Register      Login
Reproduction, Fertility and Development Reproduction, Fertility and Development Society
Vertebrate reproductive science and technology
RESEARCH ARTICLE

Dimethylsulfoxide and conjugated linoleic acids affect bovine embryo development in vitro

Hanna Stinshoff A C E , Sandra Wilkening A , Ana Hanstedt A B , Heinrich Bollwein A D and Christine Wrenzycki A B C
+ Author Affiliations
- Author Affiliations

A Clinic for Cattle, University of Veterinary Medicine, Hannover, Foundation, Bischofsholer Damm 15, 30173 Hannover, Germany.

B Unit for Reproductive Medicine, University of Veterinary Medicine, Hannover, Foundation, Bünteweg 15, 30559 Hannover, Germany.

C Present address: Clinic for Veterinary Obstetrics, Gynecology and Andrology, Frankfurter Str. 106, 35392 Giessen, Germany.

D Present address: Clinic for Reproductive Medicine, Large Animal Department, Vetsuisse-Fakultät Universität Zürich, Winterthurerstrasse 260, 8057 Zürich, Switzerland.

E Corresponding author. Email: hanna.stinshoff@vetmed.uni-giessen.de

Reproduction, Fertility and Development 26(4) 502-510 https://doi.org/10.1071/RD12372
Submitted: 15 November 2012  Accepted: 13 March 2013   Published: 11 April 2013

Abstract

Conjugated linoleic acids (CLA) are employed to overcome the bovine periparturitional negative energy balance. Especially of interest are trans10,cis12 -linoleic acid (t10c12-CLA) and cis9,trans11-linoleic acid (c9t11-CLA). Their impact on embryonic development, though, is not clear. Here, effects of both above-mentioned CLA on bovine in vitro-produced embryos were assessed. Zygotes (n = 2098) were allocated to one of seven groups: cultured with 50 or 100 µM of either c9t11-CLA or t10c12-CLA, with 14 or 28 mM DMSO or without supplement (control). Messenger RNA analysis of target gene transcripts (IGF1R, IGFBP2, IGFBP4, CPT2, ACAA1, ACAA2, FASN, SCD) via RT-qPCR was performed in single blastocysts. Cleavage rates did not differ, whereas development rates were decreased in both t10c12-supplemented groups in comparison to the unsupplemented group (31.7% ± 2.2 control vs 20.2% ± 2.0 50 µM t10c12 vs 21.0% ± 2.8 100 µM t10c12). Compared with the unsupplemented group, SCD was expressed at a lower level in embryos cultured with 50 µM c9t11-CLA. The relative amount of several transcripts was increased in embryos cultured with 14 mM DMSO in comparison to those that developed in the presence of 50 µM t10c12-CLA (IGFBP2, ACAA1, CPT2, FASN, SCD) or 50 µM c9t11-CLA (IGF1R, IGFBP2, ACAA1, CPT2, FASN, SCD). The molecular analyses show that CLA influence embryonic fat metabolism.

Additional keywords: CLA, DMSO, gene expression, high yielding dairy cows, IVP, negative energy balance.


References

Adashi, E. Y. (1993). Intraovarian regulation: the proposed role of insulin-like growth factors. Ann. N. Y. Acad. Sci. 687, 10–12.
Intraovarian regulation: the proposed role of insulin-like growth factors.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK3sXltlWmu7o%3D&md5=d09ea1ebd6a29081595ffd9ec532d6fcCAS | 8323164PubMed |

Applied Biosystems User Bulletin (1997, updated 2001). ABI Prism 770 Sequence Detection System. Applied Biosystems User Bulletin 2, 1–36.

