In vitro maturation of goat oocytes selected using Brilliant cresyl blue staining

The present study was conducted to assess the developmental competence of goat oocytes selected using Brilliant cresyl blue (BCB) staining. Goat ovaries were collected from the slaughtered animals with unknown reproductive history. The oocytes retrieved by aspiration technique were selected based on morphology and subjected to BCB staining. Brilliant cresyl blue staining is based on the activity of glucose-6-phospahte dehydrogenase (G6PDH) enzyme synthesised by the oocytes. The cytoplasm remains blue in oocytes that have finished the growth phase (BCB+) while the growing oocytes remain colourless (BCB-). The stained and unstained oocytes were subjected to in vitro maturation separately to assess cumulus cell expansion index and polar body extrusion. A total of 206 culture grade oocytes were subjected to study, out of which, 76.75 ± 2.38 per cent of oocytes showed positive to BCB staining and 23.21 ± 2.38 per cent were negatively stained. Significantly higher maturation rate was observed in BCB+(92.89 ± 2.37%) oocytes than BCB-(29.72 ± 2.46%). The present study concluded that BCB staining can be used for selecting goat oocytes with good cytoplasmic maturation for further in vitro embryo production

Effect of adding growth factors during in vitro maturation on the developmental potentials of ewe oocytes selected by brilliant cresyl blue staining

Aim: Several factors had been concerned with the developmental competence of the sheep oocyte. This study aims to investigate the effect of adding growth factors (insulin-like growth factor 1 [IGF-1] and epidermal growth factor [EGF]) in the maturation medium of ewe oocytes selected based on brilliant cresyl blue (BCB) screening on in vitro maturation (IVM), fertilization, and pre-implantation embryo development. Materials and Methods: Cumulus-oocyte complexes (COCs) were obtained from the ovaries of slaughtered ewes by either aspiration or slicing techniques. COCs were in vitro matured in a medium containing IGF-1 and EGF (control group). For BCB screening, oocytes were stained and divided into BCB+ oocytes that matured in the same maturation conditions without adding growth factors (Group 2) or in the presence of growth factors (Group 3), and BCB– oocytes that matured in medium without growth factors (Group 4) or with growth factors (Group 5). Results: The supplementation of the maturation medium with growth factors during IVM of (BCB+) oocytes resulted in a significant increase in nuclear maturation rate (90.9%), fertilization rate (75.6%), and embryo developmental rates (60.0%, 46.7%, and 33.3% for cleavage, morula, and blastocyst, respectively). Conclusion: Culturing BCB+ oocytes in a maturation medium containing both EGF and IGF-1 showed a significant improvement in nuclear maturation, fertilization, and pre-implantation embryo development in vitro.

Brilliant cresyl blue enhanced optoacoustic imaging enables non-destructive imaging of mammalian ovarian follicles for artificial reproduction

In the field of reproductive biology, there is a strong need for a suitable tool capable of non-destructive evaluation of oocyte viability and function. We studied the application of brilliant cresyl blue (BCB) as an intra-vital exogenous contrast agent using multispectral optoacoustic tomography (MSOT) for visualization of porcine ovarian follicles. The technique provided excellent molecular sensitivity, enabling the selection of competent oocytes without disrupting the follicles. We further conducted in vitro embryo culture, molecular analysis (real-time and reverse transcriptase polymerase chain reaction) and DNA fragmentation analysis to comprehensively establish the safety of BCB-enhanced MSOT imaging in monitoring oocyte viability. Overall, the experimental results suggest that the method offers a significant advance in the use of contrast agents and molecular imaging for reproductive studies. Our technique improves the accurate prediction of ovarian reserve significantly and, once standardized for in vivo imaging, could provide an effective tool for clinical infertility management.

Effects of Myo-inositol on Physiological and Reproductive Traits through Blood Parameters, Oocyte Quality and Embryo Transfer in Mice

Background: The beneficial effects of myo-inositol to mammals on physiological and reproductive functions and treatments of malfunctions have been reported. The aims of present study were to assess the efficacy of myo-inositol supplementation on physiological and reproductive performances through blood parameters, oocyte quality and embryo transfer in mice. Methods: Seventy-five virgin female mice were classified into three equal groups; control group (T1) versus two myo-inositol groups (T2 receive 30 mmol/l and T3 receive 60 mmol/l). Changes of body temperature, blood oxygen and glucose, heart rate, blood parameters (RBCs, WBCs, hematocrit, glucose and total protein), oocyte quality (cumulus enclosed, diameter and brilliant cresyl blue stain) and reproductive performances (litter weight and size) were determined. Result: The results indicated that myo-inositol supplementation resulted in significant increase in values of RBCs, WBCs, hematocrit and total protein in addition to significant hypoglycemia. The quality of oocytes in case of cumulus enclosed, diameter and brilliant cresyl blue staining and reproductive performances were improved due to myo-inositol supplementation. Furthermore, myo-inositol supplementation effectively alleviated hypothermia and hyperglycemia following general anaesthesia.