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A male factor, commonly associated with poor semen quality, is revealed in about 50% of infertile couples. CFTR gene (Cystic Fibrosis Transmembrane Conduction Regulator) variants are one of the common genetic causes of azoospermia-related male infertility. Notably, the spectrum and frequency of pathogenic CFTR variants vary between populations and geographical regions. In this work, we made an attempt to evaluate the allele frequency (AF) of 12 common CFTR variants in infertile Russian men and healthy individuals from different districts of Russia. Because of the limited number of population-based studies on Russian individuals, we characterized the population AFs based on data from the Registry of Russian cystic fibrosis (CF) patients. In addition to the CF patient registry, we estimated the local frequencies of the same set of variants based on the results of genotyping of CF patients in local biocollections (from St. Petersburg and Yugra regions). AFs of common CFTR variants calculated based on registry and biocollection data showed good concordance with directly measured population AFs. The estimated region-specific frequencies of CFTR variants allowed us to uncover statistically significant regional differences in the frequencies of the F508del (c.1521_1523del; p.Phe508del) and CFTRdele2,3(21kb) (c.54-5940_273+10250del21kb; p.Ser18ArgfsX) variants. The data from population-based studies confirmed previous observations that F508del, CFTRdele2,3(21kb), and L138ins (c.413_415dup; p.Leu138dup)variants are the most abundant among infertile patients, and their frequencies are significantly lower in healthy individuals and should be taken into account during genetic monitoring of the reproductive health of Russian individuals.

We examined a cohort of 93 cystic fibrosis (CF) male patients who were pancreatic-sufficient (PS-CF; n=40) or pancreatic-insufficient (PI-CF; n = 53). Complex semen examination was performed, including standard semen analysis, quantitative karyological analysis (QKA) of immature germ cells (IGCs), transmission electronic microscopy (TEM), biochemical analysis, and sperm DNA fragmentation by terminal deoxynucleotidyl transferase-mediated dUTP nickend labeling (TUNEL) assay. Azoospermia was diagnosed in 83 (89.2%) patients. The other 10 (10.8%) patients were found to be nonazoospermic and showed various spermatological diagnoses (asthenozoospermia, n = 2; asthenoteratozoospermia, n = 3; oligoasthenozoospermia, n = 1; oligoasthenoteratozoospermia, n = 3; and normozoospermia, n = 1) with no specific morphological abnormalities. Oligospermia was detected in 89.2% azoospermic and 30.0% nonazoospermic patients. Low seminal pH (<7.0) was found in 74 (89.2%) of 83 azoospermic patients. Moderate leukocytospermia (2.0 × 10 6 -2.2 × 10 6 ml -1 ) was revealed in 2.4% azoospermic and 40.0% nonazoospermic semen samples. The signs of partial meiotic arrest at prophase I were found in 4 of 6 nonazoospermic patients examined by QKA of IGCs. The content of fructose and citrate was low in oligospermic and normal in nonoligospermic semen samples. An increased percentage (>30%) of spermatozoa with noncondensed ("immature") chromatin was revealed in 2 of 6 nonazoospermic semen samples analyzed by TEM., (Copyright © 2023 Copyright: © The Author(s)(2023).)

Various pathogenic factors can lead to oogenesis failure and seriously affect both female reproductive health and fertility. Genetic factors play an important role in folliculogenesis and oocyte maturation but still need to be clarified. Oocyte maturation is a well-organized complex process, regulated by a large number of genes. Pathogenic variants in these genes as well as aneuploidy, defects in mitochondrial genome, and other genetic and epigenetic factors can result in unexplained infertility, early pregnancy loss, and recurrent failures of IVF/ICSI programs due to poor ovarian response to stimulation, oocyte maturation arrest, poor gamete quality, fertilization failure, or early embryonic developmental arrest. In this paper, we review the main genes, as well as provide a description of the defects in the mitochondrial genome, associated with female infertility.

