The prognostic role of the extent of Y microdeletion on spermatogenesis and maturity of Sertoli cells

Substantial involvement of the Y chromosome in sexual development and spermatogenesis has been demonstrated. Over the last decade, varying extent of Y chromosome microdeletions have been identified among infertile patients with azoospermia or oligozoospermia. These microdeletions were clustered in three main regions named AZFa, AZFb, and AZFc. Analysis of the Y chromosome microdeletion was found to be of prognostic value in cases of infertility, both in terms of clinical management as well as for understanding the aetiology of the spermatogenesis impairment. However, the accumulated data are difficult to analyse, due to the variable extent of these deletions, the different sequence-tagged sites (STS) used to detect the microdeletions, and the non-uniformity of the histological terminology used by different investigators. This debate discusses the chances of finding testicular spermatozoa in men with a varying extent of Y chromosome microdeletions. The genotype and germ cell findings in men with AZFa microdeletions as well as those that include more than a single AZF region are reviewed, as is the effect of Y chromosome AZF microdeletions on the maturity of the Sertoli cells.     

Y染色体微缺失父子间垂直遗传分析

目的对Y染色体微缺失男性不育患者家系分析,探讨Y染色体微缺失父子间的自然垂直遗传。方法调查12例Y染色体无精子症因子(azoospermia factor,AZF)微缺失不育患者直系男性家族成员,取外周血抽提DNA进行改良多重PCR,绘制男子不育家系系谱图。结果12例家系中2例存在AZFc微缺失的家族遗传性。10例AZF微缺失患者仅本人存在缺失,没有家族遗传性。结论Y染色体AZFc微缺失有生育子代可能,并将这种微缺失自然垂直遗传给男性后代,且相同遗传类型可有不同的临床表型。     

Klinefelter综合征患者Y染色体AZF微缺失分析

目的观察Klinefelter综合征患者Y染色体AZF微缺失发生情况。方法12例Klinefelter综合征患者ICSI/IVF等辅助受孕前进行睾丸细针穿刺吸液细胞学检查及Y染色体AZF微缺失分析。确定8个实验用序列标签位点（STS）,分别是：sY84、sY86、sY127、sY134、sY152、sY153、sY254、sY255,并以X/Y连锁锌指蛋白基因（ZFX/Y）为内对照进行多重PCR筛查AZF微缺失。结果睾丸细针穿刺吸液细胞学检查显示,3例（25.0%,3/12）可见到极少量形态较完整的精子及各级生精细胞、精子细胞,7例（58.3%,7/12）仅见少量生精细胞及精子细胞,2例（16.7%,2/12）仅见支持细胞,未见生精细胞及精子。12例Klinefelter综合征患者共检测出AZF微缺失2例分别为AZFa＋AZFc区缺失和AZFb＋AZFc区缺失;对照组32例样本未检出AZF基因微缺失。KS患者AZF微缺失检出率与对照组比较有显著差异（χ^2=5.587,P=0.018）。结论Klinefelter综合征患者存在Y染色体长臂AZF微缺失,缺失率为16.7%。     

Y chromosome microdeletions and germinal mosaicism in infertile males

Molecular deletions of the Y chromosome long arm are a frequent cause of male infertility. Because these deletions are thought to be inherited from fathers without Y chromosome deletions, the question arises as to whether their relatively high incidence in the male population could be due to the existence of a mosaicism in somatic and/or germinal paternal cells. This study included a total of 181 infertile men, among whom 18 were found to have an abnormal karyotype. In the other 163, polymerase chain reaction (PCR) analysis detected nine (5.5%) Y chromosome microdeletions. Blood, spermatozoa or testicular cells from 47 men (27 oligozoospermia, 20 azoospermia), including six Y-deleted patients, were screened for mosaicism using double target fluorescence in-situ hybridization (FISH) with Y centromeric and deleted in azoospermia (DAZ) gene-specific probes. Results indicated that: (i) percentages of double (intact Y chromosome) or single (deleted Y chromosome) fluorescent signals by FISH were in agreement with PCR data, thus demonstrating the reliability of the method; and (ii) a weak germ cell mosaicism was found in only two oligozoospermic patients, carrying 1.97 and 4.13% respectively of spermatozoa with a deleted Y chromosome. Further studies on larger populations are needed to evaluate precisely the incidence of Y deletion mosaicisms in infertile men.     

