Purpose: To investigate whether sterile filtered light paraffin oil (SPO) overlaying is superior to washed light mineral oil (WMO) in supporting the in vitro developmental competence of bovine follicular oocytes. In addition, the effects of the two types of oil overlaying were compared with oil overlaying plus co-culture (CC) on bovine embryo development in vitro.
Methods: Bovine follicular oocytes retrieved from abattoir-derived ovary were in vitro matured, fertilized and cultured in 50 μL drops overlayed with WMO or SPO and were subsequently evaluated for development rates. In second experiment, day 2 embryos grown under WMO overlaying were further cultured for 6 days in the presence (WMO+CC and SPO+CC) or absence of adult ear skin fibroblast-based co-culture system overlaid with WMO or SPO. Blastocysts from each group were evaluated for total nuclei number or were further cultured for 48 h to evaluate post-hatching development.
Results: SPO overlaying resulted in significant higher (p < 0.05) development rate to morula (44.8% versus 30.6%) and blastocyst (32.8% versus 21.7%) than WMO. Also, treatment of the day 2 embryo cultures with SPO overlaying or oil plus CC (WMO+CC or SPO+CC groups) reached significantly higher development rates from the morula stage compared to embryo cultures treated with the WMO overlaying (p < 0.05). However, the development rates of the SPO treatment group (morula: 72.7%; blastocyst: 53.1%) were slightly high compared to development of the culture treated with WMO+CC (69.6 and 50.4%, respectively). This similar developmental competence pattern was also observed in cell number and embryo hatching rate.
Conclusion: SPO overlaying is superior to WMO and WMO+CC in supporting in vitro development of bovine embryos. The development rates are further enhanced when embryos are cultured in co-culture system overlaid with SPO. Thus, this data suggest that overlaying oil can significantly influence the pre-implantation embryo development in vitro.
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This study was supported by a grant (01-PJ10-PG8-01EC01-0010) of the Korea Health 21 R&D Project, Ministry of Health & Welfare, Republic of Korea.
Bavister BD, Rose-Hellekant TA, Pinyopummintr T: Development of in vitro matured/in vitro fertilized bovine embryos into morulae and blastocysts in defined culture media. Theriogenology 1992;37:127–146CrossRefGoogle Scholar
Rief S, Sinowatz F, Stojkovic M, Einspanier R, Wolf E, Prelle K: Effects of a novel co-culture system on development, metabolism and gene expression of bovine embryos produced in vitro. Reproduction 2002;124:543–556PubMedCrossRefGoogle Scholar
Fukuda Y, Ichikawa M, Naito K, et al.: Birth of normal calves resulting from bovine oocytes matured, fertilized, and cultured with cumulus cells in vitro up to the blastocyst stage. Biol Reprod 1990;42:114–119PubMedCrossRefGoogle Scholar
Lee ST, Cho MY, Kim EJ, Kim TM, Lee CK, Han JY, Lim JM: Renovation of a drop embryo cultures system by using refined mineral oil and the effect of glucose and/or hemoglobin added to a serum-free medium. J Vet Med Sci 2004;66(1):63–66PubMedCrossRefGoogle Scholar
Fukui Y, McGowan LT, James RW: Factors affecting the in vitro development to blastocysts of bovine oocytes matured and fertilized in vitro. J Reprod Fertil 1991;92:125–131PubMedCrossRefGoogle Scholar
Hwu Y, Lee RK, Chen C, Su J, Chen Y, Lin S: Development of hatching blastocysts from immature human oocytes following in-vitro maturation and fertilization using a co-culture system. Hum Reprod 1998;13:1916–1921PubMedCrossRefGoogle Scholar
Fukui Y, Lee ES, Araki N: Effect of medium renewal during culture in two different culture systems on development to blastocysts from in vitro produced early bovine embryos. J Anim Sci 1996;74:2752–2758PubMedGoogle Scholar
Miller KF, Goldberg JM, Collins RL: Covering embryo cultures with mineral oil alters embryo growth by acting as a sink for an embryo toxic substance. J Assist Reprod Genet 1994;11:342–345PubMedCrossRefGoogle Scholar
Van Soom A, Langendonckt A, Mahmoudzadeh AR, Deluyker H, Dessy F, de Kruif A: Effect of oil quality on in vitro embryonic development in the bovine. Theriogenology 1994;41:325CrossRefGoogle Scholar
Erbach GT, Bhatnagar P, Baltz JM, Biggers JD: Zinc is a possible toxic contaminant of silicone in microdrop cultures of preimplantation mouse embryos. Hum Reprod 1995;10:3248–3254PubMedGoogle Scholar
Borque C, Pintado B, Garcia P, Sanchez R: Effect of washing oil on in vitro development of mouse embryos. Theriogenology 1996;45:206CrossRefGoogle Scholar
Provo MB, Herr C: Washed paraffin oil becomes toxic to mouse embryos upon exposure to sunlight. Theriogenology 1998;49:214CrossRefGoogle Scholar
Van Soom A, Mahmoudzadeh AR, Christophe A, Ysebaert MT, de Kruif A: Silicone oil used in microdrop culture can affect bovine embryonic development and freezability. Reprod Domest Anim 2001;36:169–176PubMedGoogle Scholar
Shimade M, Kawano N, Terada T: Delay of nuclear maturation and reduction in developmental competence of pig oocytes after mineral oil overlay of in vitro maturation media. Reproduction 2002;124:557–564CrossRefGoogle Scholar
Rosenkrans CF Jr, Zeng GQ, Mcnamara GT, Schoff PK, First NL: Development of bovine embryos in vitro as affected by energy substrates. Biol Reprod 1993;49:459–462PubMedCrossRefGoogle Scholar
Thouas GA, Korfiatis NA, French AJ, Tervit HR: Simplified technique for differential staining of inner cell mass and trophectoderm cells of mouse and bovine blastocysts. Reprod Biomed Online 2000;3:25–29CrossRefGoogle Scholar