Oligoasthenoteratozoospermic Patients Research Articles

Novel and recurrent hemizygous variants in BCORL1 cause oligoasthenoteratozoospermia by interfering transcription.

Oligoasthenoteratozoospermia (OAT) is a common cause of male infertility, of which the causes remain largely unknown. Recently, BCORL1 was identified as a contributor to male infertility from non-obstructive azoospermia (NOA) to OAT. To identify novel and hotspot variants in BCORL1 from infertile men with OAT and reveal their outcomes of assisted reproductive treatments (ARTs). Forty-six infertile men characterized by OAT were recruited from 2017 to 2022. Variants in OAT patients were identified by whole-exome sequencing (WES) and verified by Sanger sequencing. Papanicolaou staining was used for sperm morphology analysis. Pathogenicity of BCORL1 variants were analyzed by bioinformatics analysis, and further confirmed in vitro by using recombinant plasmids and cells. Meanwhile, ARTs were performed on these patients to investigate the appropriate clinical treatment strategy. We identified a novel hemizygous missense variant (NM_021946: c.G4171A; p.G1391R) and a recurrent variant (NM_021946: c.T2615G; p.V872G) in BCORL1 from four OAT patients. Notably, routine semen assessment and Papanicolaou staining revealed a special OAT phenotype of patients with BCORL1 variants, whose rare mature sperm characterized by acephalic and abnormal acrosome. Pathogenicity analysis showed the interaction between BCORL1 with histone deacetylases (HDACs) were disrupted after variance, accompanied with epigenetic alterations and finally the orderly transcriptions of spermatogenetic genes were interfering. Besides, clinical record presented the poor outcomes of ARTs in these patients with BCORL1 variants. Our findings further expand the variant spectrum of BCORL1 related to OAT, and provide new evidences that BCORL1 acts as an important transcriptional regulator, participating in epigenetic regulation and directing the expression of key genes throughout spermatogenesis. The outcomes of ARTs will facilitate the genetic counseling and clinical treatment of infertile men with BCORL1 variants in the future.

The therapeutic effect and metabolic mechanism analysis of Guilingji on idiopathic oligo-asthenoteratozoospermia

IntroductionGuilingji, a famous traditional Chinese medicine (TCM) formula, has been used to combat aging and male sexual dysfunction in China for centuries. To date, there has been little evidence-based clinical research on the use of Guilingji to treat idiopathic oligo-asthenoteratozoospermia (OAT), and the therapeutic mechanism from a metabolic perspective needs to be investigated further. MethodsThis was a multicenter, double-blind, randomized controlled clinical study of 240 patients with idiopathic OAT recruited from four hospitals between January 2020 and January 2022. Patients were randomly assigned in a 1꞉1 ratio to receive oral Guilingji capsules or placebo for 12 weeks. The total progressive motile sperm count (TPMSC) was considered the primary outcome, and the other sperm parameters, seminal plasma parameters and serum hormones were considered the secondary outcome. A nontargeted metabolomics analysis of serum from OAT patients before and after Guilingji administration was performed by HPLC–MS to identify key metabolites. Furthermore, we used a rat model to show spermatogenesis phenotypes to validate the effect of the key metabolites screened from the patients. ResultsAt weeks 4, 8 and 12, TPMSC and other sperm parameters were significantly improved in the Guilingji group compared with the placebo group (P < 0.05 for all comparisons). At week 4, superoxide dismutase (SOD) and acrosomal enzyme activity of seminal plasma were significantly elevated in the Guilingji group compared with the placebo group, while reactive oxygen species (ROS) levels were significantly reduced (P < 0.05). Lactate dehydrogenase-X (LDHX) levels appeared to be significantly increased after 12 weeks continuous medication compared with Placebo group (P = 0.032). The metabolomics analysis of serum from OAT patients before and after Guilingji administration showed that the glucose-6-phosphate (G6P) concentration in patients' serum was significantly elevated after Guilingji treatment. Compared to the control, when Kidney-Yang deficiency model rats were treated with Guilingji or its key intermediate metabolite G6P, their sperm concentration and spermatozoic activity were improved similarly, and their structural damage of rat's testicular and epididymal tissues were recovered. ConclusionThis study provided valuable clinical evidence for the utility of Guilingji as a treatment for OAT. These findings thus demonstrate that G6P is involved in the therapeutic mechanism of Guilingji in OAT treatment based on clinical and rat intervention studies.

