The relationship between DNA fragmentation and the intensity of morphologically abnormal human spermatozoa

Objective:To determine the relationship between teratozoospermia and sperm DNA fragmentation(SDF)in the human ejaculate.Methods:This retrospective study included 100 normozoospermic men as a control cohort(abnormal forms>14%),210 patients with a high level of abnormal forms(≤4%)and 65 patients presenting with a moderate level of abnormal forms(>4%to≤14%)based on the World Health Organization definitions.Sperm morphology was assessed using bright field microscopy.Sperm DNA fragmentation was assessed using the sperm chromatin dispersion assay.Non-parametric analyses were conducted to examine the relationship between abnormal sperm morphology and sperm DNA fragmentation;receiver operating characteristic(ROC)analyses were conducted to assess sensitivity and specificity of this relationship.Results:A correlation analysis revealed that the higher the proportion of abnormal spermatozoa in the ejaculate,the higher the level of SDF(Spearman's Rho=-0.230;P<0.001).Significant differences in the proportion of SDF were found when all cohorts were compared(P<0.001);these significant differences were also retained when the different cohorts were compared pairwise.ROC analysis showed a moderate but significant predictive value for SDF to differentiate patients with different levels of teratozoospemia.Conclusions:Although analysis of a more continuous range of values for teratozoospermia would help further clarify any causal relationship with SDF,there is clearly a synergistic or coincident affiliation between these variables that needs to be acknowledged by the clinician when interpreting the spermiogram.

Resolution of sperm quality impairment following SARS-CoV-2 infection:A prospective study

Objective:To investigate the length of time required to resolve COVID-19 effects on semen quality and DNA integrity.Methods:A prospective cohort study was conducted among 42 men who tested positive for SARS-CoV-2 and underwent semen analysis at baseline and four months’post-recovery.Semen samples were collected and evaluated for macroscopic and microscopic parameters,sperm chromatin maturation,and DNA fragmentation.Results:The mean age of participants was 37(±7)years,and 14%had normozoospermia at baseline.After a four-month recovery from COVID-19,48%of patients had normozoospermia.Sperm count,motility,and morphology increased significantly,while sperm DNA fragmentation and sperm chromatin maturation decreased significantly post-recovery from COVID-19.Conclusions:Sperm parameters improve after a four-month recovery from COVID-19.The findings indicate significant improvements in sperm count,motility,morphology,DNA fragmentation,and chromatin maturation after a four-month recovery period.

Mechanism of action of Wuzi Yanzong pill in the treatment of oligoasthenozoospermia in rats determined via serum metabolomics

Objective:To investigate the mechanism of action of Wuzi Yanzong pill(WYP)in rats with oligoasthenozoospermia(OAZ)via metabolomics and to provide a possible basis for improving this WYP-based treatment.Methods:A rat model of OAZ was established by treating male SpragueeDawley rats with glucosides from Tripterygium wilfordii Hook.F.Seventy-two rats were randomly divided into six groups:control,L-carnitine(positive control),model,and low-,medium-,and high-dose WYP groups.Rats in the experimental groups were treated with WYP for 4 weeks.At the end of the treatment period,sperm cell quality(density,motility,and viability)was assessed using a semen analysis system,mitochondrial membrane potential(MMP)was assessed using flow cytometry,and testicular injury was assessed using hematoxylin and eosin staining to validate the therapeutic effect of WYP in OAZ.Further,serum metabolomics-based analysis was performed using high-performance liquid chromatography-mass spectrometry to identify differential metabolic pathways and possible mechanisms of action of WYP in OAZ treatment.Results:A rat model of OAZ was considered successfully-established after comparing the quality of spermatozoa in the model group to that in the control group.WYP-M and WYP-H treatments significantly improved sperm cell density,motility,and viability compared with those in the model group(all P<.05).Compared with the model group,both WYP-M and WYP-H treatments increased MMP values(P=.006 and P=.021 respectively),while there was no significant difference in the L-carnitine group.L-carnitine and WYP administration reversed damage to the testes to varying degrees compared with that in the model group.Further,44 differential metabolites and four metabolic pathways,especially autophagy pathway,related to OAZ were identified via metabolomics.Conclusions:WYP improves sperm cell quality and MMP in OAZ primarily via autophagy regulation.These findings can be employed to improve the efficacy of WYP in humans.

