Y染色体基因微缺失与男性不育的关系

目的 :探讨男性不育患者尤其是特发性无精子症、严重少精子症及双侧输精管缺如与 Y染色体基因 (无精子因子 ,AZF)微缺失的关系。方法 :对 97例男性不育患者及 2 0例正常男性采用多重聚合酶链反应法进行基因微缺失检测。结果 :36例特发性无精症患者中存在 3例缺失 ,占 8.33% ;1 4例双侧输精管缺如患者存在 2例缺失 ,占 1 4.2 9% ;2 7例严重少精症患者中存在 2例缺失 ,占 7.41 %。2 0例精子数正常的男性不育患者及 2 0例正常男性对照无 AZF缺失。缺失以 AZFa,AZFc区为主 ,AZFb区无缺失。结论 :Y染色体 AZF微缺失可能是导致男性特发性无精症、少精症的原因之一 ,双侧输精管缺如患者也存在 Y染色体的基因微缺失     

无精症、少精症患者中AZF缺失的检测

目的:探讨AZF缺失与男性无精症、少精症之间的关系。方法:采用多重PCR方法,对84例汉族无精症和少精症患者的AZF区4个STS位点:AZFa亚区的sY86、AZFb亚区的sY127、AZFc亚区的sY254和sY255进行了缺失检测。用PCR-SSCP方法对其中的48例作了sY254点突变检测。并对DAZ基因在人不同组织的表达进行了研究。结果:3例无精症患者同时具有位于AZFc亚区中DAZ基因的sY254和sY255位点缺失。48例患者未见sY254有点突变。只在睾丸组织中检测到DAZ基因的表达。结论:部分汉族无精症患者可能与AZFc的缺失有关。     

精子发生障碍患者Y染色体微缺失分子诊断

目的:通过建立的Y染色体微缺失筛查方法了解生精障碍与Y染色体微缺失的关系。方法:同时采用多重PCR-凝胶电泳技术和多重PCR-液态芯片技术对原发性无精症、少精症患者和精液常规正常男性对照进行Y染色体微缺失筛查。结果:42例无精症、少精症患者中,6例有AZF区域STS位点或基因的缺失,总缺失率为14.3%,AZFc/DAZ区发生微缺失的频率较高,AZFa区发生微缺失的频率较低。结论:本研究建立的Y染色体微缺失多重PCR-MASA检测系统具有相对高通量、简便、快速、特异性强、敏感性高等特点;染色体微缺失是导致男性精子发生障碍的重要原因之一,AZF的候选基因在精子发生过程中可能起重要作用。     

精子发生相关基因的分子遗传学研究进展

特发性无精、严重少精症的分子遗传学研究越来越广泛深入。近年来,控制精子发生的相关基因,主要包括位于Y染色体AZFa区的USP9Y和DBY、AZFb区的RBM基因、AZFc区的DAZ基因,和位于常染色体的DAZLA基因等在分子遗传学研究方面均取得了可喜的进展。对这方面的研究进展及其展望进行了综述。     

男性不育中Y染色体微缺失的研究进展

近年来对男性不育的遗传学因素的广泛研究显示Y染色体微缺失是导致不同程度生精障碍从而引起男性不育的第二大遗传学病因。无精子症因子区(AZF区)由近至远包含3个不同的亚区:AZFa、AZFb和AZFc,不同缺失类型的表型不同。目前常采用PCR法进行Y染色体微缺失的检测,其缺点是准确度低、特异性差、耗时。而基因芯片技术虽能克服上述缺点,但目前成本过高。通过检测能预测患者男性后代的遗传风险,有助于患者选择辅助治疗的方式。虽然Y染色体微缺失的严重不育患者能通过辅助生殖技术成功获得后代,但有可能将遗传缺陷传给男性后代,使之获得相同的Yq微缺失和不育。     

Y染色体微缺失与辅助生殖技术关系的研究进展

Y染色体是男性特有的染色体,其长臂上的无精子因子(AZF)区域具与男性不育密切相关的基因,目前将该区域分为AZFa、AZFb、AZFc和AZFd4个区域。AZF缺失是导致男性不育的重要因素之一,可以通过辅助生殖技术(ART)遗传给下一代引起不育。研究Y染色体微缺失分类与表型关系,可以为临床治疗各种男性不育症提供分子或细胞水平的依据。Y染色体微缺失发生频率存在种族差异性;目前Y染色体微缺失的检测方法仍然以多重PCR为主;对于ICSI助孕的男性后代是否会出现新发Y染色体微缺失仍然存在争论。     

