Antibody to Hsp70 alters response of rabbit preimplantation embryos to hyperthermia in vitro

The aim of the study was to determine a role of Hsp70 in the response of rabbit preimplantation embryos to hyperthermia (HT) in vitro. The embryos were cultured at standard (ST, 37.5 °C, control) or elevated (HT, 41.5 °C) temperature for 6 h. In half of the embryos from both groups, Hsp70 was blocked by the addition of an antibody against Hsp70 into the medium at stages either prior to (≤8 cell) or after (morula) developmental block (DB). After 6 h, the HT embryos were post-cultured at 37.5 °C for 16–20 h and then evaluated for developmental stages, apoptosis (TUNEL, T), cell number and quality of actin cytoskeleton. The exposure of embryos to 41.5 °C either before or after DB did not affect blastocysts (Bl) rate, T-index and embryo cell number. Blockage of Hsp70 before DB resulted in no embryos at the Bl stage. From the Hsp70-blocked embryos after DB, only 27.6% (ST + anti-Hsp70) and 22.6% (HT + anti-Hsp70) of blastocysts were formed, versus 71.4% (ST) and 69.7% (HT) in the non-Hsp70-blocked group. T-index of Hsp70-blocked embryos prior to DB increased from 6.8% (ST) to 28.6% (ST + anti-Hsp70) and from 6.1% (HT) to 29.6% (HT + anti-Hsp70), whilst the cell number dropped from 111 ± 8.1 (ST) to 20.5 ± 2.42 (ST + anti-Hsp70) and from 106.8 ± 8.4 (HT) to 15.4 ± 1.34 (HT + anti-Hsp70). In contrast, in the Hsp70-blocked embryos after DB, neither T-index nor cell number was changed comparing to the non-blocked. Damages to the actin cytoskeleton, like intracytoplasmic clumps or loss of actin filaments, were only minimally caused by hyperthermia but mostly were a result of Hsp70 blockage. In conclusion, the embryos at stages before the genome activation (GA) appear to be more sensitive to HT. The resistance of pre-GA embryos to HT is related to Hsp70, whilst after GA, Hsp70 is probably not the only factor of thermotolerance and the existence of an additional protective mechanism(s) is hypothesized.