Objective: Assessment of leukocyte activity as the main source of ROS in seminal fluid of patients with normospermia and pathospermia using an original protocol based on the kinetic chemiluminescence method and adapted for semen analysis., Materials and Methods: A prospective study was attended by 95 men of reproductive age who applied to the Research Center of Medical Genetics (Moscow) for semen analysis. The material for the study were samples of native ejaculate. Chemiluminescent measurements were performed on a Lum-1200 chemiluminometer (DISoft, Russia) using the original method., Results: Both in amplitude of basal and stimulated response, between the "normozoospermia", "pathozoospermia" and "pathozoospermia + leucospermia" groups, significant differences were obtained in the level of ROS production by leukocytes (p<0.05): the median level of basal chemiluminescence, normalized on the count of leukocytes was 0.13, 0.71 and 1.78, respectively; the median level of stimulated chemiluminescence normalized to the number of leukocytes was 0.62, 2.14, 5.94, respectively. The level of stimulated response did not exceed 0.5 arb. units in normozoospermic samples. In pathospermic groups, the level of stimulated response was low in about a third of semen samples, it was moderate in one third of patients, and high in one third of patients., Conclusions: A protocol previously developed for blood analysis was adapted to analyze the total ROS level produced by leukocytes in seminal fluid. In the groups of "pathozoospermia", "pathozoospermia + leucospermia", the level of basal ROS production by leukocytes was about 5 and 15 times higher than in the "normozoospermia" group, the level of stimulated ROS production was 3.5 and 9.5 times; this indicates oxidative stress, including with a normal number of leukocytes.

Only a few studies have described sperm chromosome intranuclear positioning changes in men with reproductive failure and an incorrect somatic karyotype. We studied the influence of Robertsonian translocations on the acrocentric chromosome positioning in human sperm cells. The basis of the analysis was the localization of NORs (nucleolar organizing regions) in sperm nuclei from three Robertsonian translocation carriers, namely, rob(13;22), rob(13;15) and rob(13;14), with a known meiotic segregation pattern. All three carriers presented with a similar percentage of genetically normal sperm cells (i.e., approximately 40%). To visualize NORs, we performed 2D-FISH with directly labelled probes. We used the linear and radial topologies of the nucleus to analyse the NORs distribution. We found an affected positioning of NORs in each case of the Robertsonian translocations. Moreover, the NORs tended to group, most often in two clusters. Both in Robertsonian carriers and control sperm cells, NORs mostly colocalized in the medial areas of the nuclei. In the case of the Roberstonian carriers, NORs were mostly concentrated in the peripheral part of the medial area, in contrast to control sperm cells in which the distribution was more dispersed towards the internal area.

Testicular microlithiasis (TM) is one of the symptoms of testicular dysgenesis syndrome (TDS). TM is particularly interesting as an informative marker of testicular germ cell tumors (TGCTs). KIT ligand gene (KITLG), BCL2 antagonist/killer 1 (BAK1), and sprouty RTK signaling antagonist 4 (SPRY4) genes are associated with a high risk of TGCTs, whereas bone morphogenetic protein 7 gene (BMP7), transforming growth factor beta receptor 3 gene (TGFBR3), and homeobox D cluster genes (HOXD) are related to TDS. Using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis, we investigated allele and genotype frequencies for KITLG (rs995030, rs1508595), SPRY4 (rs4624820, rs6897876), BAK1 (rs210138), BMP7 (rs388286), TGFBR3 (rs12082710), and HOXD (rs17198432) in 142 TGCT patients, 137 TM patients, and 153 fertile men (control group). We found significant differences in the KITLG GG&#95;rs995030 genotype in TM (P = 0.01) and TGCT patients (P = 0.0005) compared with the control. We also revealed strong associations between KITLG&#95;rs1508595 and TM (G allele, P = 0.003; GG genotype, P = 0.01) and between KITLG&#95;rs1508595 and TGCTs (G allele, P = 0.0001; GG genotype, P = 0.0007). Moreover, there was a significant difference in BMP7&#95;rs388286 between the TGCT group and the control (T allele, P = 0.00004; TT genotype, P = 0.00006) and between the TM group and the control (T allele, P = 0.04). HOXD also demonstrated a strong association with TGCTs (rs17198432 A allele, P = 0.0001; AA genotype, P = 0.001). Furthermore, significant differences were found between the TGCT group and the control in the BAK1&#95;rs210138 G allele (P = 0.03) and the GG genotype (P = 0.01). KITLG and BMP7 genes, associated with the development of TGCTs, may also be related to TM. In summary, the KITLG GG&#95;rs995030, GG&#95;rs1508595, BMP7 TT&#95;rs388286, HOXD AA&#95;rs17198432, and BAK1 GG&#95;rs210138 genotypes were associated with a high risk of TGCT development., Competing Interests: None

