Live motile sperm sorting device for enhanced sperm-fertilization competency: comparative analysis with density-gradient centrifugation and microfluidic sperm sorting.

PURPOSE: A live motile sperm sorting device (LensHooke® CA0) developed to prevent the deleterious effects of centrifugation was evaluated comparatively with conventional density-gradient centrifugation (DGC) and microfluidic-based device (Zymot) in sperm selection. METHODS: Semen samples from 239 men were collected. CA0 under different incubation intervals (5, 10, 30, and 60 min) and temperatures (20, 25, and 37℃) was conducted. The sperm quality in CA0-, DGC-, and Zymot-processed samples was then comparatively evaluated. Semen parameters included concentration, motility, morphology, motion kinematics, DNA fragmentation index (DFI), and the rate of acrosome-reacted sperm (AR). RESULTS: Total motility and motile sperm concentration increased in a time- and temperature-dependent manner and the total motility peaked for 30 min at 37℃. In paired analysis, CA0 showed significantly higher total motility (94.0%), progressive motility (90.8%), rapid progressive motility (83.6%), normal morphology (10.3%), and lower DFI (2.4%) and AR (4.7%) than the other two methods in normozoospermic samples (all p < 0.05). For non-normozoospermic samples, CA0 had significantly better results than the other two methods (total motility 89.2%, progressive motility 80.4%, rapid progressive motility 74.2%, normal morphology 8.5%, DFI 4.0%, and AR 4.0%; all p < 0.05). CONCLUSION: CA0 yielded spermatozoa with enhanced sperm fertilization potentials; DFI was minimized in samples processed by CA0. CA0 was effective for both normal and abnormal semen samples due to its consistent selection efficiency.

A Study of Thyroid Profile and Lipid Profile in Patients with Chronic Kidney Disease with or without Hemodialysis in a Tertiary Care Hospital.

BACKGROUND: Chronic kidney disease (CKD), due to increasing frequency and prevalence, has become one of the leading public health issues. The Kidney Disease Outcome Quality Initiative (KDOQI) defines CKD as kidney injury or a reduction in the glomerular filtration rate (GFR) to <60 mL/minute/1.73 m2 for at least 3 months. This study aims to compare the effects of decreased renal function on thyroid profile and lipid profile in CKD patients. MATERIALS AND METHODS: This is a prospective cross-sectional observational study conducted among the patients attending Outpatient Department/Inpatient Department (OPD/IPD) at the School of Medical Sciences & Research, Sharda Hospital, Greater Noida, Uttar Pradesh, India, in known cases of CKD, irrespective of the treatment/stage of CKD. All patients of >18 years of age with CKD were included in the study. RESULT: A total of 200 patients who met the inclusion criteria were included after obtaining detailed informed consent, of which 100 were cases and 100 were controls. The mean age of patients in the study was 47.74 years, with the mean age in patients with CKD 52 years, and the control was 43 years. The mean level of triglycerides (TGs) was significantly higher among the cases, and the high-density lipoprotein (HDL) was significantly lower among cases compared to controls (p < 0.05). Pearson's correlation between thyroid-stimulating hormone (TSH) with creatinine showed a weak albeit significant positive association (r = 0.200; p < 0.05). CONCLUSION: Our study shows a higher incidence of alteration in thyroid profile and dyslipidemia among the patients with CKD compared to controls. There is a necessary need to screen routinely for hypothyroidism and dyslipidemia among patients with CKD. Importantly, thyroid hormone levels and their effects on the progression of CKD have not been studied exhaustively.

Male infertility.

It is estimated that infertility affects 8-12% of couples globally, with a male factor being a primary or contributing cause in approximately 50% of couples. Causes of male subfertility vary highly, but can be related to congenital, acquired, or idiopathic factors that impair spermatogenesis. Many health conditions can affect male fertility, which underscores the need for a thorough evaluation of patients to identify treatable or reversible lifestyle factors or medical conditions. Although semen analysis remains the cornerstone for evaluating male infertility, advanced diagnostic tests to investigate sperm quality and function have been developed to improve diagnosis and management. The use of assisted reproductive techniques has also substantially improved the ability of couples with infertility to have biological children. This Seminar aims to provide a comprehensive overview of the assessment and management of men with infertility, along with current controversies and future endeavours.

