The motility-based swim-up technique separates bull sperm based on differences in metabolic rates and tail length

Swim-up is a sperm purification method that is being used daily in andrology labs around the world as a simple step for in vitro sperm selection. This method accumulates the most motile sperm in the upper fraction and leaves sperm with low or no motility in the lower fraction. However, the underlying reasons are not fully understood. In this article, we compare metabolic rate, motility and sperm tail length of bovine sperm cells of the upper and lower fraction. The metabolic assay platform reveals oxygen consumption rates and extracellular acidification rates simultaneously and thereby delivers the metabolic rates in real time. Our study confirms that the upper fraction of bull sperm has not only improved motility compared to the cells in the lower fraction but also shows higher metabolic rates and longer flagella. This pattern was consistent across media of two different levels of viscosity. We conclude that the motility-based separation of the swim-up technique is also reflected in underlying metabolic differences. Metabolic assays could serve as additional or alternative, label-free method to evaluate sperm quality.

[1]  A. Girling,et al.  Critical evaluation of methylcellulose as an alternative medium in sperm migration tests. , 2002, Human reproduction.

[2]  R. Rikmenspoel,et al.  Diffusion of ATP in sperm flagella. , 1970, Journal of theoretical biology.

[3]  E. C. Slater,et al.  The use of oligomycin as an inhibitor of oxidative phosphorylation. , 1961, Journal of biochemistry.

[4]  J. Gadea,et al.  Incubation of boar spermatozoa in viscous media by addition of methylcellulose improves sperm quality and penetration rates during in vitro fertilization. , 2017, Theriogenology.

[5]  S. Ratnam,et al.  Effect of sperm motility on human in vitro fertilization. , 1989, Archives of andrology.

[6]  Robert S. Balaban,et al.  Mitochondria, Oxidants, and Aging , 2005, Cell.

[7]  K. Reinhardt,et al.  Sperm metabolism is altered during storage by female insects: evidence from two-photon autofluorescence lifetime measurements in bedbugs , 2015, Journal of The Royal Society Interface.

[8]  W. Holt,et al.  The long and the short of sperm selection in vitro and in vivo: swim‐up techniques select for the longer and faster swimming mammalian sperm , 2010, Journal of evolutionary biology.

[9]  W. Ford,et al.  Glycolysis and sperm motility: does a spoonful of sugar help the flagellum go round? , 2006, Human reproduction update.

[10]  M. Teitell,et al.  Measuring energy metabolism in cultured cells, including human pluripotent stem cells and differentiated cells , 2012, Nature Protocols.

[11]  Charlie Chandsawangbhuwana,et al.  Comparison of glycolysis and oxidative phosphorylation as energy sources for mammalian sperm motility, using the combination of fluorescence imaging, laser tweezers, and real‐time automated tracking and trapping , 2008, Journal of cellular physiology.

[12]  H. Leese,et al.  Adenosine triphosphate production by bovine spermatozoa and its relationship to semen fertilizing ability. , 2008, Journal of andrology.

[13]  A. Agarwal,et al.  Oxidative phosphorylation versus glycolysis: what fuel do spermatozoa use? , 2014, Asian journal of andrology.

[14]  T. Hyakutake,et al.  Effects of surrounding fluid on motility of hyperactivated bovine sperm. , 2018, Journal of biomechanics.

[15]  E. Roldan,et al.  Differences in ATP Generation Via Glycolysis and Oxidative Phosphorylation and Relationships with Sperm Motility in Mouse Species* , 2015, The Journal of Biological Chemistry.

[16]  D. Smith,et al.  Sperm motility: is viscosity fundamental to progress? , 2011, Molecular human reproduction.

[17]  D. Sinton,et al.  Microfluidic assessment of swimming media for motility-based sperm selection. , 2015, Biomicrofluidics.

[18]  D F Katz,et al.  Relationships of mammalian sperm motility and morphology to hydrodynamic aspects of cell function. , 1981, Biology of reproduction.

[19]  J. Evers,et al.  Female subfertility , 2002, The Lancet.

[20]  [Laboratory manual of the WHO for the examination of human semen and sperm-cervical mucus interaction]. , 2001, Annali dell'Istituto superiore di sanita.

[21]  P. Brennan,et al.  Sperm storage: distinguishing selective processes and evaluating criteria. , 2015, Trends in ecology & evolution.

[22]  C. Yates,et al.  Male-factor infertility and in vitro fertilization , 1987, Journal of in Vitro Fertilization and Embryo Transfer.

