The goal was to understand the role of heat shock at the zygote stage in causing infertility. Culture at 40°C reduced the percentage of inseminated oocytes that became a morula or blastocyst by d 6 or that were a blastocyst at d 8. An additional experiment was done to test whether effects of heat shock occur early in development or at the time of morula formation. Exposure to 40°C for 24 h decreased development to the blastocyst stage if exposure was at the zygote stage [8 to 32 h postinsemination (hpi)] but not if exposure occurred at the morula stage (116 to 140 hpi). To test effect of oxygen concentration, inseminated oocytes were cultured at 40°C for 12 or 24 h in either air (20.95% O₂; high oxygen) or a 5% (vol/vol) O₂ environment (low oxygen) that approximates the partial oxygen pressure of the reproductive tract. Blastocyst development was reduced by 40°C for 12 or 24 h under both atmospheres and was higher for embryos cultured in low oxygen than for embryos cultured in high oxygen. Examination of cell numbers at 72 hpi indicated that heat shock reduced developmental potential of embryos by reducing competence to complete cleavage divisions after first cleavage. Changes in expression of genes involved in heat shock and oxidative stress were measured to determine whether zygotes are more susceptible to heat shock because of reduced capacity for transcription. Heat shock was performed for 24 h at the 1-cell stage (expression examined in 2-cell embryos) or at d 5 (examined in morulae). Heat shock increased amounts of steady-state mRNA for HSPA1A but not for HSP90AA, SOD1, or CAT. We observed a tendency for a stage × temperature interaction for HSPA1A because the difference in expression between 38.5 and 40°C was greater for morulae than for 2-cell embryos. The amount of HSPA1A mRNA was less for morulae that were heat shocked than for 2-cell embryos cultured at 38.5°C. Heat shock at a temperature and oxygen tension similar to those seen in vivo can disrupt developmental competence of bovine zygotes. Increased susceptibility of the early embryo compared with the morula to heat shock was not due to reduced HSPA1A mRNA because amounts were higher for 2-cell embryos than for morulae.

Type 5 acid phosphatase (ACP5; also known as tartrate-resistant acid phosphatase or uteroferrin) is a metalloprotein secreted by the endometrial glandular epithelium of pigs, mares, sheep, and water buffalo. In this paper, we describe the phylogenetic distribution of endometrial expression of ACP5 and demonstrate that endometrial expression arose early in evolution (i.e., before divergence of prototherian and therian mammals ~166 million years ago). To determine expression of ACP5 in the pregnant endometrium, RNA was isolated from rhesus, mouse, rat, dog, sheep, cow, horse, armadillo, opossum, and duck-billed platypus. Results from RT-PCR and RNA-Seq experiments confirmed that ACP5 is expressed in all species examined. ACP5 was also demonstrated immunochemically in endometrium of rhesus, marmoset, sheep, cow, goat, and opossum. Alignment of inferred amino acid sequences shows a high conservation of ACP5 throughout speciation, with species-specific differences most extensive in the N-terminal and C-terminal regions of the protein. Analysis by Selecton indicated that most of the sites in ACP5 are undergoing purifying selection, and no sites undergoing positive selection were found. In conclusion, endometrial expression of ACP5 is a common feature in all orders of mammals and has been subjected to purifying selection. Expression of ACP5 in the uterus predates the divergence of therians and prototherians. ACP5 is an evolutionary conserved gene that likely exerts a common function important for pregnancy in mammals using a wide range of reproductive strategies.

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