Male infertility might be clearly associated with aberrant DNA methylation patterns in human spermatozoa. An association between oxidative stress and the global methylation status of the sperm genome has also been suggested. The aim of the present study was to determine whether the global sperm DNA methylation status was affected in the spermatozoa of carriers of chromosome structural aberrations. The relationships between the 5-methylcytosine (m 5 C) levels in spermatozoa and chromatin integrity status were evaluated. The study patients comprised male carriers of chromosome structural aberrations with reproductive failure (n = 24), and the controls comprised normozoospermic sperm volunteers (n = 23). The global m 5 C level was measured using thin-layer chromatography (TLC) and immunofluorescence (IF) techniques. The sperm chromatin integrity was assessed using aniline blue (AB) staining and TUNEL assay. The mean m 5 C levels were similar between the investigated chromosome structural aberrations carriers (P) and controls (K). However, sperm chromatin integrity tests revealed significantly higher values in chromosomal rearrangement carriers than in controls (P < 0.05). Although the potential relationship between sperm chromatin integrity status and sperm DNA fragmentation and the m 5 C level juxtaposed in both analyzed groups (P vs K) was represented in a clearly opposite manner, the low chromatin integrity might be associated with the high hypomethylation status of the sperm DNA observed in carriers of chromosome structural aberrations.

Testicular cancer is the most common form of solid cancer in young men. Testicular cancer is represented by testicular germ cell tumors (TGCTs) derived from embryonic stem cells with different degrees of differentiation in about 95% of cases. The development of these tumors is related to the formation of a pool of male germ cells and gametogenesis. Clinical factors that are predisposed to the development of germ-cell tumors include cryptorchidism and testicular microlithiasis, as well as infertility associated with the gr/gr deletion within the AZFс locus. KITLG, SPRY4, and BAK1 genes affect the development of the testes and gametogenesis; mutations and polymorphisms of these genes lead to a significant increase in the risk of the TGCT development. To determine the relationship between gene polymorphisms and the development of TGCTs, we developed a system for detection and studied the allele and genotype frequencies of the KITLG (rs995030, rs1508595), SPRY4 (rs4624820, rs6897876), and BAK1 (rs210138) genes in fertile men, patients with TGCTs, and patients with infertility that have the AZFс deletion. A significant association of rs995030 of the KITLG gene with the development of TGCTs (p = 0.029 for the allele G, p = 0.0124 for the genotype GG) was revealed. Significant differences in the frequencies of the studied polymorphisms in patients with the AZFc deletion and the control group of fertile men were not found. We showed significant differences in the frequencies for the combination of all high-risk polymorphisms in the control group, patients with the AZFc deletion and patients with TGCTs (p (TGCTs-AZF-control) = 0.0207). A fivefold increase in the frequency of the combination of all genotypes in the TGCT group (p = 0.0116; OR = 5.25 [1.44-19.15]) and 3.7-fold increase was identified in patients with the AZFc deletion (p = 0.045; OR = 3.69 [1.11-12.29]) were revealed. The genotyping of patients with infertility caused by the AZFc deletion can be used to identify individuals with an increased risk of TGCTs.

Objective: We report a male patient with ovotesticular disorder of sex development (OTDSD), resulting from structurally abnormal Y chromosome., Case Report: A 3-year-old boy was admitted to the Surgical Pediatric Department for masculinizing reconstruction. He had a clitorophallus, bifid scrotum, perineal hypospadias and bilateral impalpable gonads. Pelvic ultrasound and laparoscopy showed a uterus and two gonads with primary ovarian follicles. Chromosome analysis detected a mos 47,XX,mar/46,XX karyotype. Complex genetic evaluation revealed that the marker was Yp isochromosome. Surgical care included a feminizing genitoplasty and separation of the gonads with total excision of testicular tissue., Conclusions: The presented case emphasizes the importance of a systematic approach to the investigation and management of the patients with ovotesticular DSD. It also raises the important issue about gender reassignment in intersex individuals in mid-childhood.