[23]  A. Agarwal,et al.  Improvement in motion characteristics and acrosome status in cryopreserved human spermatozoa by swim-up processing before freezing. , 2000, Human reproduction.

[24]  C. Mukai,et al.  Glycolysis Plays a Major Role for Adenosine Triphosphate Supplementation in Mouse Sperm Flagellar Movement , 2004, Biology of reproduction.

[25]  P. Vercellini,et al.  Age-related infertility and unexplained infertility: an intricate clinical dilemma. , 2016, Human reproduction.

[26]  R. Vishwanath,et al.  Measurements of bovine sperm velocities under true anaerobic and aerobic conditions. , 1999, Animal reproduction science.

[27]  D. Amir,et al.  The effect of high sperm concentrations on the rates of respiration and fructolysis by ram spermatozoa. , 1967, Journal of reproduction and fertility.

[28]  E. Kommisrud,et al.  Relationship between post‐thaw adenosine triphosphate content, motility and viability in cryopreserved bovine semen applying two different preservation methods , 2018, Reproduction in domestic animals = Zuchthygiene.

[29]  Justin Hanes,et al.  Rapid transport of large polymeric nanoparticles in fresh undiluted human mucus , 2007, Proceedings of the National Academy of Sciences.

[30]  R. Foote,et al.  Quantification of bovine sperm separation by a swim-up method. Relationship to sperm motility, integrity of acrosomes, sperm migration in polyacrylamide gel and fertility. , 1987, Journal of andrology.

[31]  R. Montgomerie,et al.  Sperm swimming speed and energetics vary with sperm competition risk in bluegill (Lepomis macrochirus) , 2004, Behavioral Ecology and Sociobiology.

[32]  Paolo Gasparini,et al.  Insights into the Binding of Phenyltiocarbamide (PTC) Agonist to Its Target Human TAS2R38 Bitter Receptor , 2010, PloS one.

[33]  E. Delpiano,et al.  Fertility testing and ICSI sperm selection by hyaluronic acid binding: clinical and genetic aspects. , 2007, Reproductive biomedicine online.

[34]  R. Aitken,et al.  Oxidative stress and male reproductive biology. , 2004, Reproduction, fertility, and development.

[35]  R. Rikmenspoel.  Movements and active moments of bull sperm flagella as a function of temperature and viscosity. , 1984, The Journal of experimental biology.

[36]  D. Bates,et al.  Fitting Linear Mixed-Effects Models Using lme4 , 2014, 1406.5823.

[37]  K. Reinhardt,et al.  An Ecology of Sperm: Sperm Diversification by Natural Selection , 2015 .

[38]  A. Møller,et al.  Sexual selection and the temporal separation of reproductive events: sperm storage data from reptiles, birds and mammals , 1993 .

[39]  D. Cumming,et al.  Sperm Preparation for Intra-uterine Insemination , 1999 .

[40]  B. T. Storey,et al.  Mammalian sperm metabolism: oxygen and sugar, friend and foe. , 2008, The International journal of developmental biology.

[41]  A. Trounson,et al.  The influence of seminal characteristics on the success rate of human in vitro fertilization. , 1984, Fertility and sterility.

[42]  K. Tremellen.  Oxidative stress and male infertility--a clinical perspective. , 2008, Human reproduction update.

[43]  T. Hyakutake,et al.  Effect of non-Newtonian fluid properties on bovine sperm motility. , 2015, Journal of biomechanics.

[44]  M. Nichi,et al.  The Stimulated Glycolytic Pathway Is Able to Maintain ATP Levels and Kinetic Patterns of Bovine Epididymal Sperm Subjected to Mitochondrial Uncoupling , 2017, Oxidative medicine and cellular longevity.

[45]  L. Simmons,et al.  Sperm Swimming Velocity Predicts Competitive Fertilization Success in the Green Swordtail Xiphophorus helleri , 2010, PloS one.

[46]  John Sibert,et al.  AD Model Builder: using automatic differentiation for statistical inference of highly parameterized complex nonlinear models , 2012, Optim. Methods Softw..

[47]  E. Gaffney,et al.  Human sperm swimming in a high viscosity mucus analogue. , 2018, Journal of theoretical biology.

[48]  L. Yogev,et al.  Separation of sperm cells by sedimentation technique is not suitable for in vitro fertilization purposes , 2009, Andrologia.

[49]  Craig F. Purchase,et al.  Technical contribution Modifications to the IMAGEJ computer assisted sperm analysis plugin greatly improve efficiency and fundamentally alter the scope of attainable data , 2012 .