Avaliação de oócitos caprinos submetidos à estresse calórico induzido durante a maturação in vitro / Evaluation of Goat Oocytes Subjected to Heat Stress Induced During Maturation In Vitro

ABSTRACT

Background: The low productivity of Northeast goat herds has been circumvented by the importation of exotic species; however, caution is needed due to the susceptibility of these breeds to the high temperatures found in this region. It is now known that the oocyte and the embryo are the primary targets of the deleterious effects induced by heat stress, causing cellular damage that triggers the cascade of apoptosis. Therefore, the purpose of this study was to evaluate the effect of thermal heat stress during in vitro maturation of oocytes and its effects on embryo production in goats. Materials, Methods & Results: The ovaries were collected in a slaughterhouse and transported to the Laboratory of Biotechnical Reproduction of UFRPE. The cumulus oophorus complexes (COCs) were collected by the technique of slicing the follicles from 2 to 6 mm in diameter, selected based on morphology and placed in a basic medium for maturation. In 10 replications, the COCs were submitted to the thermal heat stress at 41°C for 0 (thermoneutrality at 39°C), 3, 6, 12, 18, and 24 h of maturation in vitro. The data was evaluated in maturation, fertilization, cleavage (D-3), stage of 8-16 cells (D4), morula (D-5), and blastocyst (D-8) after fertilization and blastocysts positive for apoptosis through the TUNEL test. For statistics, the results were expressed as mean and standard deviation. Also considering the measurements addressed in percentages, a comparison of variances was carried out, F-test for variances to the level of signifi cance 5% (P < 0.05). Then, a t-test to compare averages was conducted, to the significance level of 5%, for equivalent variances or distinct variances, according to what was observed in the F-test for variances. A signifi cant difference (P < 0.05) was observed during all time periods studied for heat stress on maturation, fertilization, D-3, D-4, and D-5. On D-8 no significant difference (P > 0.05) was observed between the periods of 3 vs 6 and 18 vs 24 h, and in the blastocysts positive by the TUNEL test for the periods of 0 vs 3, 3 vs 6, 12 vs 18, and 18 vs 24 h of heat stress. Discussion: When applying a thermal shock that produces damage to the oocyte maturation in vitro, the characteristic membrane, chromatin configuration, and meiotic spindles are changed, and thus, the developmental potential of oocytes after fertilization is compromised. It was observed in this study that there was a gradual reduction in the number of oocytes as the time of exposure to heat shock increased, reflecting directly on each stage of IVP embryos. These stages are most vulnerable during maturation in vivo (ovulation), fertilization, within two days after fertilization, and in the first division of cleavage, as evidenced in this study in vitro after heat stress, reducing the number of blastocysts. This suggests that apoptosis can be induced in pre-implantation of embryos exposed to maternal hyperthermia. Moreover, the degree of apoptosis experienced by IVP embryos generally reflects the severity of thermal shock. In this study, the percentage of cells that were TUNEL positive increased with the prolongation of thermal shock. Induction of apoptosis was time dependent and the number of apoptotic cells increased proportionally after 6, 12, 18, and 24 h of exposure. Under the conditions observed in this study, the results indicate that the time in which the oocyte is exposed to heat stress during maturation in vitro is of great importance for embryonic development and their level of apoptotic cells.