In vitro maturation of caprine follicular oocytes

Abstract

This study was designed to analyse the effect of three oocyte retrieval methods, aspiration, slicing and puncture on the yield of different quality grades of oocytes and to evaluate the in vitro maturation rate of different grades of caprine oocytes. One hundred and thirty eight ovaries of Malabari goats and its crossbreds collected from the slaughter house were subjected to the oocyte retrieval methods. The oocytes harvested were graded based on the number of cumulus cell layers and ooplasm character into A, B, C and poor quality grades. Oocytes of A, B and C grades were subjected to maturation for 24 h in TCM-199 medium under standard culture conditions. Average yield of COC per ovary by aspiration, slicing and puncture was 3.93 ± 0.11, 4.44 ± 0.06 and 3.59 ± 0.07 respectively. Yield was significantly higher in slicing method than aspiration and puncture. The percentage yield of A, B, C and poor quality grades of oocytes by aspiration method was 26.74 per cent, 26.62 per cent, 24.77 per cent and 21.87 per cent respectively. Mean yield of oocytes of each quality grade by the same method were 1.05 ± 0.05, 1.05 ± 0.08, 0.98 ± 0.07 and 0.86 ± 0.04 respectively. Slicing yielded 23.08 per cent A class, 28.15 per cent B class, 24.44 per cent C class and 23.96 per cent poor quality oocytes. Mean yield of oocytes per ovary in these classes by slicing method were 1.04 ± 0.04, 1.25 ± 0.05, 1.08 ± 0.07 and 1.06 ± 0.06 respectively. Percentage yield of A, B, C and poor quality oocytes by puncture was 29.01 per cent, 30.26 per cent, 22.07 per cent and 18.66 per cent respectively. Mean yield per ovary by puncture method was 1.04 ± 0.04, 1.08 ± 0.03, 0.79 ± 0.04 and 0.67 ± 0.03 for A, B, C and poor quality oocytes respectively. No significant difference was observed in the yield of A, B and C class oocytes between aspiration, slicing and puncture. Yield of poor quality oocytes were significantly more in slicing method. The cumulus expansion rate of A class oocytes obtained by aspiration, slicing and puncture was 77.75 per cent, 69.70 per cent and 71.49 per cent respectively. Class C oocytes exhibited a cumulus expansion rate of 63.48 per cent, 51.37 per cent and 63.39 per cent respectively when collected by aspiration, slicing and puncture method. Class C oocytes obtained by aspiration, slicing and puncture when subjected to in vitro maturation exhibited a cumulus expansion rate of 39.17, 32.57 and 37.29 per cent respectively. Retrieval method was found to have no significant effect on cumulus expansion potential of caprine oocytes, whereas the COC morphology had significant effect on cumulus expansion potential. Nuclear maturation rate of A class oocytes collected by aspiration, slicing and puncture method were respectively 40, 30 and 50 per cent and polar body extrusion rate was 30, 20 and 30 per cent respectively. Class B oocytes exhibited nuclear maturation rate of 20, 10 and 30 per cent and polar body extrusion rate of 10, 10 and 20 per cent respectively by aspiration, slicing and puncture. Ten, 10 and 20 per cent of C class oocytes retrieved by aspiration, slicing and puncture exhibited nuclear maturation and 10 per cent polar body extrusion was observed in C class oocytes retrieved by puncture. None of the C class oocytes collected by aspiration or slicing exhibited polar body extrusion. This study proved that slicing is a better method than aspiration or puncture for retrieval of oocytes from caprine ovaries as it yielded more number of oocytes per ovary. Retrieval methods had no significant effect, whereas COC morphology was found to have significant effect on cumulus expansion, nuclear maturation and polar body extrusion rates of different grades of oocytes