男性不育患者Y染色体AZF基因微缺失检测

目的探讨原发性无精子症、严重少精子症及少精子症患者与Y染色体无精子因子（azoospermia factor,AZF）区微缺失的关系。方法采用多重PCR方法对对照组192例已正常生育男性和实验组448例男性不育患者进行AZF区域内的15个序列标签位点（STS）的检测。结果对照组未发现AZF基因微缺失,实验组448例患者检测出五种AZF微缺失类型共41例,总缺失率为9.2%（41/448）,其中无精子症、严重少精子症和少精子症患者的缺失率分别为12.0%（19/158）、10.8%（17/157）、3.8%（5/133）,无精子症和严重少精子症患者Y染色体AZF微缺失率明显高于少精子症组,差别有统计学意义（P〈0.05）。使用15个STS位点进行检测其检出率较利用欧洲男科学会（European Academy of Andrology,EAA）推荐的6个STS位点提高约14%（5/36）。结论AZF微缺失是引起原发性无精子症、严重少精子症和少精子症的重要原因之一;增加STS位点检测数有利于提高AZF微缺失的检出率。     

The incidence and possible relevance of Y-linked microdeletions in babies born after intracytoplasmic sperm injection and their infertile fathers

Microdeletions linked to deletion intervals 5 and 6 of the Ychromosome have been associated with male factor infertility.Members from at least two gene families lie in the region containingazoospermia factor (AZF), namely YRRM and DAZ. With the adventof intracytoplasmic sperm injection (ICSI), it is possible formen with severe male factor infertility to produce a child.The genetic consequences of such a procedure have been questioned.This report describes the first study of a population (32 couples)of infertile fathers and their sons born after ICSI. The objectiveswere firstly to determine the incidence and map location ofY chromosome microdeletions and to compare the frequencies withother population studies involving severe male factor infertility,and secondly to formulate a working hypothesis concerning developmentalaetiology of Y chromosome microdeletions. The incidence of microdeletionsin the ICSI population was shown to be 9.4% (within the range9–18% reported for populations of severe male factor infertilitypatients). Microdeletions in two out of three affected fatherisonpairs mapped in the region between AZFb and AZFc and the thirdinvolved a large microdeletion in AZFb and AZFc. Of three affectedfather/son pairs, microdeletions were detected in the bloodof one infertile propositus father and three babies. Assumingthat the gonomes of the ICSI-derived babies are direct reflectionsof those of their fathers' germ lines, it is possible that twoof three infertile fathers were mosaic for intact Y and microdeletedY chromosomes. In such cases, the developmental aetiology ofthe microdeletion may be due to a de-novo microdeletion arisingas a post-zygotic mitotic error in the infertile propositusfather, thus producing a mosaic individual who may or may nottransmit the deletion to his ICSI-derived sons depending onthe extent of primordial germ cell mosaicism. In one of threeaffected fathers, the microdeletion detected in his blood wasalso detected in his ICSI-derived son. In this case the de-novoevent giving rise to the microdeletion may have occurred dueto a post- (or pre-) meiotic error in the germ line of thisfather's normally fertile father (i.e. the ICSI-derived baby'sgrandfather). aetiology/intracytoplasmic sperm injection/Y-chromosome microdeletions     

Prognostic value of Y deletion analysis: How reliable is the outcome of Y deletion analysis in providing a sound prognosis?

Y chromosomal microdeletions at the azoospermia factor (AZF) locus have been implicated as one of the major causes of idiopathic male infertility. The availability of intracytoplasmic sperm injection (ICSI) in treating a variety of male infertility has raised the risk of the transmission of Y microdeletions from father to son. In many IVF centres, Y microdeletion analysis has been used as a diagnostic tool for genetic counselling of infertile couples. Presently, the only prognosis that can be derived from Y microdeletion analysis is that the affected male offspring would benefit from proper clinical management of their infertility. Prognoses based on the pattern of Y microdeletions in relation to phenotype are rather subjective and inconclusive because of insufficient data to derive a definitive correlation whose significance can be determined by statistical analysis. Standardization of the number and choice of sequence-tagged sites (STS), whose deletions result in defective spermatogenesis, for the polymerase chain reaction (PCR) analysis of Y microdeletions would enhance its reliability in the interpretation of the results which is crucial for therapeutic decision-making. Furthermore, in-depth understanding of the gene functions in male infertility, especially at the AZF locus, would contribute greatly to the quality of the prognostic value of Y microdeletion analysis.     

