Elucidating the impact of Y chromosome microdeletions and altered gene expression on male fertility in assisted reproduction.

BACKGROUND: Genetic abnormalities like Y chromosome microdeletions are implicated in male infertility. This study investigated the association of azoospermia factor (AZF) region microdeletions with unsuccessful assisted reproductive techniques (ART), including in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI). METHODS: This cross-sectional analysis study examined 80 Iranian oligospermic men (mean age 34 years) with prior failed ICSI and IVF cycles (IR.IAU.TNB.REC.1401.041). Semen analysis evaluated quantity/quality parameters based on World Health Organization guidelines. Participants were stratified by sperm DNA fragmentation (SDF) levels into: control (SDF < 15%, n = 20), mild elevation (15% ≤ SDF ≤ 30%, n = 60), and high (SDF > 30%, n = 20). Multiplex PCR mapped AZF microdeletions in the high SDF group. The AZF-associated genes were selected by RNA Seq analysis, and the candidate genes were checked for expression level by real-time PCR. RESULTS: High SDF individuals exhibited poorer semen metrics, including 69% lower sperm concentration (P = 0.04) than those without SDF. Of this subset, 45% (9/20 men) harboured predominately AZF microdeletions. Men with AZF microdeletions showed higher SDF (32% vs 21%, P = 0.02) and altered AZF-associated genes expression. As USP9Y 3-fold, UTY 1.3-fold, and BPY2 1-fold revealed up-regulation, while IQCF1 8-fold, CDY 6.5-fold, DAZ 6-fold, and DDX3Y 1-fold underwent down-regulation. The PAWP gene was also down-regulated (5.7-fold, P = 0.029) in the IVF/ICSI failure group. CONCLUSION: AZF microdeletions significantly impact male infertility and ART outcomes. High SDF individuals exhibited poorer semen metrics, with 45% AZF microdeletions. These microdeletions altered AZF-associated genes expression, affecting fertility mediator PAWP independently. Dual AZF and SDF screening enables personalized management in severe male infertility, potentially explaining IVF/ICSI failures.

M1AP interacts with the mammalian ZZS complex and promotes male meiotic recombination.

Following meiotic recombination, each pair of homologous chromosomes acquires at least one crossover, which ensures accurate chromosome segregation and allows reciprocal exchange of genetic information. Recombination failure often leads to meiotic arrest, impairing fertility, but the molecular basis of recombination remains elusive. Here, we report a homozygous M1AP splicing mutation (c.1074 + 2T > C) in patients with severe oligozoospermia owing to meiotic metaphase I arrest. The mutation abolishes M1AP foci on the chromosome axes, resulting in decreased recombination intermediates and crossovers in male mouse models. M1AP interacts with the mammalian ZZS (an acronym for yeast proteins Zip2-Zip4-Spo16) complex components, SHOC1, TEX11, and SPO16. M1AP localizes to chromosomal axes in a SPO16-dependent manner and colocalizes with TEX11. Ablation of M1AP does not alter SHOC1 localization but reduces the recruitment of TEX11 to recombination intermediates. M1AP shows cytoplasmic localization in fetal oocytes and is dispensable for fertility and crossover formation in female mice. Our study provides the first evidence that M1AP acts as a copartner of the ZZS complex to promote crossover formation and meiotic progression in males.

Identification of novel homozygous missense and deletion mutations manifesting oligospermia infertility in Kashmiri population.

BACKGROUND: Human male infertility has a lot of known molecular components that have an accurate diagnosis, such as Y chromosome deletion and monogenic causes. Only 4% of all infertile males are diagnosed with genetic causes, while 60-70% of infertile men remain without an accurate diagnosis and are classified as unexplained. Oligospermia is a major cause of human male infertility. Its etiology and pathogenesis are linked to genetic abnormalities. The majority of genetic causes related to human male infertility remain unclear. RESULTS: Generally, we found a significant association between the specific type of disease and gender (p = 0.003), and the regression value (R2 ) for this association was 0.75. Association of the type of disease with body mass index was not significant (p = 0.34). There was no statistically significant difference (p = 0.40) among disease types with patients occupations. All explored mutations are listed for primary and secondary infertility in relation to the oligospermia condition. p.Arg286X is the outcome of a mismatch mutation in which the nucleotide change resulted in the substitution of Arg (arginine) amino acid with X (any amino acid) at position 286 in the Hyal3 gene of primary infertile patients having oligospermia. In primary infertile patients with the p.Arg286X mutation, a frameshift deletion mutation was also found just after the 25 nucleotide sequences of the Hyal3 genes of the second mutated exon. This deletion mutation was only detected in patients with primary infertility and was not found in people with secondary infertility or healthy controls. The other mutations in secondary infertile patients with oligospermia were: p.Lys168Ser, replacement of lysine (Lys) with serine (Ser) at position 168; p.Lys168The, replacement of lysine (Lys) with threonine (The) at position 168; p.His113X, substitution of histidine (His) with an unknown amino acid (X) at position 113; p.Pro162X, substitution of proline (Pro) with an unknown amino acid (X) at position 162; and p.Phe157X, phenylalanine (Phe) substitution with an unknown amino acid (X) at position 157. CONCLUSION: This study clarifies the site of novel mismatch and frameshift deletion mutations in the Hyal3 gene in primary infertile oligospermia patients.

