Advanced methods in reproductive medicine: Application of optical nanoscopy, artificial intelligence-assisted quantitative phase microscopy and mitochondrial DNA copy numbers to assess human sperm cells

Declined fertility rate and population is a matter of serious concern, especially in the developed nations. Assisted Reproductive Technologies (ART), including in vitro fertilization (IVF), have provided great hope for infertility treatment and maintaining population growth and social structure. With the help of ART, more than 8 million babies have already been born so far. Despite the worldwide expansion of ART, there is a number of open questions on the IVF success rates. Male factors for infertility contribute equally as female factors, however, male infertility is primarily focused on the “semen quality”. Therefore, the search of new semen parameters for male fertility evaluation and the exploration of the optimal method of sperm selection in IVF have been included among the top 10 research priorities for male infertility and medically assisted reproduction. The development of imaging systems coupled with image processing by Artificial Intelligence (AI) could be the revolutionary step for semen quality analysis and sperm cell selection in IVF procedures. For this work, we applied optical nanoscopy technology for the analysis of human spermatozoa, i.e., label-based Structured Illumination Microscopy (SIM) and non-invasive Quantitative Phase Microscopy (QPM). The SIM results demonstrated a prominent contrast and resolution enhancement for subcellular structures of living sperm cells, especially for mitochondria-containing midpiece, where features around 100 nm length-scale were resolved. Further, non-labeled QPM combined with machine learning technique revealed the association between gradual progressive motility loss and the morphology changes of the sperm head after external exposure to various concentrations of hydrogen peroxide. Moreover, to recognize healthy and stress-affected sperm cells, we applied Deep Neural Networks (DNNs) to QPM images achieving an accuracy of 85.6% on a dataset of 10,163 interferometric images of sperm cells. Additionally, we summarized the evidence from published literature regarding the association between mitochondrial DNA copy numbers (mtDNAcn) and semen quality. To conclude, we set up the high-resolution imaging of living human sperm cells with a remarkable level of subcellular structural details provided by SIM. Next, the morphological changes of sperm heads resulting from peroxidation have been revealed by QPM, which may not be explored by microscopy currently used in IVF settings. Besides, the implementation of DNNs for QPM image processing appears to be a promising tool in the automated classification and selection of sperm cells during IVF procedures. Moreover, the results of our meta-analysis showed an association of mtDNAcn in human sperm cells and semen quality, which seems to be a relevant sperm parameter for routine clinical practice in male fertility assessment

Proper use and impact of ‘Computer Assisted Semen Analysis’ technique on semen evaluation of farm animals

Σκοπός της παρούσας ανασκόπησης είναι η παρουσίαση της λειτουργίας του αυτόματου αναλυτή σπέρματος υποβοηθούμενου από ηλεκτρονικό υπολογιστή (CASA) και των εφαρμογών του στον τομέα της αναπαραγωγής των παραγωγικών ζώων, ώστε να γίνει ευρέως αντιληπτή η συμβολή του ως εργαλείο πρόγνωσης της γονιμότητας των αρσενικών ζώων Παρατίθενται οι προϋποθέσεις ορθής λειτουργίας του αναλυτή, οι παράγοντες που επηρεάζουν αυτήν και η αξιολόγηση των εκτιμούμενων παραμέτρων, με ιδιαίτερη έμφαση στην κινητικότητα των σπερματοζωαρίων. Ο αναλυτής CASA αποτελεί σημαντικό εφόδιο για τους ασχολούμενους με τη διαχείριση υγείας των παραγωγικών ζώων, ωστόσο απαιτείται τήρηση των κανόνων για ορθή εκτίμηση των δειγμάτων και για αξιοπιστία και δυνατότητα σύγκρισης των αποτελεσμάτων.Objective of this review is to present the use of ‘Computer Assisted Semen Analysis’ technique application in farm animal health management, in particular the steps for accurate semen evaluation and the impact of the method in predicting male animal fertility under field conditions. Requirements for proper use of the equipment, factors affecting the evaluation process and the role of the estimated parametres for fertility under field conditions are described. Special reference is made in sperm motility evaluation. It is concluded that the method is an effective and efficient tool for semen evaluation, provided good practices are strictly applied and adhered to, by means of which valid results may be obtained

