Loss of Nuclear/DNA Integrity in Mouse Epididymal Spermatozoa after Short-Term Exposure to Low Doses of Dibutyl Phthalate or Bisphenol AF and Its Mitigation by Oral Antioxidant Supplementation.

Routine exposure to chemicals omnipresent in the environment, particularly the so-called endocrine-disrupting chemicals (EDCs), has been associated with decreased sperm quality and increased anomalies in testis. The decline in semen quality and testicular abnormalities have been attributed to the disruption of endocrine signaling as well as oxidative stress. The present study set out to examine the effect of short-term exposure of two common EDCs widely used in the plastic industry: Dibutyl Phthalate (DBP) and Bisphenol AF (BPAF). Our research objective was to focus on the post-testicular compartment of the epididymis, where spermatozoa acquire their functional capacity and are stored. The data obtained indicated no significant effect for either chemicals on sperm viability, motility or acrosome integrity. Neither of the EDCs had a noticeable effect on the structures of the testis and epididymis. However, substantial impact on the integrity of the sperm nucleus and DNA structure was evidenced by a significant increase in nuclear decondensation and DNA base oxidation. The damage observed was postulated to arise from the pro-oxidant properties of the EDCs generating excess of reactive oxygen species (ROS) and triggering a state of oxidative stress. This hypothesis was confirmed when the observed damage was largely blocked by co-administering EDCs with an evidenced-based antioxidant formulation.

OXIDATIVE STRESS AND REPRODUCTIVE FUNCTION: Sperm telomeres, oxidative stress, and infertility.

In brief: Oxidative stress is recognized as an underlying driving factor of both telomere dysfunction and human subfertility/infertility. This review briefly reassesses telomere integrity as a fertility biomarker before proposing a novel, mechanistic rationale for the role of oxidative stress in the seemingly paradoxical lengthening of sperm telomeres with aging. Abstract: The maintenance of redox balance in the male reproductive tract is critical to sperm health and function. Physiological levels of reactive oxygen species (ROS) promote sperm capacitation, while excess ROS exposure, or depleted antioxidant defenses, yields a state of oxidative stress which disrupts their fertilizing capacity and DNA structural integrity. The guanine moiety is the most readily oxidized of the four DNA bases and gets converted to the mutagenic lesion 8-hydroxy-deoxyguanosine (8-OHdG). Numerous studies have also confirmed oxidative stress as a driving factor behind accelerated telomere shortening and dysfunction. Although a clear consensus has not been reached, clinical studies also appear to associate telomere integrity with fertility outcomes in the assisted reproductive technology setting. Intriguingly, while sperm cellular and molecular characteristics make them more susceptible to oxidative insult than any other cell type, they are also the only cell type in which telomere lengthening accompanies aging. This article focuses on the oxidative stress response pathways to propose a mechanism for the explanation of this apparent paradox.

Rapid impact of COVID-19 infection on semen quality: a case report.

A unique opportunity to conduct a longitudinal analysis of semen quality in a male subject immediately before, during and after COVID-19 infection, has revealed new insights into the impact of this virus on male reproductive function. A moderate COVID infection that did not require hospitalization resulted in a state of azoospermia that persisted for 4 weeks. Given that the duration of spermatogenesis and epididymal sperm maturation in the human is 78 days, we calculate that a viral attack on the germ line was initiated at or before the patient was symptomatic and may have been signalled by a sudden reduction in sperm count and motility, several weeks earlier. Before the virus had been fully cleared, reinitiation of spermatogenesis occurred as evidenced by spermatozoa reappearing in the ejaculate exhibiting high levels of motility but significant levels of oxidative DNA damage as measured by a modified 8-OHdG assay protocol. These unique data indicate that even a moderate COVID-19 infection is capable of rapidly inducing a state of azoospermia that rapidly reverses as the infection wanes.

Male Infertility and Oxidative Stress: A Focus on the Underlying Mechanisms.

Reactive oxygen species (ROS) play a critical role in defining the functional competence of human spermatozoa. When generated in moderate amounts, ROS promote sperm capacitation by facilitating cholesterol efflux from the plasma membrane, enhancing cAMP generation, inducing cytoplasmic alkalinization, increasing intracellular calcium levels, and stimulating the protein phosphorylation events that drive the attainment of a capacitated state. However, when ROS generation is excessive and/or the antioxidant defences of the reproductive system are compromised, a state of oxidative stress may be induced that disrupts the fertilizing capacity of the spermatozoa and the structural integrity of their DNA. This article focusses on the sources of ROS within this system and examines the circumstances under which the adequacy of antioxidant protection might become a limiting factor. Seminal leukocyte contamination can contribute to oxidative stress in the ejaculate while, in the germ line, the dysregulation of electron transport in the sperm mitochondria, elevated NADPH oxidase activity, or the excessive stimulation of amino acid oxidase action are all potential contributors to oxidative stress. A knowledge of the mechanisms responsible for creating such stress within the human ejaculate is essential in order to develop better antioxidant strategies that avoid the unintentional creation of its reductive counterpart.