Standards in semen examination: publishing reproducible and reliable data based on high-quality methodology.

Biomedical science is rapidly developing in terms of more transparency, openness and reproducibility of scientific publications. This is even more important for all studies that are based on results from basic semen examination. Recently two concordant documents have been published: the 6th edition of the WHO Laboratory Manual for the Examination and Processing of Human Semen, and the International Standard ISO 23162:2021. With these tools, we propose that authors should be instructed to follow these laboratory methods in order to publish studies in peer-reviewed journals, preferable by using a checklist as suggested in an Appendix to this article.

Predictive value of seminal oxidation-reduction potential analysis for reproductive outcomes of ICSI.

RESEARCH QUESTION: Is seminal oxidation-reduction potential (ORP) clinically relevant to reproductive outcome? DESIGN: Prospective observational study including a total of 144 couples who had an intracytoplasmic sperm injection (ICSI) cycle between June 2018 and December 2020. The study included patients undergoing fresh ICSI cycles with autologous gametes. Cycles that had day 3 embryo transfers and cryopreservation cycles were excluded. There was no restriction on patients with severe male infertility; couples with unexplained infertility and unexplained male infertility were included, those with azoospermia were excluded. Semen analysis, seminal ORP as determined by means of the MiOXSYS system, sperm DNA fragmentation (SDF) and reproductive outcomes (fertilization, blastocyst development, clinical pregnancy and live birth) were determined. RESULTS: Seminal ORP was significantly negatively correlated with fertilization rate (r = -0.267; P = 0.0012), blastocyst development rate (r = -0.432; P < 0.0001), implantation/clinical pregnancy (r = -0.305; P = 0.0003) and live birth (r = -0.366; P < 0.0001). Receiver operating characteristic curve analysis showed significant predictive power for ORP for fertilization (≥80%; area under the curve [AUC] 0.652; P = 0.0012), blastocyst development rate (≥60%; AUC 0.794; P < 0.0001), implantation/clinical pregnancy (AUC 0.680; P = 0.0002) and live birth (AUC 0.728; P < 0.0001). Comparable results were obtained for SDF (fertilization: AUC 0.678; blastocyst development: AUC 0.777; implantation/clinical pregnancy: AUC 0.665; live birth: AUC 0.723). Normal sperm morphology showed the lowest predictive power for all reproductive outcome parameters. With male age as confounding factor, ORP (cut-off value of 0.51 mV/106 sperm/ml) has significant (P < 0.04667) effects on odds ratios for all reproductive outcome parameters. Multivariate logistic regression to investigate potential seminal and female confounding factors revealed that seminal ORP significantly (P < 0.0039; P < 0.0130) affects reproductive outcome. CONCLUSION: Seminal ORP is relevant for good fertilization, blastocyst development, implantation, clinical pregnancy and live birth.

Unraveling the Molecular Impact of Sperm DNA Damage on Human Reproduction.

Semen analysis is the cornerstone in the investigation of fertility status of male partner. However, more advanced tests have emerged including the analysis of sperm chromatin integrity and DNA damage as markers of semen quality. This is of particular interest, as preserving the genetic information is essential to achieve a successful reproductive event. Moreover, the presence of unrepaired DNA lesions can affect cellular functions, resulting in the onset of pathological conditions associated with male infertility, and the transmission of diseases to the offspring. Hence, in this chapter, we aim to review the main factors leading to sperm DNA damage, along with the different types of damage which can occur. Furthermore, molecular mechanisms involved in DNA repair during spermatogenesis or after fertilization of the oocyte are described, and the laboratory techniques currently used in diagnostics and research, for the analysis of sperm DNA damage are also presented. Finally, the impact of sperm DNA damage on reproductive outcomes such as fertilization and pregnancy rates will be discussed with a focus on animal and human studies, along with the identification of new markers of sperm chromatin integrity.

Role of Infection and Leukocytes in Male Infertility.

