Journal of Animal Reproduction and Biotechnology 2019; 34(4): 300-304
Published online December 31, 2019
Copyright © The Korean Society of Animal Reproduction and Biotechnology.
Sung-Sik Kang, Ui-Hyung Kim, Seok-Dong Lee, Myung-Suk Lee, Man-Hye Han and Sang-Rae Cho*
Hanwoo Research Institute, NIAS, RDA, Pyeongchang 25340, Korea
Correspondence to: Sang-Rae Cho
This study was conducted to examine the oocyte recovery efficiency through having an OPU session once and twice a week. Also, the oocyte recovery efficiency was examined by using OPU after two and three months of rest period. Six cows were used for oocytes collection and were randomly divided into two groups. In experiment 1, OPU sessions were conducted once and twice a week to collect oocytes. The collected oocytes between once and twice OPU groups were classified into four groups (grade 1, 2, 3 and 4) according to the quality of cumulus cells and ooplasm. Based on the result, the percentage of collected oocytes per aspirated follicle number was similar between once and twice OPU session groups (65.5 ± 1.9 and 68.7 ± 1.4 vs.). However, the percentage of grade 1 oocytes from the twice OPU session group was significantly high compared with that of the once a week OPU session group (25.3 ± 0.9 and 32.5 ± 1.2% vs. once and twice session group, respectively,
Keywords: COCs, Hanwoo, OPU
The multiple ovulation and embryo transfer method (MOET) is an important reproductive tool in dairy and beef cattle industry. The improvement of genetically valuable cow and elite bulls will be accelerated by using the MOET method. Recently, the tool for embryo production system in vivo using MOET has been changed with ovum pick up (OPU) with ultrasound guided probe, and in
In the present study, six Hanwoo cows or heifers were used in Hanwoo Research Institute, National Institute of Animal Science, Rural Development Administration (RDA), Republic of Korea. Hanwoo cows with normal estrus cycle and normal body condition score (2.5-3.0) were used as a donor. Formulated concentrate (3 kg/day) and hay (rice straw, 6 kg/day) were fed to the individual Hanwoo cows. The chemical composition of formulated concentrates to donor cows are shown below: 12.4% moisture, 13.6% crude protein, 1.5% ether extract, 9.34% crude fiber, crude ash 4.79%, 0.72% calcium, and 0.53% P. chemical composition of hay were as below: 11.64% moisture, 5.91% crude protein, 0.23% ether extract, 7.48% crude fiber, 4.44 % crude ash, 0.17% calcium and 0.53% P.
COCs were collected by using OPU procedure once and twice a week. The donor cows were restrained by using a restraining frame, 7 mg Xylazine HCL 0.2 mL (Rompun, Bayel corp.), injected in the jugular vein, and was given caudal epidural anesthesia with 5 mL of 2% lidocaine. The ovarian follicles were seen by using ultrasound-guided probe (7.5 MHz convex scanner, MyLabTM30-VETGOLD, Esaote, Genova, Italy). Follicles with the size of 2 to 8 mm were aspirated by using probe with 19G needle. The vacuum pressure was maintained with 60-80 mmHg (10-15 mL/min). To prevent coagulation of blood and oocyte, HEPES solution with 10 IU/mL heparin was used for aspiration and collection COCs. The aspirated follicular fluids were collected by using 50 mL tube and were transferred to the laboratory. Recovered COCs were classified to grade 1 to 4 under stereomicroscope by compactness and thickness of cumulus cells. COCs with grade 1 have normal morphological ooplasm and compactness cumulus cells. While COCs with grade 2 showed some cumulus cells were detached and cumulus cells were normal. The COCs with grade 3 showed degenerated cumulus cells and abnormal ooplasm, and COCs with grade 4 showed that almost cumulus cells were degenerated and detached.