Apoptotic M540 bodies present in human semen interfere with flow cytometry-assisted assessment of sperm DNA fragmentation and oxidation.

BACKGROUND: The use of flow cytometry (FC) to evaluate sperm DNA fragmentation via deoxynucleotidyl transferase terminal fluorescein dUTP nick-end labeling (TUNEL) has shown inconsistencies compared with conventional fluorescent microscopic analyses. It has been hypothesized that the observed discrepancies could be attributed to the presence of apoptotic bodies that can be labeled with merocyanine 540, the so-called M540 bodies. In order to verify this hypothesis and determine the accuracy of our in-house FC-assisted evaluation of spermatozoa parameters, we used FC to evaluate both the fragmentation of sperm DNA using the TUNEL assay and the oxidation of sperm DNA using the 8-OHdG assay on semen samples with or without M540 bodies. RESULTS: We show that the presence of M540 bodies lead to underestimation of both the level of sperm DNA fragmentation and sperm DNA oxidation when using FC assisted detection systems. We also observed that this situation is particularly pertinent in semen samples classified as abnormal with respect to the routine WHO semen evaluation as they appear to contain more M540 bodies than normal samples. CONCLUSIONS: We conclude that M540 bodies interfere with both FC-conducted assays designed to evaluate sperm nuclear/DNA integrity. Exclusion of these contaminants in unprepared semen samples should be performed in order to correctly appreciate the true level of sperm DNA/nuclear damage which is known to be a critical male factor for reproductive success.

RéSUMé: CONTEXTE: L'utilisation de la cytométrie en flux (CF) pour évaluer la fragmentation de l'ADN des spermatozoïdes via la technique TUNEL (Terminal transferase dUTP nick-end labelling) a montré des incohérences par rapport aux analyses conventionnelles par microscopie fluorescente. L'hypothèse a été émise que les divergences observées pourraient être attribuées à la présence de corps apoptotiques qui peuvent être marqués à la mérocyanine 540 (corps M540). Afin de vérifier cette hypothèse et de déterminer la précision de notre évaluation interne des paramètres des spermatozoïdes, nous avons mesuré par CF à la fois la fragmentation de l'ADN des spermatozoïdes en utilisant le test TUNEL et l'oxydation de l'ADN des spermatozoïdes en utilisant le test 8-OHdG sur des échantillons de sperme avec ou sans corps M540. RéSULTATS: Nous montrons que la présence des corps M540 entraîne une sous-estimation du niveau de fragmentation et d'oxydation de l'ADN des spermatozoïdes lors de l'utilisation de systèmes de détection assistée par CF. Nous avons également observé que cette situation est exacerbée dans les échantillons de sperme classés comme anormaux (selon les standards de l'OMS), car ces derniers semblent contenir plus de corps M540 que les échantillons normaux. CONCLUSIONS: Nous concluons que les corps M540 interfèrent avec les deux tests conduits par CF et conçus pour évaluer l'intégrité nucléaire des spermatozoïdes. L'exclusion de ces contaminants dans les échantillons de sperme non préparés devrait être considérée afin d'apprécier correctement le véritable niveau de dommages au noyau spermatique qui est connu pour être un facteur critique pour le succès reproductif.

Varicocoele and oxidative stress: New perspectives from animal and human studies.

BACKGROUND: Varicocoele (VCL), one of the main causes of male subfertility, negatively affects testicular function. Due to limited access to human testicular tissue, animal model studies have been used to evaluate molecular and, recently, epigenetic changes attributed to pathophysiology induced by VCL. OBJECTIVES: This review aims to provide an update on the latest findings regarding the link between VCL-induced biochemical stress and molecular changes in germ cells and spermatozoa. Endocrine and antioxidant status, testicular chaperone-specific hemostasis failure, altered testicular ion balance, metabolic disorders, and altered carbon cycling during spermatogenesis are among the many features that will be presented. DISCUSSION: Literature review coupled with our own findings suggests that ionic imbalance, hypoxia, hyperthermia, and altered blood flow could lead to severe chronic oxidative and nitrosative stress in patients with VCL leading to defective spermatogenesis and impairment of the integrity of all sperm cell components and compartments down to the epigenetic information they carry. CONCLUSION: Since oxidative stress is an important feature of the reproductive pathology of VCL, therapeutic strategies such as the administration of appropriate antioxidants could be undertaken as a complementary non-invasive treatment line.