ICSI, vitrification, testicular tissue, oocytes, bovine, fertilization.
Objective. To determine the differences in the embryo development of bovine oocytes fertilized with frozen/thawing (F/T) spermatozoa or with the intracytoplasmic sperm injection (ICSI) of F/T, spermatozoa from fresh testicular tissue (FTT), and cryopreserved testicular tissue (CTT), using three spermatozoa membrane-destabilizing agents.
Methodology. Four treatments were used. Treatment (TRT-1): In vitro fertilization (IVF) with F/T. TRT-2: ICSI with F/T. TRT-3: ICSI with FTT. TRT-4: ICSI with CTT. The spermatozoa membranes were destabilized with Triton X-100 (TX), Lysolecithin (LL), and Heparin-Glutathione (Hep-GSH). Embryonic division was recorded at 48 h and grade 1 and 2 blastocysts (BL) were recorded 8 days (D8) after the fertilization. The means were compared using Fisher’s least significant difference method.
Results. At D8, the blastocysts formation between ICSI treatments (F/T 13 ± 3, FTT 6 ± 3, and CTT 6 ± 3, p>0.05) were lower than control (IVF 23 ± 5). There was a lower cleavage at 48 h using Hep-GSH than when LL and TX were used (35 ± 5 vs 50± 5 and 56± 5, p<0.05). Embryo division at 48 h obtained better results with the ICSI + F/T and LL treatment, while the highest blastocyst percentage at D8 was obtained using TX.
Conclusions. Blastocysts can be produced through ICSI, using spermatozoa from fresh or cryopreserved testicular tissue. The spermatozoa treated with TX and LL produced a higher percentage of BL than the spermatozoa treated with Hep-GSH. Further experiments should be carried out using spermatozoa obtained from different sources, in order to improve embryo development after the ICSI.