Avery, B., and Greve, T. (2000). Effects of ethanol and dimethylsulphoxide on nuclear and cytoplasmic maturation of bovine cumulus–oocyte complexes. Mol. Reprod. Dev. 55, 438–445.
Effects of ethanol and dimethylsulphoxide on nuclear and cytoplasmic maturation of bovine cumulus–oocyte complexes.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3cXhslOksLc%3D&md5=1c21b21751ba38546bb060468d0f89ceCAS | 10694752PubMed |

Bauman, D. E., Harvatine, K. J., and Lock, A. L. (2011). Nutrigenomics, rumen-derived bioactive fatty acids and the regulation of milk-fat synthesis. Annu. Rev. Nutr. 31, 299–319.
Nutrigenomics, rumen-derived bioactive fatty acids and the regulation of milk-fat synthesis.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3MXhtFais73F&md5=2b2c8c39bd172b0325c664bbbd07cfb7CAS | 21568706PubMed |

Baumgard, L. H., Corl, B. A., Dwyer, D. A., Saebo, A., and Bauman, D. E. (2000). Identification of the conjugated linoleic acid isomer that inhibits milk-fat synthesis. Am. J. Physiol. Regul. Integr. Comp. Physiol. 278, R179–R184.
| 1:CAS:528:DC%2BD3cXht1GgtrY%3D&md5=182cd7c3a6b29e6d189968bcab176735CAS | 10644637PubMed |

Baumgard, L. H., Matitashvili, E., Corl, B. A., Dwyer, D. A., and Bauman, D. E. (2002). Trans-10, cis-12 conjugated linoleic acid decreases lipogenic rates and expression of genes involved in milk-lipid synthesis in dairy cows. J. Dairy Sci. 85, 2155–2163.
Trans-10, cis-12 conjugated linoleic acid decreases lipogenic rates and expression of genes involved in milk-lipid synthesis in dairy cows.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD38XnsV2nsrs%3D&md5=8f2b349bdd314b77f9d6f598cf5dc24aCAS | 12362447PubMed |

Bernard, L., Leroux, C., and Chilliard, Y. (2008). Expression and nutritional regulation of lipogenic genes in the ruminant lactating mammary gland. Adv. Exp. Med. Biol. 606, 67–108.
Expression and nutritional regulation of lipogenic genes in the ruminant lactating mammary gland.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3cXhtleqsr3N&md5=fa7282f3ad1a0b7e116264f01472a9c7CAS | 18183925PubMed |

Block, J., Hansen, P. J., Loureiro, B., and Bonilla, L. (2011). Improving post-transfer survival of bovine embryos produced in vitro: actions of insulin-like growth factor-1, colony stimulating factor-2 and hyaluronan. Theriogenology 76, 1602–1609.
Improving post-transfer survival of bovine embryos produced in vitro: actions of insulin-like growth factor-1, colony stimulating factor-2 and hyaluronan.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3MXhsVanurzL&md5=d328cfa7af783145c703b10dd05a2e8aCAS | 21890189PubMed |

Childs, S., Hennessy, A. A., Sreenan, J. M., Wathes, D. C., Cheng, Z., Stanton, C., Diskin, M. G., and Kenny, D. A. (2008). Effect of level of dietary n-3 polyunsaturated fatty acid supplementation on systemic and tissue fatty acid concentrations and on selected reproductive variables in cattle. Theriogenology 70, 595–611.
Effect of level of dietary n-3 polyunsaturated fatty acid supplementation on systemic and tissue fatty acid concentrations and on selected reproductive variables in cattle.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1cXpsVCrtbs%3D&md5=aad79f18152e51f093ed9584f0c307f6CAS | 18514298PubMed |

de Veth, M. J., Griinari, J. M., Pfeiffer, A. M., and Bauman, D. E. (2004). Effect of CLA on milk-fat synthesis in dairy cows: comparison of inhibition by methyl esters and free fatty acids, and relationships among studies. Lipids 39, 365–372.
Effect of CLA on milk-fat synthesis in dairy cows: comparison of inhibition by methyl esters and free fatty acids, and relationships among studies.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2cXmtFWmur0%3D&md5=450a65dff489d1ac99280028439d0f4aCAS | 15357024PubMed |

Dovolou, E., Clemente, M., Amiridis, G. S., Messinis, I. E., Kallitsaris, A., Gutierrez-Adan, A., and Rizos, D. (2011). Effects of guaiazulene on in vitro bovine embryo production and on mRNA transcripts related to embryo quality. Reprod. Domest. Anim. 46, 862–869.
Effects of guaiazulene on in vitro bovine embryo production and on mRNA transcripts related to embryo quality.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3MXhtlemtr3F&md5=63b32caccb9da949518508643c2a1d70CAS | 21323756PubMed |