Chromatin condensation but not DNA integrity of pig sperm is greater in the sperm-rich fraction

Background Protamination and condensation of sperm chromatin as well as DNA integrity play an essential role during fertilization and embryo development.In some mammals,like pigs,ejaculates are emitted in three separate fractions:pre-sperm,sperm-rich(SRF)and post sperm-rich(PSRF).These fractions are known to vary in volume,sperm concentration and quality,as well as in the origin and composition of seminal plasma(SP),with differences being also observed within the SRF one.Yet,whether disparities in the DNA integrity and chromatin condensation and pro-tamination of their sperm exist has not been interrogated.Results This study determined chromatin protamination(Chromomycin A3 test,CMA_(3)),condensation(Dibromobi-mane test,DBB),and DNA integrity(Comet assay)in the pig sperm contained in the first 10 m L of the SRF(SRF-P1),the remaining portion of the sperm-rich fraction(SRF-P2),and the post sperm-rich fraction(PSRF).While chromatin protamination was found to be similar between the different ejaculate fractions(P>0.05),chromatin condensation was seen to be greater in SRF-P1 and SRF-P2 than in the PSRF(P=0.018 and P=0.004,respectively).Regarding DNA integrity,no differences between fractions were observed(P>0.05).As the SRF-P1 has the highest sperm concentra-tion and ejaculate fractions are known to differ in antioxidant composition,the oxidative stress index(OSi)in SP,calcu-lated as total oxidant activity divided by total antioxidant capacity,was tested and confirmed to be higher in the SRF-P1 than in SRF-P2 and PSRF(0.42±0.06 vs.0.23±0.09 and 0.08±0.00,respectively;P<0.01);this index,in addition,was observed to be correlated to the sperm concentration of each fraction(Rs=0.973;P<0.001).Conclusion While sperm DNA integrity was not found to differ between ejaculate fractions,SRF-P1 and SRF-P2 were observed to exhibit greater chromatin condensation than the PSRF.This could be related to the OSi of each fraction.

Sperm function, mitochondrial activity and in vivo fertility are associated to their mitochondrial DNA content in pigs

Background Despite their low abundance in sperm, mitochondria have diverse functions in this cell type, includ-ing energy production, signalling and calcium regulation. In humans, sperm mitochondrial DNA content(mtDNAc) has been reported to be negatively linked to sperm function and fertility. Yet, the association between mtDNAc and sperm function in livestock remains unexplored. For this reason, this study aimed to shed some light on the link between mtDNAc and sperm function and fertilising potential in pigs. A qPCR method for mtDNAc quantification was optimised for pig sperm, and the association of this parameter with sperm motility, kinematics, mitochondrial activity, and fertility was subsequently interrogated.Results First, the q PCR method was found to be sensitive and efficient for mtDNAc quantification in pig sperm. By using this technique, mtDNAc was observed to be associated to sperm motility, mitochondrial activity and in vivo, but not in vitro, fertility outcomes. Specifically, sperm with low mtDNAc were seen to exhibit greater motility but decreased mitochondrial activity and intracellular reactive oxygen species. Interestingly, samples with lower mtD-NAc showed higher conception and farrowing rates, but similar in vitro fertilisation rates and embryo development, when compared to those with greater mtDNAc.Conclusions These findings enrich our comprehension of the association of mtDNAc with sperm biology, and lay the foundation for future research into employing this parameter as a molecular predictor for sperm function and fer-tility in livestock.