Seminal calbindin 2 level in azoospermia and oligoasthenoteratozoospermia and its correlation with seminal and hormonal parameters.

We aimed to assess seminal calbindin 2 (CALB 2) expression in men with different semen parameters as well as its correlation with reproductive hormones in azoospermic patients and different semen parameters in oligoasthenoteratozoospermic patients. CALB 2 is also known as calretinin and 29 kDa calbindin. This prospective study was performed on 96 cases from the andrology outpatient clinic divided into 3 groups as follows: group 1 including 32 non obstructive azoospermic (NOA) patients, group 2 including 32 patients with oligoasthenoteratozoospermia (OAT), and Group 3 including normozoospermic individuals as controls. Semen analysis and estimation of seminal CALB 2 concentrations by enzyme linked immunosorbent assay (ELISA) technique were performed for all participants. Reproductive hormones were measured in nonobstructive NOA patients. The mean seminal CALB 2 level was higher in OAT patients compared to NOA patients and controls (7.8 ± 1.30 ng/ml, 7.3 ± 0.80 and 7.4 ± 1.0, respectively). Furthermore, the study had shown strong positive correlations between CALB 2 and sperm normal forms in controls and OAT patients. In contrast, there was no significant correlation between seminal CALB 2 and any of the reproductive hormones measured in NOA patients. Seminal CALB 2 may play a role in increasing the abnormal forms in OAT patients.

The Association between Clusterin Sialylation Degree and Levels of Oxidative-Antioxidant Balance Markers in Seminal Plasmas and Blood Sera of Male Partners with Abnormal Sperm Parameters.

Nearly 30% of infertility cases are caused by male factor. This study aimed at checking the associations between the sialylation degree of glycoprotein clusterin (CLU) and levels of oxidative–antioxidant balance markers in infertile men. Using lectin-ELISA with biotinylated lectins specific to α2,6-linked (Sambucus nigra agglutinin, SNA) and α2,3-linked (Maackia amurensis agglutinin, MAA) sialic acid (SA), the CLU sialylation in 132 seminal plasmas (SP) and 91 blood sera (BS) were analyzed. Oxidative–antioxidant status was measured by determining Sirtuin-3 (SIRT3), Sirtuin-5 (SIRT5), total antioxidant status (TAS), and ferric reducing antioxidant power (FRAP) levels. We indicate that multiple sperm disorders are associated with decreased expression of MAA-reactive SA in SP. Decreased SP SIRT3 concentrations may be associated with teratozoospermia and oligoasthenoteratozoospermia. ROC curve and cluster analysis revealed that SP relative reactivity of CLU glycans with MAA, the value of MAA/SNA ratio, and SIRT3 and SIRT5 concentrations may constitute an additional set of markers differentiating infertile oligoasthenoteratozoospermic patients (OAT) from normozoospermic (N), asthenoteratozoospermic (AT) and teratozoospermic (T). The multinomial logistic regression analysis confirmed the potential utility of SIRT3 determinations for differentiation between N and OAT groups as well as between N and T groups for SIRT3 and SIRT5. For BS, based on ROC curve and cluster analysis, relative reactivities of CLU glycans with SNA, MAA, SIRT3 and FRAP concentrations may be useful in the differentiation of normozoospermic patients from those with sperm disorders. The multinomial logistic regression analysis showed that the SNA relative reactivity with CLU glycans significantly differentiated the N group from AT, OAT and T groups, and FRAP concentrations significantly differed between N and AT groups, which additionally confirms the potential utility of these biomarkers in the differentiation of infertile patients with abnormal sperm parameters. The knowledge about associations between examined parameters may also influence future research aimed at seeking new male infertility therapies.

Positive Effect of a New Combination of Antioxidants and Natural Hormone Stimulants for the Treatment of Oligoasthenoteratozoospermia.