Use of empirical medical therapies for idiopathic male infertility in Australia and New Zealand.

OBJECTIVE: Idiopathic male infertility is common, yet there is no approved treatment. This study aimed to understand practice patterns towards empirical medical therapy (EMT) for idiopathic male infertility in Australia and New Zealand (NZ). DESIGN: Clinical members of the Endocrine Society of Australia, Fertility Society of Australia & NZ, and Urological Society of Australia & NZ were invited to complete a survey. Questions included demographics, EMT practice habits, and thoughts regarding infertility case scenarios. Unadjusted group differences between specialists, those with and without additional training in male infertility, and frequency of managing it were evaluated. RESULTS: Overall, 147 of 2340 members participated (6.3%); majority were endocrinologists and gynaecologists. Participants were experienced; 35% had completed additional training in male infertility and 36.2% reported they frequently manage male infertility. Gynaecologists were more likely to manage male infertility and attend education courses than endocrinologists and urologists. Beliefs about the effect of EMT on sperm concentration and pregnancy did not differ between speciality types. Many respondents considered all patient scenarios suitable for EMT. Of medications, hCG and clomiphene were selected most. Two respondents indicated they would use testosterone to treat male infertility. CONCLUSIONS: This study demonstrates common use of EMT in Australia and NZ for idiopathic male infertility. The breadth of responses reflects a lack of consensus within the current literature, highlighting the need for further research to clarify their role in the management of idiopathic male infertility.

Evaluation of telomere length, reactive oxygen species, and apoptosis in spermatozoa of patients with oligospermia.

50% of cases of infertility are caused by male factor, which acquired or congenital problems may bring on. Male infertility can be caused by oligospermia and asthenozoospermia, which are common. Since the same mutations that cause azoospermia in some people also cause oligozoospermia in others, oligozoospermia may be thought of as a less severe form of azoospermia. Studies have demonstrated telomere length, catalase activity, super oxide dismutase (SOD), and DNA fragmentation can be influential factors for male infertility. The amount of apoptosis, oxidative stress factors, telomere length, and DNA fragmentation were some aspects of healthy sperm that we chose to look into in this study and compare to oligospermia individuals. Oligospermia patients (n = 24) and fertile men (n = 27) semen samples were collected, and the apoptosis rate of sperms in both groups was analyzed (Flow cytometry). Also, gene expression of apoptotic and antiapoptotic markers and telomere length were examined (real-time polymerase chain reaction). The sperm DNA fragmentation kit was used to determine DNA fragmentation and to evaluate catalase and SOD activity; the specific kits and methods were utilized. Higher expression levels of caspase3 (p = .0042), caspase8 (p = .0145), caspase9 (p = .0275), and BAX (p = .0202) mRNA were observed in patients who had oligospermia. In contrast, lower mRNA expression of BCL-2 (p = .0009) was detected in this group. In addition, telomere length was decreased in the oligospermia group (p < .0001) compared to the health group. Moreover, the frequency of apoptosis is induced in patients (p = .0026). The catalase activity is low (p = .0008), but the SOD activity is high (p = .0015) in the patient group. As a result of our findings, we may list the sperm cell apoptosis rate, telomere length, the degree of sperm DNA fragmentation, and lastly, the measurement of significant and efficient oxidative stress markers like SOD and catalase in semen plasma among the principal diagnostic characteristics for oligospermia. Future studies will be better able to treat oligospermia by showing whether these indicators are rising or falling.

Safety evaluation of single-sperm cryopreservation technique applied in intracytoplasmic sperm injection.

