Heat shock protein 70 gene expression in equine blastocysts after exposure of oocytes to high temperatures in vitro or in vivo after exercise of donor mares

Heat above homeothermy can be detrimental to embryonic development, and cells may produce heat shock proteins to try to mitigate these effects. The authors examined the developmental competence of equine oocytes after a single heat exposure (42 degrees C, 2 or 4 h) during early or late stages of in vitro maturation. Rates of nuclear maturation, cleavage after intracytoplasmic sperm injection, and advanced embryonic development (morula or blastocyst) were compared to those for unexposed controls. Concentrations of heat shock protein 70 (HSPA1A) mRNA were determined by real-time RT-PCR in resulting blastocysts, and were compared to those for embryos derived in vivo from control or exercised mares. Exposure of oocytes to heat at the onset of in vitro maturation did not affect any measured end point. However, exposure to 42 degrees C late in maturation culture reduced rates of oocyte nuclear maturation for both the 2 h (43/105 (43%) compared to control 70/103 (68%); P < 0.01), and 4 h (47/106 (44%) compared to control 60/103 (59%); P < 0.05) groups. Additionally, late heat exposure reduced development to morulae and blastocyst stages after intracytoplasmic sperm injection (ICSI; 18/89 (20%) compared to control 43/128 (34%); P < 0.05). Seven days after oocyte heat exposure, resultant blastocysts had a higher abundance of HSPA1A gene transcripts, relative to those for 18S rRNA. In vitro-produced embryos and lower-quality in vivo-produced embryos had significantly higher relative HSPA1A mRNA (lower 18S rRNA) concentrations than did higher-quality in vivo-produced embryos. The authors concluded that equine oocytes were sensitive to heat during late in vitro maturation, and responded to thermal shock with an increased ratio of HSPA1A:18S gene expression that was measurable in the resulting blastocyst. Embryos produced in vitro (including controls) had increased levels of HSPA1A mRNA relative to 18S rRNA compared to in vivo-produced embryos, suggesting a response to environmental insult.