A novel homozygous frameshift mutation in MNS1 associated with severe oligoasthenoteratozoospermia in humans

Oligoasthenoteratozoospermia(OAT)refers to the combination of various sperm abnormalities,including a decreased sperm count,reduced motility,and abnormal sperm morphology.Only a few genetic causes have been shown to be associated with OAT.Herein,we identified a novel homozygous frameshift mutation in meiosis-specific nuclear structural 1(MNS1;NM_018365:c.603_604insG:p.Lys202Glufs*6)by whole-exome sequencing in an OAT proband from a consanguineous Chinese family.Subsequent variant screening identified four additional heterozygous MNS1 variants in 6/219 infertile individuals with oligoasthenospermia,but no MNS1 variants were observed among 223 fertile controls.Immunostaining analysis showed MNS1 to be normally located in the whole-sperm flagella,but was absent in the proband's sperm.Expression analysis by Western blot also confirmed that MNS1 was absent in the proband's sperm.Abnormal flagellum morphology and ultrastructural disturbances in outer doublet microtubules were observed in the proband's sperm.A total of three intracytoplasmic sperm injection cycles were carried out for the proband's wife,but they all failed to lead to a successful pregnancy.Overall,this is the first study to report a loss-of-function mutation in MNS1 causing OAT in a Han Chinese patient.     

FSH dosage effect on conventional sperm parameters:a meta-analysis of randomized controlled studies

Follicle-stimulating hormone(FSH)represents a therapeutic option in normogonadotropic patients with idiopathic oligozoospermia.The aim of this review was to evaluate the possible dose-and drug-dependent efficacy of FSH treatment on conventional sperm parameters.We performed a comprehensive systematic review via a meta-analysis of all available randomized controlled trials,in which FSH administration was compared with placebo or no treatment when administered to normogonadotropic patients with idiopathic oligozoospermia.Of the 971 articles that were retrieved,5 were fin ally included,in eluding a total of 372 patie nts and 294 controls.Overall,FSH treatment was effective in ameliorating the sperm concentration,total count,progressive motility,but not normal forms.On the basis of the weekly dosage,the studies were classified into those using low(175-262.5 IU per week),intermediate(350-525 IU per week),and high(700-1050 IU per week)doses.At low doses,FSH improved only sperm motility.At intermediate doses,FSH ameliorated sperm concentration and morphology.Total sperm count and progressive motility showed a trend toward the increase.At high doses,FSH increased sperm concentration,total sperm count,and progressive motility.Sperm morphology showed a trend toward the increase.Finally,both highly purified FSH(hpFSH)and recombinant human FSH(rhFSH)improved sperm concentration,total sperm count,progressive motility,but not morphology.No different efficacy was observed between these two preparations.This meta-analysis provides evidence in favor of high FSH doses.The FSH efficacy was not related to the preparation type(recombinant vs highly purified).Further studies are needed to evaluate the effectiveness of Iong-standing treatment regimes.     

Management of primary ciliary dyskinesia/Kartagener＇s syndrome in infertile male patients and current progress in defining the underlying genetic mechanism

Kartagener＇s syndrome （KS） is an autosomal recessive genetic disease accounting for approximately 50% of the cases of primary ciliary dyskinesia （PCD）. As it is accompanied by many complications, PCD/KS severely affects the patient＇s quality of life. Therapeutic approaches for PCD/KS aim to enhance prevention, facilitate rapid definitive diagnosis, avoid misdiagnosis, maintain active treatment, control infection and postpone the development of lesions. In male patients, sperm flagella may show impairment in or complete absence of the ability to swing, which ultimately results in male infertility. Assisted reproductive technology will certainly benefit such patients. For PCD/KS patients with completely immotile sperm, intracytoplasmic sperm injection may be very important and even indispensable. Considering the number of PCD/KS susceptibility genes and mutations that are being identified, more extensive genetic screening is indispensable in patients with these diseases. Moreover, further studies into the potential molecular mechanisms of these diseases are required. In this review, we summarize the available information on various aspects of this disease in order to delineate the therapeutic objectives more clearly, and clarify the efficacy of assisted reproductive technology as a means of treatment for patients with PCD/KS-associated infertility.     