Microfluidic thermotaxic selection of highlymotile sperm and in vitro fertilization

The selection of the most motile and functionally competent sperm is an essential basis for in vitro fertilization(IVF)and normal embryonic development.Widely adopted clinical approaches for sperm sample processing intensely rely on centrifugation and wash steps that may induce mechanical damage and oxidative stress to sperm.Although a few microfluidic sperm sorting devices may avoid these adverse effects by exploiting intrinsic guidance mechanisms of sperm swimming,none of these approaches have been fully validated by clinical-grade assessment criteria.In this study,a microfluidic sperm sorting device that enables the selection of highly motile and functional sperm via their intrinsic thermotaxis is presented.Bioinspired by the temperature microenvironment in the fallopian tube during natural sperm selection,a microfluidic device with controllable temperature gradients along the sperm separation channel was designed and fabricated.This study investigated the optimal temperature conditions for human sperm selection and fully characterized thermotaxis-selected sperm with 45 human sperm samples.Results indicated that a temperature range of 35–36.5℃along the separation channel significantly improves human sperm motility rate((85.25±6.28)%vs.(60.72±1.37)%;P=0.0484),increases normal sperm morphology rate((16.42±1.43)%vs.(12.55±0.88)%;P<0.0001),and reduces DNA fragmentation((7.44±0.79)%vs.(10.36±0.72)%;P=0.0485)compared to the nonthermotaxis group.Sperm thermotaxis is species-specific,and selected mouse sperm displayed the highest motility in response to a temperature range of 36–37.5℃along the separation channel.Furthermore,IVF experiments indicated that the selected sperm permitted an increased fertilization rate and improved embryonic development from zygote to blastocyst.This microfluidic thermotaxic selection approach will be translated into clinical practice to improve the IVF success rate for patients with oligozoospermia and asthenozoospermia.

Protective effect of co-enzyme Q10 on testicular tissue and sperm parameters in adult male rats treated with Sunset Yellow FCF

Objective:To determine the protective effect of co-enzyme Q10(CoQ10)on testicular tissue and sperm parameters in male rats treated with SunsetYellow FCF.Methods:Sixty male Sprague-Dawley rats were randomly divided into 6 groups of the control,CoQ10(10 mg/kg/day),low dose of Sunset Yellow(2.5 mg/kg),high dose of Sunset Yellow(70 mg/kg),low dose of Sunset Yellow(2.5 mg/kg)plus CoQ10,and high dose of Sunset Yellow(70 mg/kg)plus CoQ10.The drugs were administered via daily oral gavages for 6 weeks.At the end of the experiment,sperm analysis,stereological and histological assessments of the testis were carried out.Results:The normal morphology(by 41.1%)and progressive spermatozoa(by 74.8%),testicle volume(by 33.4%),lumen volume(by 38.3%),interstitial tissue volume(by 44.7%),seminiferous tubule volume(by 40.7%),and number of spermatogonia(by 53.9%)and Leydig cells(by 70.7%)reduced in the rats that received high doses of Sunset Yellow in comparison to the control group.Nonetheless,all these alterations were recovered by CoQ10 treatment in the CoQ10 plus high dose of Sunset Yellow group.Furthermore,low doses of Sunset Yellow did not affect different parameters of the testis and sperm.Conclusions:CoQ10 could,to some extent,prevent structural changes of the testis induced by the high dose of SunsetYellow.

Human circBOULE RNAs as potential biomarkers for sperm quality and male infertility

Reliable molecular biomarkers to predict fertility remain scarce.The current study investigated the potential of testis-specific circBOULE RNAs as biomarkers for male infertility and sperm quality.Using reverse transcription-PCR and real-time reverse transcription-PCR assays,we identified seven circular RNAs from the human BOULE gene in human sperm.We observed that the expression level of circEx3-6 was significantly reduced in asthenozoospermia,while the expression levels of both circEx2-6 and circEx2-7 were decreased in terato-zoospermia,compared with the controls.Furthermore,we demonstrated that the expression level of circEx2-6 was negatively correlated with the sperm DNA fragmentation index,and the expression level of circEx2-7 was correlated with both fertilization and cleavage rates in those treated with the assisted reproductive technologies.Further functional analyses in a transgenic fly model supported the roles of circBOULE RNAs in sperm development and human male fertility.Collectively,our findings support that sperm circBOULE RNAs may serve as diagnostic biomarkers for assessing sperm motility and DNA quality.Therefore,clinical application and significance of sperm circBOULE RNAs in the assisted reproductive technologies warrant further investigation.