Oligoasthenoteratozoospermia (OAT) accounts for about 90% of male infertility; in many cases this disorder may be associated with oxidative stress, a condition that decreases the success of fertilization. Therefore, the empirical treatment of male infertility is often based on the use of antioxidants. The aim of the present study was to assess the effectiveness of three months’ administration of a new nutraceutical preparation on hormone profile, sperm parameters and fertilization capability in men undergoing in vitro fertilization (IVF). A total of 36 OAT patients were daily treated for 3 months with a dose of a formulation containing: Inositol, L-Carnitine, Vitamins C, D, E, Coenzyme Q10 and Selenium. Selected parameters were analysed before (T0) and after (T1) treatment, and IVF outcomes were evaluated. We observed an improvement of sperm concentration, motility, morphology and vitality; blood level of testosterone also showed an increase. A significant increase of fertilization rate was detected in 14 couples, whose male partner were treated with the nutraceutical preparation. The present results indicate that a formulation containing antioxidant and energy supply substances was effective in the treatment of sperm alterations and led to significant recovery of fertilizing capacity.

In vitro effects of pentoxifylline and coenzyme Q10 on the sperm of oligoasthenoteratozoospermia patients

The effect of in-vitro sperm incubation with Pentoxifylline (PTX) and Coenzyme Q10 (CoQ10) in Oligoasthenoteratozoospermia (OAT) patients was evaluated. Semen samples were obtained from men with Normozoospermia and men with OAT. Motile sperm from the two groups were subdivided into four subgroups: (i) without incubation with PTX + CoQ10; (ii) incubation with PTX; (iii) Incubation with CoQ10; and (iv) incubation with a combination of PTX + CoQ10. Then, sperm parameters, chromatin, DNA and membrane integrity, protamine deficiency, apoptosis, mitochondrial activity, sperm chromatin dispersion test (SCD), hypo-osmotic swelling test (HOS), chromomycin A3 (CMA3), Terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL), and diaminobenzidine (DAB) assays were evaluated, respectively. Sperm incubated with CoQ10 and a combination of CoQ10 and PTX resulted in a significant increase in the sperm parameters. Also, a significant decrease was noted with a combination of PTX and CoQ10 in normal men. There was a significant difference between CoQ10 treated and CoQ10 + PTX treated groups in comparison with the OAT group in the percentage of the DNA fragmentation, sperm apoptosis, AB+, HOS test + and sperm mitochondrial activity. Incubated sperm with CoQ10, PTX, and in combination with each other can improve sperm parameters in OAT patients.

KIF3B gene silent variant leading to sperm morphology and motility defects and male infertility†.

Sperm structural and functi onal defects are leading causes of male infertility. Patients with immotile sperm disorders suffer from axoneme failure and show a significant reduction in sperm count. The kinesin family member 3B (KIF3B) is one of the genes involved in the proper formation of sperm with a critical role in intraflagellar and intramanchette transport. A part of exon 2 and exons 3-5 of the KIF3B encodes a protein coiled-coil domain that interacts with intraflagellar transport 20 (IFT20) from the intraflagellar transport protein complex. In the present study, the coding region of KIF3B coiled-coil domain was assessed in 88 oligoasthenoteratozoospermic (OAT) patients, and the protein expression was evaluated in the mature spermatozoa of the case and control groups using immunocytochemistry and western blotting. According to the results, there was no genetic variation in the exons 3-5 of the KIF3B, but a new A>T variant was identified within the exon 2 in 30 patients, where nothing was detected in the control group. In contrast to healthy individuals, significantly reduced protein expression was observable in oligoasthenoteratozoospermic patients carrying variation where protein organization was disarranged, especially in the principal piece and midpiece of the sperm tail. Besides, the protein expression level was lower in the patients' samples compared to that of the control group. According to the results of the present study the KIF3B gene variation as well as lower protein expression leads to defects in sperm morphology and motility and consequently to male infertility.