毛细管电泳用于Y染色体微缺失的研究

目的:探讨毛细管电泳在研究男性不育中特发性无精子症和射出精液严重少精子症与Y染色体无精子因子(azoospermiafactor,AZF)缺失的作用。方法:应用多重PCR技术对4例无精子症和29例严重少精子症患者的外周血细胞中Y染色体AZF所在的11.23区的15个位点进行扩增,分别用琼脂糖凝胶电泳和毛细管电泳分离扩增产物,并以6例正常生育男性和2例女性为对照。结果:毛细管电泳发现4例无精子症中3例发生缺失,29例严重少精子症患者中6例发生缺失。其中6例同为SY254(C)、SY242(C)、SY255(C)、SY239(C)、SY152(D)缺失。6例正常生育男性毛细管电泳未发现Y染色体微缺失,而凝胶电泳有2个条带模糊,难以判断。结论:毛细管电泳可提高AZF检测准确性。Y染色体AZFc/DAZ的缺失可能是引起无精子和严重少精子并造成男性不育的重要原因之一。     

Y染色体长臂缺失及不分离不育男性1例报道

目的:报道1例Y染色体长臂缺失合并不分离的男性无精子症患者。方法:常规染色体核型分析,荧光原位杂交以确定核型。PCR-STSs检测以确定Y染色体断裂点,并行睾丸活检。结果:细胞遗传学和FISH证实患者为嵌合体,核型为45,X/46,X,del(Y)/47,X,del(Y)del(Y)。分别占27%,68%,5%。C带显示患者Yq12全部丢失。PCR-STSs检测AZFa存在,AZFb和AZFc区域全部丢失,断裂点位于sY88和sY95之间及sY88以下。睾丸病理显示精曲小管中只有支持细胞,没有生精细胞。未见卵巢组织。结论:患者无精子症、睾丸体积小与病理结果一致,其原因是由于Yq11.2的缺失。     

无精子因子AZFc微缺失的相关研究进展

Y染色体无精子因子(azoospermia factor,AZF)缺失是男性不育的重要原因,该缺失可导致男性生精障碍甚至是无精子症。AZFc缺失在AZF所有缺失类型中最为常见的,加上AZFc区域在Y染色体上的基因结构复杂,被发现有多种缺失类型,因此AZFc微缺失男性的临床表现也多样,从精子密度正常、少精到无精都可能发生。随着辅助生殖技术的发展,因AZFc缺失而严重少弱精的患者可通过辅助生育方式将这种染色体的缺陷遗传给男性子代并可能导致子代缺失的扩大;并且,因严重少弱精而通过ICSI治疗的男性,其子代AZFc微缺失的发生风险明显增加。     

Y染色体单倍群与精子发生相关性

Y染色体上存在众多精子发生的相关基因。在Y染色体进化过程中形成可稳定遗传的碱基突变位点:SNP位点。根据Y染色体SNP位点不同,把全球人群分为153个单倍群(haplogroup)。大量研究显示,Y染色体无精子因子(azoospermia factor,AZF)区基因重组引起的缺失是导致男性不育的重要原因。常见的缺失类型为AZFa、AZFb、AZFc、AZFbc、AZFabc,以及AZFc区的部分缺失:gr/gr缺失、b2/b3缺失。目前研究表明,Y染色体单倍群不但与精子发生相关,而且与Y染色体微缺失相关,即某些单倍群易发生Y染色体微缺失而导致男性不育。     

Microdeletions within the azoospermia factor subregions of the Y chromosome in patients with idiopathic azoospermia.

