N-acetyl-L-cysteine Improves the Developmental Competence of Bovine Oocytes and Embryos Cultured In Vitro by Attenuating Oxidative Damage and Apoptosis
Abstract
:1. Introduction
2. Materials and Methods
2.1. Chemicals
2.2. Oocyte Collection and In Vitro Maturation
2.3. In Vitro Fertilization (IVF) and In Vitro Culture (IVC)
2.4. Total RNA Extraction and Real-Time RT-PCR
2.5. Measurement of Intracellular ROS, GSH Levels and Mitochondrial Membrane Potential
2.6. Cell Proliferation Assay
2.7. Total Antioxidant Capability Measurement
2.8. Apoptosis Assays and Total Blastocyst Cell Counting
2.9. Blastocyst Outgrowth Assay
2.10. Statistical Analyses
3. Results
3.1. Effects of NAC Supplementation in IVM Medium on the Expression of NFKB1 Gene in Bovine Oocytes
3.2. Effects of NAC Supplementation in IVM Medium on Oocyte Maturation and Embryo Development
3.3. Effects of NAC Supplementation in IVM Medium on Bovine Oocyte Quality
3.4. Effects of NAC Supplementation in IVM Medium on Bovine Cumulus Cells Quality
3.5. Effects of NAC Supplementation in IVC Medium on Bovine Embryo Development
3.6. Effects of NAC Supplementation in IVC Medium on Bovine Embryo Quality
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Gene | Primer Sequences (5′-3′) | Accession Number | Product Size (bp) |
---|---|---|---|
Bax | GGCTGGACATTGGACTTCCTTC | NM_173894 | 112 |
TGGTCACTGTCTGCCATGTGG | |||
Bcl-2 | TGGATGACCGAGTACCTGAA | NM_001166486.1 | 123 |
GAGACAGCCAGGAGAAATCAAA | |||
SOD1 | CCACGTCCATCAGTTTGGAGA | NM_174615.2 | 92 |
CTTTTGGCCCACCGTGTTTT | |||
CAT | CTGGGACCCAACTATCTCCA | NM_001035386.2 | 148 |
GATGCTCGGGAGCACTAAAG | |||
NFKB1 | TGGCGGAATTACCTTCCATAC | DQ464067 | 110 |
CATCACTCTTGCCACAACTTTC | |||
GAPDH | CCCAGAATATCATCCCTGCT | NM_001034034 | 185 |
CTGCTTCACCACCTTCTTGA |
Group | No. (%, ± SEM) of Mature/Retrieved Oocytes | No. of Oocytes Used for IVF | No. (%, ± SEM) of IVF Embryos Developed to | |||
---|---|---|---|---|---|---|
2-Cell | Top-Quality 2-Cell 1 | Blastocyst | Hatched Embryo | |||
IVM medium (Control) | 236/299 | 219 | 185 | 163 | 69 | 38 |
(79.03 ± 0.22) b | (83.56 ± 0.40) b | (88.30 ± 0.65) b | (37.15 ± 1.99) b,2 | (54.77 ± 0.14) b | ||
IVM + Lcys (0.6 mM) | 182/220 | 250 | 231 | 206 | 90 | 51 |
(82.76 ± 1.08) a | (91.42 ± 1.59) a | (89.26 ± 1.48) b | (38.97 ± 1.88) ab | (57.22 ± 1.04) b | ||
IVM + NAC (0.1 mM) | 178/231 | 233 | 192 | 166 | 72 | 40 |
(77.10 ± 0.18) bc | (82.74 ± 1.97) b | (86.71 ± 0.97) b | (37.73 ± 0.80) ab | (55.94 ± 1.56) b | ||
IVM + NAC (1.0 mM) | 167/201 | 246 | 211 | 195 | 88 | 53 |
(83.06 ± 1.85) a | (85.86 ± 0.46) b | (92.93 ± 1.38) a | (41.69 ± 0.63) a | (60.65 ± 0.44) a | ||
IVM + NAC (10.0 mM) | 188/248 | 205 | 160 | 132 | 59 | 12 |
(75.69 ± 0.55) c | (78.59 ± 1.03) c | (82.28 ± 1.39) c | (37.19 ± 0.26) b | (19.42 ± 1.49) c |
Oocytes Treated | No. of Zygotes for Culture | No. (%, ± SEM) of Embryos Developed to | ||
---|---|---|---|---|
2-Cell | Blastocyst | Hatched Embryo | ||
Basic CR1aa (Control) | 398 | 361 (90.25 ± 0.26) a,1 | 137 (38.20 ± 0.43) c | 80 (58.44 ± 1.18) b |
CR1aa + NAC (0~3 dpf) | 346 | 284 (81.46 ± 1.29) b | 127 (44.46 ± 0.42) b | 72 (57.20 ± 2.05) b |
CR1aa + NAC (3~6 dpf) | 295 | 263 (87.26 ± 1.82) a | 130 (49.72 ± 0.72) a | 86 (65.85 ± 3.23) a |
CR1aa + NAC (6~9 dpf) | 342 | 315 (90.93 ± 0.76) a | 151 (48.06 ± 1.84) a | 90 (59.44 ± 1.67) ab |
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Sun, W.-S.; Jang, H.; Park, M.-R.; Oh, K.B.; Lee, H.; Hwang, S.; Xu, L.-J.; Hwang, I.-S.; Lee, J.-W. N-acetyl-L-cysteine Improves the Developmental Competence of Bovine Oocytes and Embryos Cultured In Vitro by Attenuating Oxidative Damage and Apoptosis. Antioxidants 2021, 10, 860. https://doi.org/10.3390/antiox10060860
Sun W-S, Jang H, Park M-R, Oh KB, Lee H, Hwang S, Xu L-J, Hwang I-S, Lee J-W. N-acetyl-L-cysteine Improves the Developmental Competence of Bovine Oocytes and Embryos Cultured In Vitro by Attenuating Oxidative Damage and Apoptosis. Antioxidants. 2021; 10(6):860. https://doi.org/10.3390/antiox10060860
Chicago/Turabian StyleSun, Wu-Sheng, Hoon Jang, Mi-Ryung Park, Keon Bong Oh, Haesun Lee, Seongsoo Hwang, Li-Jie Xu, In-Sul Hwang, and Jeong-Woong Lee. 2021. "N-acetyl-L-cysteine Improves the Developmental Competence of Bovine Oocytes and Embryos Cultured In Vitro by Attenuating Oxidative Damage and Apoptosis" Antioxidants 10, no. 6: 860. https://doi.org/10.3390/antiox10060860
APA StyleSun, W.-S., Jang, H., Park, M.-R., Oh, K. B., Lee, H., Hwang, S., Xu, L.-J., Hwang, I.-S., & Lee, J.-W. (2021). N-acetyl-L-cysteine Improves the Developmental Competence of Bovine Oocytes and Embryos Cultured In Vitro by Attenuating Oxidative Damage and Apoptosis. Antioxidants, 10(6), 860. https://doi.org/10.3390/antiox10060860