Eckert, J., and Niemann, H. (1995). In vitro maturation, fertilization and culture to blastocysts of bovine oocytes in protein-free media. Theriogenology 43, 1211–1225.
In vitro maturation, fertilization and culture to blastocysts of bovine oocytes in protein-free media.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD28zgtVCltg%3D%3D&md5=658c59925569f49b37d3ed2fb8c9af26CAS | 16727707PubMed |

Ferguson, E. M., and Leese, H. J. (1999). Triglyceride content of bovine oocytes and early embryos. J. Reprod. Fertil. 116, 373–378.
Triglyceride content of bovine oocytes and early embryos.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK1MXkslOltbc%3D&md5=1a1797e271c04cc7e69c0e31412f7fb5CAS | 10615263PubMed |

Han, L. Q., Pang, K., Li, H. J., Zhu, S. B., Wang, L. F., Wang, Y. B., Yang, G. Q., and Yang, G. Y. (2012). Conjugated linoleic acid-induced milk-fat reduction associated with depressed expression of lipogenic genes in lactating Holstein mammary glands. Genet. Mol. Res. 11, 4754–4764.
| 1:CAS:528:DC%2BC3sXhtlGjtr4%3D&md5=1e007e56240975ec1eb5fbcab6292f7dCAS | 23079976PubMed |

Holm, P., Booth, P. J., Schmidt, M. H., Greve, T., and Callesen, H. (1999). High bovine blastocyst development in a static in vitro-production system using SOFaa medium supplemented with sodium citrate and myo-inositol with or without serum proteins. Theriogenology 52, 683–700.
High bovine blastocyst development in a static in vitro-production system using SOFaa medium supplemented with sodium citrate and myo-inositol with or without serum proteins.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD3c7pvVGnsw%3D%3D&md5=bda1bd1463ad693633a4028a58f68438CAS | 10734366PubMed |

Hutchinson, I. A., Hennessy, A. A., Dewhurst, R. J., Evans, A. C., Lonergan, P., and Butler, S. T. (2012). The effect of strategic supplementation with trans-10,cis-12 conjugated linoleic acid on the milk production, oestrous cycle characteristics and reproductive performance of lactating dairy cattle. J. Dairy Sci. 95, 2442–2451.
The effect of strategic supplementation with trans-10,cis-12 conjugated linoleic acid on the milk production, oestrous cycle characteristics and reproductive performance of lactating dairy cattle.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC38XlvFylsr0%3D&md5=835939215db9317828b3479b52b85de4CAS | 22541471PubMed |

Kasai, M., and Mukaida, T. (2004). Cryopreservation of animal and human embryos by vitrification. Reprod. Biomed. Online 9, 164–170.
Cryopreservation of animal and human embryos by vitrification.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2cXnt1Gjt7k%3D&md5=8d989e477db3af1621309ce72912c02dCAS | 15333245PubMed |

Kastrop, P. M., Bevers, M. M., Destree, O. H., and Kruip, T. A. (1990). Analysis of protein synthesis in morphologically classified bovine follicular oocytes before and after maturation in vitro. Mol. Reprod. Dev. 26, 222–226.
Analysis of protein synthesis in morphologically classified bovine follicular oocytes before and after maturation in vitro.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK3cXlt12rt7g%3D&md5=41e41890f830b3405e743f5658269a8fCAS | 2375875PubMed |

Kuzmany, A., Havlicek, V., Wrenzycki, C., Wilkening, S., Brem, G., and Besenfelder, U. (2011). Expression of mRNA, before and after freezing, in bovine blastocysts cultured under different conditions. Theriogenology 75, 482–494.
Expression of mRNA, before and after freezing, in bovine blastocysts cultured under different conditions.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3MXjs1Knuw%3D%3D&md5=3852d8355bdf40ec150e74d3b353cd01CAS | 21144573PubMed |

Lapa, M., Marques, C. C., Alves, S. P., Vasques, M. I., Baptista, M. C., Carvalhais, I., Silva Pereira, M., Horta, A. E., Bessa, R. J., and Pereira, R. M. (2011). Effect of trans-10 cis-12 conjugated linoleic acid on bovine oocyte competence and fatty-acid composition. Reprod. Domest. Anim. 46, 904–910.
Effect of trans-10 cis-12 conjugated linoleic acid on bovine oocyte competence and fatty-acid composition.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3MXhtlemtrvM&md5=48f88517ebf95b5a2de109e2752456a1CAS | 21366717PubMed |