OBJECTIVE: To examine the patterns of submicroscopic DNA deletions in the AZF (AZoospermia Factor) subregions of the Y chromosome in patients with idiopathic azoospermia. DESIGN: Controlled clinical study. SETTING: University-based infertility clinic. PATIENT(S): Infertile men (n = 40) with nonobstructive, idiopathic azoospermia. The control group consisted of proven fathers (n = 14) and healthy women (n = 4). INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Semen analysis; polymerase chain reaction amplification of the 37 loci spanning the AZFa, AZFb, and AZFc subregions of the Y chromosome; serum FSH, LH, and testosterone levels; and testicular histologic analysis. RESULT(S): Testicular histologic analysis of the subjects revealed Sertoli cell-only syndrome (n = 36) and spermatogenic arrest (n = 4). Microdeletions of the Y chromosome were found in eight (20%) of the patients with azoospermia. All eight affected patients had interstitial microdeletions within the AZFc subregion. Patients with Sertoli cell-only syndrome had additional microdeletions in regions distal to DAZ (Deleted in Azoospermia), although DAZ deletion was observed in seven of the eight affected patients. In five patients, microdeletions were found in the AZFb region containing RBM (RNA Binding Motif). CONCLUSION(S): Our results add to the evidence supporting the current suggestion that there is a cause-and-effect relation between Yq11 microdeletions in the AZF region and azoospermia.     

Screening for Y chromosome microdeletions in assisted reproductive techniques

In case of infertility due to azoospermia, clinical, sonographical and biological examinations suggest obstructive or non obstructive causes. In cases of non obstructive azoospermia, genomic microdeletions must be determined particularly in the Y chromosome long arm, as well as autosomal abnormality. A constitutional karyotype must also be done. The so-called Y AZFa, AZFb and AZFc zones could be partially or totally absent. Genotype is mostly correlated with histology. Thus, when large AZFa and AZFb microdeletions are detected there is theoretically no chance to find testicular spermatozoa. If only AZFc microdeletions are present, testicular biopsy is possible with a good chance of mature spermatozoa retrieval before microinjection, and AZFc microdeletions are also often present (10%) in cases of severe oligospermia. Couples must be informed of genomic deletions and a genetic counseling is essential to explain the potential childhood risks after assisted reproductive techniques. This problem has been discussed by the French "High Authority of Health". It recommends determination of these Y microdeletions when oligozoospermia is severe (lower than one million spermatozoa per milliliter).     

Detection of Azoospermic Factor Genes in Chinese Men with Azoospermia or Severe Oligozoospermia

Purpose: We investigated the prevalence of deletions in the azoospermic factor (AZF) region of chromosome Yq11 in Chinese men with infertility due to idiopathic azoospermia or severe oligozoospermia. The DAZ gene cluster was also examined for mutations. Methods: Sixty-eight men with azoospermia or severe oligozoospermia taking part in an intracytoplasmic sperm injection program were recruited. Four loci specific for AZFa, AZFb, and AZFc were amplified from genomic DNA via polymerase chain reaction to determine whether deletions were present in the AZF region. Direct DNA sequencing of amplified products was also performed to look for mutations or polymorphism from exon 2 to exon 6 of the DAZ gene cluster. Results: Six (9%) of the 68 patients had AZF deletions. None had mutations in exons 2 to 6 of DAZ. Conclusions: The prevalence of AZF deletions in our study was similar to those in Western reports, as was the lack of DAZ mutations.     

非梗阻性无精子症和隐匿精子症与睾丸体积、血FSH和AZF基因微缺失的相关性分析

目的：探讨非梗阻性无精子症和隐匿精子症与睾丸体积、血FSH和AZF基因微缺失的相关性。方法：161例男性不育患者分为非梗阻性无精子症（A组,n=86）、隐匿精子症（B组,n=49）、严重少精子症（c组,n=13）和其它患者（D组,n=13,包括死精子症4例,梗阻性无精子症3例,正常精子1例,其他少弱精子症5例）,进行睾丸体积、血FSH水平和彳砑基因微缺失检测,AZF基因微缺失检测位点包括AZFa（sY84、sY86）、AZFb（sYl27、sYl34）、AZFcd（sY254、sY255、sYl57、sYl45和sYl52）。结果：4组睾丸体积〈12ml的比例分别为73．26％（63／86）、34．69％（17／49）、7．69％（1／13）、30．77％（4／13）;4组FSH水平升高1倍以上的比例分别为67．44％（58／86）、32．65％（16／49）、15．38％（2／13）和O．OO％（o／13）,组间均有极显著统计学差异（P〈O．001）;4组4ZF基因微缺失率分别为15．12％（13／86）．18．37％（9／49）、0．oo％（o／13）和0．00％（0／13）,组间均无统计学差异（P〉0．05）,其中A组爿zn基因缺失1例,AZF（b＋c＋d）缺失5例,AZF（c＋d）基因缺失7例,B组9例均AZFC和（或MZFd基因缺失,c组和D组均未见AZF基因缺失。结论：随着生精功能障碍程度的加重,血FSH水平升高和睾丸体积缩小趋势明显,而与AZF缺失几率无高度相关性;但在非梗阻性无精子症和隐匿精子症组中AZF基因缺失几率仍然偏高,AZFa和AZFb缺失大多出现在非梗阻性无精子症组中,而隐匿精子症组以AZFc和AZFd缺失为主。     

