Abstract This study aimed to evaluate the effect of two Embryo Manipulation Solutions (EMS and EMS supplemented) in maintenance of the viability of embryos, initially using structures derived from mice (first phase). Next, the efficiency of these solutions in routines of bovine embryo transfer was evaluated (second stage). Mice embryos were used in the stages of early blastocyst, and compact morula grades I and II. These embryos were initially randomly distributed and maintained for four hours in three solutions: Modified phosphate buffered saline (PBS; Control); EMS (treatment 1), and EMS supplemented (treatment 2). Subsequently, they were cultured in TCM 199 medium and evaluated in terms of total number of cells, morphometric characteristics, ultra structural aspects, detection of cell apoptosis, and quantification of Hsp70.3 gene expression. In the second phase, these same solutions were tested in the transfer of quality I and II bovine embryos (excellent and good). These embryos were transferred fresh to 58 recipients. The results showed that the total number of cells in embryos expanded blastocyst (ExB), the number of apoptotic cells, the cell, nuclear, nucleolar diameter and the nucleus/nucleolus ratio was similar among the treatments. The pregnancy rate shown on second phase was also similar. However, the EMS supplemented expressed more Hsp70.3 than EMS. The expression of Hsp70.3 was also greater for embryos in EMS than that of EMS supplemented. The McII embryos, EMS and EMS supplemented samples also expressed more Hsp70.3 compared to control embryos. In conclusion, the tested solutions can be used in routine embryo transfer techniques, replacing modified PBS solution as an effective media in maintaining embryo viability.

O objetivo deste estudo foi investigar a interação do FSH humano (10 ng/mL) com T4 (20 ng/mL) na sobrevivência, ativação e crescimento de folículos pré-antrais cultivados in vitro, por um período de longa duração (28 dias). Fragmentos de tecido ovariano não cultivado e cultivados foram processados para histologia clássica e microscopia eletrônica de transmissão. Os resultados mostraram uma redução na taxa de sobrevivência em todos os meios testados (um a 28 dias) quando comparado ao controle fresco. Entretanto, o tratamento com T4/hFSH por sete dias de cultivo resultou em taxa semelhante ao controle. Os meios testados por 28 dias reduziram o percentual de folículos primordiais em todos os períodos de cultivo. Contudo, T4/hFSH no dia um de cultivo manteve-se semelhante ao controle fresco. Nenhum dos meios foi capaz de manter o percentual dos folículos em desenvolvimento. Observou-se que o diâmetro folicular cultivado no meio com T4/hFSH manteve-se semelhante ao controle fresco. As análises ultraestruturais confirmaram a integridade de folículos cultivados por sete dias em meio suplementado com T4/hFSH.  Em conclusão, o meio com T4/hFSH é capaz de manter a sobrevivência, promover a ativação e a integridade ultraestrutural de folículos pré-antrais caprinos, por até sete dias.

The aim of this study was to investigate the interaction of human FSH (10ng/ml) with T4 (20ng/mL) on survival, activation and growth of preantral follicles cultured in vitro for 28 days. Fragments of non-cultured and cultured ovarian tissue were processed for classic histology and transmission electron microscopy. The results showed a reduction in the survival rate in all the media tested (one to 28 days) when compared to the fresh control. However the treatment with T4/hFSH for seven days of culture maintained the rate similar to the control. The media tested by one and 28 days reduced the percentage of primordial follicles in all periods of culture. However, T4/hFSH on day one of culture remained similar to the fresh control. None of the media were able to keep the percentage of the developing follicles. It was observed that the follicular diameter in the medium with T4/hFSH remained similar to the fresh control. The ultrastructural analysis confirmed the integrity of follicles cultured for seven days in a medium supplemented with T4/hFSH. In conclusion, the medium with T4/hFSH is able to maintain the survival, promote the activation, and the ultrastructural integrity of caprine preantral follicles for until seven days.