Novel association between sperm deformity index and oxidative stress-induced DNA damage in infertile male patients

Aim:To investigate the impact of abnormal sperm morphology using the sperm deformity index (SDI) on reactive oxygen species (ROS) production and its correlation with sperm DNA damage.Methods:Semen samples were collected from men undergoing infertility screening (n=7) and healthy donors (n=6).Mature spermatozoa were isolated and incubated with 5 mmol/L β-nicotinamide adenine dinucleotide phosphate (NADPH) for up to 24 h to induce ROS.Sperm morphology was evaluated using strict Tygerberg's criteria and the SDI.ROS levels and DNA damage were assessed using chemiluminescence and terminal deoxynucleotidyl transferase-mediated fluorescein- dUTP nick end labeling (TUNEL) assays,respectively.Results:SDI values (median [interquartiles]) were higher in patients than donors (2 [1.8,2.1] vs.1.53 [1.52,1.58],P=0.008).Aliquots treated with NADPH showed higher ROS levels (1.22 [0.30,1.87] vs.0.39 [0.10,0.57],P=0.03) and higher incidence of DNA damage than those not treated (10 [4.69,24.85] vs.3.85 [2.58,5.10],P=0.008).Higher DNA damage was also seen following 24 h of incubation in patients compared to donors.SDI correlated with the percentage increase in sperm DNA damage following incubation for 24 h in samples treated with NADPH (r=0.7,P=0.008) and controls (r=0.58,P=0.04). Conclusion:SDI may be a useful tool in identifying potential infertile males with abnormal prevalence of oxidative stress (OS)-induced DNA damage.NADPH plays a role in ROS-mediated sperm DNA damage,which appears to be more evident in infertile patients with semen samples containing a high incidence of morphologically abnormal spermatozoa.

Second to fourth digit ratio (2D:4D) and testosterone in men

Aim: To investigate the relationship between 2D:4D and testosterone in men attending an infertility clinic and men drawn from the general population. Methods: Data on 2D:4D and testosterone from two samples were collected: (1) 43 men attending an infertility clinic, and (2) 51 men drawn from the general population without regard to fertility. Results: In sample (1) there were negative associations between 2D:4D and testicular function, and men with lower 2D:4D in their right compared to left hand had higher testosterone levels than men with higher 2D:4D in their right compared to left hand. Sample (2) showed no significant associations between 2D:4D or side differences in 2E>:4D and testosterone. Conclusion: Adult levels of testosterone may be related to aspects of 2D:4D in samples which contain men with compromised testicular function, but not in men from normative samples. Associations between 2D:4D and fertility-associated traits probably arise from early organisational effects of testosterone rather than from activational effects of current testosterone.

Obesity leads to higher risk of sperm DNA damage in infertile patients

There has been a growing interest over the past few years in the impact of male nutrition on fertility. Infertility has been linked to male overweight or obesity, and conventional semen parameter values seem to be altered in case of high body mass index （BMI）. A few studies assessing the impact of BMI on sperm DNA integrity have been published, but they did not lead to a strong consensus. Our objective was to explore further the relationship between sperm DNA integrity and BMI, through a 3-year multicentre study. Three hundred and thirty male partners in subfertile couples were included. Using the terminal uridine nick-end labelling （TUNEL） assay, we observed an increased rate of sDerm DNA damage in obese men （odds ratio （95% confidence interval）： 2.5 （1.2-5.1））.

Predictive value of hypo-osmotic swelling test to identifyviable non-motile sperm

Aim: To determine the predictive value of the hypo-osmotic swelling (HOS) test to identify viable, non-motile sperm. Methods: Semen samples from 20 men with severe asthenozoospermia underwent traditional seminal analysis, eosin-nigrosin (EN) staining and the HOS test. A further EN stain was then performed on a HOS pre-treated aliquot and a total of 2000 further sperm examined. Results: The median sperm density was 5.1 million/mL (IQR 4.3-13.1) and the median motility was 3.0 % (IQR 0-7). Seven samples showed complete asthenozoospermia. Initial EN staining showed 59 % viability (range 48-69) despite the poor standard parameters and 47 % (range 33-61) in the complete asthenozoospermia subgroup. The HOS test showed 49.9 % reacted overall (range 40-59) and 41.7 % (range 22-61) in the complete asthenozoospermia subgroup. The combined HOS/EN stain showed the positive predictive value of the HOS test to identify viable sperm was 84.2 % overall and 79.7 % in the complete asthenozoospermia subgroup. Conclusion: The HOS test can effectively predict sperm viability in patients with severe and complete asthenozoospermia.

