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Published-Ahead-of-Print June 28, 2006, DOI:10.2164/jandrol.106.000109

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Comparative Study of Four Different Sperm Washing Methods Using Apoptotic Markers in Ram Spermatozoa

Elena Marti , Rosaura Perez-Pe , Teresa Muino-Blanco , and Jose A. Cebrian-Perez *

* To whom correspondence should be addressed. E-mail: pcebrian{at}unizar.es.

The accurate measurement of the semen fertilizing potential is of great importance in determining the acceptability of processed semen for breeding purposes. A good sperm preparation technique requires not only providing a sample with a high viability and motility, but also taking into account other parameters such as the capacitation and apoptotic state which could comprise the ability to fertilize the oocyte. In this study, we investigate the effects of the application of four sperm preparation techniques (a dextran/swim-up procedure, discontinuous Percoll density gradient centrifugation, sucrose washing and filtration) on ram sperm quality parameters. Besides the evaluation of viability and the capacitation state, we also analyse the apoptotic status of the outcome sperm samples assessing the phosphatidylserine translocation and caspase-3 and -7 activity. This is the first report, to our knowledge, that evidences the presence of active caspases in ram sperm. The obtained results confirm the higher capacity of the dextran/swim-up procedure to select non-apoptotic spermatozoa, in addition to viable and non-capacitated sperm, compared with the other sperm preparation methods. This should be considered in order to enhance results by means of AI techniques in ovine reproduction.


Key words: Annexin V • Caspase Activity • Swim-up






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