The frequency and spectrum of mutations and the IVS8- T polymorphism of the CFTR gene have been studied in a sample of 963 in Russian infertile men. Mutations have been found in 48 out of 1926 analyzed chromosomes (2.5%) in the heterozygous state (n = 46) and in the compound heterozygote L138ins/N1303K (n = 1/n = 1). A CFTR gene mutation was combined with the 5T allele (mutCFTR/5T) in 11 patients. The following mutations have been found: F508del (n = 18), CFTRdele2,3 (21kb) (n = 9), W1282X (n = 7), 2143delT (n = 4), 3849+10kbC>T (n = 2), L138ins (n = 2), 1677delTA (n = 1), 2184insA (n = 1), 3821delT (n = 1), G542X (n = 1), N1303K (n = 1), and R334W (n = 1). The F508del mutation is the most frequent; it has been detected in 37.5% of the affected chromosomes. The total proportion of four mutations (F508del, CFTRdele2,3 (21kb), W1282X, and 2143delT) is about 79% of all mutations found. The 5T allele has been found in 10.9% infertile men and 4.8% of control men. Significant differences in the frequency of the IVS8-5T variant of the CFTR gene have been found between these groups (p = 0.005), as well as between infertile patients without mutations and control men (p = 0.019). In total, the mutations and/or 5T allele have been found in 14.6% of the patients examined. These data indicate increased frequencies of the mutations of the CFTR gene and its allele variant IVS8-5T in Russian infertile men.

We report on a 37-year-old XX male with complex hidden X chromosomal mosaicism. The patient had fully mature male genitalia with hypoplastic testes descended in the scrotum and no sign of undervirilization. Hormonal examination demonstrated hypergonadotropic hypogonadism, semen analysis showed severe oligoasthenoteratozoospermia. In situ hybridization revealed the presence of 3 SRY-positive cell lines bearing 1, 2 or 3 X chromosomes. Skewed inactivation of the paternal SRY-bearing X chromosome was detected by molecular analysis of the androgen receptor gene., ((c) 2009 S. Karger AG, Basel.)

We report on a 45,X male with hydrocephaly, lobar holoprosencephaly and ichthyosis. In situ hybridization and molecular analysis have demonstrated the presence of a mosaic SRY-bearing derivative X chromosome that included Yp and heterochromatic Yq fragments., (Copyright (c) 2008 Wiley-Liss, Inc.)

Molecular genetic analysis was performed for 26 phenotypically male patients lacking the Y chromosome in the karyotype. The sex-determining region Y (SRY) gene was found in 77% of the patients. PCR analysis of Y-specific loci in the 17 SRY-positive patients revealed Yp fragments varying in size in 16 cases and cryptic mosaicism (or chimerism) for the Y chromosome in one case. The frequencies of class I, II, and III (Yp+)XX sex reversals were 18.75, 25.25, and 56%, respectively. All of the class III (Yp+)XX sex-reversed patients had a 3.5-Mb paracentric inversion flanked by inverted repeats 3 (IR3) on the short arm of the Y chromosome.

Deletions of Y chromosome AZF locus were analyzed during a large-scale andrological and genetic examination of 810 infertile men. The search for Yq microdeletions was carried out according to the standard EAA/EMQN guidelines. The breakpoints were mapped for the deletions in AZF locus. The Y chromosome macro- and microdeletions were detected in 61 (7.5%) infertile men. The frequencies of AZF deletions during azoospermia and severe oligozoospermia amounted to 12.2 and 8.1 %, respectively. On the whole, the frequencies of Yq microdeletions and the genophenotypic correlations characteristic of various AZF deletion types comply with the relevant published data. However, spermatozoids in the ejaculate sediment of men with completely deleted AZFa region or AZFb+c deletions (from solitary spermatozoids to several dozens) were detected for the first time. It was demonstrated that the breakpoints were localized between AZFa and AZFb regions proximally to AZFb+c microdeletions for the majority of cytogenetically detectable deletions in the Y chromosome long arm. This indicates that the mechanisms underlying Yq macro- and microdeletions are somewhat different. The issues related to the role of Y chromosome deletions in the origins of monosomy for X chromosome and X/XY mosaicism are discussed.

Literature data on genetic control of hormonal regulation of sexual differentiation and reproduction system development in humans are reviewed. The conditions caused by mutations of genes for gonadoliberin, gonadotorpin, and anti-Mullerian hormone, as well as genes controlling biosynthesis of sexual steroids and the receptors for the latter, are considered.

The current literature results on genetic control of sex differentiation and morphogenesis of the human reproductive system are reviewed. Several examples of the nosologic forms caused by mutations in the genes analyzed are considered.