Green tea extract as a cryoprotectant additive to preserve the motility and DNA integrity of human spermatozoa

Cryopreservation impairs sperm quality and functions,including motility and DNA integrity.Antioxidant additives in sperm freezing media have previously brought improvements in postthawed sperm quality.Green tea extract(GTE)is widely considered as an excellent antioxidant,and its beneficial role has been proven in other human cells.This study aims to evaluate the GTE as a potential additive in cryopreservation media of human spermatozoa.In part one,the semen of 20 normozoospermic men was used to optimize the concentration of GTE that maintains sperm motility and DNA integrity against oxidative stress,induced by hydrogen peroxide(H₂O₂).Spermatozoa were treated with GTE at different concentrations before incubation with H₂O₂.In part two,the semen of 45 patients was cryopreserved with or without 1.0 ng ml^-1GTE.After 2 weeks,the semen was thawed,and the effect on sperm motility and DNA fragmentation was observed.Our data showed that GTE significantly protected sperm motility and DNA integrity against oxidative stress induced by H₂O₂when added at a final concentration of 1.0 ng ml^-1.We found that the addition of 1.0 ng ml^-1GTE to cryopreservation media significantly increased sperm motility and DNA integrity(both P<0.05).More interestingly,patients with high sperm DNA damage benefited similarly from the GTE supplementation.However,there was no significant change in the reactive oxygen species(ROS)level.In conclusion,supplementing sperm freezing media with GTE has a significant protective effect on human sperm motility and DNA integrity,which may be of clinical interest.     

Genetic and epigenetic risks of intracytoplasmic sperm injection method

Pregnancies achieved by assisted reproduction technologies,particularly by intracytoplasmic sperm injection(ICSI)procedures,are susceptible to genetic risks inherent to the male population treated with ICSI and additionalrisks inherent to this innovative procedure.The documented,as well as the theoretical,risks are discussed in thepresent review study.These risks mainly represent thatconsequences of the genetic abnormalities underlying malesubfertility(or infertility)and might become stimulators for the development of novel approaches and applications inthe treatment of infertility.In addition,risks with a polygenic background appearing at birth as congenital anomaliesand other theoretical or stochastic risks are discussed.Recent data suggest that assisted reproductive technologymight also affect epigenetic characteristics of the male gamete,the female gamete,or might have an impact on earlyembryogenesis.It might be also associated with an increased risk for genomic imprinting abnormalities.(Asian JAndrol 2006 Nov;8:643-673)     

Comparative study of the effects of three semen preparation media on semen analysis,DNA damage and protamine deficiency,and the correlation between DNA integrity and sperm parameters

Semen samples collected from 28 male partners of infertile couples were divided into three equal aliquots and prepared with three selected media,such as PureSperm （Nidacon,Gothenburg,Sweden）,Sil-Select Plus^TM （Fertipro,Beemem,Belgium） and SpermGrad^TM（Vitrolife,Gothenburg,Sweden）. The differences in mean percentages of semen parameters were assessed by repeated measures analysis. Correlations of sperm DNA damage,as measured by terminal deoxynucleotidyl transferase dUTP nick end labeling （TUNEL） assay,and of protamine deficiency,as measured by chromomycin A3 （CMA3） staining with sperm parameters,were determined by Pearson＇s correlation. After preparation with all three media,sperm concentrations decreased （P〈0.05） while percentages of sperm with normal morphology increased （P〈0.05）. Percentages of sperm motility,rapid motility and progressive motile concentration （PMC） increased （P〈0.05） for each ofthese parameters,PureSperm preparation gave the best results （P〈0.05）. The percentage of DNA damage decreased in the PureSperm and Sil-Select Plus preparations （17.9% and 31.3%,respectively,P〈0.05） and increased in the SpermGrad preparation （56.3%,P〈0.05）. Protamine deficiency also decreased in all three kinds of media,59.3%,47.7% and 40.3% for PureSperm,Sil-Select Plus and SpermGrad preparations,respectively （P〈0.05）. The percentage of DNA-damaged sperm was negatively correlated with the percentages of sperm motility,rapid motility and PMC,but was positively correlated with static motility （P〈0.05）. This comparative study and correlation analysis revealed that PureSperm preparation yielded sperm with the best motility and the lowest percentage of protamine deficiency. The Sil-Select Plus preparation yielded sperm with the lowest amount of DNA damage. The SpermGrad preparation had a high percentage of sperm with normal morphology,but also had the highest percentage of sperm with DNA damage. Sperm DNA damage was correlated with percentages of sperm motility,rapid motility,static motility and PMC.     

Sperm DNA damage and its clinical relevance in assessing reproductive outcome

The routine examination of semen, which assesses sperm concentration, percentage motility and morphology,does not identify subtle defects in sperm chromatin architecture. The focus on the genomic integrity of the male gamete has intensified recently due to the growing concern that genetic diseases may be transmitted via assisted reproductive techniques (ART). Accordingly, the intent of this review is to describe the details of the informationpertaining to mitochondfial/nuclear sperm DNA damage with an emphasis on its clinical significance and its relationship with male infertility. Assessment of sperm DNA damage appears to be a potential tool for evaluating semen samples prior to their use in ART. Testing DNA integrity may help select spermatozoa with intact DNA or with the least amount of DNA damage for use in assisted conception. In turn, this may alleviate the financial, social and emotional problems associated with failed ART attempts.     