Pig Reproductive and Respiratory Syndrome Virus (PRRSV) Does Not Attach to Boar Sperm;It Affects Only the Velocity Pattern and the Mobility Pattern

The purpose of the study was to evaluate the sperm viability of semen infected with PRRSV viral particles, observing the effect of the Virus on the motility of boar sperm. The work was carried out at the FMVZ-BUAP Genetics and Reproduction Laboratory. 5 stallions were used. Each sample contained 1 × 106 sperm, the PRRS virus strain was ATCC-VR-2332 (0, 102, 104 and 106 copies of RNA/mL in triplicate), it was observed daily at the CASA;Hamilton Thorne®. Cells with MT (P < 0.05) on days 1, 3, 5, 7 and 10 of evaluation with 201 ± 7.3, 167 ± 10.1, 165 ± 14.6, 134 ± 8.2 and 120 ± 8.8, respectively. The % MP between control and virus concentrations (P ≥ 0.05). The LCV on day 1 and 7 PI at 10X2 and 10X6 (P < 0.05) vs control. In the Correlation Matrix, where it is observed that there is a correlation between VSL and VAP, VSL and VCL, VCL and ALH, VAP with ALH. There is a correlation of VSL and ALH, STR and ALH. In this study there were (P ≤ 0.01) in the VCL, in the concentrations (102) 162.81 ± 10.65 and (106) 177.12 ± 5.77 vs 193.04 ± 4.62 of control. This indicates that altering these parameters would be related to fertility and the PRRS virus affects the LCV. Regarding the VSL, it was observed that the sperm infected with viruses 102, 104 and 106 of 48.00 ± 3.38, 49.88 ± 1.83 and 50.55 ± 2.24 Vs. 56.66 ± 1.68 of control respectively, the control would have greater possibilities of fertilizing the oocyte. In this study, it was found (P ≤ 0.01) in the VAP with 102 of 77.26 ± 5.16, 104 with 83.35 ± 2.41 and 106 with 81.29 ± 3.14 vs the control with 90.56 ± 2.07. Regarding the ALH there is (P < 0.05) a 104 with 8.70 ± .26 and 106 with 9.64 ± 0.23 vs control 8.50 ± 0.27. The presence of different concentrations of PRRSV in boar semen induces changes in different types of sperm motility. Infection of ejaculates with the PRRS virus affects sperm motility on days 1, 3, 5, 7, and 10 post-infections.

Low Level of Cystic Fibrosis Transmembrane Conductance Regulator Is Associated with Human Sperm Autophagy and Vitality

Low sperm motility is one of the main causes of male infertility. Cystic fibrosis transmembrane conductance regulator (CFTR, an anion channel protein) is related to the progressive motility of sperm. CFTR disruptor CFTRinh-172 or forskolin (FSK) in this study were used to treat human sperm separately, and the rates of sperm autophagy and progressive motility, mitochondrial membrane potential (MMP) and ATP concentration, and the expression levels of related factors were detected to explore their relationship. It was showed that sperms treated with CFTRinh-172 or FSK reduced the levels of cAMP, CFTR and PKA, but increased sperm autophagy rate, expression levels of AMPK and LC3B. However, reactive oxygen species content had no significant difference. It was indicated that low level of CFTR performed with cAMP and its downstream effectors such as PKA and AMPK to regulate mitochondrial structure and function, leading to increased autophagy rate and reduced vitality of sperm.

Laboratory and Clinical Outcomes of Single Sperm Cryopreservation in Patients Underwent Micro-TESE

