Genetic disorders and male infertility

Abstract Background At present, one out of six couples is infertile, and in 50% of cases, infertility is attributed to male infertility factors. Genetic abnormalities are found in 10%‐20% of patients showing severe spermatogenesis disorders, including non‐obstructive azoospermia. Methods Literatures covering the relationship between male infertility and genetic disorders or chromosomal abnormalities were studied and summarized. Main findings (Results) Genetic disorders, including Klinefelter syndrome, balanced reciprocal translocation, Robertsonian translocation, structural abnormalities in Y chromosome, XX male, azoospermic factor (AZF) deletions, and congenital bilateral absence of vas deferens were summarized and discussed from a practical point of view. Among them, understanding on AZF deletions significantly changed owing to advanced elucidation of their pathogenesis. Due to its technical progress, AZF deletion test can reveal their delicate variations and predict the condition of spermatogenesis. Thirty‐nine candidate genes possibly responsible for azoospermia have been identified in the last 10 years owing to the advances in genome sequencing technologies. Conclusion Genetic testing for chromosomes and AZF deletions should be examined in cases of severe oligozoospermia and azoospermia. Genetic counseling should be offered before and after genetic testing.


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KURODA et Al. on its causes and reveal their pathogenic details. In cases of non-obstructive azoospermia (NOA) and severe oligo-astheno-tetrazoospermia, chromosomal or genetic disorders were confirmed in 15%-20% of patients. 4 A recent review reported that genetic disorders could possibly explain at least some of these idiopathic cases. 5 In fact, in clinical practice, Y chromosome microdeletion analysis has become routine for patients with severe oligozoospermia and azoospermia. Y chromosome microdeletions indicate genomic deletions in the region of azoospermic factor (AZF) spreading on the Y chromosome. The deletion of AZF is currently the only predictor of spermatogenic condition and success rate of micro-testicular sperm extraction (micro-TESE) in patients with azoospermia. In addition, owing to recent progress in genome-analyzing technologies, especially in the last 10 years, studies have identified many genetic variations which associated with male infertility. 6 Next-generation sequencing technologies have made a particularly significant contribution to the search for candidate genes. 7,8 Moreover, not only genomic but also epigenetic mechanisms have been recently investigated. [9][10][11] Epigenetics regulates gene expression and genome stability without altering DNA sequence via reversible modifications of chromatin in either DNA or histones and, in some cases, both DNA and histones. 9 To date, genetic testing for chromosomal abnormalities and AZF deletions can provide important information to doctors and patients for decision making. However, no specific genes for any subgroup of "idiopathic" infertility have been identified and the exact relationships between genetics and impaired spermatogenesis remain mostly unclear. Nonetheless, the unveiling pathophysiology of male infertility through a genetic approach has a certain potential to contribute to an increased pregnancy rate in the era of artificial reproductive technology. In this review, we highlight literatures covering the relationship between male infertility and genetic disorders or chromosomal abnormalities.

| Chromosomal analysis
Chromosomal disorders are confirmed in 5% of patients with severe oligozoospermia and in 10%-15% of patients with azoospermia. 12,13 Usually, a lymphocyte culture (72 hours) is performed to analyze the chromosomes. In routine analysis, 20 cells are analyzed. In cases of chromosomal mosaicism or chromosomal abnormalities, 30 cells are analyzed. 14 Table 1 shows the chromosome abnormalities in individuals with male infertility. Klinefelter syndrome (KS) is the most common sex chromosome disorder responsible for male infertility. 15 Its karyotype has two or more X chromosomes in males; 47,XXY is the most common karyotype. Symptoms are typically more severe if three or more X chromosomes are present (48,XXXY or 49,XXXXY). 16 The prevalence of KS was reported to be approximately 1 in 1000 newborn males during the 1970s and 1980s. 17,18 In 1990, Danish registry studies described the prevalence of KS to be 153-173 in every 100 000 newborn males. 19 Recent studies reported that the prevalence was increasing, and 1 in 500-600 newborn males had KS. [20][21][22] Crawford stated that this change may be due to increasing awareness and optimization of diagnostic methods. 21 Semen analysis of non-mosaic KS patients usually shows azoospermia, while ejaculated spermatozoa are sometimes confirmed in patients with mosaic KS (46,XY/47,XXY). In cases of azoospermic KS, the sperm retrieval rate (SRR) with micro-TESE was reported to be between 40% and 70%, [23][24][25][26][27][28] which was higher than those in unexplained NOA patients, that were reported to be between 31% and 42.9%. 28,29 Chromosomal translocations are the most common structural disorders in men with a frequency of 1.23 per 1000, 30 and their prevalence is 10 times greater in the infertile population. 31 51  For the detection of AZFa, AZFb, and AZFc, PCR primers should at least include sY14(SRY), ZFX/ZFY, sY84, sY86, sY127, sY134, sY254, and sY255. 50 Attention must be paid to whether different primers were used by researchers for the diagnosis of AZF deletions.
The AZFa region spans 1100 kb and contains only two genes, USP9Y and DDX3Y. This region also contains retroviral sequences such as HERVyq1 and HERVyq2 that have been acquired in humans through the evolutionary process. Between these same directional retroviral sequences which are flanking AZFa, homologous recombination could occur resulting in the deletion of AZFa. [54][55][56] The AZFb region spans 6.2 Mb and contains 32 gene copies and transcription units including HSFY and RPS4Y2. 53 Again, based on the palindrome structure, the AZFb deletion is located between P5 and proximal P1, which is supposed to occur by homologous recombination between the palindromes. 57