Sperm DNA fragmentation in Chinese couples with unexplained recurrent pregnancy loss

We aimed to study the association between sperm DNA fragmentation and recurrent pregnancy loss(RPL)in the Chinese population via a retrospective observational study of Chinese couples who had experienced RPL between May 2013 and August 2018.The study population included 461 men from couples with RPL and 411 men from a control group(couples with clinical pregnancy via in v/tro fertiIization owing to female causes).Routine semen analysis,sperm chromatin analysis,and microscopic(high-power)morphological analysis were performed using semen samples.Semen samples were assessed for volume,sperm count,and motility.The sperm DNA fragmentation index(DFI)was calculated,and the median DFI was obtained.Men were categorized as having normal(37.8%;DFI<15.0%),moderate(33.6%;15.0%相似文献   

Focus on intracytoplasmic morphologically selected sperm injection （IMSI）. a mini-review

Intracytoplasmic sperm injection （ICSI） is the recommended treatment in many cases of male-factor infertility. Several studies have demonstrated a positive correlation between optimal sperm morphology and positive ICSI outcomes. In fact, spermatozoa with severe abnormalities of the head are well documented to be associated with low fertilisation, implantation and pregnancy rates. However, a spermatozoon which is classified as ＇normal＇ by microscopic observation at low magnification could contain ultrastructural defects that impair both the fertilisation process and embryonic development. The intracytoplasmic morphologically selected sperm injection （IMSI） procedure changed the perception of how a spermatozoon suitable for injection should appear. Sperm selection is carried out at x 6000 magnification, allowing improved assessment of the sperm nucleus. Currently, standardized clinical indications for I MSI are lacking and the candidates are selected on the grounds of their medical history or of a careful analysis of the sperm suspension. Further prospective randomized studies are needed to confirm the advantages of IMSI in specific groups of patients. In addition to providing a brief overview of the IMSI procedure, this study aims to review the literature, which explains the theoretical basis and the clinical outcomes of this technique. Several reports show that IMSI is associated with improved implantation and clinical pregnancy rates as well as lower abortion rates when compared to ICSI. Although a possible correlation between the sperm＇s abnormal nucleus shape, increased DNA fragmentation and negative laboratory and clinical outcomes has been long investigated, the results are conflicting.     

Function of seminal vesicles and their role on male fertility

The present review has been designed to update the recent developments on the function of seminal vesicles and their role on male fertility. It is indicated that the true corrected fructose level is a simple method for the assessment of the seminal vesicular function. Measttrement of seminal fructose used universally as a marker of the seminal vesicle function is not an appropriate approach due to its inverse relationship with the sperm count. The llue corrected fructose defined as [ log. motile sperm concentration ] multiplied by [ seminal fructose concentration ] has been shown to be a better marker of the seminal vesicle function. Seminal vesicular secretion is important for semen coagulation, sperm motility, and stability of sperm chromatin and suppression of the immune activity in the female reproductive tract. In conclusion, the function of seminal vesicle is important for fertility. Parameters as sperm motility, sperm chromatin stability, and immuno-protection may be changed in case of its hypofunction.     

Clinical significance of the low normal sperm morphology value as proposed in the fifth edition of the WHO Laboratory Manual for the Examination and Processing of Human Semen

The very low cut-off value for sperm morphology of 4% morphologically normal spermatozoa, as proposed in the new edition of the World Health Organization （WHO） manual on semen analysis, is in agreement with recently published values and reflects the trend of a decline in reported mean values for normal sperm morphology. The reduced value for morphologically normal spermatozoa over the years may be due to several factors. The first is the introduction of strict criteria for the evaluation of sperm morphology. Other reasons may include the introduction of additional criteria for sperm morphology abnormalities and the suggested decrease in semen parameters because of increasing negative environmental influences. Although on its own the newly proposed very low normal value may not provide the strong predictive value for a males＇ fertility potential, as originally reported for sperm morphology evaluated according to strict criteria, a good predictive value can still be obtained if the holistic, strict approach for sperm morphology evaluation is followed together with additional sperm morphology parameters now available, because certain morphology patterns and sperm abnormalities are now known to be of strong prognostic value. In addition, better international standardization of the technical methodology, consensus on the interpretation of sperm morphology evaluation criteria and standardized international external quality control （EQC） schemes, are of utmost importance to maintain the good predictive value of sperm morphology.     