Y-chromosome microdeletion and phenotype in cytogenetically normal men with idiopathic azoospermia

OBJECTIVE: To determine the prevalence of microdeletions of the long arm of chromosome Y within the AZFa, AZFb, and AZFc subregions in patients with idiopathic azoospermia, and then correlate the microdeletions with clinical phenotypes to determine the most important subregion for screening. DESIGN: Controlled clinical study. SETTING: Male infertility clinic, Kobe University Hospital. PATIENT(S): Among 89 consecutive azoospermic patients, those whose infertility was related to known hereditary, endocrine, or obstructive causes or a cytogenetic abnormality were excluded; 54 remaining patients were studied using a polymerase chain reaction (PCR). Of these patients, 33 had Sertoli cell only syndrome, 10 had maturation arrest, and 11 had hypospermatogenesis. INTERVENTION(S): Blood and semen samples and testicular biopsies were obtained from all of the participants. MAIN OUTCOME MEASURE(S): We performed semen analysis, polymerase chain amplification of 28 DNA loci on the long arm of the Y chromosome involving the DAZ (deleted in azoospermia), and measured the plasma FSH, LH, testosterone, prolactin, and estradiol levels. RESULT(S): Microdeletions were detected in 14 of the 54 patients (nine with Sertoli cell only, three with maturation arrest, and two with hypospermatogenesis). Most microdeletions involved AZFb or AZFc. Patients with hypospermatogenesis or maturation arrest showed deletion only in AZFc. The DAZ gene was deleted in four patients with Sertoli cell only and one patient with maturation arrest. The RBM gene was deleted in two patients with Sertoli cell only who had particularly large deletions, but in no patients with arrest or hypospermatogenesis. CONCLUSION(S): Cytogenetically azoospermic patients should be examined for microdeletions before undertaking assisted reproduction. AZFc may be the most important subregion to screen. In addition, intact AZFa and AZFb subregions may be important for the presence of germ cells.     

Genetic screening for chromosomal abnormalities and Y chromosome microdeletions in Chinese infertile men

Purposes

To investigate the frequency and type of both chromosomal abnormalities and Y chromosome microdeletions and analyze their association with defective spermatogenesis in Chinese infertile men.

Methods

This is a single center study. Karyotyping using G-banding and screening for Y chromosome microdeletion by multiplex polymerase chain reactio(PCR)were performed in 200 controls and 1,333 infertile men, including 945 patients with non-obstructive azoospermia and 388 patients with severe oligozoospermia.

Results

Out of 1,333 infertile patients, 154(11.55%) presented chromosomal abnormalities. Of these, 139 of 945 (14.71%) were from the azoospermic and 15 of 388 (3.87%) from the severe oligozoospermic patient groups. The incidence of sex chromosomal abnormalities in men with azoospermia was 11.53% compared with 1.03% in men with severe oligozoospermia (P < 0.01). Also 144 of 1,333(10.80%) patients presented Y chromosome microdeletions. The incidence of azoospermia factor(AZF) microdeletion was 11.75% and 8.51% in patients with azoospermia and severe oligozoospermia respectively. Deletion of AZFc was the most common and deletions in AZFa or AZFab or AZFabc were found in azoospermic men. In addition, 34 patients had chromosomal abnormalities among the 144 patients with Y chromosome microdeletions. No chromosomal abnormality and microdeletion in AZF region were detected in controls.

Conclusions

There was a high incidence (19.80%) of chromosomal abnormalities and Y chromosomal microdeletions in Chinese infertile males with azoospermia or severe oligozoospermia. These findings strongly suggest that genetic screening should be advised to infertile men before starting assisted reproductive treatments.