P–072 Pregnancy rate in male factor infertility with oligoastheneteratozoospermia - evaluation of Letrozole and Coenzyme Q10 supplementation on sperm parameter

Abstract Study question Male infertility due to idiopathic oligoasthenoteratozoospermia- Does combining Letrozole as antiestrogenic with Coenzyme Q10 as antioxidant give better pregnancy rate ? Summary answer Combination of Co enzyme Q10 with Letrozole can significantly improve semen parameters and outcome of clinical pregnancy rate in idiopathic oligoasthenoteratozoospermic patients. What is known already Elevated levels of reactive oxygen species(ROS) are a major cause of idiopathic male factor infertility which results in sperm membrane lipid peroxidation, DNA damage and apoptosis leading to decrease sperm viability and motility. Antioxidant like Coenzyme Q10 have been used empiricallyin the treatment of oligoasthenozoospermia based on its ability to reverse oxidative stress and sperm dysfunction. Aromatase inhibitor like Letrozolehave been used in idiopathic male infertility by reducing estrogenic effect on spermatogenesis and reducing feedback inhibition of hypothalamopituatarygonadal axis. Thus a therapeutic strategy would need to use supplements to increase sperm energy metabilism, minimise free radical damage. Study design, size, duration Study design: prospective comperative clinical study Primary purpose: treatmenr Size: 60 infertile male attending OPD of SHRISTI HEALTHCARE diagnosed as idiopathic oligoasthenoteratozoospermia Duration: from March2018 to February 2020 Primary outcome: improvement in sperm count, motility and morphology after treatment Secondary outcome: clinical pregnancy rate and live birth rate. Participants/materials, setting, methods Exclusion criteria: Smoker, drug and alcohol abuse, medical treatment with gonadotropin and steroids, varicocele.60 patients were randomisedinto 3 groups. Gr A(N = 20) received Letrozole 2.5mg/day + Co enzyme Q10 300mg/day for 3 months, Gr B(N = 20) received Letrozole 2.5mg/day for 3 months, and Gr C(N = 20) received Coenzyme Q10 300mg/day for 3 months. History taking, general examination, semen analysis, sr.FSH,LH, Testesteron, E2 and scrotal duplex were done for all patients. Main results and the role of chance After treatment, Gr A as compared to Gr B and C showed significant imprivement in all 3 parameters of semen eg sperm count( 3.15±3.38 - 20.9±2.11, p &amp;lt; 0.001), sperm motility( 5.25±3.25 - 42.85±3.30, p &amp;lt; 0.001), sperm morphology( 2.26±7.81 - 25.89±7.05, p &amp;lt; 0.001). Improvement in sperm count and morphology was seen in Gr B(Letrozole gr) but not in sperm motility whereas Gr C ( Co enzyme Q10 gr)showed significant improvement in sperm motility and morphology but not in sperm count. 10 pregnancies occured during follow up period of 1 yr. Clinical pregnancy rate was 30%in Gr A(6/20), 5% in Gr B(1/20), AND 15% in Gr C( 3/20). Live birth rate was 83% in Gr A(5/6), 33.3% inGr C(1/3) whereas sponteneous abortion occured in Gr B pregnancy. Limitations, reasons for caution Limitation of my study was the small sample sizewhich could have some bias in outcome. I did not evaluate DNA fragmentation and level of ROS. Latest evidences report that evaluating ROS can be a diagnostic tool in predictingthe best responder to supplementation. Wider implications of the findings: Majority of studies had investigated the effect of antioxidant and aromatase inhibitor on semen parameter but few concluded their effect on live birth rate. Assisted reproductive techniques are expensive and not universally available, so any pharmacological agent with satisfactory effectiveness should be considered as 1st line treatment of oligoasthenoteratozoospermia. Trial registration number Not applicable

Biallelic mutations in KATNAL2 cause male infertility due to oligo-astheno-teratozoospermia.

Oligo-astheno-teratozoospermia (OAT) is a common cause of male infertility, and most of idiopathic OAT patients are thought to be caused by genetic defects. Here, we recruited 38 primary infertile patients with the OAT phenotype and 40 adult men with proven fertility for genetic analysis and identified biallelic mutations of KATNAL2 by whole-exome sequencing in two cases. F013/II:1, from a consanguineous family, carried the KATNAL2 c.328C > T:p.Arg110X homozygous mutations. The other carried c.55A > G: p.Lys19Glu and c.169C > T: p Arg57Trp biallelic mutations. None of the KATNAL2 variants were found in the 40 adult men with proven fertility. The spermatozoa from patients with KATNAL2 biallelic mutations exhibited conspicuous defects in maturation, head morphology, and the structure of mitochondrial sheaths and flagella. KATNAL2 was mainly expressed in the pericentriolar material and mitochondrial sheath of the spermatozoa from control subjects, but it was undetectable in the spermatozoa from the patients. Furthermore, Katnal2 null male mice were infertile and displayed an OAT phenotype. Our results proved that the biallelic mutations in KATNAL2 cause male infertility and OAT in humans for the first time, to our knowledge, which could enrich the genetic defect spectrum of OAT and be beneficial for its accurate genetic screening and clinical diagnosis.