For men with severe oligozoospermia, sperm cryopreservation can preserve surgically obtained sperm. How to cryopreserve single sperm in men is still a hot topic in assisted reproduction technology. Aim to analyze the laboratory and pregnancy outcomes of single sperm cryopreservation group, we retrospectively selected 38 cycles underwent single sperm cryopreservation and thawing as the study group and 618 cycles underwent conventional sperm cryopreservation and thawing as the control group, which were performed in the reproductive medicine center of the Sixth Affiliated Hospital, Sun Yatsen University, from April 2014 to October 2023. All the sperm came from microdissection testicular sperm extraction (micro-TESE), and performed intracytoplasmic sperm injection (ICSI) for fertilization. Zygotes were cultured to Day 3 embryo, which were freshly transferred to female uterus. Surplus embryos were cultured to blastosphere and cryopreserved. There was no statistical difference in female/male age, female BMI, infertility duration and female basal sex hormone (FSH, LH E2, AMH), No. of oocytes retrieved per cycle, No. of ICSI oocytes per cycle and No. of embryos transferred per cycle between the two groups (P > 0.05). No significant difference was found in two-pronuclear oocyte fertilization rate (59.23% VS 58.84%), Day 3 available embryo rate (61.81% VS 63.55%), Day 3 good-quality embryo rate (45.73% VS 50.27%), blastocyst formation rate (47.83% VS 49.46%), the implantation rate (47.37% VS 52.16%), clinical pregnancy rate (36.84% VS 47.18%), miscarriage rate (14.29% VS 12.68%) and live birth rate (85.71% VS 81.70%) between two groups (P > 0.05). In conclusion, single-sperm cryopreservation was the optimal method to preserve sperm after micro-TESE. It can increase the utilization of each sperm and lead to clinical pregnancy.

Tales of the Tail and Sperm Head Aches --Changing concepts on the prognostic significance of sperm pathologies affecting the head, neck and tail

This article presents an update on the variable prognostic significance of different sperm pathologies in patients with severe male factor infertility due to morphology and motility disorders. Severe asthenozoospermia is one of the leading causes of male infertility as spermatozoa cannot reach the oocyte and/or penetrate normally. Identifying structural causes of sperm immotility was of great concern before the advent of intracytoplasmic sperm injection （ICSI）, because immotility was the limiting factor in the treatment of these patients. In these cases, in vitro methods are used to identify live spermatozoa or stimulate sperm motility to avoid selection of non-viable cells. With these advances, fertilization and pregnancy results have improved dramatically. The identification of genetic phenotypes in asthenozoospermia is important to adequately inform patients of treatment outcomes and risks. The one sperm characteristic that seriously affects fertility prognosis is teratozoospermia, primarily sperm head and neck anomalies. Defects of chromatin condensation and acrosomal hypoplasia are the two most common abnormalities in severe teratozoospermia. The introduction of microscopic methods to select spermatozoa and the development of new ones to evaluate sperm quality before ICSI will assure that ultrastructural identification ofsperm pathologies will not only be of academic interest, but will also be an essential tool to inform treatment choice. Herein, we review the differential roles played by sperm components in normal fertilization and early embryo development and explore how assisted reproductive technologies have modified our concepts on the prognostic significance of sperm pathologies affecting the head, neck, mid-piece and tail.

Determination of double-and single-stranded DNA breaks in bovine sperm is predictive of their fertilizing capacity

Background:The analysis of chromatin integrity has become an important determinant of sperm quality.In frozenthawed bovine sperm,neither the sequence of post-thaw injury events nor the dynamics of different types of sperm DNA breaks are well understood.The aim of the present work was to describe such sperm degradation aftermath focusing on DNA damage dynamics,and to assess if this parameter can predict pregnancy rates in cattle.Results:A total of 75 cryopreserved ejaculates from 25 Holstein bulls were evaluated at two post-thawing periods(0-2 h and 2-4 h),analyzing global and double-stranded DNA damage through alkaline and neutral Comet assays,chromatin deprotamination and decondensation,sperm motility,viability,acrosomal status,and intracellular levels of total ROS,superoxides and calcium.Insemination of 59,605 females was conducted using sperm from the same bulls,thus obtaining the non-return to estrus rates after 90 d(NRR).Results showed an increased rate of double-stranded breaks in the first period(0-2 h:1.29±1.01%/h vs.2-4 h:0.13±1.37%/h;P<0.01),whereas the rate of sperm with moderate+high single-stranded breaks was higher in the second period(0-2 h:3.52±7.77%/h vs.2-4h:21.06±11.69%/h;P<0.0001).Regarding sperm physiology,viability decrease rate was different between the two periods(0-2 h:-4.49±1.79%/h vs.2-4 h:-2.50±3.39%/h;P=0.032),but the progressive motility decrease rate was constant throughout post-thawing incubation(0-2 h:-4.70±3.42%/h vs.2-4 h:-1.89±2.97%/h;P>0.05).Finally,whereas no correlations between bull fertility and any dynamic parameter were found,there were correlations between the NRR and the basal percentage of highly-damaged sperm assessed with the alkaline Comet(Rs=-0.563,P=0.003),between NRR and basal progressive motility(Rs=0.511,P=0.009),and between NRR and sperm with high ROS at 4 h post-thaw(Rs=0.564,P=0.003).Conclusion:The statistically significant correlations found between intracellular ROS,sperm viability,sperm motility,DNA damage and chromatin deprotamination suggested a sequence of events all driven by oxidative stress,where viability and motility would be affected first and sperm chromatin would be altered at a later stage,thus suggesting that bovine sperm should be used for fertilization within 2 h post-thaw.Fertility correlations supported that the assessment of global DNA damage through the Comet assay may help predict bull fertility.

