Klinefelter syndrome (KS), marked by testicular heterogeneity, provides a rationale for testicular sperm extraction in azoospermic men, yet large-scale evaluations of predictive and reproductive outcomes remain limited. This retrospective study included 79 karyotype-confirmed KS patients who underwent conventional testicular sperm extraction (cTESE) and microdissection testicular sperm extraction (micro-TESE) between February 2001 and August 2024. Clinical, hormonal, histological, embryological, and reproductive data were analyzed, with successful sperm retrieval (SR) defined as the presence of at least one spermatozoon. SR was achieved in 40.5% of patients and was significantly associated with larger testicular volumes (left: P = 0.018; right: P = 0.028), lower follicle-stimulating hormone (mean ± standard deviation [s.d.]: 29.06 ± 14.49 IU l -1vs 38.22 ± 16.97 IU l -1 ; P = 0.016), and lower luteinizing hormone levels (mean ± s.d.: 16.41 ± 7.46 IU l -1vs 20.65 ± 9.59 IU l -1 ; P = 0.047). These parameters also correlated with the number of frozen sperm vials. Among those with SR, 46 embryo transfer cycles were conducted, involving 79 embryos. Fertilization, cleavage, and blastulation rates were 50.3%, 80.7%, and 40.2%, respectively. Good-quality embryos were observed in 45.1% on day 3 and 55.0% on day 5, with clinical pregnancy and live birth rates of 39.1% and 39.5% per transfer. These findings indicate that favorable hormonal profiles and testicular characteristics are predictive of SR success and that reproductive outcomes among KS patients with sperm retrieval are comparable to those in non-KS populations. Overall, the results support the continued use of testicular sperm extraction as a viable path to genetic parenthood in this population.

BACKGROUND:Cryopreservation is widely used in assisted reproductive technologies. While fresh sperm undergoes gradual time-dependent deterioration, it remains unclear whether thawed sperm exhibits a more accelerated decline. OBJECTIVES:To directly compare the rate of deterioration in sperm motility, vitality, and DNA fragmentation between fresh washed and thawed sperm samples over time. MATERIALS AND METHODS:This prospective study included semen samples from 50 males. Samples were split into two groups: Washed (freshly washed sperm) and Thawed (washed, cryopreserved for at least 2 weeks, and thawed). Sperm parameters, including motility, vitality, and DNA fragmentation index (DFI), were assessed immediately after processing (Time 1) and again after 75 min incubation at room temperature (Time 2). Additionally, control experiments tested whether cryoprotectant exposure alone could account for deterioration by comparing washed samples incubated with washing versus freezing medium (10 patient samples), and by assessing post-thaw washing (10 donor samples). RESULTS:Total sperm motility declined significantly more in thawed samples (29 ± 16%) compared to fresh washed samples (17 ± 9%, p < 0.0001). Vitality similarly deteriorated more in thawed samples (21 ± 14%) versus fresh washed samples (7 ± 5%, p < 0.0001). DNA fragmentation increased significantly only in thawed samples (p = 0.0331), reaching clinically critical levels (mean DFI 34 ± 13% at Time 2), compared to fresh samples which remained within normal range (12 ± 5%). Motility grade transitions differed markedly, with thawed samples showing direct transitions from Grade A motility to immotility, unlike fresh washed samples, which transitioned gradually from Grade A to Grade B. In additional control experiments, cryoprotectant exposure alone did not induce deterioration, and post-thaw washing did not improve metrics. DISCUSSION:Thawed sperm exhibited accelerated deterioration across all measured parameters, highlighting cumulative stress from cryopreservation. The rapid decline underscores the need to minimize the interval between thawing and insemination. CONCLUSION:Thawed spermatozoa demonstrate significantly greater susceptibility to time-dependent deterioration compared to fresh washed samples, advocating for immediate use post-thaw to optimize reproductive outcomes.