Male infertility is considered as a multifactorial complex reproductive illness, and male urogenital infection and inflammation are crucial etiologies contributing up to 35% of all cases. Mostly triggered by sexually transmitted diseases and uropathogens, chronic manifestation of such infection may cause irreversible infertility in the male. Male urogenital infection involves bacterial, viral, protozoal, and fungal infections many of which remain asymptomatic most of the time and are passed to the sexual partner leading to fertilization failure, pregnancy loss, and even development of illness in the offspring. The abundance of leukocytes in semen can be used as an indicator of urogenital infection. Its contribution in male infertility can be as high as 30% and the clinical condition is referred to as leukocytospermia. Seminal bacterial load together with increased leukocytes contribute to the impairment of male fertility parameters such as, sperm motility, DNA integrity, acrosome reaction, and damage sperm molecular structure. Pathophysiology of bacteriospermia-induced impairment of male infertility is probably mediated by the involvement of bacterial pathogens in the intrinsic apoptotic pathway resulting in sperm death, whereas that of seminal leukocytes operates through excessive generation of ROS. Although the application of antibiotics forms the frontline therapeutic approach, the growing resistance to antibiotics poses a concern in the management of microbes-induced male urogenital infection. Complementary and alternative medicine may offer additional management options in combating such infections. On the other hand, both broad spectrum antibiotics and antioxidant therapy have showed promising results in the management of infertile men with leukocytospermia. Use of herbal medicine may also play a promising role in the management of such patients. However, recent molecular biology techniques have noted the association of elevated levels of IL-8 with both the Chlamydial infection of the male urogenital tract as well as the clinical condition of leukocytospermia. On the basis of such common pathogenesis, further research involving advanced molecular techniques may pave the way towards the development of better diagnostic tools in the clinical management of male urogenital infection and leukocytospermia.

Interplay of Oxidants and Antioxidants in Mammalian Embryo Culture System.

One principal purpose of assisted reproductive technology (ART) is to produce viable and good quality embryos. However, a variety of environmental factors may induce epigenetic changes in the embryo. Moreover, laboratory conditions including the culture media may also affect embryo development. Therefore, media change is an important factor in maintaining proper oxidant/antioxidant balance during embryo culture. Alterations in the oxidant/antioxidant balance are related to various cellular responses such as an increase in the level of reactive oxygen species (ROS) and consequent lipid peroxidation (LPO), DNA damage, and apoptosis. The current study focuses on the role of external factors on embryo culture and the ability of antioxidants to enhance in vitro fertilization (IVF) outcomes. Indeed, an optimization of media culture by the addition of enzymatic and nonenzymatic antioxidants in animal models and human embryos in ART has been updated in this study, with an emphasis on comparing the available results and their possible reasons.

A systemic review and meta-analysis exploring the predictors of sperm retrieval in patients with non-obstructive azoospermia and chromosomal abnormalities.

To identify the most prevalent chromosomal abnormalities in patients with non-obstructive azoospermia (NOA), consolidate their surgical sperm retrieval (SSR) rates and determine the significant predictors of positive SSR in this patient population. A systematic review and meta-analysis were performed following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. Fifty-three studies including 2965 patients were identified through searching the PubMed database. Klinefelter Syndrome (KS) was the most prevalent chromosomal abnormality reported in 2239 cases (75.5%). Azoospermia factor c (AZFc) microdeletions were the second most common (18.6%), but men with these deletions had higher SSR rates than patients with KS (41.95% with AZFc vs. 38.63% with KS). When examining predictors of SSR in KS patients, younger age was a significant predictor of positive SSR in patients undergoing microsurgical testicular sperm extraction (micro-TESE). Higher testosterone was a favourable predictor in those undergoing micro-TESE and conventional TESE. Lower luteinizing hormone (LH) and follicular stimulating hormone (FSH) values were significantly associated with positive SSR with testicular sperm aspiration (TESA). No parameter predicted SSR rates in patients with AZFc microdeletions. Overall, genetic abnormalities have significant implications on SSR success in patients with NOA.

Effect of redo varicocelectomy on semen parameters and pregnancy outcome: An original report and meta-analysis.