Bull spermatozoa selected by thermotaxis exhibit high DNA integrity,specific head morphometry,and improve ICSI outcome

Background Sperm migration by thermotaxis is a guidance mechanism that operates along the oviduct and it has proved to be a valid method for selecting spermatozoa with low DNA fragmentation(SDF)in mice,humans,and stallions.This study aimed to analyse if bull spermatozoa could be selected by thermotaxis and to assess their quality in terms of SDF as well as determine the presence of a specific sperm subpopulation based on sperm morphometry and assess their fertilizing capacity by ICSI.Methods We used frozen-thawed sperm from 6 bulls and sperm selection by thermotaxis was performed with TALP medium supplemented with 25 mmol/L of HEPES and 5 mmol/L of caffeine.In these conditions,sperm selection was achieved,obtaining a net thermotaxis of 3.6%.Subsequently,we analysed the SDF of the migrated and not-migrated spermatozoa using the neutral COMET assay,and we evaluated the size of the sperm head using Hemacolor■ staining with Motic Images Plus 3 software.Additionally,migrated and not-migrated spermatozoa by thermotaxis were used to fertilize bovine in vitro matured(IVM)oocytes by ICSI,a very inefficient procedure in cattle that is only successful when the oocyte is artificially activated.Results The results showed lower SDF(χ^(2),P<0.001,13.3%reduction,n=8)and lower head size parameters(length and width,P<0.01;and perimeter and area,P<0.001;n=4)in those spermatozoa migrated in comparison to those not-migrated.The distribution of sperm subpopulations structure varied between groups,highlighting cluster 2,characterized by spermatozoa with small head size,and high ellipticity and elongated heads,as the most abundant in the thermotaxis migrated group.When performed ICSI(without oocyte artificial activation)with the thermotactic sperm,the blastocyst rate was 32.2%±9.3%in the group microinjected with the thermotactic spermatozoa vs.8.3%±7.8%in the group of not-migrated sperm(χ^(2),P<0.05).Conclusion Our results showed that bull sperm selection by thermotaxis has a much higher DNA integrity,small and elongated head size parameters,and different sperm subpopulation structure than the not-selected spermatozoa.Additionally,we evidenced that thermotactic spermatozoa improve ICSI success rates.

Bacteriospermia among smallholder artificial insemination boars in the Philippines and potential associated factors

Objective:To determine the prevalence of bacteriospermia,the bacterial load,and the potential factors associated with bacterial contamination in boar semen collected by local smallholder artificial insemination operators.Methods:Fifteen individual raw semen samples were collected from locally available artificial insemination boars owned by different smallholder boar operators within the 5th district of Leyte,Philippines and were subjected to standard bacteriological culture and identification,including a survey of potentially associated factors.Prevalence and bacterial count were determined accordingly,while boar characteristics and collection practices were clustered following agglomerative hierarchical clustering technique.Results:One hundred percent contamination with a bacterial count of(2.01±0.38)×10^(3) CFU/mL was observed.At least 73.33%of the samples were positive for Bacillus spp.,while other identified isolates included Enterobacter spp.,Staphylococcus spp.,E.coli,Pseudomonas spp.,Citrobacter spp.,and Klebsiella spp.Conclusions:Despite the high prevalence of bacteriospermia,the bacterial count is low.Nevertheless,on-farm practices on boar health and management,semen collection,and sanitation as well as the enhancement of basic protocols to control contamination should be conscientiously considered in smallholder artificial insemination operation.