Intracytoplasmic sperm injection (ICSI) is a technique that directly injects a single sperm into the cytoplasm of mature oocytes. Here, we explored the safety of single-sperm cryopreservation applied in ICSI. This retrospective study enrolled 186 couples undergoing ICSI-assisted pregnancy. Subjects were allocated to the fresh sperm (group A)/single-sperm cryopreservation (group B) groups based on sperm type, with their clinical baseline/pathological data documented. We used ICSI-compliant sperm for subsequent in vitro fertilization and followed up on all subjects. The recovery rate/cryosurvival rate/sperm motility of both groups, the pregnancy/outcome of women receiving embryo transfer, and the delivery mode/neonatal-related information of women with successful deliveries were recorded. The clinical pregnancy rate, cumulative clinical pregnancy rate, abortion rate, ectopic pregnancy rate, premature delivery rate, live birth delivery rate, neonatal birth defect rate, and average birth weight were analyzed. The two groups showed no significant differences in age, body mass index, ovulation induction regimen, sex hormone [anti-Müllerian hormone (AMH)/follicle-stimulating hormone (FSH)/luteinizing hormone (LH)] levels, or oocyte retrieval cycles. The sperm recovery rate (51.72%-100.00%) and resuscitation rate (62.09% ± 16.67%) in group B were higher; the sperm motility in the two groups demonstrated no significant difference and met the ICSI requirements. Group B exhibited an increased fertilization rate, decreased abortion rate, and increased safety versus group A. Compared with fresh sperm, the application of single-sperm cryopreservation in ICSI sensibly improved the fertilization rate and reduced the abortion rate, showing higher safety.

Idiopathic secondary azoospermia occurrence in men with oligospermia over time.

PURPOSE: To investigate the occurrence of idiopathic secondary azoospermia (ISA) in men with oligospermia over time and identify risk factors for ISA in this population. METHODS: This was a retrospective cohort study conducted in a university-affiliated male infertility clinic. A total of 1056 oligospermic men (concentration < 15 million/ml (M/ml) and no azoospermia) with at least two SA done between 2000 and 2019 were included. The primary outcome was the occurrence of ISA by oligospermia severity. RESULTS: In the entire cohort, 31 patients (2.9%) eventually became azoospermic with time. The ≤ 1 M/ml extremely severe oligospermia (ESO) group (283 patients) had significantly higher rates of ISA in each time period compared to the 1-5 M/ml severe oligospermia (SO) (310 patients) and 5-15 M/ml mild oligospermia (MO) (463 patients) groups (p < 0.05 for all comparisons), with rates of 21.1% in the ESO, 4.8% in the SO, and 0% in the MO group (p = 0.02) after 3-5 years, reaching 32% after 5 years in the ESO group compared to no cases in the other two groups (p = 0.006). Parameters shown to predict ISA were initial concentration < 1 M/ml (OR 22.12, p < 0.001) and time interval of > 3 and 5 years (OR 4.83 and 6.84, p = 0.009 and < 0.001, respectively), whereas testosterone levels were negatively associated with ISA (OR 0.88, p = 0.03). CONCLUSIONS: Men with ≤ 1 M/ml, especially those with low testosterone levels, have a dramatically increased chance of becoming azoospermic with time. Therefore, sperm banking should be recommended in these cases. Men with a sperm concentration above 1 M/ml have low chances of becoming azoospermic, even after 3 or more years.

Modeling methods for busulfan-induced oligospermia and asthenozoospermia in mice: a systematic review and meta-analysis.

OBJECTIVE: Modeling methods for busulfan-induced oligoasthenozoospermia are controversial. We aimed to systematically review the modeling method of busulfan-induced oligospermia and asthenozoospermia, and analyze changes in various evaluation indicators at different busulfan doses over time. METHODS: We searched the Cochrane Library, PubMed databases, Web of Science, the Chinese National Knowledge Infrastructure, and the Chinese Biomedical Literature Service System until April 9, 2022. Animal experiments of busulfan-induced spermatogenesis dysfunction were included and screened. The model mortality and parameters of the evaluation indicators were subjected to meta-analysis. RESULTS: Twenty-nine animal studies were included (control/model: 669/1829). The mortality of mice increased with busulfan dose. Significant spermatogenesis impairment occurred within 5 weeks, regardless of busulfan dose (10-40 mg/kg). Testicular weight (weighted mean difference [WMD]: - 0.04, 95% CI: - 0.05, - 0.03), testicular index (WMD: - 2.10, 95% CI: - 2.43, - 1.76), and Johnsen score (WMD: - 4.67, 95% CI: - 5.99, - 3.35) were significantly decreased. The pooled sperm counts of the model group were reduced by 32.8 × 106/ml (WMD: - 32.8, 95% CI: - 44.34, - 21.28), and sperm motility decreased by 37% (WMD: - 0.37, 95% CI: - 0.47, - 0.27). Sperm counts decreased slightly (WMD: - 3.03, 95% CI: - 3.42, - 2.64) in an intratesticular injection of low-dose busulfan (4 - 6 mg/kg), and the model almost returned to normal after one seminiferous cycle. CONCLUSION: The model using low-dose busulfan (10 - 20 mg/kg) returned to normal after 10 - 15 weeks. However, in some spermatogenesis cycles, testicular weight reduction and testicular spermatogenic function damage were not proportional to busulfan dose. Sperm counts and motility results in different studies had significant heterogeneity. Standard protocols for sperm assessment in animal models were needed to reduce heterogeneity between studies.

