Cleavage ability of in vitro embryos of bali cattle based on different reproductive status of ovary at 48 hours after fertilization process

Abstract

This study aims to determine the ability of in vitro embryo cleavage of Bali cattle 48 hours after the fertilization process based on different ovarian reproductive status. Ovaries from abattoirs are grouped into four categories: ovaries without corpus luteum and dominant follicles (-CL and -FD), without corpus luteum and with dominant follicles (-CL and + FD), with corpus luteum and without dominant follicle (+ CL and -FD), and with corpus luteum and dominant follicles (+ CL and + FD). After that, oocytes are collected by slicing and only oocytes with a cytoplasmic are homongent and have compact cumulus cells used in the study. The selected oocytes are then matured in an incubator of 5% CO2, a temperature of 38.5 °C for 24 hours. After maturation, in vitro fertilization was carried out using fertilization media with a final concentration of spermatozoa 1.5 x 106 spermatozoa/ml. Oocytes 5-6 after fertilization were washed 3 times using CR1aa media then cultured in 5% CO2 incubator, temperature 38.5 °C for 48 hours. After 48 hours the culture was evaluated for the ability of embryo cleavage. The results showed that the number of embryos capable of cleave to stage 2 cells were not significantly different (P>0.05) in all treatments (1.64 ± 0.38% vs 8.00 ± 0.90% vs. 4.85 ± 0.96% vs. 8.22 ± 1.86%), while the number of embryos capable of cleave reached stage 4 cells (11.48 ± 0.82% vs 24.00 ± 2.64% vs. 24.27 ± 2.23% vs. 10.96 ± 0.69%), 8 cells (11.48 ± 1.15% vs 8.00 ± 1.07% vs. 26.21 ± 2.79 % vs 15.07 ± 1.40%), 16 cells (0.00 ± 0.00% vs 1.33 ± 0.38% vs. 4.85 ± 0.76% vs. 1.37 ± 0.38%), and the ability of embryos cleavage (24.59 ± 11.60% vs 41.33 ± 24.32% vs. 60.19 ± 22.45 % vs. 35.62 ± 10.83%) was significantly higher (P<0.05) in the ovaries with corpus luteum and without dominant follicles (+ CL and – FD). Conclusion, the ability of embryo division to reach 4 cells, 8 cells, and 16 cells after 48 hours culture was higher in the ovaries with corpus luteum and without dominant follicles (+ CL and – FD).

Habitat Preferences of Wild Mammalian Species around River Chenab in Sialkot, Gujrat and Gujranwala Districts, Punjab, Pakistan

“…The semen used was obtained from the same bull. It was thawed for 20 seconds at 37 o C, was centrifuged for 5 min at 1500 rpm 2 times, split the supernatant and spermatozoa, and then semen was added with fertilization media so that the final concentration of spermatozoa was 1.5×10 6 cells/ ml (Hall and Glaze, 2014;Hasbi et al, 2020). After that, prepared four drops on a petri dish (80 μL/drop) covered by mineral oil (Sigma Chemical Co. St. Louis MO, USA), equilibrated for 30 min.…”

“…In addition, different culturalsystems can also affect embryonic development (Abd El-Aziz et al, 2016). In vitro production of Bali cattle embryos resulted in zygote cultures which show varying rates of embryonic cell division indicated by the occurrence of 2-, 4-, 8-and 16-cell embryos on the second day or 48 h after culture (Sonjaya and Hasbi, 2019;Hasbi et al, 2020). The in vitro embryo culture system has not been able to be properly conditioned to resemble in vivo so that it can lead to the increase of reactive oxygen species (Agarwal et al, 2006).…”

Apoptosis in Bali Cattle Embryo Cells Produced In Vitro

“…Bali cattle semen was frozen in straw by centrifuging the 0.25 mL semen for 5 minutes at 1500 rpm twice, then the supernatant and spermatozoa were separated (Hasbi et al, 2020). The semen was added with fertilization media (Suzuki et al, 2000) so that the final concentration of spermatozoa was 1.5 x 10 6 cells/mL (Hasbi et al, 2020). Then, four drops (80 μL/drop) were placed into a Petri dish covered with mineral oil (Sigma-Aldrich, USA), and equilibrated for 30 minutes.…”

“…After fertilization for 5-6 hours, oocytes were washed twice using CR1aa culture media, transferred into an 80-μL drop of CR1aa culture media following modification by Sagirkaya et al (2006) and Somfai et al (2010) with some modifications. In the next step, 5 mg/mL BSA and 2.5% FBS were added, oocytes were covered with mineral oil (Sigma Chemical Company, USA), and cultured in a 5% CO 2 incubator at the temperature of 38.5°C for 48 hours (Hasbi et al, 2020). On the second day of culture, classification was carried out based on the stages of division.…”

“…Cell death during this process has been reported with unclear interpretations ranging from pathological phenomena to an integral part of normal blastocyst development (Betts and King, 2001). Preliminary studies on the embryonic development of Bali cattle cultured in vitro have different developments on the same culture day (Hasbi et al, 2020). In order to find out the differences, it is necessary to measure DOI: https://dx.doi.org/10.54203/scil.2022.wvj57 the fragmentation of DNA in embryonic cells.…”