Detection of Y chromosome microdeletions in AZF region by liquid chip technology]

OBJECTIVE: To establish a liquid chip technology to detect Y chromosome microdeletions in Chinese infertile males with azoospermia or oligozoospermia. METHODS: Multiplex PCR and liquid chip technology were used to detect the Y chromosome microdeletions in AZF region in 178 infertile patients with azoospermia and 134 infertile patients with oligozoospermia as well as 40 fertile control men. RESULTS: Forty out of 312 patients (12.8%) were found to have deletions in AZF region. The microdeletion frequency was 14%(25/178) in the azoospermic group, 9.6%(11/114) in the oligospermic and 20%(4/20) in the severe oligospermic group. CONCLUSION: The authors developed a high-throughput, fast and simple assay to screen the AZF region microdeletions of Y chromosome.     

Transmission of Y chromosomal microdeletions from father to son through intracytoplasmic sperm injection

 We conducted chromosomal analysis of three male infants fathered by severe oligozoospermic males with Y chromosomal microdeletions through intracytoplasmic sperm injection (ICSI). Two of the infants had the same Y chromosomal microdeletions as their fathers. The third infant also had a Y chromosomal microdeletion, which was longer than that found in his father. The results confirm that Y chromosomal microdeletions are transmitted from a father to a son via ICSI and also suggest that the microdeletions may be expanded during such transmission. Genetic counseling for infertile couples contemplating ICSI is important if the male carries Y chromosomal microdeletions. Received: January 11, 2002 / Accepted: May 19, 2002     

180例无精子症或严重少精子症不育男性的Y染色体微缺失检测

目的探讨不育男性无精子症或严重少精子症与Y染色体微缺失之间的关系.方法利用9个Y染色体特异序列标签位点,以多重PCR法检测无精子症或严重少精子症患者的Y染色体微缺失情况.结果 180例无精子症或严重少精子症患者中共检出Y染色体微缺失15例,缺失率为8.3%.精液正常者(对照组)20例未发现Y染色体微缺失.9例Y染色体微缺失的无精子症患者睾丸细胞学检查均未发现精子.结论 Y染色体微缺失是造成男性精子发生障碍的常见病因之一.     

200例男性不育患者的Y染色体微缺失检测

目的探讨男性不育患者与Y染色体微缺失之间的关系。方法利用15个Y染色体特异序列标签位点,以多重PCR法检测男性不育患者的Y染色体微缺失情况。结果 200例男性不育患者中共检出Y染色体微缺失7例,缺失率为3.5%。其中单纯A;ZFc缺失2例,缺失率为1%(2/200);A;ZFb缺失率为3例,缺失率为1.5%(3/200);单纯A;ZFa缺失2例,缺失率为1%(2/200),尚未发现联合缺失或大片段缺失患者。精液正常者(对照组)30例未发现Y染色体微缺失。结论 Y染色体微缺失是造成男性不育的常见病因之一。     

Lack of association between Y chromosome haplogroups and male infertility in Japanese men

The Y chromosome carries several genes involved in spermatogenesis, which are distributed in three regions in the euchromatic part of the long arm, called AZFa (azoospermia factor a), AZFb, and AZFc. Microdeletions in these regions have been seen in 10-15% of sterile males with azoospermia or severe oligozoospermia. The relatively high de novo occurrence of these microdeletion events might be due to particular chromosome arrangements associated with certain Y chromosome haplogroups. To test whether there is any association between Y chromosome types and male infertility, we studied a sample of 84 Japanese oligozoospermic or azoospermic males. The patients were analyzed for the presence of Yq microdeletions and also typed with a battery of unique event polymorphisms (UEPs) to define their Y haplogroups. Six of the infertile patients presented likely pathological microdeletions detectable with the sequence tagged sites (STS) markers used. There was no significant association between Y chromosome haplogroups and the microdeletions. We also compared the Y haplogroup frequencies in our subset sample of 51 idiopathic azoospermia patients with 57 fertile control Japanese males, and did not observe any significant differences. Contrary to previous reports, our data suggest that Y microdeletions and other molecular events causally associated with male infertility in Japan occur independently of the Y chromosome background.     

