Development of a computational tool to detect and classify human spermatozoa with hyperactivated motility

ARIAS RODOLFO; GARCIA ALEJANDRA; CASTAÑEDA VÍCTOR; LANDI PUJOL LUCRECIA; MATA MARTINEZ ESPERANZA; HÄRTEL STEFFEN; VARGAS SUSANA; ALE CHILET LUCAS; DE BLAS, GERARDO ANDRÉS

Mendoza

Simposio; XIV Jornadas de Investigación FCM. II Simposio de Medicina Traslacional; 2019

Facultad de Ciencias Médica. UNCuyo

The spermatozoon?s function is fertilization, a primordial event for the generation of a new individual and species propagation. To complete it, the mammalian sperm has a long process of preparation that occurs during its transit through the female genital tract. Four imbricated and essential processes that occur prior to fertilization can be defined: chemotaxis, capacitation, hyperactivation (HA) and acrosome reaction (AR). HA is a movement pattern characterized by a vigorous flagellum movement that results in an asymmetric flagellar curvature, fundamental for fertilization success since it allows the sperm to detach from the oviduct wall, penetrate the cumulus cells and oocyte?s zona pellucida. In addition, spermatozoon is the only mammalian cell whose function for which it was generated complies outside the organism that gave rise to it. Fertility has decreased in recent decades and 50% of cases are due to male factor, due to the decrease in quantity and quality of sperm. Currently the spermogram is the only clinical test that evaluates seminal quality, however this is not enough to achieve a conclusive diagnosis. Often the analysis of sperm quality is carried out visually and manually, but may be susceptible to errors inherent to the nature of the procedure, the results are subjective and lacking in reproducibility. In order to solve these difficulties, commercial systems of computer-assisted seminal analysis (CASA) have been developed to automate the process and solve these drawbacks. However, its acquisition cost is very high. In this sense, the TeleAnálisis and Translational Research Laboratory (LaTIT, in spanish), in collaboration with the Center of Digital Spermograms Assisted by Internet (CEDAI, in spanish) of the Faculty of Medicine, University of Chile; and through the use of the software developed by CEDAI, we have made computerized spermograms with high quality standards and results reproducibility. Considering this tool and the need to improve the diagnosis of male infertility, we set out to develop new tests allowing evaluation of sperm functionality and thereby improving diagnosis. In this project we proposed to add to the CEDAI software an algorithm that evaluates sperm functionality through the measurement of hyperactivated motility (HM) as a capacitation marker. To achieve our goal we work with semen samples from normal donors according to WHO criteria. The sperm fraction of greater vitality and motility was separated by swim-up technique and subsequently incubated in capacitating conditions for 6 hours. Capacitated and non-capacitated samples were observed by light field microscopy and 50 fps image sequences were taken and analyzed by CEDAI software. From the software?s data obtained, we counted cells with HM parameters [curvilinear velocity (VCL) > 150μm / sec, linearity (LIN) < 50% and an amplitude of lateral head movement (ALH) > 7μm] (Figure 1), we analyzed in total an average of 419 sperm per condition. We were able to observe that non-capacitated sample lacks sperms with MH; while after 6 hours of capacitation, 7% of motile sperms presented HM. Later, we visually evaluated the videos paying attention to the sperms that presented HM and correlated them with the results of the software (Figure 3). We observed high correlation between the results obtained with the software and the HM pattern in the videos. Based on these results, our CEDAI-UChile collaborators are working on the development of the mathematical algorithm for the software to perform the classification and automatic quantification of sperm with MH. This tool will allow urologists and andrologists to add a test to the spermogram to improve the diagnosis of male infertility.