Changing sperm donors after unsuccessful intrauterine insemination (IUI) cycles is a common yet understudied practice. This study evaluates whether switching sperm donors impacts the number of IUI cycles required to achieve pregnancy. This retrospective cohort study analyzed 312 women undergoing donor sperm IUI at Lis Maternity Hospital, Tel Aviv Sourasky Medical Center, from 1992 to 2020. Participants were divided into two groups: Group A (conceived using only one donor) and Group B (switched donors after initial unsuccessful attempts). The primary outcome was the number of IUI cycles until pregnancy. Statistical analyses included t-tests, ANOVA, and multivariate analysis of covariance (MANCOVA). Women in Group A required fewer cycles (mean 3.78 ± 1.90) to achieve pregnancy compared to Group B (mean 6.07 ± 2.95, P < .001). However, after switching donors, the mean number of cycles needed in Group B (2.23 ± 1.61) was significantly lower than the total cycles required by Group A (P < .001). Cumulative live birth rates were higher in Group A (50.5

What factors predict successful microsurgical sperm extraction in azoospermic Klinefelter Syndrome (KS) patients, and what are their chances of achieving parenthood? Age, follicle-stimulating hormone and testosterone influence the sperm-retrieval rate (SRR). Once retrieved, sperm achieved pregnancy and live birth rates comparable to or exceeding non-KS patients. Testicular heterogeneity, a hallmark of KS, supports the use of microsurgical sperm extraction for azoospermic patients seeking fertility treatment. When feasible, early fertility preservation is crucial. Despite its clinical significance, there have been only few large-scale studies evaluating predictive factors for successful sperm-retrieval (SR) in microsurgical sperm extraction procedures of KS patients, and even fewer have assessed embryological and obstetrical outcomes. This retrospective study included 79 KS patients, who underwent microsurgical sperm extraction at a single university-affiliated medical center between January 2001 and June 2024. Patients' data were collected from electronic medical records and telephone questionnaires assessing demographic, histological, embryological and obstetrical outcomes. Patients with successful and unsuccessful sperm retrieval were compared, and available embryological and obstetrical data are presented. All patients were diagnosed with KS via karyotype after infertility referral. Sperm was retrieved using conventional testicular sperm extraction (cTESE) or microsurgical testicular sperm extraction (mTESE), with positive-SR defined as the retrieval of at least one spermatozoon. Retrieved sperm was frozen and later thawed for use in intracytoplasmic sperm injection (ICSI) with oocyte from the female partner. Group comparisons were conducted using Pearson’s χ2, Student’s T-test or Mann-Whitney U test, with α = 0.05. The mean patient age was 32.18 ± 6.19 years. Sperm retrieval was successful in 32 KS patients (Sperm retrieval rate, SRR = 41%). Success was associated with larger left testicular volume (2 [1.5-2.5] vs. 1.5 [1.5-2] cm, p = 0.038), a higher prevalence of mixed atrophy histology (65.6% vs. 12.8%, p < 0.001), and lower luteinizing hormone (LH) levels (16.41 ± 7.46 vs. 20.65 ± 9.59 IU/L, p = 0.047). Subgroup analysis demonstrated optimal SRRs in patients younger than 31 years (48.7%), follicle-stimulating hormone (FSH) <27.8 IU/L (51.6%) and testosterone >11 nmol/L (45.2%). A mean of 9.13 ± 5.68 sperm straws were frozen, and this negatively correlated with both age (r=-0.338, p = 0.05) and FSH (r=-0.343, p = 0.05). Seventy-nine embryo transfers were performed. Embryological and obstetrical outcomes were comparable to rates reported in the literature, with implantation rates of 52.25% ± 38.99%, and clinical pregnancy rates of 39.11% ± 28.64%. Notably, live birth rates of 39.57% ± 28.43%, exceeded rates reported for non-KS patients undergoing ICSI. Potential recall bias may arise from the retrospective design and reliance on telephone questionnaires. The small sample size and single-center data limit generalizability. Descriptive reporting of embryological and obstetrical outcomes prevents definitive conclusions about their broader significance. We provided valuable insights into microsurgical SR in KS patients, highlighting rarely reported obstetrical outcomes. Higher SRRs linked with younger age, lower FSH, and higher testosterone. Implantation, pregnancy, and live birth rates matched or exceeded reported rates of non-KS patients, underscoring the intact potential of retrieved sperm in these cases. No