Associations of semen quality with pyrethroids in semen and urine samples of men with oligozoospermia: A cross-sectional study in Samsun, Turkey.

Pyrethroids (PYRs) may act as endocrine disrupters and lead to infertility. The aim of the study was to analyze the levels of anti-androgenic PYRs (cypermethrin, deltamethrin, and permethrin) and 3-phenoxy benzoic acid (3-PBA), a general metabolite of PYRs, in both semen and urine samples of men with oligozoospermia. The PYRs and 3-PBA metabolite levels in the semen and urine samples of the men were analyzed through GC-MS. The results indicated that the levels of PYRs in the semen samples of the infertile group were significantly higher than those of the fertile group. It was determined that cypermethrin exposure was associated with changes in sperm count and total sperm motility, while permethrin, deltamethrin, and 3-PBA levels were associated with changes in sperm morphology. It was determined that there was a significant negative correlation between semen deltamethrin levels and sperm morphology and sperm count. In addition, exposure of these patients to deltamethrin (range; 1.53-8.02 µg/l) and having farmer parents were determined to increase the risk of infertility. In conclusion, the findings of this study showed that exposure to environmental PYRs may adversely affect semen quality, especially in terms of sperm morphology, in men with oligozoospermia.

The Loss-Function of KNL1 Causes Oligospermia and Asthenospermia in Mice by Affecting the Assembly and Separation of the Spindle through Flow Cytometry and Immunofluorescence.

KNL1 (kinetochore scaffold 1) has attracted much attention as one of the assembly elements of the outer kinetochore, and the functions of its different domains have been gradually revealed, most of which are associated with cancers, but few links have been made between KNL1 and male fertility. Here, we first linked KNL1 to male reproductive health and the loss-function of KNL1 resulted in oligospermia and asthenospermia in mice (an 86.5% decrease in total sperm number and an 82.4% increase in static sperm number, respectively) through CASA (computer-aided sperm analysis). Moreover, we introduced an ingenious method to pinpoint the abnormal stage in the spermatogenic cycle using flow cytometry combined with immunofluorescence. Results showed that 49.5% haploid sperm was reduced and 53.2% diploid sperm was increased after the function of KNL1 was lost. Spermatocytes arrest was identified at the meiotic prophase I of spermatogenesis, which was induced by the abnormal assembly and separation of the spindle. In conclusion, we established an association between KNL1 and male fertility, providing a guide for future genetic counseling regarding oligospermia and asthenospermia, and a powerful method for further exploring spermatogenic dysfunction by utilizing flow cytometry and immunofluorescence.

Protective Effects of Chia Seeds and Omega-3 Fatty Acid against Cyclophosphamide-Induced Oligospermia in Male Wistar Rats: Potential Risks of Adverse Drug Interaction with Chia Seeds.

Objectives: The aim of this study was to investigate whether chia (Salvia hispanica) seeds, which are rich in omega-3 fatty acids, amino acids, and vitamins with antioxidant properties, can mitigate the negative effects on male reproductive function caused by cyclophosphamide, a frequently used chemotherapeutic agent. Methods: Male wistar rats are divided into seven groups (n=6). All groups except the normal control (NC) received cyclophosphamide (30mg/kg, i.p.) for the first 5 days. The standard group received clomiphene citrate (0.25 mg/kg, p.o.). Treatment groups T1%, T5%, T10%, and ω-3 received 1%, 5%, and 10% chia seeds in the diet, and 880 mg/kg omega-3 fatty acid (p.o) respectively for 15 days. The effect on the reproductive system was evaluated by analysis of epididymal sperm characteristics, biochemical parameters, and serum testosterone level. Results: Clomiphene citrate improved oligospermia via hormone mediated effect. Chia seeds and omega-3 fatty acid treatment also showed improvement in reproductive parameters including oxidative stress and histological features of the testes. Omega-3 fatty acid treatment was more effective for the prevention of cyclophosphamide toxicity on testes as compared to chia seeds. Nasal bleeding was noted in several animals subjected to chia seed treatment. This occurrence might be attributed to chia seeds' impact on coagulation and/or platelet function, potentially heightened due to chemotherapy associated bone marrow suppression. Conclusions: In our study, chia seeds as well as omega-3 fatty acid treatment were found to be protective against cyclophosphamide-induced reproductive toxicity in rats. However, the adverse effect of hemorrhage associated with drug interaction of chia seeds with cytotoxic chemotherapeutic drugs needs careful attention and further investigation.