Leese, H. J. (2002). Quiet please, do not disturb: a hypothesis of embryo metabolism and viability. Bioessays 24, 845–849.
Quiet please, do not disturb: a hypothesis of embryo metabolism and viability.Crossref | GoogleScholarGoogle Scholar | 12210521PubMed |

Leese, H. J., Baumann, C. G., Brison, D. R., McEvoy, T. G., and Sturmey, R. G. (2008). Metabolism of the viable mammalian embryo: quietness revisited. Mol. Hum. Reprod. 14, 667–672.
Metabolism of the viable mammalian embryo: quietness revisited.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1MXltV2gsA%3D%3D&md5=9d036ceba35f2b76b7f7b7f53ba983d2CAS | 19019836PubMed |

Leone, V. A., Worzalla, S. P., and Cook, M. E. (2010). Evidence that maternal conjugated linoleic acid negatively affects lipid uptake in late-stage chick embryos resulting in increased embryonic mortality. Poult. Sci. 89, 621–632.
Evidence that maternal conjugated linoleic acid negatively affects lipid uptake in late-stage chick embryos resulting in increased embryonic mortality.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3cXkvFGhtrg%3D&md5=c93ae75b4f72942c9b756dc7480f7d7bCAS | 20308393PubMed |

Leroy, J. L., Opsomer, G., De Vliegher, S., Vanholder, T., Goossens, L., Geldhof, A., Bols, P. E., de Kruif, A., and Van Soom, A. (2005a). Comparison of embryo quality in high-yielding dairy cows, in dairy heifers and in beef cows. Theriogenology 64, 2022–2036.
Comparison of embryo quality in high-yielding dairy cows, in dairy heifers and in beef cows.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD2MnitlejtQ%3D%3D&md5=03606f79c6c4ae0610aea21607fa035aCAS | 15936067PubMed |

Leroy, J. L., Vanholder, T., Mateusen, B., Christophe, A., Opsomer, G., de Kruif, A., Genicot, G., and Van Soom, A. (2005b). Non-esterified fatty acids in follicular fluid of dairy cows and their effect on developmental capacity of bovine oocytes in vitro. Reproduction 130, 485–495.
Non-esterified fatty acids in follicular fluid of dairy cows and their effect on developmental capacity of bovine oocytes in vitro.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2MXhtFent7rJ&md5=cd82fd88938bb4bfbef1f4cb6ee5e020CAS | 16183866PubMed |

Livak, K. J., and Schmittgen, T. D. (2001). Analysis of relative gene expression data using real-time quantitative PCR and the 2(–Delta Delta C(T)) Method. Methods 25, 402–408.
Analysis of relative gene expression data using real-time quantitative PCR and the 2(–Delta Delta C(T)) Method.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD38XhtFelt7s%3D&md5=9cad4ff7133d540452bb50aaaa4ededfCAS | 11846609PubMed |

Lonergan, P., Gutiérrez-Adán, A., Pintado, B., Fair, T., Ward, F., Fuente, J. D., and Boland, M. (2000). Relationship between time of first cleavage and the expression of IGF-I growth factor, its receptor and two housekeeping genes in bovine two-cell embryos and blastocysts produced in vitro. Mol. Reprod. Dev. 57, 146–152.
Relationship between time of first cleavage and the expression of IGF-I growth factor, its receptor and two housekeeping genes in bovine two-cell embryos and blastocysts produced in vitro.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3cXmsVeisb0%3D&md5=167149a0f360a7ee88a79b838fdc546cCAS | 10984414PubMed |

Marei, W. F., Wathes, D. C., and Fouladi-Nashta, A. A. (2010). Impact of linoleic acid on bovine oocyte maturation and embryo development. Reproduction 139, 979–988.
Impact of linoleic acid on bovine oocyte maturation and embryo development.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3cXns12qsLY%3D&md5=4ae3e076f816f501fab3be94536e7b5dCAS | 20215338PubMed |