Two subsequent seminal productions: A good strategy to treat very severe oligoasthenoteratozoospermic infertile couples

Sexual abstinence is considered one of the several factors that influence sperm quality. Recent studies show that a shortening of the abstinence period could be beneficial mostly in oligoasthenoteratozoospermic (OAT) patients. Retrospective study to verify the efficacy of a second semen sample after a short abstinence to treat severe OAT infertile patients. 127 couples treated between May 2014 and May 2018 were divided into two groups. Study Group 1 (75 cycles): severe OAT characteristics: count <0.2 × 106 /mL no progressive motility; count ≥0.2 × 106 /mL and no total or progressive motility; 0% normal morphology; a second semen sample was requested after abstinence of 2 h. Control Group 0 (52 cycles): normozoospermic or mild OAT; only one sample was requested. Intracytoplasmic sperm injection was utilized in all cases. All semen parameters were significantly different between Group 0 vs both samples of Group 1 (p < 0.001), excluding volume between Group 0 and 1st sample of Group 1 (p = 0.682). The comparison between 1st and 2nd samples from Group 1 showed significant differences in volume, total and progressive motility and morphology (p < 0.001, p < 0.001, p < 0.020) but not in total sperm count (p = 0.970). Fertilization, pregnancy rate/transfer, implantation and miscarriage rates were 85.9% and 61.1% (p < 0.001), 30.6% and 35.8% (p = 0.700), 17.5% and 24.0 (p = 0.292), 20.0% and 25.0% (p = 0.017) in Group 0 and Group 1 respectively. The results show that a short abstinence in severe OAT patients allows us to obtain spermatozoa with better motility. The request for a second semen sample in couples with extreme semen parameters is a valid and simple strategy that helps to achieve the same probability of pregnancy compared to a Control Group. Furthermore, it allows us to utilize fresh spermatozoa avoiding the need to resort to cryopreserved reserves or testicular surgery.

Comparison of physiological-ICSI (PICSI) with ICSI in cases of moderate to severe oligoasthenoteratozoospermia (OAT) − a randomized control study

Objective: To compare the efficiency of physiological intracytoplasmic sperm injection (PICSI) over intracytoplasmic sperm injection (ICSI) in Oligoasthenoteratozoospermic patients. Design: Randomized control study. Setting: KJIVF and laparoscopy center Delhi. Methods: From September 2019 to May 2020, 45 patients of male factor felled under our criteria of OAT were divided into two groups of PICSI and ICSI. Sperm selection was performed under high magnification in both the groups and results were compared. Outcome measure: Primary outcome: Fertilization rate, cleavage rate, blastulation, and utility rate; Secondary outcome: Clinical pregnancy rates and miscarriage rates. Results: 20 PICSI and 25 ICSI treatments were performed and observed that the rate of blastulation and embryo utility shows significant difference in PICSI group (P = 0.013). The pregnancy percentage was also better in PICSI but implantation, fertilization, cleavage; clinical pregnancy rates were clinically comparable in both the groups but not statistically significant. Conclusion: The outcomes of our study are independent of male factor and it only depends on intervention whether it was PICSI or ICSI. The blastulation rate and embryo utility shows statistically significant difference between PICSI and ICSI groups. Hence, we are slowly progressing toward the superiority of PICSI over ICSI but enough evidences are still not available.