Correlation of oxidation reduction potential and total motile sperm count:its utility in the evaluation of male fertility potential

Oxidative stress(OS)is detri me ntal to sperm fun ctio ns,and the oxidation reductio n pote ntial(ORP)is a good measure of OS as it considers the balance betwee n oxidants and reducta nts.Total motile sperm count(TMSC)is viewed as the single most im porta nt semen analysis parameter that can predict male infertility severity,and its correlation with ORP has never been undertaken.The objectives of this study were to assess the correlation between ORP and TMSC,to identify the ORP cutoff value based on the TMSC result,and to compare this cutoff value with previously reported ORP cutoff values in literature.One thousand one hundred and sixty-eight infertile patients and 100 fertile controls were enrolled.Demographic and semen data of the participants were retrieved and analyzed.Wilcox on's rank-sum test compared variables betwee n infertile men and fertile controls;Spearman's correlation assessed the static ORP(sORP)-TMSC relationship for the whole sample and among each group individually.Using a 20×10^6TMSC threshold,receiver operator characteristic(ROC)analysis determined the sORP cutoff associated with the highest predictive values.TMSC was significantly negatively correlated with sORP across all participants(r=0.86,P<0.001),among infertile patients(r=0.729,P<0.001),and among fertile controls(r=0.53,P<0.001).A 20-million TMSC threshold determined an sORP cutoff value of 2.34 mV/106sperm/ml to be associated with 82.9%sensitivity,82.8%specificity,91.5%positive predictive value(PPV),68.5%negative predictive value(NPV),and 82.9%overall accuracy.Compared with previously reported cutoff values in searched literature,the 2.34 mV/10^6sperm/ml cutoff value identified in our study yielded the highest overall diagnostic accuracy in the evaluation of in fertile men.     

Clinical outcomes of microdissection testicular sperm extraction-intracytoplasmic sperm injection with fresh or cryopreserved sperm in patients with nonobstructive azoospermia

We performed this study to evaluate the clinical outcomes of microdissection testicular sperm extraction-intracytoplasmic sperm injection(micro-TESE-ICSI)treatment that used fresh or cryopreserved sperm in patients with nonobstructive azoospermia(NOA).A total of 338 NOA patients with 344 consecutive cycles received treatment in the reproductive medicine center of Peking University Third Hospital in Beijing,China,from January 2014 to December 2017.Fresh oocytes and fresh sperm were used in 222 patients with 234 cycles(Group A).Fresh oocytes and cryopreserved sperm were used in 116 patients with 110 cycles(Group B).We compared patient characteristics,embryonic development,and pregnancy outcomes between Groups A and B.There was no statistical difference in the patient characteristics,and no differences were observed with fertilization or quality embryo rates between Groups A and B.The rates of clinical pregnancy and live birth were both higher for Group A than those for Group B(both P<0.05).In conclusion,fresh testicular sperm appears to produce better ICSI outcomes than cryopreserved testicular sperm in patients with NOA.     

The human acrosome reaction

We developed tests of sperm-oocyte interaction: sperm-zona binding, zona-induced acrosome reaction, spermzona penetration and sperm-oolemma binding, using oocytes which failed to fertilise in clinical in vitro fertilization (IVF). Although oocyte defects contribute to failure of sperm oocyte interaction, rarely are all oocytes from one woman affected. Low or zero fertilization in standard IVFwas usually caused by sperm abnormalities. Poor sperm-zona pellucida binding was frequently associated with failure of standard IVF and obvious defects of sperm motility or morphology. The size and shape of the acrosome is particularly important for sperm binding to the oocyte. The proportion of acrosome intact sperm in the insemination medium was related to the IVF rate. Inducing the acrosome reaction with a calcium ionophore reduced sperm-zona binding. Blocking acrosome dispersal with an acrosin inhibitor prevented spermzona penetration. Sperm-zona penetration was even more highly related to IVF rates than was sperm-zona binding. Some patients had low or zero fertilization rates with standard IVF but normal sperm by conventional tests and normal sperm-zona binding. Few of their sperm underwent the acrosome reaction on the surface of the zona and none penetrated the zona. In contrast, fertilization and pregnancy rates were high with intracytoplasmic sperm injection. We call thiscondition defective zona pellucida induced acrosome reaction. Discovery of the nature of the abnormalities in the signal transduction and effector pathways of the human zona pellucida induced acrosome reaction should result in simpler tests and treatments for the patients and also provide new leads for contraceptive development.     

