Embryo development after ICSI, using spermatozoa from bovine testicular tissue treated with three membrane-destabilizing agents

Abstract

ABSTRACT

Objective: To determine differences in embryo development of bovine oocytes fertilized by frozen/thawed spermatozoa (F/T), or by intracytoplasmic sperm injection (ICSI) using F/T or spermatozoa from fresh (FTT) or cryopreserved testicular tissue (CTT) using three spermatozoa membrane destabilizers. Methods. Treatment (TRT) 1- In vitro fertilization (FIV) with F/T, TRT-2 ICSI with F/T, TRT-3 ICSI with FTT, TRT-4 ICSI with CTT. The spermatozoa membranes were destabilized using Triton X-100 (TX), Lysolecithin (LL) or Heparin--Glutathione (Hep-GSH). Embryo cleavage at 48 h and grade 1 and 2 blastocyst on day 8 post fertilization were recorded. The comparison among main effect means were analyzed based on the least significant difference of Fisher. Results. At D8 there was no difference in percentage of blastocyst formation among ICSI TRTs (F/T 13 ± 3, FTT 6 ± 3 and CTT 6 ± 3 p>0.05), but they were lower than control (FIV 23 ± 5). With Hep-GSH destabilizer, there was a lower cleavage at 48 h than the LL and TX (35± 5, vs 50± 5 and 56± 5 p<0.05). Cleavage at 48 h was better for the ICSI with F/T and LL, while for D8, the best percentage to blastocyst was for TX. Conclusion. It is possible to produce blastocysts using ICSI with spermatozoa obtained from fresh or cryopreserved testicular tissue. Sperm cells treated with TX or LL produced more BL than those treated with Hep-GSH. More experiments using spermatozoa obtained from different sources are necessary to improve embryo development after ICSI.

Keywords:  ICSI, Vitrification, Testicular tissue, Oocytes, Bovine, Fertilization.