Cryoprotective effect of sericin on re-freezing of bull spermatozoa and subsequent embryonic development after in vitro fertilization

Abstract

Sericin, a protein produced by silk worm, has been used to improve quality of frozen-thawed spermatozoa in several species. However, the cryoprotective effect of sericin on re-freezing of spermatozoa has not been reported. This study was conducted to investigate the putative cryoprotective effect of sericin on re-freezing of bull spermatozoa followed by an assessment of in vitro fertilizing capacity of sericin-supplemented re-frozen spermatozoa. In experiment 1, the effect of 0.5% sericin supplemented in semen extender on re-freezing of spermatozoa was examined. Highly motile frozen-thawed spermatozoa were prepared with Percoll discontinuous gradient centrifugation prior to re-freezing. Refrozen-thawed spermatozoa were examined for motility, viability, membrane integrity, and acrosome integrity. In experiment 2, the effect of sericinsupplemented re-frozen spermatozoa on fertilizing capacity and subsequent embryonic development were determined using in vitro embryo production approach. Bovine oocytes were fertilized with three groups spermatozoa, re-frozen/control, re-frozen/sericin, and once frozen. The results showed that the total motility of re-frozen thawed spermatozoa in the sericin supplemented group was higher (53.9 % vs. 41.5%, p<0.05) than that of the control group whereas viability, membrane integrity, and acrosome integrity were unaffected. The proportion of cleaved oocytes in re-frozen/sericin-supplemented group was comparable to those in the re-frozen/control and once frozen groups. Additionally, the percentages of blastocyst development from cleaved oocytes were similar among the three groups. In conclusion, the results of this study demonstrated that pretreatment of frozen-thawed spermatozoa with 0.5% sericin improved motility after re-frozen thawed without affecting the in vitro fertilizing capacity and subsequent embryonic development.