Recurrence following varicocelectomy is an important cause of treatment failure and persistence of subnormal semen parameters. This original study was combined with a systemic review and meta-analysis aiming to evaluate the efficacy of redo varicocelectomy on male fertility potential and pregnancy outcome. The retrospective study included 32 patients who underwent microsurgical subinguinal varicocelectomy for patients with recurrent varicocele. Changes in semen parameters and hormone profiles before and after surgery were compared. The literature review followed the Preferred Reporting Items for Systematic Reviews and Meta-Analyses and included seven articles in addition to our original report. Results of the original study revealed statistically significant improvements in sperm concentration, progressive motility, total motile sperm count and normal morphology following redo varicocelectomy. The meta-analysis results echoed those reported in our original study and depicted significant improvements in sperm concentration (mean difference [MD] = +20.281 million/ml, p < 0.001), total motility (MD = +9.659%, p = 0.001), total motile sperm count (MD = +23.258 million sperm, p < 0.001) and normal morphology (MD = +4.460%, p < 0.001). Overall pregnancy outcome was reported in seven studies with a rate of 34.6%. No significant changes were noted in any of the collected hormone results both in this original report and in the meta-analysis. In conclusion, redo varicocelectomy has a beneficial role on male fertility potential and can be offered for men with recurrent varicocele as directed by their individual clinical condition.

ANXA2, SP17, SERPINA5, PRDX2 genes, and sperm DNA fragmentation differentially represented in male partners of infertile couples with normal and abnormal sperm parameters.

This study aims to evaluate the expression of genes associated with the fertilisation potential and embryo development, sperm DNA fragmentation (SDF), and acrosome reaction in male partners of infertile couples with different sperm parameters compared to fertile men. First, male partners of infertile couples with abnormal (N = 25) and normal sperm parameters (N = 25), and fertile men (N = 10) were included in experimental groups I, II, and controls respectively. The mRNA levels of the Annexin A2 (ANXA2), Sperm protein 17 (SP17), Plasma serine protease inhibitor (SERPINA5), and Peroxiredoxin-2 (PRDX2) genes and SDF were evaluated. To evaluate the maturity of the sperm and oxidative stress, the acrosome reaction, the lipid peroxidation, and total antioxidant were measured. As result, SP17 showed a significantly lower expression in both experimental groups. SERPINA5 was significantly down-regulated in experimental group I that was aligned with the low rate of acrosome reaction. Significant overexpression of PRDX2 was found between experimental group II and controls. Significant higher rates of SDF were seen in both experimental groups compared to the controls. Finally, our data suggest that differentially gene expression of SP17 is a potential diagnostic biomarker in infertile men either with normal or abnormal sperm parameters. SDF is one of the causes of male infertility, independent of the sperm parameters.

In vitro ameliorative effects of ellagic acid on vitality, motility and DNA quality in human spermatozoa.

Oxidative stress (OS) plays a significant role in the etiology of male infertility, resulting in the impairment of male reproduction. This condition, characterized by an imbalance in the levels of oxidizing and antioxidant species in the seminal fluid, has a harmful impact on sperm functions and DNA integrity. The present study aimed to evaluate the anti-genotoxic action of ellagic acid, a polyphenolic molecule of natural origin having a powerful antigenotoxic, anti-inflammatory and antiproliferative role. An OS condition was induced in vitro by incubating normozoospermic human semen samples in benzene for 45, 60 and 90 min. DNA integrity was evaluated by terminal deoxynucleotidyl transferase dUTP nick end labeling assay, RAPD-PCR was performed to calculate the genome template stability, while the percentage of intracellular reactive oxygen species (ROS) was assessed by the 2', 7'-dichlorofluorescein assay. Our results showed that ellagic acid has a consistent protective effect on DNA integrity, as well as on sperm vitality and motility, by counteracting generation of intracellular ROS. The results of this study suggest ellagic acid as a suitable molecule to protect sperm DNA from oxidative stress, with a potentially significant translational impact on the management of the male infertility.

Environmental contaminants and male infertility: Effects and mechanisms.