[Six-sequence-tagged site (STS) versus eight-STS scheme for detection of Y chromosome microdeletions].

OBJECTIVE: To compare the six-sequence-tagged site (STS) with the eight-STS scheme in the detection of Y chromosome microdeletions. METHODS: Using real-time quantitative PCR, we compared the results of the six-STS (sY84, sY86, sY127, sY134, sY254, sY255) scheme with those of the eight-STS (sY84, sY86, sY127, sY134, sY254, sY255, sY145, sY152) scheme in detecting Y chromosome microdeletions. RESULTS: No statistically significant difference was found in the detection rate of the deletion of the azoospermia factor (AZF) regions between the six-STS and eight-STS methods (9.34% ï¼»575/6177ï¼½ vs 8.85% ï¼»542/6122ï¼½, P > 0.05). CONCLUSION: Though the eight-STS scheme increased the detection of AZFd, its detection rate of the AZF region deletion was not significantly different from that of the six-STS method. From the perspectives of experimental operation, economic cost and clinical strategy guidance, the six-STS is better than the eight-STS scheme for the detection of Y chromosome microdeletions.

Molecular diversity and phenotypic pleiotropy of ancient genomic regulatory loci derived from human endogenous retrovirus type H (HERVH) promoter LTR7 and HERVK promoter LTR5_Hs and their contemporary impacts on pathophysiology of Modern Humans.

Timelines of population-level effects of viruses on humans varied from the evolutionary scale of million years to contemporary spread of viral infections. Correspondingly, these events are exemplified by: (i) emergence of human endogenous retroviruses (HERVs) from ancient germline infections leading to stable integration of viral genomes into human chromosomes; and (ii) wide-spread viral infections reaching a global pandemic state such as the COVID-19 pandemic. Despite significant efforts, understanding of HERV's roles in governance of genomic regulatory networks, their impacts on primate evolution and development of human-specific physiological and pathological phenotypic traits remains limited. Remarkably, present analyses revealed that expression of a dominant majority of genes (1696 of 1944 genes; 87%) constituting high-confidence down-steam regulatory targets of defined HERV loci was significantly altered in cells infected with the SARS-CoV-2 coronavirus, a pathogen causing the global COVID-19 pandemic. This study focused on defined sub-sets of DNA sequences derived from HERVs that are expressed at specific stages of human preimplantation embryogenesis and exert regulatory actions essential for self-renewal and pluripotency. Evolutionary histories of LTR7/HERVH and LTR5_Hs/HERVK were charted based on evidence of the earliest presence and expansion of highly conserved (HC) LTR sequences. Sequence conservation analyses of most recent releases 17 primate species' genomes revealed that LTR7/HERVH have entered germlines of primates in Africa after the separation of the New World Monkey lineage, while LTR5_Hs/HERVK successfully colonized primates' germlines after the segregation of Gibbons' species. Subsequently, both LTR7 and LTR5_Hs undergo a marked ~ fourfold-fivefold expansion in genomes of Great Apes. Timelines of quantitative expansion of both LTR7 and LTR5_Hs loci during evolution of Great Apes appear to replicate the consensus evolutionary sequence of increasing cognitive and behavioral complexities of non-human primates, which seems particularly striking for LTR7 loci and 11 distinct LTR7 subfamilies. Consistent with previous reports, identified in this study, 351 human-specific (HS) insertions of LTR7 (175 loci) and LTR5_Hs (176 loci) regulatory sequences have been linked to genes implicated in establishment and maintenance of naïve and primed pluripotent states and preimplantation embryogenesis phenotypes. Unexpectedly, HS-LTRs manifest regulatory connectivity to genes encoding markers of 12 distinct cells' populations of fetal gonads, as well as genes implicated in physiology and pathology of human spermatogenesis, including Y-linked spermatogenic failure, oligo- and azoospermia. Granular interrogations of genes linked with 11 distinct LTR7 subfamilies revealed that mammalian offspring survival (MOS) genes seem to remain one of consistent regulatory targets throughout ~ 30 MYA of the divergent evolution of LTR7 loci. Differential GSEA of MOS versus non-MOS genes identified clearly discernable dominant enrichment patterns of phenotypic traits affected by MOS genes linked with LTR7 (562 MOS genes) and LTR5_Hs (126 MOS genes) regulatory loci across the large panel of genomics and proteomics databases reflecting a broad spectrum of human physiological and pathological traits. GSEA of LTR7-linked MOS genes identified more than 2200 significantly enriched records of human common and rare diseases and gene signatures of 466 significantly enriched records of Human Phenotype Ontology traits, including Autosomal Dominant (92 genes) and Autosomal Recessive (93 genes) Inheritance. LTR7 regulatory elements appear linked with genes implicated in functional and morphological features of central nervous system, including synaptic transmission and protein-protein interactions at synapses, as well as gene signatures differentially regulated in cells of distinct neurodevelopmental stages and morphologically diverse cell types residing and functioning in human brain. These include Neural Stem/Precursor cells, Radial Glia cells, Bergman Glia cells, Pyramidal cells, Tanycytes, Immature neurons, Interneurons, Trigeminal neurons, GABAergic neurons, and Glutamatergic neurons. GSEA of LTR7-linked genes identified significantly enriched gene sets encoding markers of more than 80 specialized types of neurons and markers of 521 human brain regions, most prominently, subiculum and dentate gyrus. Identification and characterization of 1944 genes comprising high-confidence down-steam regulatory targets of LTR7 and/or LTR5_Hs loci validated and extended these observations by documenting marked enrichments for genes implicated in neoplasm metastasis, intellectual disability, autism, multiple cancer types, Alzheimer's, schizophrenia, and other brain disorders. Overall, genes representing down-stream regulatory targets of ancient retroviral LTRs exert the apparently cooperative and exceedingly broad phenotypic impacts on human physiology and pathology. This is exemplified by altered expression of 93% high-confidence LTR targets in cells infected by contemporary viruses, revealing a convergence of virus-inflicted aberrations on genomic regulatory circuitry governed by ancient retroviral LTR elements and interference with human cells' differentiation programs.