Study question Does thawed sperm deteriorate more rapidly than washed sperm over time? Summary answer Thawed sperm deteriorates more rapidly than washed sperm over time, with significant declines in motility and vitality and a greater increase in DNA fragmentation. What is known already Intrauterine insemination (IUI) treatments are performed using either washed sperm from a partner or thawed donor sperm. While most procedures occur on-site, some patients opt for off-site insemination. In such cases, thawed sperm samples are prepared in the laboratory and transported to the physician for the procedure. It is known that human spermatozoa is effected by time. However the magnitude of decline over time in thawed human spermatozoa compared to fresh sperm remains unknown. This raises the question regarding the effect of time on different parameters of thawed sperm, compared to fresh washed sperm. Study design, size, duration This prospective study was conducted between 2021-2024 and analyzed sperm samples from 50 men referred for semen analysis. Participants/materials, setting, methods Sperm samples from 50 men were collected, following, each sample sperm was subdivided and analyzed in two comparable ways to answer the main study question. Samples were liquefied for 20 minutes at 37 °C and assessed for motility, vitality, and DNA fragmentation (DFI). Each ejaculate was subdivided into two groups: (A) washing only, (B) washed, frozen, and thawed. Sperm quality was evaluated immediately after preparation (Time 1) and again 75 minutes later (Time 2). Main results and the role of chance Time significantly impacted sperm quality, with thawed sperm showing greater deterioration than washed sperm. Motility declined by 16.87% ± 8.58 in washed sperm versus 28.7% ± 15.55 in thawed sperm (P < 0.001). Vitality dropped by 7.04% ± 5.47 in washed samples and 21.02% ± 13.73 in thawed samples (P < 0.001). DFI increased significantly in both groups (P < 0.0001), with -60.62% ± 65.54 in washed samples and -65.16% ± 57.46 in thawed samples. Notably, thawed samples showed a steeper and more detrimental rise in DFI compared to washed samples (P = 0.0221), emphasizing the adverse impact of thawing on DNA integrity. Further analysis of motility grades revealed that the reduction in grade A motility was similar between the two groups, averaging approximately 32% (P = 0.3). However, washed sperm primarily transitioned from Grade A to Grade B, while thawed sperm showed a sharper decline, shifting to Grade C. Limitations, reasons for caution This study was limited by a single measurement (75 minutes), further research should explore additional time intervals. Additionally, while sperm quality is a viable and important index, it does not fully attest to clinical testing such as pregnancy outcome which should be incorporated in future investigations. Wider implications of the findings Thawed sperm deteriorates more rapidly than washed sperm, emphasizing the importance of minimizing delays between thawing and intrauterine insemination (IUI). These findings may inform clinical guidelines for sperm handling to optimize fertility treatment success. Trial registration number No

Quantitative and qualitative spermatogenic impairments are major causes of men's infertility. Although in vitro fertilization (IVF) is effec-tive, some couples persistently fail to conceive. To identify causal variants in patients with severe male infertility factor and repeated IVF failures, we sequenced the exome of two consanguineous family members who underwent several failed IVF cycles and were diagnosed with low sperm count and motility. We identified a rare homozygous nonsense mutation in a previously uncharacterized gene, RNF212B, as the causative variant. Recurrence was identified in another unrelated, infertile patient who also faced repeated failed IVF treatments. scRNA-seq demonstrated meiosis-specific expression of RNF212B. Sequence analysis located a protein domain known to be associated with aneuploidy, which can explain multiple IVF failures. Accordingly, FISH analysis revealed a high aneuploidy rate in the patients' sperm cells and their IVF embryos. Finally, inactivation of the Drosophila orthologs significantly reduced male fertility. Given that members of the evolutionary conserved RNF212 gene family are involved in meiotic recombination and crossover matura-tion, our findings indicate a critical role of RNF212B in meiosis, genome stability, and in human fertility. Since recombination is completely absent in Drosophila males, our findings may indicate an additional unrelated role for the RNF212-like paralogs in spermato-genesis.