Changes in women’s facial skin color over the ovulatory cycle are not detectable by the human visual system

Human ovulation is not advertised, as it is in several primate species, by conspicuous sexual swellings. However, there is increasing evidence that the attractiveness of women’s body odor, voice, and facial appearance peak during the fertile phase of their ovulatory cycle. Cycle effects on facial attractiveness may be underpinned by changes in facial skin color, but it is not clear if skin color varies cyclically in humans or if any changes are detectable. To test these questions we photographed women daily for at least one cycle. Changes in facial skin redness and luminance were then quantified by mapping the digital images to human long, medium, and shortwave visual receptors. We find cyclic variation in skin redness, but not luminance. Redness decreases rapidly after menstrual onset, increases in the days before ovulation, and remains high through the luteal phase. However, we also show that this variation is unlikely to be detectable by the human visual system. We conclude that changes in skin color are not responsible for the effects of the ovulatory cycle on women’s attractiveness

Impact of Nanotechnology-Based Semen Purification on Reproduction of Gilts and Developmental Performance of Offspring

Semen contain a heterogeneous population of viable and non-viable (damaged) spermatozoa. Proportions of non-viable spermatozoa interfere with male fertility, with available techniques unable to selectively remove prior to breeding. Nanobiotechnology may allow removal, enriching semen with high quality spermatozoa for improved productivity. Here, we applied double nanopurification with boar semen using functionalized magnetic nanoparticles. Non-viable and viable spermatozoa were magnetically separated and verified through various microscopy imaging. Viable (nanopurified) spermatozoa showed no additional damages. Nanopurification did not interfere with sperm motility and viability, with beneficial effects on motion parameters. Nanopurified spermatozoa maintained fertility following insemination, with resulting offspring indicating no impaired growth or health performance. Pork quality was unaffected showing comparable characteristics to the control. In summary, the use of magnetic nanopurification in boar spermatozoa showed sperm viability and fertility improvements with successful offspring performance. This study shows promise for large-scale commercial applications to enhance male fertility and offspring performance

The role of staining techniques in seminological analysis of mammalian semen

The most important semen parameters are the concentration, motility and morphology of sperm cells. Sperm morphology is regarded as the most reliable parameter for predicting fertility in males. A problem in evaluating sperm morphology and morphometry is the lack of standardization of staining techniques. The staining procedure and reagents used can significantly affect the morphometric parameters of the sperm cell. The use of stains with different pH or osmotic pressure, as well as the duration of the procedure, may influence the shape and size of the sperm, and thus the result of the morphological evaluation of the semen. It is necessary to develop an evaluation procedure for sperm morphology and morphometry that will minimize the changes in the structure of the evaluated semen in relation to the native semen

The Role of Cytometry for Male Fertility Assessment in Toxicology

Infertility is nowadays a major concern, affecting approximately 8–12% of the couples and the male factor accounts for about 50% of the cases. Occupational and/or environmental exposure to heavy metals and other pollutants is the main cause of male infertility. Lead, cadmium and chromium are heavy metals widely used in industry and quite persistent in the environment, raising major concerns over the possible effects on the reproductive health of workers and the general population. Sperm DNA integrity is essential for the accurate transmission of paternal genetic information, and normal sperm chromatin structure is important for sperm fertilizing ability. Flow cytometry can be to characterize multiple physical characteristics of the population of spermatozoa in the sperm, including sperm concentration, viability, mitochondrial mass and function, acrosome integrity, capacitation, membrane fluidity, DNA content and status, etc. This chapter elucidates the role of cytometry in the study of male fertility under toxicological insult by pollutants such as chromium, cadmium and lead. Some representative examples are presented using in vivo studies with rodents. In addition, complementary techniques to cytometry and future perspectives will be mentioned in an interdisciplinary point of view to gain knowledge on this subject