RESUMO
The technique of laparoscopic oocyte aspiration has been increasingly used in animals; however, there are few records of its use in buffaloes. To describe this technique, six suckling Murrah buffaloes aged between 3 and 5 months were used. Three laparoscopic ovum pick-ups were performed in each animal, with intervals of 15 days between surgeries, completing a total of 18 procedures. The technique used three surgical ports with optics and a high-definition video camera. The introduction of the first portal and insufflation of the abdomen was performed through the open technique, with aspiration using a 20 G needle transabdominally and a vacuum pump calibrated at 50 mmHg. The mean complete surgical time from anesthesia to the removal of the animal from the litter was 49 ± 9.8 min. There were 27.8% cases of insufflation on the wrong side of the omentum. The oocyte recovery rate of 60.3% remained within the normal range. However, the rate of viable oocytes recovered was low, with only 40.8% of those recovered undergoing in vitro embryo production (IVEP). These data demonstrate that this simple, minimally invasive technique is an excellent reproductive tool for the genetic improvement of buffalo species.
RESUMO
PURPOSE: Despite advances in the composition of defined embryo culture media, co-culture with somatic cells is still used for bovine in vitro embryo production (IVEP) in many laboratories worldwide. Granulosa cells are most often used for this purpose, although recent work suggests that co-culture with stem cells of adult or embryonic origin or their derived biomaterials may improve mouse, cattle, and pig embryo development. MATERIALS AND METHODS: In experiment 1, in vitro produced bovine embryos were co-cultured in the presence of two concentrations of bovine adipose tissue-derived mesenchymal cells (b-ATMSCs; 103 and 104 cells/mL), in b-ATMSC preconditioned medium (SOF-Cond), or SOF alone (control). In experiment 2, co-culture with 104 b-ATMSCs/mL was compared to the traditional granulosa cell co-culture system (Gran). RESULTS: In experiment 1, co-culture with 104 b-ATMSCs/mL improved blastocyst rates in comparison to conditioned and control media (p < 0.05). Despite that it did not show difference with 103 b-ATMSCs/mL (p = 0.051), group 104 b-ATMSCs/mL yielded higher results of blastocyst production. In experiment 2, when compared to group Gran, co-culture with 104 b-ATMSCs/mL improved not only blastocyst rates but also quality as assessed by increased total cell numbers and mRNA expression levels for POU5F1 and G6PDH (p < 0.05). CONCLUSIONS: Co-culture of bovine embryos with b-ATMSCs was more beneficial than the traditional co-culture system with granulosa cells. We speculate that the microenvironmental modulatory potential of MSCs, by means of soluble substances and exosome secretions, could be responsible for the positive effects observed. Further experiments must be done to evaluate if this beneficial effect in vitro also translates to an increase in offspring following embryo transfer. Moreover, this study provides an interesting platform to study the basic requirements during preimplantation embryo development, which, in turn, may aid the improvement of embryo culture protocols in bovine and other species.