The escalating prevalence of male infertility and decreasing trend in sperm quality have been correlated with rapid industrialisation and the associated discharge of an excess of synthetic substances into the environment. Humans are inevitably exposed to these ubiquitously distributed environmental contaminants, which possess the ability to intervene with the growth and function of male reproductive organs. Several epidemiological reports have correlated the blood and seminal levels of environmental contaminants with poor sperm quality. Numerous in vivo and in vitro studies have been conducted to investigate the effect of various environmental contaminants on spermatogenesis, steroidogenesis, Sertoli cells, blood-testis barrier, epididymis and sperm functions. The reported reprotoxic effects include alterations in the spermatogenic cycle, increased germ cell apoptosis, inhibition of steroidogenesis, decreased Leydig cell viability, impairment of Sertoli cell structure and function, altered expression of steroid receptors, increased permeability of blood-testis barrier, induction of peroxidative and epigenetic alterations in spermatozoa resulting in poor sperm quality and function. In light of recent scientific reports, this review discusses the effects of environmental contaminants on the male reproductive function and the possible mechanisms of action.

Diagnostic value of advanced semen analysis in evaluation of male infertility.

Conventional semen analysis is the standard of care to initially evaluate the fertility status of a male patient. However, it has some limitations and among these are failure to correctly identify the aetiology underlying fertility problems, intra- and inter-observer variability and incomplete information about sperm function. Considering these drawbacks, advanced semen tests have been developed to assess male infertility, including sperm function tests, oxidative stress (OS) and sperm DNA fragmentation (SDF) tests. This review illustrates the commonly utilised sperm function techniques, along with the assays used to assess SDF and OS and their diagnostic value.

Obesity and male infertility: Mechanisms and management.

Obesity is considered a global health problem affecting more than a third of the population. Complications of obesity include cardiovascular diseases, type 2 diabetes mellitus, malignancy (including prostatic cancer), neurodegeneration and accelerated ageing. In males, these further include erectile dysfunction, poor semen quality and subclinical prostatitis. Although poorly understood, important mediators of obesity that may influence the male reproductive system include hyperinsulinemia, hyperleptinemia, chronic inflammation and oxidative stress. Obesity is known to disrupt male fertility and the reproduction potential, particularly through alteration in the hypothalamic-pituitary-gonadal axis, disruption of testicular steroidogenesis and metabolic dysregulation, including insulin, cytokines and adipokines. Importantly, obesity and its underlying mediators result in a negative impact on semen parameters, including sperm concentration, motility, viability and normal morphology. Moreover, obesity inhibits chromatin condensation, DNA fragmentation, increases apoptosis and epigenetic changes that can be transferred to the offspring. This review discusses the impact of obesity on the male reproductive system and fertility, including associated mechanisms. Furthermore, weight management strategies, lifestyle changes, prescription medication, and complementary and alternative medicine in the management of obesity-induced subfertility is discussed.

Total antioxidant capacity-Relevance, methods and clinical implications.

Oxidative stress is pre-empted by an adequate level of antioxidants, which scavenge oxidants when they are produced in excess by different sources, including leukocytes and immature spermatozoa. Enzymatic antioxidants, such as superoxide dismutase, catalase and glutathione peroxidase, and several non-enzymatic antioxidants (proteins, vitamins and minerals), working as oxidant scavengers and cofactors of enzymatic antioxidants have been identified in seminal plasma. The total antioxidant capacity (TAC) is a diagnostic test that can be utilised in the male infertility workup. TAC measures the amount of total antioxidants in seminal plasma. Therefore, it provides an assessment of the reductive potential in seminal plasma. Several studies have investigated the diagnostic application of TAC in various andrology conditions. There is substantial evidence in the literature to show that infertile patients have lower seminal TAC in comparison with fertile men. Moreover, there is a positive correlation between TAC and seminal parameters, such as sperm concentration, motility and morphology. Evaluation of TAC together with reactive oxygen species (ROS) and sperm DNA fragmentation index (DFI) may be beneficial in the diagnosis of male infertility.

Diagnostic value of routine semen analysis in clinical andrology.

