Fig 1.
Development of yak cumulus-oocyte complexes (COCs) and embryos produced in vitro at different stages, which were used to calculate cumulus expansion, oocyte maturation and embryonic development rates.
(a): Immature COCs; (b): Mature COCs in a group without 17β-estradiol (E2); (c): Mature COCs in the 10−4 mM E2 group; (d): Mature COCs in the fulvestrant group; (e) and (f): Mature oocytes and stained with DAPI; the arrows indicate the first polar body, which was used to evaluate oocyte maturation in the different treatment groups; (g): Cleaved embryo; (h): Blastocyst. Size of Bar was noted in each figure.
Table 1.
Primer sequences and cycling conditions used in real-time PCR.
Fig 2.
Expansion areas of cumulus–oocyte complexes (COCs) matured for 22 h in media supplemented with different concentrations of 17β-estradiol (E2) or 2.9 nM fulvestrant.
Different letters on the bars indicate values that differed significantly (P < 0.05).
Table 2.
Effects of E2 on maturation of yak oocytes.
Fig 3.
Relative mRNA expression of GDF9, FGF10 and BMP15 in mature cumulus-oocyte complexes (COCs).
Expression levels are shown as relative quantities and were analysed using real-time polymerase chain reaction. β-Actin was used to normalise each gene, and mature COCs without 17β-estradiol (E2) were used as calibrators. (a), (b) and (c) represent the relative levels of GDF9, FGF10 and BMP15 mRNA in mature COCs. Significant differences between different groups are indicated by different letters on the bars (P < 0.05).
Fig 4.
Relative abundances of GDF9, FGF10 and BMP15 proteins in mature cumulus-oocyte complexes (COCs).
(a): Expression levels of GDF9, FGF10 and BMP15 proteins as detected by western blotting in mature COCs in different treatment groups. (b), (c) and (d) represent the relative abundances of GDF9, FGF10 and BMP15 proteins in mature COCs, respectively. Expression levels are shown as relative quantities. β-Actin was used to normalise each protein, and mature COCs without 17β-estradiol (E2) were used as calibrators. Significant differences between different groups are indicated by different letters on the bars (P < 0.05).
Fig 5.
Relative mRNA expression levels of HAS2, PTGS2, PTX3 and TNFAIP6 in mature cumulus-oocyte complexes (COCs).
Expression levels are shown as relative quantities and were analysed using real-time polymerase chain reaction. β-Actin was used to normalise gene expression, and mature COCs without 17β-estradiol (E2) were used as calibrators. (a), (b), (c) and (d) represent the relative levels of HAS2, PTGS2, PTX3, and TNFAIP6 mRNA in mature COCs, respectively. Significant differences between different groups are indicated by different letters on the bars (P < 0.05).
Fig 6.
Relative abundances of HAS2, PTGS2, PTX3 and TNFAIP6 proteins in mature cumulus-oocyte complexes (COCs).
(a): Expression levels of HAS2, PTGS2, PTX3 and TNFAIP6 proteins as determined by western blot in mature COCs in different treatment groups. (b), (c), (d) and (e) represent the relative abundances of HAS2, PTGS2, PTX3, and TNFAIP6 proteins in mature COCs, respectively. Expression levels are shown as relative quantities. β-Actin was used to normalise each protein, and mature COCs without 17β-estradiol (E2) were used as calibrators. Significant differences between different groups are indicated by different letters on the bars (P < 0.05).
Table 3.
Developmental competence of yak IVF embryos from different groups.
Fig 7.
Total cell numbers and their allocation to the inner cell mass (ICM) and trophectoderm (TE) lineages of yak blastocysts in different treatment groups.
TE cells stained with CDX2 appear red, and total cell nuclei stained with 4′,6-diamidino-2-phenylindole appear blue. Bar = 100 μm in panels.
Table 4.
Total cell numbers and their allocation to the ICM and TE lineages of yak blastocysts from different groups.