Comparison of physiological-ICSI (PICSI) with ICSI in cases of moderate to severe oligoasthenoteratozoospermia (OAT) − a randomized control study

Objective: To compare the efficiency of physiological intracytoplasmic sperm injection (PICSI) over intracytoplasmic sperm injection (ICSI) in Oligoasthenoteratozoospermic patients. Design: Randomized control study. Setting: KJIVF and laparoscopy center Delhi. Methods: From September 2019 to May 2020, 45 patients of male factor felled under our criteria of OAT were divided into two groups of PICSI and ICSI. Sperm selection was performed under high magnification in both the groups and results were compared. Outcome measure: Primary outcome: Fertilization rate, cleavage rate, blastulation, and utility rate; Secondary outcome: Clinical pregnancy rates and miscarriage rates. Results: 20 PICSI and 25 ICSI treatments were performed and observed that the rate of blastulation and embryo utility shows significant difference in PICSI group (P = 0.013). The pregnancy percentage was also better in PICSI but implantation, fertilization, cleavage; clinical pregnancy rates were clinically comparable in both the groups but not statistically significant. Conclusion: The outcomes of our study are independent of male factor and it only depends on intervention whether it was PICSI or ICSI. The blastulation rate and embryo utility shows statistically significant difference between PICSI and ICSI groups. Hence, we are slowly progressing toward the superiority of PICSI over ICSI but enough evidences are still not available.

Frequency of Sperm Aneuploidy in Oligoasthenoteratozoospermic (OAT) Patients by Comprehensive Chromosome Screening: A Proof of Concept.

Background:Embryonic aneuploidy usually results in implantation failure and miscarriage. Considering significantly high frequency of sperm aneuploidy reported in oligoasthenoteratozoospermia (OAT) using fluorescence in situ hybridization (FISH) in limited number of chromosomes and lack of comprehensive chromosome screening (CCS) in OAT, the aim of this study was applying CCS in OAT sperm and comparison of the results with FISH findings.Methods:Five OAT patients with normal blood karyotypes and history of implantation failure were included. The successfully amplified samples, each containing two sperm, were analyzed by array comparative genomic hybridization (aCGH). FISH was utilized mainly depending on the aneuploidies found by aCGH to assess their frequencies in total sperm population.Results:In aCGH for 30 sperm, aneuploidy was found in 66% of samples. Following the study of 4300 sperm by FISH, an average of 55.46% aneuploidy was observed. No pregnancy was resulted with normal partners.Conclusion:Using aCGH, some abnormalities were observed that are not typically considered in sperm FISH studies. Despite small sample size of the comprehensive study, like other similar studies, the frequency of aneuploidies was considerable and similar to FISH. Aneuploidies revealed by aCGH at single sperm resolution were different from sperm population detected by FISH. Considering high frequency of aneuploidy in OATs sperm, preimplantation genetic testing for aneuploidy (PGT-A) can be used for in transfer of chromosomally normal embryos.

Decreased prostate‐specific membrane antigen levels in the seminal plasma of oligoasthenoteratozoospermic men

Prostate-specific membrane antigen (PSMA) is a transmembrane glycoprotein with glutamate carboxypeptidase activity. However, its precise function in the prostate, prostasomes and seminal plasma with regard to male fertility remains unknown. This study was conducted to investigate the seminal plasma PSMA levels in fertile men and patients with oligoasthenoteratozoospermia (OAT) and to analyse its association with sperm parameters. Twenty fertile men and twenty patients admitted at the urology clinic of our institution with the diagnosis of OAT were included in the study. Following semen analysis, seminal plasma was isolated from semen ejaculates. PSMA concentrations in the seminal plasma were determined by ELISA. The correlations between seminal PSMA concentrations and semen parameters were statistically analysed. Seminal plasma PSMA concentration was significantly lower in OAT patients compared to fertile controls (p<.01). In fertile men, PSMA concentration was significantly correlated with the sperm concentration (r=-.481, p<.05), whereas in the patient group no statistically significant correlation was found between the sperm parameters and seminal PSMA level. This is the first study in the literature to investigate PSMA levels in the seminal plasma from infertile men. Decreased levels of seminal plasma PSMA might suggest a role for compromised prostasome function in the pathogenesis of OAT syndrome.

Meiotic epigenetic factor PRDM9 impacts sperm quality of hybrid mice.

