Evaluation of quality and gene expression of goat embryos produced in vivo and in vitro after cryopreservation.

In the present study, we aimed to identify morphological and molecular changes of in vivo and in vitro-produced goat embryos submitted to cryopreservation. In vivo embryos were recovered by transcervical technique from superovulated goats, whereas in vitro produced embryos were produced from ovaries collected at a slaughterhouse. Embryos were frozen by two-steps slow freezing method, which is defined as freezing to -32 °C followed by transfer to liquid nitrogen. Morphological evaluation of embryos was carried out by assessing blastocoel re-expansion rate and the total number of blastomeres. The expression profile of candidate genes related to thermal and oxidative stress, apoptosis, epigenetic, and implantation control was measured using RT-qPCR based SYBR Green system. In silico analyses were performed to identify conserved genes in goat species and protein-protein interaction networks were created. In vivo-produced embryos showed greater blastocoel re-expansion and more blastomere cells (P < 0.05). The expression level of CTP2 and HSP90 genes from in vitro cryopreserved embryos was higher than their in vivo counterparts. Unlikely, no significant difference was observed in the transcription level of SOD gene between groups. The high similarity of CPT2 and HSP90 proteins to their orthologs among mammals indicates that they share conserved functions. In summary, cryopreservation negatively affects the morphology and viability of goat embryos produced in vitro and changes the CPT2 and HSP90 gene expression likely in response to the in vitro production process.

Occurrence, morphology, and morphometry of follicles containing multiple oocytes in FecGE mutant Santa Inês ewes.

This study was conducted to characterize the morphology and morphometry of follicles containing multiple oocytes (MOFs) and determine the association with the FecGE mutation in Santa Inês ewes. Based on the genotypes, 65 ewes were characterized as being homozygous wild-type (n = 25; FecG+/+), heterozygous mutant (n = 27, FecG+/E), and homozygous mutant (n = 13, FecGE/E). The variables evaluated were follicle developmental stage, number of oocytes per follicle, morphology, and morphometry of MOFs. The FecGE mutation did not affect the frequency of MOFs (P > 0.05) (3.0 % in FecG+/+; 3.3 % in FecG+/E; and 3.5 % in FecGE/E). The greater viability (P < 0.05) of MOFs was identified in transitory stage of the FecGE/E (95.0 %) and FecG+/E (90.9 %) when compared to the FecG+/+ genotype (73.3 %). Furthermore, the morphology of transitory follicles with two oocytes was the variable and when evaluated was the most reliable determinant for predicting which ewes had an FecGE mutation. In conclusion, the FecGE mutation did not affect the frequency of MOFs. The ewes with FecGE mutation had a greater frequency of morphologically normal MOFs in the transitory stage. Furthermore, the ewes with the FecGE mutation had a greater likelihood of having MOFs containing two morphologically normal oocytes.

Ovarian and follicular variables used to determine ewes with different FecGE genotypes.

Based on ovarian and follicular variables, there was determination of ewes with different FecGE genotypes. Based on the FecGE genotype, 65 Santa Inês ewes were assigned to three experimental groups: homozygous wild-type (n = 25; FecG+/+), mutant heterozygous (n = 27; FecG+/E) and mutant homozygous (n = 13; FecGE/E). The ewe's ovaries were weighed and measured, then the follicles (oocyte, nucleus and nucleolus) were histologically evaluated for morphometry and morphology. Morphologically normal follicles, in the primordial and transitional stages, explained 70.18% of the variability morphological characteristics between mutant and wild-type ewes. Conducting the morphometric evaluation resulted in a more precise determination of the genotype groups when there was assessment of the primordial and secondary follicular developmental stages. The diameter of the oocyte and the oocyte nucleus of the primordial follicles explained 36.76% of the variability in follicular morphology between ewes with the mutation and those with the wildtype group. Similarly, the core diameter of oocytes in secondary follicles explained 10.63% of the variability in follicular morphology among FecGE/E, FecG+/E and FecG+/+ ewes. Thus, morphologically normal follicles in the primordial and transitional stages of development are the variables that allow for a more precise differentiation of Santa Inês ewes with the FecGE mutation. These variables may be evaluated to make more efficient the adoption of biotechniques that when conducted there is utilisation of follicles in the initial developmental stages as a physiological basis for classifying whether specific follicles are useful when conducting the techniques.