Androgen receptors are expressed in a variety of human fetal extragenital tissues： an immunohistochemical study

Aim： To investigate the expression of androgen receptors in the extragenital tissues of developing human embryo. Methods： Using immunohistochemistry, we investigated the distribution of androgen receptor （AR） in the extragenital tissues of paraffin-embedded tissue sections of first trimester （8-12 weeks gestation） human embryos. Gender was determined by polymerized chain reaction. Results： There were no differences in the expression and distribution of AR in male and female embryos at any stage of gestation. AR expression was seen in the thymus gland. The bronchial epithelium of the lungs showed intense positive staining with surrounding stroma negative. Furthermore, positive staining for androgen receptor was exhibited in the spinal cord with a few positive cells in the surrounding tissues. Cardiac valves also showed strong positive staining but with faint reactivity of the surrounding cardiac muscle. There was no staining in kidney, adrenal, liver or bowel. Conclusion： This study demonstrates that immunoreactive AR protein is present in a wide variety of human first trimester fetal tissues and shows the potential for androgen affecting tissues, which are mostly not considered to be androgen dependent. Moreover, it implies that androgen might act as atrophic factor and affect the early development of these organs rather than simply sexual differentiation.

Androgen receptor gene polymorphism and sex hormones in elderly men:the Tromsøstudy

The aim of this study was to examine whether CAG/GGN repeats are significant modulators of serum concentrations of total and free testosterone(T)as well as of luteinizing hormone(LH)in elderly men.Sixty-nine 60-to 80-year-old men with subnormal T levels(≤11.0 nmol L^(-1))and 104 men with normal T levels taking part in a nested case-control study were used for these analyses.Sex hormones were measured and free T was calculated.The CAG and GGN polymorphisms in the androgen receptor gene were determined by polymerase chain reaction and subsequent direct sequencing.There were no differences in the CAG and GGN repeat lengths between the groups.In cross-sectional analyses of the whole cohort,total and free T were positively associated with CAG length(all P<0.05)before,but not after,waist circumference or body mass index was added to the model.CAG repeat lengths were weakly,but not independently,associated with total and free T.These findings indicate that when clinically evaluating T and LH levels in elderly men,the CAG and GGN repeat lengths do not need to be taken into consideration.

腹腔镜子宫肌瘤切除术后的产科及分娩结局

Objective: To assess the risks and outcome of pregnancies and deliveries after laparoscopic myomectomy (LM). Design: Retrospective study. Setting: Center of Reconstructive Pelvic Endosurgery, Reproductive Medicine Unit, S. Orsola-Malpighi Hospital, University of Bologna, Bologna, Italy. Patient(s): A total of 514 patients of fertile age that underwent LM at the Center were selected. Intervention(s): All the surgical procedures were performed using the same technique employing a vertical uterine incision and avoiding the use of electrosurgery. Main Outcome Measure(s): Number and outcome of pregnancies achieved after surgery, abortion rate, preterm delivery, gesta tional age, malpresentation, spontaneous or cesarean delivery, and postpartum hemorrhage. We also paid particular attention to the occurrence of uterine rupture. Result(s): A total of 158 pregnancies were achieved. There were 43 (27.2% )spontaneous abortions, 4 (2.6% ) ectopic pregnancies, and 1 (0.6% ) therapeutic abortion. Only 27 patients (25.5% ) had vaginal deliveries, whereas 79 (74.5% ) underwent cesarean section. No instances of uterine rupture were recorded. Conclusion(s): Our preliminary results confirmed that LM, performed by an expert surgeon, can restore reproductive capacity, allowing patients to have a successful pregnancy.

Up-regulation of NKG2A Inhibitory Receptor on Circulating NK Cells Contributes to Transfusion-induced Immunodepression in Patients with β-thalassemia Major

Accumulating evidence has shown that allogeneic blood transfusions can induce significant immunosuppression in recipients, and thereby increase the risk of postoperative infection and/or tumor relapse. Although it is well known that natural killer（NK） cells are responsible for the immunodepression effects of transfusion, the underlying mechanisms remain obscure. In this study, we investigated the role of NK cells in transfusion-induced immunodepression in β-thalassemia major. The proportion of circulating NK cells and the expression of NK receptors（NKG2A, CD158 a, NKP30, NKP46 and NKG2D） as well as CD107 a were detected by multicolor flow cytometry. IFN-γ production by circulating NK cells was detected by intracellular cytokine staining. Our results showed that the proportion and cytotoxicity（CD107a expression） of circulating NK cells in transfusion-dependent β-thalassemia major patients were remarkably lower than those of β-thalassemia minor patients or healthy volunteers. Expression of NKG2 A inhibitory receptor on circulating NK cells in patients with β-thalassemia major was remarkably up-regulated, but there were no significant differences in the expression levels of NKP30, NKP46, NKG2 D, CD158 a and IFN-γ. These results indicate NKG2 A inhibitory receptor may play a key role in transfusion-induced immunodepression of NK cells in patients with β-thalassemia major.

