Changes of the Protein CoAlation Pattern in Response to Oxidative Stress and Capacitation in Human Spermatozoa

The spermatozoa have limited antioxidant defences, a high polyunsaturated fatty acids content and the impossibility of synthesizing proteins, thus being susceptible to oxidative stress. High levels of reactive oxygen species (ROS) harm human spermatozoa, promoting oxidative damage to sperm lipids, proteins and DNA, leading to infertility. Coenzyme A (CoA) is a key metabolic integrator in all living cells. Recently, CoA was shown to function as a major cellular antioxidant mediated by a covalent modification of surface-exposed cysteines by CoA (protein CoAlation) under oxidative or metabolic stresses. Here, the profile of protein CoAlation was examined in sperm capacitation and in human spermatozoa treated with different oxidizing agents (hydrogen peroxide, (H2O2), diamide and tert-butyl hydroperoxide (t-BHP). Sperm viability and motility were also investigated. We found that H2O2 and diamide produced the highest levels of protein CoAlation and the greatest reduction of sperm motility without impairing viability. Protein CoAlation levels are regulated by 2-Cys peroxiredoxins (PRDXs). Capacitated spermatozoa showed lower levels of protein CoAlation than non-capacitation cells. This study is the first to demonstrate that PRDXs regulate protein CoAlation, which is part of the antioxidant response of human spermatozoa and participates in the redox regulation associated with sperm capacitation

Sperm DNA methylome abnormalities occur both pre- and post-treatment in men with Hodgkin disease and testicular cancer

Combination chemotherapy has contributed to increased survival from Hodgkin disease (HD) and testicular cancer (TC). However, questions concerning the quality of spermatozoa after treatment have arisen. While studies have shown evidence of DNA damage and aneuploidy in spermatozoa years following anticancer treatment, the sperm epigenome has received little attention. Our objectives here were to determine the impact of HD and TC, as well as their treatments, on sperm DNA methylation. Semen samples were collected from community controls (CC) and from men undergoing treatment for HD or TC, both before initiation of chemotherapy and at multiple times post-treatment. Sperm DNA methylation was assessed using genome-wide and locus-specific approaches. Imprinted gene methylation was not affected in the sperm of HD or TC men, before or after treatment. Prior to treatment, using Illumina HumanMethylation450 BeadChip (450 K) arrays, a subset of 500 probes was able to distinguish sperm samples from TC, HD and CC subjects; differences between groups persisted post-treatment. Comparing altered sperm methylation between HD or TC patients versus CC men, twice as many sites were affected in TC versus HD men; for both groups, the most affected CpGs were hypomethylated. For TC patients, the promoter region of GDF2 contained the largest region of differential methylation. To assess alterations in DNA methylation over time/post-chemotherapy, serial samples from individual patients were compared. With restriction landmark genome scanning and 450 K array analyses, some patients who underwent chemotherapy showed increased alterations in DNA methylation, up to 2 to 3 years post-treatment, when compared to the CC cohort. Similarly, a higher-resolution human sperm-specific assay that includes assessment of environmentally sensitive regions, or "dynamic sites," also demonstrated persistently altered sperm DNA methylation in cancer patients post-treatment and suggested preferential susceptibility of "dynamic" CpG sites. Distinct sperm DNA methylation signatures were present pre-treatment in men with HD and TC and may help explain increases in birth defects reported in recent clinical studies. Epigenetic defects in spermatozoa of some cancer survivors were evident even up to 2 years post-treatment. Abnormalities in the sperm epigenome both pre- and post-chemotherapy may contribute to detrimental effects on future reproductive health. The online version contains supplementary material available at 10.1186/s13148-022-01417-1

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.Peer reviewe

Absence of PEROXIREDOXIN 6 is Associated with Low Sperm Chromatin Quality and Subfertility in Mice Challenged with Oxidative Stress

<p>This poster was presented at Gordon Research Conference on Fertilization and Activation of Development in July 2013. </p

Improving standard practices in studies using results from basic human semen examination

: The purpose of this article is to provide an explanation of the background behind a checklist that declares the laboratory methods used in a scientific study. It focuses primarily on implementing laboratory procedures to yield reliable results in basic semen examinations. While the World Health Organization (WHO) and international standards provide recommendations for basic semen examination, manuscripts submitted to Andrology frequently lack transparency regarding the specific techniques used. In addition, the terminology used for semen examination results often fails to provide a clear definition of the groups under study. Furthermore, the WHO's reference limits are often misinterpreted as strict boundaries between fertility and infertility. It is important to note that valid clinical andrological diagnoses and treatments cannot rely solely on semen examination results; they require proper laboratory procedures as a foundation for diagnosing and treating male patients. Therefore, scientific journals should promote the adoption of robust laboratory practices and an accurate definition of patient groups. A checklist can facilitate the design of high-quality studies and the creation of informative publications. Further, it can help journals assess submitted manuscripts and improve the overall quality of their publications

Effects of Heparan sulfate acetyl-CoA: Alpha-glucosaminide N-acetyltransferase (HGSNAT) inactivation on the structure and function of epithelial and immune cells of the testis and epididymis and sperm parameters in adult mice.

Heparan sulfate (HS), an abundant component of the apical cell surface and basement membrane, belongs to the glycosaminoglycan family of carbohydrates covalently linked to proteins called heparan sulfate proteoglycans. After endocytosis, HS is degraded in the lysosome by several enzymes, including heparan-alpha-glucosaminide N-acetyltransferase (HGSNAT), and in its absence causes Mucopolysaccharidosis III type C (Sanfilippo type C). Since endocytosis occurs in epithelial cells of the testis and epididymis, we examined the morphological effects of Hgsnat inactivation in these organs. In the testis, Hgsnat knockout (Hgsnat-Geo) mice revealed statistically significant decrease in tubule and epithelial profile area of seminiferous tubules. Electron microscopy (EM) analysis revealed cross-sectional tubule profiles with normal and moderately to severely altered appearances. Abnormalities in Sertoli cells and blood-testis barrier and the absence of germ cells in some tubules were noted along with altered morphology of sperm, sperm motility parameters and a reduction in fertilization rates in vitro. Along with quantitatively increased epithelial and tubular profile areas in the epididymis, EM demonstrated significant accumulations of electrolucent lysosomes in the caput-cauda regions that were reactive for cathepsin D and prosaposin antibodies. Lysosomes with similar storage materials were also found in basal, clear and myoid cells. In the mid/basal region of the epithelium of caput-cauda regions of KO mice, large vacuolated cells, unreactive for cytokeratin 5, a basal cell marker, were identified morphologically as epididymal mononuclear phagocytes (eMPs). The cytoplasm of the eMPs was occupied by a gigantic lysosome suggesting an active role of these cells in removing debris from the epithelium. Some eMPs were found in proximity to T-lymphocytes, a feature of dendritic cells. Taken together, our results reveal that upon Hgsnat inactivation, morphological alterations occur to the testis affecting sperm morphology and motility parameters and abnormal lysosomes in epididymal epithelial cells, indicative of a lysosomal storage disease