如（CBAVD）患者和 21 名非 CBAVD 患者。健康个体和输精管结扎者作为对照。
所有参与者均检测到 TGM4。与诊断一致，DDX4 在所有 MA 或 iSCO 患者中均有检测到，但在大多数 cSCO（n=21，75.0%）或非 CBAVD（n=18，85.7%）患者中未检测到，且在所有输精管结扎者或 CBAVD 患者中均未检测到。在其他 7 名 cSCO 患者和 3 名非 CBAVD 患者中检测到 DDX4，提示睾丸中存在生精细胞，分别存在不完全梗阻。在 3 名双侧精囊缺失的 CBAVD 患者和 2 名精液量低的非 CBAVD 患者中未检测到 SEMG1。
这些结果表明，cfs-mRNA 具有高灵敏度和代表性，可作为非侵入性生物标志物，用于鉴定无精子症中是否存在生精细胞或完全梗阻。
