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Abstract：
OBJECTIVE: Cystic fibrosis transmembrane conductance regulator (CFTR) and adhesion G protein-coupled receptor G2 (ADGRG2) have been identified as the main pathogenic genes in congenital bilateral absence of the vas deferens (CBAVD), which is an important cause of obstructive azoospermia. This study aimed to identify the disease-causing gene in two brothers with CBAVD from a Chinese consanguineous family and reveal the intracytoplasmic sperm injection (ICSI) outcomes in these patients.
METHODS: Whole-exome sequencing and Sanger sequencing were used to identify the candidate pathogenic genes. Real-time polymerase chain reaction, immunohistochemistry, and immunofluorescence were used to assess the expression of the mutant gene. Moreover, the ICSI results from both patients were retrospectively reviewed.
RESULTS: A novel hemizygous loss-of-function mutation (c.G118T: p.Glu40*) in ADGRG2 was identified in both patients with CBAVD. This mutation is absent from the human genome databases and causes an early translational termination in the third exon of ADGRG2. Expression analyses showed that both the ADGRG2 mRNA and the corresponding protein were undetectable in the proximal epididymal tissue of ADGRG2-mutated patients. ADGRG2 expression was restricted to the apical membranes of non-ciliated epithelia in human efferent ducts, which was consistent with a previous report in mice. Both ADGRG2-mutated patients had normal spermatogenesis and had successful clinical outcomes following ICSI.
CONCLUSIONS: Our study verifies the pathogenic role of ADGRG2 in X-linked CBAVD and broadens the spectrum of ADGRG2 mutations. In addition, we found positive ICSI outcomes in the two ADGRG2-mutated CBAVD patients.
METHODS: Whole-exome sequencing and Sanger sequencing were used to identify the candidate pathogenic genes. Real-time polymerase chain reaction, immunohistochemistry, and immunofluorescence were used to assess the expression of the mutant gene. Moreover, the ICSI results from both patients were retrospectively reviewed.
RESULTS: A novel hemizygous loss-of-function mutation (c.G118T: p.Glu40*) in ADGRG2 was identified in both patients with CBAVD. This mutation is absent from the human genome databases and causes an early translational termination in the third exon of ADGRG2. Expression analyses showed that both the ADGRG2 mRNA and the corresponding protein were undetectable in the proximal epididymal tissue of ADGRG2-mutated patients. ADGRG2 expression was restricted to the apical membranes of non-ciliated epithelia in human efferent ducts, which was consistent with a previous report in mice. Both ADGRG2-mutated patients had normal spermatogenesis and had successful clinical outcomes following ICSI.
CONCLUSIONS: Our study verifies the pathogenic role of ADGRG2 in X-linked CBAVD and broadens the spectrum of ADGRG2 mutations. In addition, we found positive ICSI outcomes in the two ADGRG2-mutated CBAVD patients.
摘要：
目的：囊性纤维化跨膜传导调节因子（CFTR）和粘附G蛋白偶联受体G2（ADGRG2）已被确定为先天性双侧输精管缺失（CBAVD）的主要致病基因。这是梗阻性无精子症的重要原因。这项研究旨在鉴定来自中国近亲家庭的两个CBAVD兄弟的致病基因，并揭示这些患者的胞浆内单精子注射（ICSI）结果。
方法：使用全外显子组测序和Sanger测序鉴定候选致病基因。实时聚合酶链反应，免疫组织化学，和免疫荧光用于评估突变基因的表达。此外，我们对2例患者的ICSI结果进行了回顾性分析.
结果：一种新的半合子功能丧失突变（c。在两名CBAVD患者中都发现了G118T：p.Glu40*）在ADGRG2中。这种突变在人类基因组数据库中不存在，并导致ADGRG2第三外显子的早期翻译终止。表达分析表明，ADGRG2mRNA和相应的蛋白在ADGRG2突变患者的近端附睾组织中均未检测到。ADGRG2的表达仅限于人输出管中无纤毛上皮的顶端膜，这与先前在小鼠中的报道一致。两名ADGRG2突变患者的精子发生正常，ICSI后临床结局成功。
结论：我们的研究验证了ADGRG2在X连锁CBAVD中的致病作用，并拓宽了ADGRG2突变的谱。此外,我们在2例ADGRG2突变的CBAVD患者中发现ICSI结果阳性.
方法：使用全外显子组测序和Sanger测序鉴定候选致病基因。实时聚合酶链反应，免疫组织化学，和免疫荧光用于评估突变基因的表达。此外，我们对2例患者的ICSI结果进行了回顾性分析.
结果：一种新的半合子功能丧失突变（c。在两名CBAVD患者中都发现了G118T：p.Glu40*）在ADGRG2中。这种突变在人类基因组数据库中不存在，并导致ADGRG2第三外显子的早期翻译终止。表达分析表明，ADGRG2mRNA和相应的蛋白在ADGRG2突变患者的近端附睾组织中均未检测到。ADGRG2的表达仅限于人输出管中无纤毛上皮的顶端膜，这与先前在小鼠中的报道一致。两名ADGRG2突变患者的精子发生正常，ICSI后临床结局成功。
结论：我们的研究验证了ADGRG2在X连锁CBAVD中的致病作用，并拓宽了ADGRG2突变的谱。此外,我们在2例ADGRG2突变的CBAVD患者中发现ICSI结果阳性.
