Criopreservación de espermatozoides de ratón: una necesidad biotecnológica para cumplir con el principio bioético de las 3rs.

NAVARRO, C.; KLINSKY LAHOZ, O.G.; SARTOR, T.; FURLAN, M.; WETTEN P.A.; MICHAUT, M.A.

Mendoza, Argentina

Congreso; XL REUNIÓN ANUAL DE LA SOCIEDAD DE BIOLOGIA DE CUYO; 2022

Sociedad de Biología de Cuyo

Sperm cryopreservation is an important technique for maintaining valuable genetic resources in biomedical research and wildlife.Research projects on reproductive biology use the mouse as an experimental model due to its short reproductive cycle. However,maintaining mice strain in an animal facility has the disadvantage of occupying large spaces with high economic and human costs. Thus,mouse sperm cryopreservation becomes a biotechnological need to reduce the drawbacks of this model and to accomplish the bioethicalprinciple of the 3R: Replacement, Reduction and Refinement. Although the cold has been used as a method of preservation since ancienttimes, it was only in 1990 that the ideal conditions to cryopreserve mouse sperm with a relatively simple cryoprotectant were found. Thiscryoprotectant solution (CPS) is composed by raffinose and skim milk. The aim of this work was to set up the sperm cryopreservationfrom CF-1 mouse strain using this CPS. The sperm were collected from caudae epididymides of 3-5 month old male mice. After killingeach mouse, both epididymides were removed and placed in a 35-mm sterile dish containing CPS (18% raffinose and 3% skim milk)equilibrated at 37 °C. Each cauda epididymis was cut three times and sperm were allowed to disperse for 3 min in the CPS solution.Immediately, 0.25 ul straws were loaded and rapidly cooled in liquid nitrogen vapor for 10 minutes. Finally, straws were plunged directlyinto liquid nitrogen for storage. To evaluate the effect of cryopreservation on sperm motility, frozen samples were rapidly thawed bytransferring them from liquid nitrogen into a 37 °C water bath for 2 min. After 30 minutes of recovering in HTF medium supplementedwith 5% BSA, in situ and progressive motility were evaluated for each sample. Results shown that this protocol yielded 3-6 % and 8-14% of mouse sperm with in situ and progressive motiliy, respectively. These results demonstrate that we have setted up the mouse spermcryopreservation protocol to reduce the breeding and slaughter of CF-1 mice males.