Maturation Capacity Of Oocytes Research Articles

Meiotic Development of Canine Oocytes from Poly-Ovular and Mono-Ovular Follicles after In Vitro Maturation

Poly-ovular follicles are defined as those with more than one oocyte present in single follicles. The occurrence frequency of this follicle type is higher in canines than that in other species. This study aimed to evaluate the in vitro meiotic maturation of dog oocytes from this follicle type in comparison to those from mono-ovular follicles of various sizes (small antral, medium antral, and large antral) considering different phases of the estrus cycle (anestrus, proestrus, estrus, and diestrus). Canine oocytes were obtained separately from the poly-ovular and mono-ovular antral follicles from the ovaries of adult females. In each experimental replicate, cumulus-oocyte complexes (COCs) from poly-ovular and mono-ovular follicles were incubated in supplemented TCM-199 at 38.5 °C and 5% CO2 for 72 h. After culturing, the meiotic development of each oocyte was evaluated using epifluorescence microscopy. Meiotic stages were classified into germinal vesicle (GV), germinal vesicle breakdown (GVBD), first metaphase (MI), and second metaphase (MII). Data were evaluated using an analysis of variance. Oocytes from poly-ovular follicles at all phases exhibited a higher (p < 0.05) percentage of oocytes arrested at the GV stage than those from mono-ovular follicles, showing the highest rate of GV in small antral follicles during anestrus. In contrast, there were no differences in MII rates (p < 0.05) in oocytes from mono-ovular and poly-ovular follicles during the estrus and diestrus phases in all sizes evaluated, with the highest MII rate in estrus. These results suggest that oocytes from poly-ovular follicles can resume meiosis at a slower rate than those from mono-ovular follicles; however, the maturation in vitro of such oocytes is possible. Furthermore, the relationship between the maturation capacity of oocytes from both poly-ovular and mono-ovular follicles depends on the ovarian cycle and follicular development.

Let-7a mimic transfection reduces chemotherapy-induced damage in a mouse ovarian transplantation model

Pharmacological approaches offer a non-invasive and promising option for fertility preservation in young female cancer patients undergoing gonadotoxic therapy. The GnRH-agonists are the only clinically available drugs in this indication, but their use and mechanisms of protection are still controversial. Recently, we have investigated new targeted drugs based on microRNA (miRNA) replacement therapy, and have identified the let-7a miRNA as candidate for fertility preservation strategies. Here, the effect of let-7a replacement during chemotherapy exposure on follicular growth and oocyte maturation capacity was investigated using a mouse ovarian-kidney transplantation model. Newborn mouse ovaries were cultured under different conditions; control, chemotherapy exposure (4-hydroperoxycyclophosphamide, 4-HC), and co-treatment with 4-HC and let-7a mimic transfection (4-HC + let-7a). The ovaries were then transplanted under the kidney capsule of recipient mice and follicular growth, survival, and oocyte in vitro maturation were assessed after 3 weeks. The results showed that the follicular pool was highest in the control group but higher in the 4-HC + let-7a group than the 4-HC group. DNA-damage/apoptosis ratios were higher in all 4-HC-exposed groups compared to control but were reduced in the 4-HC + let-7a group. In addition, the post-transplantation oocyte in vitro maturation rate was higher in the 4-HC + let-7a group compared to the 4-HC group, suggesting better oocyte quality. These results provide new information regarding the beneficial effects of let-7a replacement against chemotherapy-induced ovarian damage and open new perspectives for future in vivo applications.

AVALIAÇÃO DE EMBRIÕES OVINOS PROVENIENTES DE OÓCITOS SUBMETIDOS A ESTRESSE CALÓRICO DURANTE A MATURAÇÃO IN VITRO

The aim of this work was evaluate the effect of heat stress during oocyte maturation on ovine in vitro embryo production. Ovaries were collected at abattoirs and oocytes retrieved from follicles ranging from 2 and 6 mm in diameter. After selection, all oocytes, in 10 replicates, were placed in in vitro maturation (IVM) during 24 hours. The oocytes were submitted to heat stress of 41º C during 3, 6, 12, 18 and 24 hours and were further transferred to 39º C in order to complete IVM, which was the temperature of maturation of control oocytes. Embryonic development was determined on days 3, 4, 5, and 8 post-fertilization. Embryo evaluation was performed as total cell count by DAPI staining and determination of positive blastomere for apoptosis by the TUNEL assay. We observed that heat stress diminishes (P &lt; 0.05) oocyte maturation capacity in accordance with exposure time of 41º C. In the group of oocytes incubated at 39 ºC, 70.70% matured, while in the groups exposed to heat stress of 41º C, only 45.28%, 35.17%, 12.30%, 9.74% and 4.60% matured, respectively, after 3, 6, 12, 18 and 24 hours of incubation. The duration of exposure to heat stress is inversely proportional (P &lt; 0.05) to embryonic developmental capacity directly proportional (P &lt; 0.05) to the number of blastocysts positive to apoptosis. However, the cleavage rate and embryonic development from 8 to 16 cells and morulae stages were affected by heat stress (P &gt; 0.05) only up to 18 hours of incubation. The results allow the conclusion that heat stress during oocyte in vitro maturation reduces the quantity and quality of ovine embryos produced in vitro determined by the high incidence of apoptosis.KEYWORDS: blastocyst; IVF; IVM; IVP.