Miyazaki, M., Dobrzyn, A., Sampath, H., Lee, S. H., Man, W. C., Chu, K., Peters, J. M., Gonzalez, F. J., and Ntambi, J. M. (2004). Reduced adiposity and liver steatosis by stearoyl-CoA desaturase deficiency are independent of peroxisome proliferator-activated receptor-alpha. J. Biol. Chem. 279, 35017–35024.
Reduced adiposity and liver steatosis by stearoyl-CoA desaturase deficiency are independent of peroxisome proliferator-activated receptor-alpha.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2cXmvFemtrc%3D&md5=91b0719dc7381057ae7d9f1c9e415ae3CAS | 15180999PubMed |

Namekawa, T., Ikeda, S., Sugimoto, M., and Kume, S. (2010). Effects of astaxanthin-containing oil on development and stress-related gene expression of bovine embryos exposed to heat stress. Reprod. Domest. Anim. 45, e387–e391.
Effects of astaxanthin-containing oil on development and stress-related gene expression of bovine embryos exposed to heat stress.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3cXhs1WrsLbE&md5=ebde9da9281ee3e2992f913eb95e3632CAS | 20210879PubMed |

Ntambi, J. M., Miyazaki, M., Stoehr, J. P., Lan, H., Kendziorski, C. M., Yandell, B. S., Song, Y., Cohen, P., Friedman, J. M., and Attie, A. D. (2002). Loss of stearoyl-CoA desaturase-1 function protects mice against adiposity. Proc. Natl. Acad. Sci. USA 99, 11482–11486.
Loss of stearoyl-CoA desaturase-1 function protects mice against adiposity.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD38XmslSls74%3D&md5=f837ceaaa7a44140f31557ab6ec457d2CAS | 12177411PubMed |

Pal, R., Mamidi, M. K., Das, A. K., and Bhonde, R. (2012). Diverse effects of dimethyl sulfoxide (DMSO) on the differentiation potential of human embryonic stem cells. Arch. Toxicol. 86, 651–661.
Diverse effects of dimethyl sulfoxide (DMSO) on the differentiation potential of human embryonic stem cells.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3MXhsV2ls7bE&md5=f1d811c68f8777d0bed2180c56e6d760CAS | 22105179PubMed |

Pereira, R. M., Baptista, M. C., Vasques, M. I., Horta, A. E., Portugal, P. V., Bessa, R. J., Silva, J. C., Pereira, M. S., and Marques, C. C. (2007). Cryosurvival of bovine blastocysts is enhanced by culture with trans-10 cis-12 conjugated linoleic acid (10t,12c CLA). Anim. Reprod. Sci. 98, 293–301.
Cryosurvival of bovine blastocysts is enhanced by culture with trans-10 cis-12 conjugated linoleic acid (10t,12c CLA).Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2sXit1Gnu7k%3D&md5=916ed2b7ffe9c3122930bf95b0bf4851CAS | 16644149PubMed |

Pereira, R. M., Carvalhais, I., Pimenta, J., Baptista, M. C., Vasques, M. I., Horta, A. E., Santos, I. C., Marques, M. R., Reis, A., Pereira, M. S., and Marques, C. C. (2008). Biopsied and vitrified bovine embryos viability is improved by trans10, cis12 conjugated linoleic acid supplementation during in vitro embryo culture. Anim. Reprod. Sci. 106, 322–332.
Biopsied and vitrified bovine embryos viability is improved by trans10, cis12 conjugated linoleic acid supplementation during in vitro embryo culture.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1cXls1yku78%3D&md5=5c94148b55c533ef88869522107e8134CAS | 17580103PubMed |

Perfield, J. W., Lock, A. L., Griinari, J. M., Saebo, A., Delmonte, P., Dwyer, D. A., and Bauman, D. E. (2007). Trans-9, cis-11 conjugated linoleic acid reduces milk-fat synthesis in lactating dairy cows. J. Dairy Sci. 90, 2211–2218.
Trans-9, cis-11 conjugated linoleic acid reduces milk-fat synthesis in lactating dairy cows.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2sXkvVSlsrc%3D&md5=403a258f8db34e07a2e9555b4b72adebCAS | 17430919PubMed |