[Biological basis of oligoasthenospermia based on the theory of "kidney governing reproduction"].

Guided by the theory of "kidney governing reproduction", ancient and present-day physicians treat male infertility mainly by tonifying the kidneys, with some innovation and development based on inheritance. Relating oligoasthenospermia (OAS), the author emphasizes "the kidney as the base and the essence chamber for use", and proposes "the deficiency of kidney essence and disability of the essence chamber" as the core pathogenesis of the disease. Kidney essence deficiency is the primary cause and essence chamber disability is the main factor for the development and progression of OAS. Disorders in the reproductive microenvironment are also important causes of OAS. Studies on the biological basis of the treatment of OAS from the kidney suggest that kidney tonification has a regulatory effect on the reproductive microenvironment. A systematic investigation into the molecular mechanism of "the deficiency of kidney essence and disability of the essence chamber" in the development and pathogenesis of OAS from the perspective of the reproductive microenvironment may provide some evidence for clinical intervention in the biological basis of OAS based on the theory of "kidney governing reproduction".

Vps13b is required for acrosome biogenesis through functions in Golgi dynamic and membrane trafficking.

The sperm acrosome is a lysosome-related organelle that develops using membrane trafficking from the Golgi apparatus as well as the endolysosomal compartment. How vesicular trafficking is regulated in spermatids to form the acrosome remains to be elucidated. VPS13B, a RAB6-interactor, was recently shown involved in endomembrane trafficking. Here, we report the generation of the first Vps13b-knockout mouse model and show that male mutant mice are infertile due to oligoasthenoteratozoospermia. This phenotype was explained by a failure of Vps13b deficient spermatids to form an acrosome. In wild-type spermatids, immunostaining of Vps13b and Rab6 revealed that they transiently locate to the acrosomal inner membrane. Spermatids lacking Vps13b did not present with the Golgi structure that characterizes wild-type spermatids and showed abnormal targeting of PNA- and Rab6-positive Golgi-derived vesicles to Eea1- and Lamp2-positive structures. Altogether, our results uncover a function of Vps13b in the regulation of the vesicular transport between Golgi apparatus, acrosome, and endolysosome.

YTHDF2 is essential for spermatogenesis and fertility by mediating a wave of transcriptional transition in spermatogenic cells.