MicroRNAs are involved in the regulation of spermatogenesis, are detected in semen and may be useful as molecular markers for predicting residual complete spermatogenesis in azoospermic men.

Klinefelter syndrome (KS) is the most prevalent genetic disorder of infertile males. This study aimed to determine in Klinefelter patients (KS) the expression levels of spermatogenic markers and testicular growth factors that might predict spermatogenesis based on conventional testicular sperm extraction (TESE). The expression levels of the pre-meiotic (OCT4, CD9, GFR-α1, α-6-INTEGRIN, SALL4, C-KIT), meiotic (CREM-1), and post-meiotic (protamine) markers, as well as the colony stimulating factor-1 (CSF-1) were examined in testicular biopsies with and without mature sperm of KS and normal karyotype of azoospermic patients (AZO) with complete spermatogenesis. In the biopsies of AZO, the expression levels (fold of expression compared to the PPI of the same sample) of OCT4 were 9.68± 7.93, CREM 42.78± 28.22, CSF-1 3.07 ± 3.19, and protamine 78498.12 ± 73214.40. Biopsies from KS included 7 with sperm and 17 without sperm. Among the biopsies with sperm, the expression levels of OCT4 were 7.27± 9.29, CREM 3.13± 7.89, CSF-1 35.5 ± 48.01, and protamine 902.97 ± 2365.92. In 14 biopsies without sperm, we found low expression levels of OCT4, CREM and CSF-1, and no expression of protamine. However, in three of the biopsies without sperm that highly expressed OCT4 and CSF-1, the expression levels of CREM-1 and protamine were high. These results may be used for further consulting with patients considering repeating conventional TESE or micro TESE and cryopreservation for possible future in-vitro spermatogenesis.

BACKGROUND:Data on who among the infertile male population may benefit from round spermatid injections (ROSI) are lacking.OBJECTIVE:To determine the probability of finding round spermatids suitable for ROSI in men with non-obstructive azoospermia (NOA) in whom no spermatozoa were retrieved at testicular sperm extraction.MATERIALS AND METHODS:Four-hundred fifty-seven consecutive men with azoospermia underwent testicular sperm extraction. Clinical examination included age, secondary sexual characteristics, testicular size, reproductive hormone estimation, karyotyping, and Y chromosome microdeletion analyses. Histologic examination was performed, and histologic classification was determined by the most advanced spermatogenetic cell identified in the combined histologic and cytologic examination.RESULTS:Of the 457 azoospermic men, 342 were diagnosed with NOA, and 148 (148/342, 43%) had mixed atrophy on histopathology and retrievable spermatozoa. No spermatozoa were found in 194/342 men with NOA (57%). Histopathology diagnosed 145/194 (75%) of them with Sertoli cell only, 45/194 (23%) with spermatocyte maturation arrest, and 4/194 (2%) with spermatid maturation arrest.CONCLUSIONS:Histopathologically identified round spermatids without spermatozoa were rare in men with NOA. Only very few of them are likely to reap the benefits of ROSI, thus presenting the need to reconsider its actual clinical value.