Can the Prediction of Intrauterine Insemination Results by Used Aniline Blue Stain (ABS) and Sperm Chromatin Dispersion (SCD) Levels?

Introduction: This study aimed to perform routine seminal fluid analysis, sperm DNA fragmentation, and sperm function tests at the chromatin maturation level and evaluate pregnancy in the patients passing intrauterine insemination before starting Intrauterine Insemination (IUI) method. Materials and Methods: In this prospective study, 111 couples who underwent Intrauterine Insemination (IUI) in unexplained infertility patients were admitted to Al-Farah IVF and assisted reproductive center in Baghdad, Iraq between November 2020 and February 2021 were evaluated. Semen fluid analysis was performed based on (WHO 4th) guiding rules. In addition, Sperm Chromatin Dispersion (halo test) and sperm maturation were performed with Aniline Blue Stain (ABS). Results: Sperm Chromatin Dispersion (SCD) groups were compared in terms of pregnancy outcome;the positive pregnancy rate was found to be above in the normal SCD groups (p = 0.0005). In addition, Aniline Blue Stain (ABS) groups were compared in the terms of pregnancy outcome;the positive pregnancy rate was found to be higher in the normal ABS group (p = 0.017). Conclusion: Our study showed that the use of DNA fragmentation (SCD) and sperm maturation tests (ABS) together with routine semen analysis in intrauterine insemination cases will make a significant contribution to the prediction of Intrauterine Insemination (IUI) increased results. So, these results indicate a defect in the effect of DNA fragmentation on the outcome of intrauterine insemination.

Intervention of Xuduan Zhongzi Formula on spermatogenesis epididymal morphological changes in a mice model of oligospermia

Objective:To investigate the intervention of Xuduan Zhongzi Formula on the epididymis structure of model of oligospermia mice.Methods:Ten of the 45 male mice were labeled as the normal group,and the remaining 35 mice were injected with chloral hydrate for five consecutive days,and the randomly selected five mice were anesthetized with chloral hydrate.The left epididymis was removed,and a few sperm were found in the epididymis under the optical microscope,indicating successful construction of the model.They were randomly divided into three groups:the normal group,the L-carnitine group and the Xuduan group with 10 rats in each group.The the L-carnitine group and the Xuduan group were administrated with equivalent dose of human solvent,and the normal group and the model group were administrated with normal saline.After 8 weeks of intragastric administration,all experimental mice were killed,and the left epididymis was extracted for sperm detection,seminal plasma biochemical detection,HE staining,and electron microscopy.Results:In the model group,enlarged epididymal epithelial cells,vacuolar degeneration of primary and basal cells,and edema of interstitial cells and vascular dilation were found.Compared with the normal group,semen concentration,activity,SOD,A-glucosidase and fructose in the model group were significantly decreased,and the difference was statistically significant(P<0.01).After eight weeks of drug intragastric treatment,the semen concentration,activity,SOD,A-glucosidase and fructose in the Xuduan group were significantly increased compared with the model group,and the difference was statistically significant(P<0.01).Conclusion:Xuduan Zhongzi Formula can significantly improve the epididymal structure,the microenvironment of epididymal sperm maturation and the stability of epididymal epithelial structure of oligospermia model mice,creating conditions for sperm maturation in the epididymis.