The dynamic and reversible regulation roles of m6A modification and the characterization of m6A readers have provided new insights into spermatogenesis at the post-transcriptional level. YTHDF2, as an m6A reader, has been reported to mediate the m6A-containing transcript decay during the mouse oocyte maturation, embryonic stem cell differentiation, neural development, and zebrafish maternal-to-zygotic transition. However, the roles of YTHDF2 in mammalian spermatogenesis are uncertain. Here, we generated germ cell-specific Ythdf2 mutants (Ythdf2-vKO) at a C57BL/6J background and demonstrated that YTHDF2 is essential for mouse spermatogenesis and fertility. Ythdf2-vKO provides oligoasthenoteratozoospermia phenotype with increased apoptosis in germ cells. High-throughput RNA-seq analysis showed that a group of mRNAs is upregulated in Ythdf2-vKO mouse testis; further analysis and MeRIP-qPCR data showed that most of the upregulated genes in Ythdf2-vKO mouse testis are modified with m6A and are YTHDF2 candidate binding genes. Interestingly, RNA-seq analysis combined with our previous single-cell transcriptomics data of mouse spermatogenesis pointed out the failure of a wave of transcript transition during the spermatogenesis of Ythdf2-vKO mice, which was confirmed by gene expression analysis using qPCR of diplotene spermatocytes and round spermatids obtained through fluorescence-activated cell sorting. Our study demonstrates the fundamental role of YTHDF2 during mouse spermatogenesis and provides a potential candidate for the diagnosis of male infertility with the oligoasthenoteratozoospermia syndrome.

An effective method for establishing animal models of azoospermia and oligospermia.

The current study aims to develop a validated animal model to predict successful spermatogenesis retrieval in azoospermia and oligospermia men. Thirty-two mice were equally divided into 4 groups: control, scrotal hyperthermia (15 times), scrotal hyperthermia group (10 times), scrotal hyperthermia group (5 times). In the scrotal hyperthermia groups, their scrotum exposed to water at a temperature of 43°C for 20 min every other day. Then, the mice were euthanised and sperm samples were collected for sperm parameters analysis, and blood samples were obtained for hormonal assay. The testis samples were taken for histopathology experiments, immunofluorescence staining and Western blot in order to examine the protein expression together with RNA extraction in order to examine the gene expression of germ cell markers. The results of sperm analysis and histopathology of testicular tissue as well as the results of gene expression and Western blot showed that hyperthermia can significantly impair spermatogenesis. In conclusion, we have developed a novel model of azoospermia and oligospermia in mouse, which uses a high temperature to suppress spermatogenesis process through demolition of germ cells subsequent cell cycle arrest and apoptosis. The model will contribute to understanding azoospermia in human, oligospermia pathophysiology and the development of treatment.

[L-carnitine-astaxanthin compound nutrients for the treatment of idiopathic oligospermia and asthenozoospermia: A multicenter clinical observation].

OBJECTIVE: To investigate the clinical effects of the L-carnitine-astaxanthin compound nutrients Menglankang (MLK) on idiopathic oligospermia (OS) and asthenospermia (AS). METHODS: This study included 73 cases of OS and 220 cases of AS treated with MLK once a bag, bid, for 3 successive months. Before and at 1, 2 and 3 months after treatment, we obtained and analyzed the semen parameters and sperm DNA fragmentation index (DFI) of the patients. RESULTS: Compared with the baseline, the OS patients showed remarkable increases after 1 and 2 months of treatment in the semen volume (ï¼»3.07 ± 1.47ï¼½ vs ï¼»3.26 ± 1.26ï¼½ and ï¼»3.30 ± 1.28ï¼½ ml), sperm concentration (ï¼»10.96 ± 6.09ï¼½ vs ï¼»16.74 ± 11.15ï¼½ and ï¼»17.56 ± 9.92ï¼½ ×106/ml, P < 0.05), total sperm count (ï¼»29.78 ± 17.48ï¼½ vs ï¼»52.98 ± 32.07ï¼½ and ï¼»57.67 ± 36.98ï¼½ ×106, P < 0.05) and the percentages of progressively motile sperm (PMS) (ï¼»39.8 ± 11.66ï¼½% vs ï¼»45.3 ± 14.03ï¼½% and ï¼»46.42 ± 10.69ï¼½%, P < 0.05) and morphologically normal sperm (MNS) (ï¼»1.71 ± 1.07ï¼½% vs ï¼»1.79 ± 0.91ï¼½% and ï¼»1.84 ± 0.96ï¼½%), and so did the AS patients in PMS (ï¼»19.23 ± 8.32ï¼½% vs ï¼»25.46 ± 13.86ï¼½% and ï¼»27.33 ± 12.88ï¼½%, P < 0.05). After 3 months of medication, the OS patients exhibited even more significant increases in the semen volume (ï¼»3.63 ± 1.39ï¼½ ml) (P < 0.05), sperm concentration (ï¼»20.56 ± 14.7ï¼½ ×106/ml) (P < 0.05), total sperm count (ï¼»66.35 ± 55.91ï¼½ ×106) (P < 0.05), PMS (ï¼»49.24 ± 13.45ï¼½%) (P < 0.05) and MNS (ï¼»2.59 ± 0.93ï¼½%) (P < 0.05), and so did the AS patients in the semen volume (ï¼»3.27 ± 1.42ï¼½ vs ï¼»3.85 ± 1.59ï¼½ ml, P < 0.05), PMS (ï¼»29.11 ± 13.58ï¼½%) (P < 0.05) and NMS (ï¼»2.01 ± 1.14ï¼½% vs ï¼»2.57 ± 1.15ï¼½%, P < 0.05). In comparison with the baseline, the sperm DFI was not significantly improved at 1 month after treatment, but remarkably decreased at 2 and 3 months in the OS patients (ï¼»25.87 ± 13.76ï¼½% vs ï¼»18.66 ± 10.83ï¼½% and ï¼»16.48 ± 11.46ï¼½%, P < 0.05) and the AS patients as well (ï¼»26.40 ± 12.28ï¼½% vs ï¼»19.35 ± 11.54ï¼½% and ï¼»15.32 ± 10.89ï¼½%, P < 0.05). CONCLUSIONS: The L-carnitine-astaxanthin compound nutrients Menglankang can significantly improve the semen quality of the patients with idiopathic oligospermia or asthenospermia.