Infertility affects one in six couples, half of which are caused by a male factor. Male infertility can be caused by both, qualitative and quantitative defects, leading to Oligo- astheno-terato-zoospermia (OAT; impairment in ejaculate sperm cell concentration, motility and morphology). Azoospermia defined as complete absence of sperm cells in the ejaculation. While hundreds of genes are involved in spermatogenesis the genetic etiology of men’s infertility remains incomplete.We identified a hemizygous stop gain pathogenic variation (PV) in the X-linked Germ Cell Nuclear Acidic Peptidase (GCNA), in an Azoospermic patient by exome sequencing. Assessment of the prevalence of pathogenic variations in this gene in infertile males by exome sequence data of 11 additional unrelated patients identified a probable hemizygous causative missense PV in GCNA in a severe OAT patient. Expression of GCNA in the patients’ testes biopsies and the stage of spermatogonial developmental arrest were determined by immunofluorescence and immunohistochemistry. The Azoospermic patient presented spermatogenic maturation arrest with an almost complete absence of early and late primary spermatocytes and thus the complete absence of sperm. GCNA is critical for genome integrity and its loss results in genomic instability and infertility in Drosophila, C. elegans, zebrafish, and mouse. PVs in GCNA appear to be incompatible with male fertility in humans as well: A stop-gain PV caused Azoospermia and a missense PV caused severe OAT with very low fertilization rates and no pregnancy in numerous IVF treatments.

OBJECTIVE:To investigate the prevalence of testicular microlithiasis and its association with sperm retrieval rates and histopathology in men with non-obstructive azoospermia.METHODS:A total of 120 men underwent scrotal ultrasonography prior to microsurgical testicular sperm extraction. Sperm retrieval rate, testicular histopathology, testicular size, reproductive hormones, karyotyping, Y chromosome microdeletion analyses, and presence of varicoceles and hydroceles were compared between men with and without testicular microlithiasis.RESULTS:The total sperm retrieval rate was 40%. Ten men with normal spermatogenesis were excluded. The remaining 110 men with non-obstructive azoospermia were analyzed and testicular microlithiasis was detected in 16 of them (14.5%). The sperm retrieval rate in that subgroup was only 6.2% (1/16) as opposed to 39.4% (37/94) in men with non-obstructive azoospermia and no evidence of microlithiasis (P = 0.009). The mean right and left testicular diameters were significantly lower in the microlithiasis group (P = 0.04). On multivariate logistic regression analysis, the presence of mictolithiasis (odds ratio 7.4, 95% confidence interval 2.3, 12.2; P = 0.01) was the only independent predictor of unsuccessful sperm retrieval. The 15 patients with microlithiasis and without successful sperm extraction were diagnosed by histopathology as having Sertoli cells only. The 16th patient with successful sperm retrieval had a histopathology of mixed atrophy and was diagnosed with Klinefelter syndrome.CONCLUSION:The presence of testicular microlithiasis is associated with low sperm retrieval rates among our cohort of men with non-obstructive azoospermia undergoing scrotal ultrasonography prior to microsurgical testicular sperm extraction. Larger, prospective studies should be conducted to confirm these findings.

Refreezing of sperm samples would provide the possibility of performing more cycles of fertility treatments. Although the effect of repeated cycles of freezing on sperm quality was studied, the effect of the length of the time interval between each freeze-thaw cycle has not been reported. Hence, we assessed the effect of incubation time on the sperm quality of thawed sperm after repeated freezing. One-hundred samples of potential sperm donations with normal sperm quality were evaluated. The fresh semen samples were analyzed and cryopreserved in liquid nitrogen until use. After thawing, the samples were divided randomly to two groups and reanalyzed for motility, vitality, and DNA fragmentation. They were incubated at room temperature and reanalyzed after either 90 min (group A) or 180 min (group B) of incubation, and once again after a repeated cycle of freezing and thawing. Our results showed that the sperm parameters of fresh samples of both groups were similar. After one freeze-thaw cycle, both groups still had comparable values. At the end of their respective incubation time periods, however, there was a significant difference in the mean values of the assessed parameters between the two groups (p < 0.01). An additional freeze-thaw cycle further exacerbated those differences, with group B undergoing an even more substantial decline (p < 0.001). Our data suggest that thawed human spermatozoa sustain a significant decline in sperm parameters in association with longer incubation time, which is further exacerbated by an additional freeze-thaw cycle.