In experiment 1, we compared the number and quality of COCs between once and twice a week OPU session. Six cows were divided into two groups. The first group conducted one OPU session in a week. In total, eight times of OPU sessions were conducted in individual donor cows. The second group conducted a twice a week OPU session. Overall, 13 to 14 OPU sessions were conducted in individual donor cows. In experiment 2, we compared the effect of rest periods of cows on the number of oocytes by using an OPU. After collecting COCS for two months, the first group with two cows took two months of rest. The second group with two cows took three months of rest. After which, the COCs were collected by using OPU twice a week. A total of 26 OPUs sessions were conducted in each group. The number of recovered COCs and the grade of COCs were examined.
The mean of COCs graded between one and two times OPU interval groups were compared by using one way ANOVA with General Linear Model. A p value of ≤ 0.05 was considered significantly different.
In the present study, we examined COCs recovery efficiency between once and twice a week OPU session groups. Moreover, the effect of resting periods was compared between two and three months after finishing the OPU procedure. In Table 1, the COCs recovery efficiency in one OPU session in a week is shown. The COCs recovery rate in cow A showed a total number of 120 aspirated follicles and 74 (61.7%) recovered COCs. The percentage of grade 1 was 24.3% (18/74) and percentage of grade was 23.0 % (17/74). In, Hanwoo B, 88 follicles were aspirated and 59 (67.0%) COCs were collected. The percentage of grade 1 was 27.1% (16/59) and percentage of grade 2 was 22% (13/59). In Hanwoo cow C, the COCs recovery efficiency was 67.8% (57/84). The percentage of grade 1 was 24.6% (14/57) and percentage of grade 2 was 31.6 % (18/57). In Table 2, COCs recovery efficiency in twice OPU sessions in a week are shown. In Hanwoo cow D, 162 follicles were aspirated and 115 COCs were collected. Its COCs recovery efficiency rate was 70.9%. The percentage of grade 1 was 30.4% (35/115) and percentage of grade 2 was 17.4% (20/115). In Hanwoo cow E, 217 follicles were aspirated and 150 COCs were recovered (69.1%). The percentage of grade 1 was 34.7 % (52/150) and percentage of grade 2 was 25.3% (38/150). In Hanwoo cow F, 271 follicles were aspirated and 179 COCs were recovered (66.1%). The percentage of grade 1 was 32.4 % (58/179) and percentage of grade was 17.3% (31/179). As shown Table 3, there was no difference in oocyte recovery rate between once and twice OPU session groups. Percentage of grade 2-4 between once and twice a week OPU session group were not significantly different. However, percentage of grade 1 of twice a week OPU session group was significantly higher than that of once a week OPU session group (68.7 ± 1.4 and 65.5 ±1.9 vs. once and twice OPU session groups,
In conclusion, twice a week OPU session is more efficient to collect good quality of oocytes from donor cows compared to once a week OPU session group. In addition, the rest period for three months will induce an increase in the total number of oocytes and decease low quality of oocytes by using OPU. Further, we should examine the embryo production and embryo quality using oocytes from donor cows with two and three rest periods.
No potential conflict of interest relevant to this article was reported.
This study was supported by 2019 RDA Fellowship Program of Hanwoo Research Institute, Rural Development Administration, Republic of Korea. This work was carried out with the support of “Cooperative Research Program for Agriculture Science & Technology Development (project title: Development of production technique for same offspring using OPU Hanwoo oocytes, Project No. PJ010291032018)” Rural Development Administration, Republic of Korea.
SS Kang, NIAS, Postdoctoral Fellow, https://orcid.org/0000-0002-9453-5377
UH Kim, NIAS, Researcher, https://orcid.org/0000-0002-2197-5080
SD Lee, NIAS, Research Assistant, https://orcid.org/0000-0002-4481-5338
MS Lee, NIAS, Research Assistant, https://orcid.org/0000-0002-9111-8535
MH Han, NIAS, Researcher, https://orcid.org/0000-0002-8104-4587
SR Cho, NIAS, Researcher, https://orcid.org/0000-0003-0209-6248