| AZF-partial deletions
As the palindrome structures in the Y chromosome have been revealed, the existence of partial deletions in the AZF region was clarified by many studies. For instance, innovative diagnostic kits using precise sequence-tagged-site markers were developed and have provided new data on partial deletions in the AZF region. 69 However, the influence of these partial deletions on spermatogenesis is still unclear.
Thus, partial deletions are not the factors that definitively favor for or against the use of artificial reproductive technology or micro-TESE for these patients. Again, it should be remembered that the deletions will be inherited to their sons.
Gr/gr deletions that involve the removal of the 1. 1030 infertile males in Japan. However, they also stated that SRR in patients with gr/gr deletion was relatively lower than that in patients without the deletion (18.8% vs 28.7%, P = .09), although the difference was not statistically significant. Therefore, its clinical significance is still controversial.

TA B L E 2
The identified genes located in autosomes and X chromosome possibly implicated in male infertility Abbreviations: NOA, non-obstructive azoospermia; OAT, oligo-astheno-teratozoospermia; OMIM, online Mendelian inheritance in man.
The b2/b3 deletion removes 1.8 Mb of the AZFc section. The mechanism of b2/b3 deletion is complicated, the b2/b3 or gr/rg deletion is followed by a gr/rg or b2/b3 inversion. 79,80 Among the Chinese population, the association of b2/b3 partial deletion with male infertility was reported in 2009. 81 On the contrary, studies in other populations did not show any association with infertility. 82,83 Yuan et al reported the natural transmission of b2/b3 sub-deletion.
They performed Y microdeletion tests for each father of four infertile male patients with complete deletions of AZFc or AZFb+c. The b2/b3 sub-deletions were found in all fathers, though the fathers are not infertile, and the sons were all born through natural delivery. 84 The b1/b3 deletion removes 1.6 Mb of the AZFc region. This deletion was defined as the loss of sY1161, sY1191, and sY1291 with the presence of other sequence-tagged sites. The mechanism of b1/ b3 deletion involves homologous recombination, possibly between sister chromatids or within a chromatid. 85 Its frequency varies in previous reports. [86][87][88][89] Due to its low frequency, the effects of b1/b3 deletion on spermatogenesis remain unclear. 80

| Congenital bilateral absence of vas deferens
Congenital bilateral absence of vas deferens (CBAVD) is one of the causes of obstructive azoospermia. It is sometimes observed as a symptom of cystic fibrosis, a genetic condition causing exocrine gland disorders. Cystic fibrosis and isolated CBAVD are autosomal recessive and are recognized as cystic fibrosis transmembrane conductance regulator (CFTR)-related diseases. 90 Yu et al 91 reported in their meta-analysis that 78% of patients with CBAVD had at least one CFTR mutation, and the 5T allele and 5T/(TG)12_13 may contribute to the increased risk of CBAVD. In Japan, due to the very low frequency of CFTR mutation, commercial-based tests for this mutation are not available.

| New candidate genes in male infertility
As described above, the AZF region of the Y chromosome contains genes that affect spermatogenesis. Genome-wide association studies during the past ten years  have brought significant improvement in genetic analysis techniques and many autosomal and X-chromosomal genes have been reported to be possibility associated with spermatogenetic disorders. Representative candidate genes reported to be associated with oligozoospermia, asthenozoospermia, and azoospermia are listed in Table 2

| CON CLUS I ON S AND FUTURE PER S PEC TIVE S
Chromosomal analysis and testing for AZF deletions should be performed in cases of severe oligozoospermia and azoospermia.
Especially in cases of azoospermia, these examinations are mandatory to consider the indication for micro-TESE. Except AZF deletions, there are no other currently available genetic markers for male infertility or for predicting the success rate of sperm retrieval in azoospermic patients, although many candidate genes that may be responsible for azoospermia have been identified over the last 10 years. Although we should recognize multifactorial aspects and genetic heterogeneity of male infertility, the potential to better define male infertility may increase in the next decade due to the advances in next-generation sequencing. Genetic counseling should be offered in pre-and post-chromosome and genetic mutation analysis.

Conflict of interest:
The authors report no conflicts of interest.
Human/Animal rights statement: This article does not contain any studies with human or animal subjects.