Pfaffl, M. W. (2001). A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res. 29, e45.
A new mathematical model for relative quantification in real-time RT-PCR.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD38nis12jtw%3D%3D&md5=db02aaf063f3b9855f4e56b814573586CAS | 11328886PubMed |

Santos, J. E., Bilby, T. R., Thatcher, W. W., Staples, C. R., and Silvestre, F. T. (2008). Long-chain fatty acids of diet as factors influencing reproduction in cattle. Reprod. Domest. Anim. 43, 23–30.
Long-chain fatty acids of diet as factors influencing reproduction in cattle.Crossref | GoogleScholarGoogle Scholar | 18638102PubMed |

Stinshoff, H., Wilkening, S., Hanstedt, A., Bruning, K., and Wrenzycki, C. (2011). Cryopreservation affects the quality of in vitro-produced bovine embryos at the molecular level. Theriogenology 76, 1433–1441.
Cryopreservation affects the quality of in vitro-produced bovine embryos at the molecular level.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3MXht12isbjN&md5=25b25db3f3ac0013f548e37bc534b4c4CAS | 21835456PubMed |

Stinshoff, H., Krienke, M., Ekhlasi-Hundrieser, M., Wilkening, S., Hanstedt, A., Frese, D., Rath, D., Bollwein, H., and Wrenzycki, C. (2012). Seminal plasma and seminal plasma proteins added to bulk-sorted sperm do not alter the mRNA expression of in vitro-produced bovine embryos. Theriogenology 78, 132–139.
Seminal plasma and seminal plasma proteins added to bulk-sorted sperm do not alter the mRNA expression of in vitro-produced bovine embryos.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC38XksFegtbo%3D&md5=23abd359e3195ba43e4a04541d4ac682CAS | 22444548PubMed |

Sturmey, R. G., Reis, A., Leese, H. J., and McEvoy, T. G. (2009). Role of fatty acids in energy provision during oocyte maturation and early embryo development. Reprod. Domest. Anim. 44, 50–58.
Role of fatty acids in energy provision during oocyte maturation and early embryo development.Crossref | GoogleScholarGoogle Scholar | 19660080PubMed |

Wolf, E., Lahm, H., Wu, M., Wanke, R., and Hoeflich, A. (2000). Effects of IGFBP-2 overexpression in vitro and in vivo. Pediatr. Nephrol. 14, 572–578.
Effects of IGFBP-2 overexpression in vitro and in vivo.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD3M%2Fjt1yntA%3D%3D&md5=cd05698c836c162e2975ebb568c3e446CAS | 10912521PubMed |

Wrenzycki, C., Herrmann, D., Carnwath, J. W., and Niemann, H. (1999). Alterations in the relative abundance of gene transcripts in preimplantation bovine embryos cultured in medium supplemented with either serum or PVA. Mol. Reprod. Dev. 53, 8–18.
Alterations in the relative abundance of gene transcripts in preimplantation bovine embryos cultured in medium supplemented with either serum or PVA.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK1MXitleju7s%3D&md5=95ae4e498261534cebd62e0aefddd5d8CAS | 10230812PubMed |

Wrenzycki, C., De Sousa, P., Overström, E. W., Duby, R. T., Herrmann, D., Watson, A. J., Niemann, H., O’Callaghan, D., and Boland, M. P. (2000). Effects of superovulated heifer diet type and quantity on relative mRNA abundances and pyruvate metabolism in recovered embryos. J. Reprod. Fertil. 118, 69–78.
| 1:CAS:528:DC%2BD3cXpsl2nsw%3D%3D&md5=e46a071399326d3d40bbee8caa73a71bCAS | 10793627PubMed |

Zeron, Y., Sklan, D., and Arav, A. (2002). Effect of polyunsaturated fatty acid supplementation on biophysical parameters and chilling sensitivity of ewe oocytes. Mol. Reprod. Dev. 61, 271–278.
Effect of polyunsaturated fatty acid supplementation on biophysical parameters and chilling sensitivity of ewe oocytes.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD38Xlt1Ghsw%3D%3D&md5=f9ddc9499baf2c0a465a61683aab42fdCAS | 11803563PubMed |