Abstract :
A critical stage in the optimization of in vitro maturation (IVM) is the selection of good
quality oocytes. There exists a relationship between the size of the cumulus investment and
the in vitro developmental ability of the cumulus–oocyte complex (COC), which provides
a basis for the selection of the COCs. This study was designed to evaluate the effect of
the number of cumulus cell layers which enclose the oocytes, on the in vitro maturation,
cytoplasm quality and cumulus expansion of the ovine oocytes. Ovaries were obtained from
an abattoir and transported to the laboratory within 1–2 h, at 37 ◦C. Oocytes (n = 535) were
recovered by means of an aspiration pump (set at a flow rate of 10 mL H2O/min), with a
disposable 20 G needle attached. Oocytes were divided into four classes (classes I to IV –
with more than 5, 3–4, 1–2 and no cumulus cell layers, respectively) and separately cultured
in a TCM199 medium for 24 h. The morphology of oocytes was evaluated following in vitro
culture (IVC) to assess cumulus expansion, cytoplasm quality (score I with a homogenous
cytoplasm and II with granulated cytoplasm) and nuclear maturation stage. The percentage
of maximum cumulus expansion for classes I to III oocytes were 53.0 ± 1.0, 36.3 ± 2.2 and
16.3 ± 1.8% respectively. The rate of meiotic resumption of oocytes in classes I to IV were
77.0 ± 2.7, 77.2 ± 1.9, 53.0 ± 2.1 and 2.7 ± 1.1% respectively. The proportion of oocytes with
a cytoplasm quality I in oocyte classes I to IV were 62.8 ± 1.5, 59.4 ± 1.2, 36.4 ± 2.1 and
0.5 ± 1.1%, respectively. Results showed that the presence of ≥3 cumulus cell layers in the
COC prior to IVM led to a better (p < 0.05) cumulus expansion, meiotic resumption and
cytoplasmic maturation rate. Thus the morphological grading of immature ovine oocytes
may be an appropriate selection criterion regarding their developmental ability.