STUDY QUESTION:Are there genetic variants that can be used for the clinical evaluation of azoospermic men?SUMMARY ANSWER:A novel homozygous frame-shift mutation in the MEIOB gene was identified in three azoospermic patients from two different families.WHAT IS KNOWN ALREADY:Up to 1% of all men have complete absence of sperm in the semen, a condition known as azoospermia. There are very few tools for determining the etiology of azoospermia and the likelihood of sperm cells in the testis. The MEIOB gene codes for a single-strand DNA binding protein required for DNA double-strand breaks repair during meiosis. MEIOB appears to be exclusively expressed in human and mouse testis, and MeioB knockout mice are azoospermic due to meiotic arrest.STUDY DESIGN, SIZE, DURATION:Two brothers with non-obstructive azoospermia (NOA) underwent whole-exome sequencing followed by comprehensive bioinformatics analyses. Candidate variations were further screened in infertile and fertile men, as well as in public and local reference databases.PARTICIPANTS/MATERIALS, SETTING, METHODS:This study included 159 infertile and 77 fertile men. The exomes of two Arab men were completely sequenced. In addition, 213 other men of the same Arab ethnicity (136 infertile and 77 fertile men) underwent restriction fragment length polymorphism (RFLP) screening, as did 21 NOA men, of other ethnicities, with testicular impairment of spermatocyte arrest. All of the infertile men underwent Y-chromosome microdeletion and CFTR gene mutation assessments. Comprehensive bioinformatics analyses were designed to uncover candidate mutations associated with azoospermia.MAIN RESULTS AND THE ROLE OF CHANCE:A novel homozygous frame-shift mutation in the MEIOB gene was identified in two brothers of Arab ethnicity. This frame-shift is predicted to result in a truncated MEIOB protein, which lacks the conserved C-terminal DNA binding domain. RFLP screening of the mutation in 157 infertile men, including 112 NOA patients of Arab ethnicity, identified an additional unrelated NOA patient with the same homozygous mutation and a similar testicular impairment. This mutation was not found in available public databases (n > 160 000), nor in the 77 proven fertile men, nor in our database of local Israeli population variations derived from exome and genome sequencing data (n = 500).LIMITATIONS, REASONS FOR CAUTION:We have thus far screened for only two specific MEIOB probable pathogenic mutations in a relatively small local cohort. Therefore, the relative incidence of MEIOB mutations in azoospermia should be further assessed in larger and diverse cohorts in order to determine the efficiency of MEIOB sequence screening for clinical evaluations.WIDER IMPLICATIONS OF THE FINDINGS:The relatively high incidence of likely NOA-causing mutations in MEIOB that was found in our cohort supports the idea that a complete screening of this gene might be beneficial for clinical evaluation of NOA patients.STUDY FUNDING/COMPETING INTEREST(S):This research was supported in part by a grant to EA from the European Research Council under the European Union's Seventh Framework Programme (FP/2007-2013)/ERC grant agreement (616088). There are no competing interests.TRIAL REGISTRATION NUMBER:N/A.

The study is aimed to describe a novel strategy that increases the accuracy and reliability of PGD in patients using sperm donation by pre-selecting the donor whose haplotype does not overlap the carrier’s one.

Purpose: Up to 1% of all men experience azoospermia, a condition of complete absence of sperm in the semen. The mechanisms and genes involved in spermatogenesis are mainly studied in model organisms, and their relevance to humans is unclear because human genetic studies are very scarce. Our objective was to uncover novel human mutations and genes causing azoospermia due to testicular meiotic maturation arrest. Methods: Affected and unaffected siblings from three families were subjected to whole-exome or whole-genome sequencing, followed by comprehensive bioinformatics analyses to identify mutations suspected to cause azoospermia. These likely mutations were further screened in azoospermic and normozoospermic men and in men proven to be fertile, as well as in a reference database of local populations. Results: We identified three novel likely causative mutations of azoospermia in three genes: MEIOB , TEX14 , and DNAH6 . These genes are associated with different meiotic processes: meiotic crossovers, daughter cell abscission, and possibly rapid prophase movements. Conclusion: The genes and pathways we identified are fundamental for delineating common causes of azoospermia originating in mutations affecting diverse meiotic processes and have great potential for accelerating approaches to diagnose, treat, and prevent infertility. Genet Med advance online publication 16 February 2017