Reduced fertility of male mouse hybrids relative to their parents, or hybrid sterility, is governed by the hybrid sterility 1 (Hst1) locus. Rescue experiments with transgenes carrying sequences within or near Hst1 manifested that Hst1 contains the gene encoding meiosis-specific histone methyltransferase PRDM9. The Prdm9 gene is responsible for partial meiotic arrest, testicular atrophy, and low sperm count in (C57BL/6J x PWD)F1 mouse hybrids. Here we report that these male hybrids suffer an additional reproductive disadvantage, decreased sperm quality, which is (i) further exacerbated by the introduction of long transgenes carrying sequences from Hst1 with incomplete Prdm9 into their genome and (ii) controlled by the Prdm9 dosage. These transgenic male hybrids displayed the features of severe oligoasthenoteratozoospermia (OAT), a human infertility syndrome characterized by a low number of spermatozoa with poor motility and morphological abnormalities. Analysis of spermiogenesis in these mice revealed acrosome detachment, aberrant elongation and condensation of the nucleus. As a result, the transgenic sperm had acrosome malformations, abnormal chromatin packaging, and fragmented DNA with elevated base oxidation, revealed by using multiple methods. Heterozygosity for one null Prdm9 allele improved meiotic progression and sperm quality of both non- and transgenic hybrids. Our results indicate that genomic analysis of OAT patients should include consideration of allelic variants in PRDM9, and our transgenic models can serve as tools to understand the diverse molecular processes that, when perturbed, can cause this disease.

Sperm Parameters before and after Swim-Up of a Second Ejaculate after a Short Period of Abstinence.

Background: Recent studies have supported the beneficial effects of a short abstinence period on sperm parameters. The aim of this study was to assess sperm motility, morphology and DNA fragmentation before and after swim-up of a second ejaculate obtained after a short abstinence period in normozoospermic men and oligo-astheno-teratozoospermic (OAT) patients. Material and methods: Semen analyses and swim-up preparations of two consecutive semen samples (collected within 1 h) were carried out in 30 normozoospermic and 35 OAT patients enrolled in an assisted reproductive technique (ART) program. Results: Compared to the first ejaculate, the second sample showed a higher percentage of spermatozoa with normal form (p < 0.01) and lower percentage of spermatozoa with DNA fragmentation (p < 0.01) in normozoospermic men, whereas a higher percentage of spermatozoa with progressive motility (p < 0.001) and normal morphology (p < 0.0001) was found in OAT patients. Swim-up separation showed a lower DNA fragmentation rate (p < 0.05) in the second ejaculate in normozoospermic men, whereas the second ejaculate of OAT patents showed an increase in normally-shaped spermatozoa (p < 0.01) and lower percentage of spermatozoa with fragmented DNA (p < 0.001) compared to the first one. Conclusions: Swim-up separation of a second ejaculate collected within 1 h might be suggested for ART procedures, especially in OAT patients.

Importance of Genetic Factors in Male Fertility Disorders, Clinical Relevance of Epigenetic Markers in Male Infertility

Male infertility is a common and complex problem affecting 1 out of 20 men. Despite extensive research in this area, in many cases, the underlying causes are unknown. Epigenetic changes control a series of processes within the body, including male fertility. Classification of infertile men using a more detailed analysis of DNA methylation patterns could reveal a new level of low rates of fertilization, implantation, or pregnancy. In this context, it seemed to us to use the techniques available to evaluate the degree of global methylation of DNA in infertile patients who have modified sperm counts, but also those who apparently do not have a clear cause of infertility. For this we used the Quest 5mC-Zymoresaerch-ELISA kit that can detect within about 5 hours the global level of genome methylation. Claims on which common illnesses have an epigenetic base are still open to speculation, but if true, it can imprint a new direction in medicine. Our data, although from a pilot study, are consistent with those in the literature. A recent study has shown that DNA methylation levels were significantly higher in oligoasthenoteratozoospermia patients than in the control group and the increase in global DNA methylation and histone retention in men with oligoasthenoteratozoospermia.

IMSI procedure improves clinical outcomes and embryo morphokinetics in patients with different aetiologies of male infertility.