Surgical Management of Severe Hidradenitis Suppurativa in Women: Outcome and Quality of Life Assessment

Background: Hidradenitis suppurativa (HS) or Acne inversa sometimes referred to as Verneuil’s disease, is a rare and debilitating condition arising in the intertriginous folds that bear apocrine glands (axillae, groin, perineum, pubis, ano-genital area). It has a deleterious impact on the life of affected persons who tend to be more depressive and poorly socially and sexually connected because of pain, malodor and stigmatization. Methods: We conducted a retrospective study in Henri Mondor Hospital in Créteil, France, in order to assess the effectiveness of surgery as a radical treatment and also assess the factors related to recurrence and the surgical and sexual satisfaction in women operated for severe HS of perineum and genitalia. Results: Fifty-five women were included. We got a recurrence rate of 23.6%. 66.7% of recurrence occurred after localized excision. 83% of patients were satisfied with surgery results and would undergo a revision surgery or recommend it. The factors significantly associated with recurrence were overweight (p = 0.0046), location in groin (p = 0.0040), localized excision (p = 0.02), total excision (p = 0.009), healing via secondary intention (p = 0.001) and disability due to HS (p = 0.006). Assessment of sexual response thanks to the FSFI showed an alteration of all the domains of sexual response but the composite index of 18.51 was below the threshold to diagnose pathologic sexual condition.

Supraumbilical Transverse Incision for Cesarean Section in Severely Obese Patients:The Experience of a French Hospital from 2009 to 2014

Aim: To describe our experience in supraumbilical cesarean deliveries in severely and morbidly obese patients (body mass index > 35 kg/m2) with a voluminous abdominal panniculus. Methods: A retrospective study in France between January 2009 and May 2014. Every woman who underwent a cesarean delivery with a supraumbilical incision was included through her digital medical record. A senior doctor made the decision for a supraumbilical skin incision after careful examination of the patient’s anatomical conditions. Results: Twenty patients were included, for a total of 21 cesarean deliveries. Their mean body mass index 50 (40 - 61.7). Nineteen of them (95.0%) had an abdominal panniculus in supine “apron” position. Twelve patients (57.1%) had a scarred uterus. During the cesarean section, 14 (66.7%) had a segmentary hysterotomy. The median operative time was 49 minutes (32 - 70). Four patients (19.1%) had a postpartum hemorrhage and 4 (19.1%) a postpartum infectious complication, none of which were severe. Conclusions: In this specific population of severely obese women undergoing cesarean procedures, variations in anatomy require each patient’s incision choice to be individualized. In women with a voluminous panniculus the supraumbilical skin incision seems to offer an adequate exposure to the peritoneal cavity and the lower uterine segment, therefore allowing to safely carry out the procedure. However a prospective study on a larger sample remains necessary to compare this technique to the classical cesarean procedure.

＂One sperm, one embryo, one baby＂？

Dear Editor, The introduction of intracytoplasmic sperm injection （ICSI） technology has revolutionised the treatment of patients with severe oligozoospermia and azoospermia. A patient is usually considered oligozoospermic if his sperm concentration is less than 20 million mL^-1. A patient is labelled azoospermic according to the World Health Organisation guideline , if, on two different occasions, no spermatozoa can be detected by high-power microscopy after the seminal fluid has been centrifuged for 15 min at a centrifugation force of 3 000 × g or greater.

Intrauterine insemination with donor sperm:only the number of motile spermatozoa inseminated influences both pregnancy and live-birth rates

Intrauterine insemination with donor sperm(IUI-D)is an assisted reproductive technology(ART)offered to couples with definitive male infertility or risk of genetic disease transmission.Here,we sought to evaluate our practice in IUI-D and identify factors that influenced the success rate.We performed a retrospective,single-center study of all IUI-D procedures performed at Lille University Medical Center(Lille,France)between January 1,2007,and December 31,2017.Single and multivariate analyses with a mixed logistic model were used to identify factors associated with clinical pregnancies and live births.We included 322 couples and 1179 IUI-D procedures.The clinical pregnancy rate was 23.5%,and the live birth rate was 18.9%per IUI-D.In a multivariate analysis,the women’s age was negatively associated with the live birth rate.The number of motile spermatozoa inseminated was the only factor associated with both clinical pregnancies and live births,with a chosen threshold of 0.75 million.The clinical pregnancy and live birth rates were,respectively,17.3%and 13.0%below the number of motile spermatozoa inseminated threshold and 25.9%and 21.0%at or above the threshold(all P=0.005).The number of motile spermatozoa inseminated was the only factor that significantly influenced both pregnancies and live-birth rates after IUI-D.Indeed,below a threshold of 0.75 million motile spermatozoa inseminated,those rates were significantly lower.Application of this number of motile spermatozoa inseminated threshold may help centers to allocate donations more effectively while maintaining reasonable waiting times for patients.