The bromodomain testis-specific (BRDT) protein belongs to the bromodomain extra-terminal (BET) family of proteins. It serves as a transcriptional regulator of gene expression during spermatogenesis, and is an essential factor for the normal spermatogenesis process. In this study, we characterized mice of several age groups who lacked the Brdt gene. The testes of Brdt mutant mice aged 8 weeks exhibited complete spermatocyte maturation arrest with a significantly increased number of apoptotic cells. The weights of the testes and accessory glands as well as the testosterone levels of the mutant mice were significantly lower compared to the normal mice. The mutant mice had delayed puberty, with normal levels of testosterone and accessory gland weights at the age of 14 and 28 weeks. The testes of the mutant mice at older ages also exhibited round spermatids. The presence of the BRDT protein was identified in the mice pituitary gland. Microarray analysis of mice pituitaries showed that 28 genes were down-regulated while 26 genes were up-regulated in the absence of the Brdt gene. Our results suggest that in addition to its critical role in the spermatogenesis process, the BRDT protein is also responsible for scheduling male puberty by regulation of the pituitary-gonad axis.

Purpose Mature sperm cells can be found in testicular specimens extracted from azoospermic men with non-mosaic Klinefelter syndrome (KS). The present study evaluates the expression of various known molecular markers of spermatogenesis in a population of men with KS and assesses the ability of those markers to predict spermatogenesis. Methods Two groups of men with non-obstructive azoospermia who underwent testicular sperm-retrieval procedures were included in the study: 31 had non-mosaic KS (KS group) and 91 had normal karyotype (NK group). Each group was subdivided into mixed atrophy (containing some mature sperm cells) or Sertoli cell only syndrome according to testicular histology and cytology observations. Semi-quantitative histological morphometric analysis (interstitial hyperplasia and hyalinization, tubules with cells and abnormal thickness of the basement membrane) and expression of spermatogenetic markers (DAZ , RBM , BOLL , and CDY1) were evaluated and compared among those subgroups. Results Clear differences in the histological morphometry and spermatogenetic marker expression were noted between the KS and NK groups. There was a significant difference in the expression of spermatogenetic markers between the subgroups of the NK group (as expected), while no difference could be discerned between the two subgroups in the KS group. Conclusion We conclude that molecular spermatogenetic markers have a pattern of expression in men with KS that is distinctively different from that of men with NK, and that it precludes and limits their use for predicting spermatogenesis in the former. It is suggested that this difference might be due to the specific highly abnormal histological morphometric parameters in KS specimens.

Bromodomain testis-specific (BRDT) protein is essential for the normal process of spermatogenesis. Mutant mice that expressed truncated BRDT had impaired testicular histology with severely reduced sperm concentration and abnormal sperm morphology, while a model of knockout Brdt mice with no BRDT protein had complete meiotic arrest. A BRDT single nucleotide polymorphism (SNP) (rs3088232) was reported as being associated with infertility in men. We assessed testicular specimens of 276 azoospermic men who underwent testicular sperm extraction to search for specimens that showed spermatogenic impairments similar to those of mutant BRDT mice. Ten similar specimens were selected for BRDT gene sequencing and they revealed three NCBI-reported SNPs (rs10783071, rs3088232 and rs10747493) variously distributed among them. Bioinformatics analysis predicted that they would not affect protein activity. Further assessment of rs3088232 frequency in a large group of non-obstructive azoospermia men and fertile controls demonstrated no significant difference between them (27.2 and 21.7% respectively; p = 0.122, Fisher's exact test). We conclude that the testicular impairments observed in the 10 specimens were not a consequence of BRDT gene mutation. The association between BRDT rs3088232 and infertility that had been reported in other studies was not supported.