Our objective was to evaluate the effect of IMSI on embryo kinetics and clinical outcomes in patients with different aetiologies of male infertility. A total of 150 couples with different aetiologies of male infertility were randomly divided into ICSI and IMSI treatment groups (n=75). ICSI was done accordingly. For IMSI group, the sperm selection was done using MSOME criteria and then injected. The zygotes were cultured in time-lapse monitoring system (TLM) for 3days. A total of 650 embryos were developed and assessed using TLM in two groups. Data showed the rate of fragmentation had significant correlation with different aetiologies (p=0.01), and the timing of s1, t4, s2 and t5 occurred significantly later in oligoasthenoteratozoospermia (OAT) patients compared with others (p<0.05). In IMSI group, there were no differences in the TLM parameters among different aetiologies (p>0.05). The rates of chemical pregnancy and implantation (37.8% and 38.2% respectively) were insignificantly higher in OAT patients compare to others (p>0.05). Also, the clinical pregnancy and live birth rates (32% and 32% respectively) were insignificantly higher in teratozoospermia (T) cases. Sperm selection with MSOME parameters and IMSI can improve the embryo morphokinetics and clinical outcomes in couples with male factor infertility, especially for OAT and T patients.

25-Hydroxy cholecalciferol, anti-Müllerian hormone, and thyroid profiles among infertile men

Background and aim We examined the relationship among vit.D3, AMH, FT3, FT4, and TSH, in addition to the serum levels of reproductive hormones (FSH, LH, prolactin, and free testosterone), in oligoasthenoteratozoospermia and azoospermia patients in a cohort of infertile men from Egypt to establish a clinical marker/cause-effect relationship. Methods This cross-sectional cohort study was carried out on 301 men (105 males with oligoasthenoteratozoospermia and 96 males with azoospermia), in addition to 100 controls. Measurements of serum vit.D3, AMH, FT3, FT4, and TSH levels, in addition to reproductive hormone assays, were performed on all included subjects, using ELISA kits. Results Overall, results showed significantly lower serum levels of vit.D3 in infertile men than in the controls, with a greater decrease observed in men with azoospermia than in oligoasthenoteratozoospermia patients, (p < .05 for all). Significantly higher serum TSH and FSH levels and significantly lower serum free testosterone levels were observed in males with azoospermia than in males with oligoasthenoteratozoospermia and the controls (p < .05 for both). There were no significant differences between the studied groups in terms of AMH, FT3 or FT4 levels. LH levels were negatively correlated with TSH levels and positively correlated with AMH levels among men with oligoasthenoteratozoospermia, while among men with azoospermia, LH levels were positively correlated with vit.D3 levels (p < .05 for all). Conclusion Decreased Vit.D3 could play a role in male infertility, in addition to abnormal thyroid function, which needs further investigation.

Sperm chromatin and DNA integrity, methyltransferase mRNA levels, and global DNA methylation in oligoasthenoteratozoospermia.

ObjectiveTo investigate sperm chromatin/DNA integrity, global DNA methylation, and DNMT mRNA transcription in men with oligoasthenoteratozoospermia (OAT) compared with normozoospermic men.MethodsSemen samples from 32 OAT patients who comprised the case group and 32 normozoospermic men who comprised the control group were isolated and purified using a standard gradient isolation procedure according to World Health Organization criteria. DNMT1, DNMT3A, and DNMT3B transcripts were then compared between groups using real-time quantitative reverse-transcription polymerase chain reaction. Global DNA methylation in sperm was determined by an enzyme-linked immunosorbent assay. Protamine deficiency and the proportion of apoptotic spermatozoa were evaluated using chromomycin A3 (CMA3), aniline blue (AB), and toluidine blue (TB) staining, as well as the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. The p-values <0.05 were considered to indicate statistical significance.ResultsSignificantly higher proportions of AB+, TB+, CMA3+, and TUNEL+ spermatozoa, as well as DNMT3A and DNMT3B transcription, were found in the OAT group. Positive correlations were detected between sperm parameters, DNA/chromatin damage, and DNMT3A and DNMT3B transcripts. Global DNA methylation was significantly higher in the OAT patients and had a significant correlation with abnormal results of all sperm chromatin integrity tests, but was not associated with DNMT1, DNMT3A, or DNMT3B expression.ConclusionOligoasthenoteratozoospermic men showed abnormal sperm parameters, abnormal chromatin/DNA integrity, and a higher global DNA methylation rate, as well as overexpression of DNMT mRNA.