Infertility is a major health issue affecting over 48.5 million couples around the world, with the male factor accounting for about 50% of the cases. The conventional semen analysis recommended by the World Health Organization (WHO) is the cornerstone in the evaluation of male fertility status. It includes macroscopic and microscopic evaluation of the ejaculate, which reflects the production of spermatozoa in the testes, the patency of the duct system and the glandular secretory activity. Evaluation of seminal fructose, sperm vitality and leucocytes (Endtz test) are useful adjuncts to semen analysis that provide information on specific clinical conditions. Though several computer-assisted sperm analysis (CASA) systems have been developed, conventional methods for semen analysis are still widely accepted in clinical practice. This review summarises the conventional techniques used in routine semen analysis and their diagnostic value in clinical andrology.

Causes and consequences of sperm mitochondrial dysfunction.

Mitochondria have multiple functions, including synthesis of adenine triphosphate, production of reactive oxygen species, calcium signalling, thermogenesis and apoptosis. Mitochondria have a significant contribution in regulating the various physiological aspects of reproductive function, from spermatogenesis up to fertilisation. Mitochondrial functionality and intact mitochondrial membrane potential are a pre-requisite for sperm motility, hyperactivation, capacitation, acrosin activity, acrosome reaction and DNA integrity. Optimal mitochondrial activity is therefore crucial for human sperm function and semen quality. However, the precise role of mitochondria in spermatozoa remains to be fully explored. Defects in sperm mitochondrial function severely impair the maintenance of energy production required for sperm motility and may be an underlying cause of asthenozoospermia. Sperm mtDNA is susceptible to oxidative damage and mutations that could compromise sperm function leading to infertility. Males with abnormal semen parameters have increased mtDNA copy number and reduced mtDNA integrity. This review discusses the role of mitochondria in sperm function, along with the causes and impact of its dysfunction on male fertility. Greater understanding of sperm mitochondrial function and its correlation with sperm quality could provide further insights into their contribution in the assessment of the infertile male.

Comparative analysis of tests used to assess sperm chromatin integrity and DNA fragmentation.

Male infertility has a complex etiology, and many times, the cause is unknown. While routine semen analysis provides an overview of basic semen parameters, such as sperm concentration, motility, viability and morphology, a significant overlap of these parameters has been reported in fertile and infertile men. Moreover, conventional semen parameters do not reveal the cellular or molecular mechanisms of sperm dysfunctions leading to infertility. Therefore, sperm functional parameters, including sperm chromatin integrity, are evaluated to provide information on subtle sperm defects that are not routinely identified. Incomplete or defective sperm chromatin condensation increases the susceptibility of the sperm DNA to oxidative damage or other factors. To evaluate sperm chromatin integrity, different methods with varying degrees of diagnostic and prognostic capabilities are available. Among these assays, SCSA, TUNEL and SCD assays are most commonly used. While these assays rather evaluate the DNA directly for damages, the aniline blue and chromomycin A3 stains test for the quality of chromatin condensation. Thus, this review discusses and compares different methods used to evaluate sperm chromatin integrity and condensation, and their inclusion in the routine evaluation of the male infertility.

Reactive oxygen species in male reproduction: A boon or a bane?

Reactive oxygen species (ROS) are free radicals derived from oxygen during normal cellular metabolism. ROS play a crucial role in the physiological processes and signalling pathways associated with male fertility. At physiological concentrations, ROS act as molecular mediators of signal transduction pathways involved in the regulation of the hypothalamic-pituitary-gonadal axis, spermatogenesis and steroidogenesis. They also trigger the morphological changes required for sperm maturation, such as DNA compaction and flagellar modification. Furthermore, ROS modulate crucial processes involved in the attainment of sperm fertilising ability such as capacitation, hyperactivation, acrosome reaction and sperm-oocyte fusion. Conversely, oxidative stress prevails when the concentration of ROS overwhelms the body's antioxidant defence. Various endogenous and exogenous factors enhance the synthesis of ROS resulting in the disruption of structural and functional integrity of spermatozoa through the induction of apoptotic pathway and oxidation of molecules, such as lipids, proteins and DNA. Therefore, maintenance of a balanced redox state is critical for normal male reproductive functions. This article discusses the dual role of ROS in male reproduction, highlighting the physiological role as well as their pathological implications on male fertility.