424例非梗阻性无精子症和严重少精子症患者的细胞与分子遗传学研究

目的探讨非梗阻性无精子症和严重少精子症患者的细胞与分子遗传学特点。方法应用染色体核型分析、Y染色体微缺失检测和荧光原位杂交（FISH）、PCR等技术对非梗阻性无精子症（n=291）和严重少精子症患者（n=133）男性不育患者（共424例）进行细胞和分子遗传学检测。结果424例患者中有98例明确为遗传异常引起的,其中66例检测到染色体畸变,44例Y染色体微缺失检测见缺失,12例患者染色体核型和微缺失检测均见异常。部分AZF缺失患者精液或睾丸中有精子,但其生精功能呈进行性下降的特点。结论男性不育最常见的遗传学病因为K linefelter综合征和Y染色体AZFc缺失。Y染色体微缺失检测对Y染色体长臂异染色质区缺失是否为多态性具有明确诊断的作用。细胞与分子遗传学检测为男性不育的诊断、治疗和预后以及ICSI治疗前遗传咨询提供重要依据。     

不育男性的AZF检测与Y染色体缺失的对照分析

目的探讨精子发生障碍的男性不育患者AZF缺失与Y染色体缺失的临床意义。方法对616例非阻塞性无精子症或少精子症患者进行AZF的检测,同时观察G显带Y染色体的形态。结果从616例患者中检测出48例患者分别为AZFa、AZFb、AZFc或AZFb+AZFc的微缺失,但显微镜下观察不到Y染色体形态改变。另外4例患者经AZF检测,2例为AZFc+sY160缺失,1例为AZFb+AZFc+sY160缺失,1例为AZFa+AZFb+AZFc+sY160缺失,显微镜下发现Yq部分或完全缺失。25例已育男性的G-显带的Y染色体和AZF也进行对照检测,均未发现AZF的缺失,但其中1例核型分析显示Y染色体长臂部分缺失,但PCR检测仅缺失sY160,即Yq12的缺失。结论Yq11.23上7Mb的缺失在细胞水平不能分辨。q11.23+q12的缺失或仅有Yq12的缺失的Y染色体显微镜下不能区分,但后者不是精子发生障碍的病因。对男性不育精子发生障碍患者,要结合细胞遗传学和AZF分子检测综合判断。     

广州地区不育男性Y染色体无精子因子微缺失的筛查

目的探讨Y染色体无精子因子(azoospermia factor,AZF)区域微缺失与原发无精、严重少精症之间的关系。方法采用多重聚合酶链反应技术对广州地区103例原发无精子症、72例原发严重少精症患者及60名正常生育男性进行AZFa、AZFb、AZFc3个区域微缺失分析。结果60名正常生育男性未发现Y染色体AZF区域微缺失,175例生精障碍患者中发现AZF微缺失19例,总缺失率为10.9%。其中11例无精症患者和4例少精症患者的缺失发生在AZFc区域,缺失率为8.6%;1例无精症患者和2例少精症患者发生AZFb、AZFc双重缺失,缺失率为1.7%;1例无精症患者发生AZFa、b、c3个区域同时微缺失,缺失率0.6%。生精障碍组与正常生育男性组比较Y染色体AZF区域微缺失率差异具有统计学意义(P<0.01)。结论Y染色体AZF区域微缺失是引起男性无精、少精子症的重要原因之一,对原发无精、少精子症患者在单精子注射之前进行微缺失筛查是必要的。     

Y chromosome microdeletions: are they implicated in teratozoospermia?

BACKGROUND: Y chromosome microdeletions are known to impair spermatogenesis. Screenings for these microdeletions are performed mostly in patients with sperm count abnormalities. METHODS: We have screened the Y chromosome of 80 infertile patients with sperm morphological abnormalities. DNA from sperm, peripheral blood or single sperm following multiple displacement amplification (MDA) was utilized to amplify 20 specific sequence-tagged sites (STS) by PCR. RESULTS: Y chromosome microdeletions were detected in sperm DNA from four of the teratozoospermic patients; while none of the 53 men with normal sperm morphology had any deletions. Two of the four patients with deletions also provided peripheral blood and a fresh semen sample. Both patients had none of the STS deleted in the peripheral blood DNA. Y chromosome microdeletion analysis in the MDA amplified SRY-positive single sperm DNA confirmed the presence of the same deletion in all 10 sperm for one patient and eight out of 10 sperm in the second patient. CONCLUSIONS: Our observations suggest that some of the teratozoospermia might be related to gonadal mosaic Y chromosome microdeletions. Gonadal mosaicism can be a source of de novo transmissions of Y chromosome microdeletions. The application of MDA can yield enough DNA from a single sperm for genetic analyses.     