Sperm collection in Black-legged Kittiwakes and characterization of sperm velocity and morphology

Background:Collecting and studying live sperm is central to many important fields of biology.Yet,a simple method to collect live sperm is lacking in wild seabird species.Here,we describe a non?invasive method to collect viable sperm samples based on a simple massage technique applied to male Black?legged Kittiwakes(Rissa tridactyla).Methods:We studied a colony breeding at Kongsfjorden,Svalbard and successfully obtained sperm samples from 32 males.With a subset of samples(n = 12 males),we compared the suitability of several extenders(0.9% NaCl,PBS,Earle's balance salt solution,Dulbecco's modified Eagle medium) in maintaining sperm alive long enough for analyses.With another 18 ejaculates,we conducted computer assisted sperm analyses using the CASA plugin for ImageJ.We provide details about the settings to be used for such analyses.Lastly,droplets from 20 ejaculates were smeared on glass slides and preserved with formalin to characterize sperm morphology in terms of total sperm length,sperm head length,midpiece length and flagellum length,and percentage of abnormal sperm.Results:With this method and under field conditions,we were able to obtain sufficient amounts of live sperm to assess traits related to sperm quality(e.g.sperm morphology,percentage of motile sperm,sperm velocity).We found that two extenders,Earle's balanced salt solution and Dulbecco modified Eagle's medium,yielded similarly good results.Additionally,we investigated whether specific behaviours were associated with successful sperm collection and whether sperm collection success depended on how long before laying sperm collection was attempted.Finally,we provide mean values for sperm morphology,sperm swimming ability and percentage of motile sperm,which may prove useful for future comparative analyses,and we report high levels of sperm abnormality and within?ejaculate variation in sperm morphology.Conclusions:We discuss the high percentage of abnormal sperm and high within?ejaculate variation in sperm morphology in light of sperm competition theory and conclude that these figures are likely due to relaxed post?cop?ulatory sexual selection,kittiwakes being strictly monogamous.Finally,we suggest that this method could be applied to other seabird species sharing similar ecology.

Microdissection testicular sperm extraction： an update

Patients with non-obstructive azoospermia （NOA） were once considered to be infertile with few treatment options due to the absence of sperm in the ejaculate. In the last two decades, the advent of intracytoplasmic sperm injection （ICSI）, and the application of various testicular sperm retrieval techniques, including fine needle aspiration （FNA）, conventional testicular sperm extraction （TESE） and microdissection testicular sperm extraction （micro-TESE） have revolutionized treatment in this group of men. Because most men with NOA will have isolated regions of spermatogenesis within the testis, studies have illustrated that sperm can be retrieved in most men with NOA, including Klinefelter＇s syndrome （KS）, prior history of chemotherapy and cryptorchidism. Micro-TESE, when compared with conventional TESE has a higher sperm retrieval rate （SRR） with fewer postoperative complications and negative effects on testicular function. In this article, we will compare the efficacy of the different procedures of sperm extraction, discuss the medical treatment and the role of testosterone optimization in men with NOA and describe the micro-TESE surgical technique. Furthermore, we will update our overall experience to allow counseling on the prognosis of sperm retrieval for the specific subsets of NOA.

Assessing human sperm morphology： top models, underdogs or biometrics？

The assessment of the percentage of spermatozoa having an ＇ideal＇ morphology using so-called strict method is the method recommended in the latest edition of the World Health Organization （WHO） laboratory manual for semen analysis. This recommendation is a result of the statistical association between ＇ideal＇ sperm morphology and fertility, and of the current general belief that sperm morphology assessment should be used primarily as a fertility tool. The notion of an ＇ideal＇ sperm morphology has persisted despite the very low percentage of such spermatozoa in the semen of fertile men, a subject of intense controversy. The detailed categorization of each abnormal spermatozoon has thus, for a long time, been considered optional and partially redundant, an idea which is reflected in the earlier editions of the WHO manual. However, several recent studies have shown the importance of carefully assessing abnormal sperm morphology for use in the diagnosis ＆infertility, to determine fertility prognosis, and for basic or public health studies. One approach, which combines videomicroscopy and computer vision, and is the only approach able to assess the continuum of sperm biometrics, has been used successfully in several recent clinical, basic and toxicology studies. In summary, the visual assessment of detailed sperm morphology--including the categorization of anomalies allowing arithmetically derived indices of teratozoospermia--and the more modern computer-based approaches, although often considered to be redundant, are in fact complementary. The choice of the most appropriate method depends on the field of investigation （clinical, research, toxicology） and the problem being addressed. Each approach has advantages as well as certain limitations, which will be discussed briefly herein.