Gene transcription profiting of astheno-and normo-zoospermic sperm subpopulations

Spermatozoa contain a repertoire of RNAs considered to be potential functional fertility biomarkers.In this study,the gene expression of human sperm subpopulations with high(F1)and low(F2)motility from healthy normozoospermic(N)and asthenozoospermic(A)individuals was evaluated using RNA microarray followed by functional genomic analysis of differentially expressed genes.Results from A-F1 versus N-F1,A-F2 versus N-F2,N-F1 versus N-F2,and A-F1 versus A-F2 comparisons showed a considerably larger set of downregulated genes in tests versus controls.Gene ontology(GO)analysis of A-F1 versus N-F1 identified 507 overrepresented biological processes(BPs),several of which are associated with sperm physiology.In addition,gene set enrichment analysis of the same contrast showed 110 BPs,36 cellular comp on ents,and 31 molecular functions,several of which are involved in sperm motility.A leadi ng・edge analysis of selected GO terms resulted in several down regulated genes encoding to dyn eins and kin esins,both related to sperm physiology.Furthermore,the predicted activation state of asthenozoospermia was increased,while fertility,cell movement of sperm,and gametogenesis were decreased.Interestingly,several downregulated genes characteristic of the canonical pathway protein ubiquitination were involved in asthenozoospermia activation.Conversely,GO analysis of A-F2 versus N-F2 did not identify overrepresented BPs,although the gene set enrichment analysis detected six enriched BPs,one cellular component,and two molecular functions.Overall,the results show differences in gene transcription between sperm subpopulations from asthenozoospermic and normozoospermic semen samples and allowed the identification of gene sets relevant to sperm physiology and reproduction.

Fluorescent probes for the detection of reactive oxygen species in human spermatozoa

Reactive oxygen species(ROS)production is a by-product of mitochondrial activity and is necessary for the acquisition of the capacitated state,a requirement for functional spermatozoa.However,an increase in oxidative stress,due to an abnormal production of ROS,has been shown to be related to loss of sperm function,highlighting the importance of an accurate detection of sperm ROS,given the specific nature of this cell.In this work,we tested a variety of commercially available fluorescent probes to detect ROS and reactive nitrogen species(RNS)in human sperm,to define their specificity.Using both flow cytometry(FC)and fluorescence microscopy(FM),we confirmed that MitoSOX™Red and dihydroethidium(DHE)detect superoxide anion(as determined using antimycin A as a positive control),while DAF-2A detects reactive nitrogen species(namely,nitric oxide).For the first time,we also report that RedoxSensor™Red CC-1,CellROX®Orange Reagent,and MitoPYl seem to be mostly sensitive to hydrogen peroxide,but not superoxide.Furthermore,mean fluorescence intensity(and not percentage of labeled cells)is the main parameter that can be reproducibly monitored using this type of methodology.

Tsc1/Tsc2 complex: A molecular target of capsaicin for protection against testicular torsion induced injury in rats

Objective: The detailed knowledge about protective effects of capsaicin(cap) and involved mechanisms against testicular torsion(TT) is still not available completely.Methods: Male Wistar rats were assigned into four major cohorts:(i) sham,(ii) TT,(iii) three subgroups subjected to TT and different doses of cap(100, 500, and 1000 μg/mL), and(iv) three subgroups of healthy animals subjected to various concentrations of cap. The animals were decapitated at 24 h after reperfusion, and the evaluation of protein expression was performed by Western blotting assay. At 72 h after reperfusion, apoptotic cell death and tissue injury were evaluated by TUNEL nuclear and H&E staining,respectively.Results: The results showed that cap administration following TT significantly increased the expression of tuberous sclerosis proteins 1 and 2(Tsc1/Tsc2) in a dose-dependent manner(P < 0.05). Cap decreased cell apoptosis at highest dose. Likewise, cap contributed to the preservation of tubular morphology and decreased tissue injury at the highest tested concentration(1000 μg/m L).Conclusion: Collectively, our findings demonstrate the validity of cap as a therapeutic agent against TT through targeting Tsc1/Tsc2 in a dose-dependent manner.