桂西地区壮族不育患者Y染色体无精子因子筛查

目的探讨桂西地区壮族不育患者Y染色体无精子因子(azoospermia factor,AZF)微缺失与原发无精子、严重少精子症之间的关系。方法采用多重聚合酶链反应技术对桂西地区52例原发无精子症、76例原发严重少精子症患者及40名正常生育男性进行4个区域15个位点微缺失分析。结果 40名正常生育男性未发现Y染色体AZF微缺失,128例生精障碍患者中发现AZF微缺失13例,总缺失率为10.2%。生精障碍组与正常生育男性组比较Y染色体AZF微缺失率差异具有统计学意义(P0.01)。结论 Y染色体AZF微缺失是男性无精子、少精子症的要重原因之一。     

208例男性不育遗传学病因分析研究

目的对208例男性不育患者进行遗传学分析,探讨染色体畸变、Y染色体微小缺失与男性不育的关系。方法采用外周血染色体核型分析技术对208例男性不育患者进行染色体核型分析,再利用改良多重PCR技术对染色体核型正常患者进行Y染色体AZF区域检测。结果 208例男性不育患者中,发现染色体异常者36例,占17.31%;Y染色体微小缺失者6例,占2.88%。结论染色体畸变及Y染色体微小缺失与男性不育关系紧密,染色体核型分析技术与Y染色体AZF检测技术相结合可提高男性不育患者的诊断水平与遗传异常检出率。     

Prognostic value of Y deletion analysis: what is the clinical prognostic value of Y chromosome microdeletion analysis?

In many centres, Y chromosome deletion analysis is still not performed routinely and if so, the results are used for genetic counselling but are not considered as having a useful prognostic value. The type of deletion (AZFa, b or c) has been proposed as a potential prognostic factor for sperm retrieval in men undergoing TESE. AZFc deletions and partial AZFb deletions are associated with sperm retrieval in approximately 50% of cases while in the case of a patient with complete AZFb deletion the probability of finding mature spermatozoa is virtually nil. Therefore the extent and position of a Y microdeletion is important (complete or partial). The prognostic value of Y chromosome deletion analysis in cases of oligozoospermia is important when one considers the progressive decrease of sperm number over time in men with AZFc deletions. Cryo-conservation of spermatozoa in these cases could avoid invasive techniques, such as TESE/ICSI, in the future. Male offspring that are conceived by ICSI or IVF techniques from father with oligozoospermia or azoospermia would also benefit from knowledge of their Y status, since the identification of the genetic defect will render future medical or surgical therapies unnecessary. Y microdeletion screening is therefore important, not only to define the aetiology of spermatogenic failure, but also because it gives precious information for a more appropriate clinical management of both the infertile male and his future male child.     

Prognostic value of Y deletion analysis. The role of current methods

Y chromosome microdeletions represent the most frequent genetic alteration in azoospermic and severely oligozoospermic men, and screening for microdeletions in AZFa, b and c are routinely performed in the major andrology and infertility centres. Since patients with Y microdeletions often require intracytoplasmic sperm injection (ICSI), the question of whether the type of the microdeletion present could have prognostic value for the presence of spermatozoa in the ejaculate or in the testes [by testicular sperm extraction (TESE)] is an interesting one. The review of the literature on this topic showed that there is still no clear genotype--phenotype relationship, i.e. similar testicular alterations may be caused by different types of microdeletions, and apparently identical microdeletions may be associated with diverse tubular damage. Even in azoospermic men, the localization of the microdeletion cannot be used as a valid prognostic parameter before TESE--ICSI to identify patients with spermatozoa in their testes. The only finding with absolute negative prognostic value is the presence of complete AZFa--c deletions, which are invariably associated with an absence of spermatozoa. Microdeletions in AZFa or AZFb seem to have promising prognostic value, but more data and gene-specific deletions have to be provided to draw clear conclusions. The absence of a clear genotype--phenotype relationship, and therefore of a prognostic value of Y deletion analysis, is probably due to the current methods used for the screening of the microdeletions. In fact, to date most centres do not use gene-specific markers but instead use anonymous primers that contribute little information to the pathogenic role of the microdeletions.