Effects of acupuncture and moxa treatment in patients with semen abnormalities

Aim: To evaluate the effect of Chinese Traditional Medicine, acupuncture and moxa treatment, on the semen quality in patients with semen abnormalities. Methods: In a prospective, controlled and blind study, nineteen patients, aged 24 years～42 years and married for 3 years～11 years without children with semen abnormalities in concentration, morphology and/or progressive motility without apparent cause, were randomized into two groups and submitted to acupuncture and moxa treatment at the therapeutic (Study Group) and the indifferent points (Control Group), respectively, for l0 weeks. Semen analyses were performed before and after the treatment course. Results:The patients of the Study Group presented a significant increase in the percentage of normal-form sperm compared to the Control Group (calculated U=16.0, critical U=17.0). Conclusion: The Chinese Traditional Medicine acupuncture and moxa techniques significantly increase the percentage of normalform sperm in infertile patients with oligoastenoteratozoospermia without apparent cause.     

Antispermatogenic activity of Morinda lucida extract in male rats

Aim： To investigate the effect of Morinda lucida Benth （Rubiaceae） on the reproductive activity of male albino rats. Methods： Two groups of rats were treated with 400 mg/（kg.d） of Morinda lucida leaf extract for 4 and 13 weeks, respectively. The control rats received the vehicle. All the treated rats had corresponding recovery groups. At the end of each experimental period, animals were killed and organ weights, sperm characteristics, serum testosterone levels, histology of the testes and fertility were assessed. Results： Morinda lucida leaf extract did not cause any changes in body and somatic organ weights, but significantly increased the testis weight （P 〈 0.05）. The sperm motility and viability, and the epididymal sperm counts of rats treated for 13 weeks were significantly reduced （P 〈 0.05）. Sperm morphological abnormalities and serum testosterone levels were significantly increased （P 〈 0.05）. There were various degrees of damage to the seminiferous tubules. The extract reduced the fertility of the treated rats by reducing the litter size. Reversal of these changes, however, occurred after a period of time. Conclusion： The extract of Morinda lucida has reversible antispermatogenic properties.     

Mountant-induced loss of Shorr-stained blue sperm tails

It has been reported that Short staining provides additional morphological information on the motility of spermatozoa in semen,by distinguishing between red and blue flagella. With our routine methods （involving mounting slides） we were unable to confirm these observations. The presence of both red- and blue-coloured sperm tails in Short-stained semen smears was apparent,however,if slides were unmounted. Only a very weak association between blue flagellar staining and immotility was observed. Stating whether a mountant was used should be reported.     

FISH studies of chromosome abnormalities in germ cells and its relevance in reproductive counseling

Chromosome abnormalities are one of the major causes of human infertility. In infertile males, abnormal karyotypes are more frequent than in the general population. Furthermore, meiotic disorders affecting the germ cell-line have been observed in men with normal somatic karyotypes consulting for infertility. In both cases, the production of unbalanced spermatozoa has been demonstrated. Basically addressed to establish reproductive risks, fluorescence in situ hybridization （FISH） on decondensed sperm heads has become the most frequently used method to evaluate the chromosomal constitution of spermatozoa in carriers of numerical sex chromosome abnormalities, carriers of structural chromosome reorganizations and infertile males with normal karyotype. The aim of this review is to present updated figures of the information obtained through sperm FISH studies with an emphasis on its clinical significance. Furthermore, the incorporation of novel FISH-based techniques （Multiplex-FISH; Multi-FISH） in male infertility studies is also discussed. （Asian J Androl 2005 Sep; 7： 227-236）     

A preliminary study of sperm identification in microdissection testicular sperm extraction samples with deep convolutional neural networks

Sperm identification and selection is an essential task when processing human testicular samples for in vitro fertilization.Locating and identifying sperm cell(s)in human testicular biopsy samples is labor intensive and time consuming.We developed a new computer-aided sperm analysis(CASA)system,which utilizes deep learning for near human-level performance on testicular sperm extraction(TESE),trained on a custom dataset.The system automates the identification of sperm in testicular biopsy samples.A dataset of 702 de-identified images from testicular biopsy samples of 30 patients was collected.Each image was normalized and passed through glare filters and diffraction correction.The data were split 80%,10%,and 10%into training,validation,and test sets,respectively.Then,a deep object detection network,composed of a feature extraction network and object detection network,was trained on this dataset.The model was benchmarked against embryologists’performance on the detection task.Our deep learning CASA system achieved a mean average precision(mAP)of 0.741,with an average recall(AR)of 0.376 on our dataset.Our proposed method can work in real time;its speed is effectively limited only by the imaging speed of the microscope.Our results indicate that deep learning-based technologies can improve the efficiency of finding sperm in testicular biopsy samples.