Preliminary Study in Immature Canine Oocytes Stained with Brilliant Cresyl Blue and Obtained From Bitches with Low and High Progesterone Serum Profiles

Abstract

This study was conducted: (i) to observe the features and levels of blue colour impregnation in morphologically selected immature canine cumulus oocyte complexes (COCs) stained with the brilliant cresyl blue (BCB) dye, as indicators of quality, and integrity of nuclear oocyte chromatin configuration before in vitro maturation (IVM); (ii) to observe the relationship between the influence of serum progesterone (SP) concentrations from ovary donors and BCB staining of immature dog oocytes. The results showed that out of 138 canine COCs, germinal vesicle (GV) stage prevailed in BCB+ oocytes at percentages of 67.4% (60/89), which were statistically higher than those observed in BCB+/- (52.2%; 23/44) and BCB- (20%; 1/5) oocytes (p = 0.023). Oocytes BCB+ were interpreted as those having completed their growth and therefore possessing the capacity to mature and develop in vitro. Ooplasm and cumulus cells (CCs) of canine oocytes were BCB staining independent. Ooplasm blue colour staining reaction varied between grown oocytes, revealing different levels of glucose-6-phosphate dehydrogenase activity among and within oocytes. Additionally, SP profile of ovary donors was not a relevant indicator for selection of oocytes screened with the BCB stain. Similar numbers of high quality oocytes were observed to be BCB+, BCB+/- and BCB- between groups of females with SP varying from 0 to 2.5 ng/ml (n = 5), and those with SP varying from 2.6 to 16.7 ng/ml (n = 4) (p = 0.680). It may be inferred that bitches with low and high SP profiles have grown oocytes in their ovaries, as determined by the BCB absorbance in their ooplasms.

Prediction of oocyte developmental competence in ovine using glucose-6-phosphate dehydrogenase (G6PDH) activity determined at retrieval time

“…One association, evidenced in a variety of species, is the inverse relationship between G6PDH-activity and oocyte diameter [13,21,23,28,30] as observed in our study. Moreover, either oocyte volume [7], or size of originating follicle [31], associated with G6PDH-activity.…”

“…Improvements in oocyte screening have been achieved in several species by application of BCB test [13,14,20,[27][28][29], a concept that we have successfully incorporated into the design of our study. To determine the validity of BCB test for in vitro ovine oocyte selection, G6PDH-activity was associated with oocyte diameter, meiotic competence and subsequent pre-implantation development.…”

Oocytes Selected Using BCB Staining Enhance Nuclear Reprogramming and the In Vivo Development of SCNT Embryos in Cattle

“…BCB staining has been used for the selection of oocytes from several mammalian species. It was reported that BCB+ oocytes yielded a significantly higher blastocyst developmental rate than the BCB− and control oocytes in pig [2], goat [3], [4], sheep [5], mouse [6], dog [7], and bovine [1], [8]–[10]. However, whether BCB+ oocytes enhance the in vivo and full-term development of SCNT embryos and why BCB+ oocytes (with finished growth) are better than BCB- (growing oocytes) or non-treated oocytes (control oocytes) is unknown.…”

Selection of Ovine Oocytes by Brilliant Cresyl Blue Staining

“…In addition, BCB stains the cytoplasm blue or colorless depending on G6PDH activity [6, 7]. Research in pigs [7–9], goats [10–12], cows [13–16], mice [17], and dogs [18] indicate that the percentage of BCB+ oocytes reaching the blastocyst stage is significantly higher than BCB− oocytes. To our knowledge, there are no reports regarding the in vitro developmental competence of sheep oocytes selected using BCB staining.…”