A genomics approach to male infertility.

PURPOSE: Male infertility remains poorly understood at the molecular level. We aimed in this study to investigate the yield of a "genomics first" approach to male infertility. METHODS: Patients with severe oligospermia and nonobstructive azoospermia were investigated using exome sequencing (ES) in parallel with the standard practice of chromosomal analysis. RESULTS: In 285 patients, 10.5% (n = 30) had evidence of chromosomal aberrations while nearly a quarter (n = 69; 24.2%) had a potential monogenic form of male infertility. The latter ranged from variants in genes previously reported to cause male infertility with or without other phenotypes in humans (24 patients; 8.4%) to those in novel candidate genes reported in this study (37 patients; 12.9%). The 33 candidate genes have biological links to male germ cell development including compatible mouse knockouts, and a few (TERB1 [CCDC79], PIWIL2, MAGEE2, and ZSWIM7) were found to be independently mutated in unrelated patients in our cohort. We also found that male infertility can be the sole or major phenotypic expression of a number of genes that are known to cause multisystemic manifestations in humans (n = 9 patients; 3.1%). CONCLUSION: The standard approach to male infertility overlooks the significant contribution of monogenic causes to this important clinical entity.

Effect of Natesto on Reproductive Hormones, Semen Parameters and Hypogonadal Symptoms: A Single Center, Open Label, Single Arm Trial.

PURPOSE: To evaluate the effect of short-acting 4.5% nasal testosterone gel (Natesto) on serum testosterone, gonadotropins, total motile sperm count, health related quality of life and sexual function. MATERIALS AND METHODS: This was a single institution, open label, single arm trial conducted between November 2017 and September 2019 at the University of Miami. Men 18 to 55 years old diagnosed with symptomatic hypogonadism (total testosterone less than 300 ng/dl on 2 occasions) were included. Men with azoospermia, vasectomy or a total motile sperm count less than 5 million were excluded. Enrolled patients were treated with Natesto, a short-acting nasal testosterone (125 µl per nostril, 11.0 mg testosterone per dose, TID) for 6 months. RESULTS: In total, 60 men were enrolled in the study. Of these, 44 and 33 patients were evaluated for testosterone at 3 and 6 months, respectively. A total of 31 patients (90.9%) reached a normal testosterone level (greater than 300 ng/dl) at 6 months. Follicle stimulating hormone and luteinizing hormone levels were maintained within the normal range in 81.8% and 72.7% of patients at 6 months, respectively. Total motile sperm count was maintained with total motile sperm count greater than 5 million over the treatment period in 88.4% of men at 3 months and 93.9% at 6 months. There were statistically significant improvements on International Index of Erectile Function sexual desire and overall satisfaction domains at 6 months. CONCLUSIONS: Natesto appears to increase testosterone while maintaining semen parameters in a majority of men. Natesto has the potential to be a safe and effective treatment for men with functional hypogonadism who wish to preserve semen parameters. Long-term studies beyond 6 months are needed before we can safely prescribe nasal testosterone gel for men interested in fertility.