Embryo development and sex ratio of in vitro-produced porcine embryos are affected by the energy substrate and hyaluronic acid added to the culture medium
Eva Torner A C , Eva Bussalleu A , M. Dolors Briz A , Marc Yeste B and Sergi Bonet AA Biotechnology of Animal and Human Reproduction (TechnoSperm), Department of Biology, Institute of Food and Agricultural Technology, University of Girona, Campus Montilivi, s/n, 17071 Girona, Spain.
B Unit of Animal Reproduction, Department of Animal Medicine and Surgery, Faculty of Veterinary Medicine, Autonomous University of Barcelona, University Campus, 08193 Bellaterra, Barcelona, Spain.
C Corresponding author. Email: eva.torner@udg.edu
Reproduction, Fertility and Development 26(4) 570-577 https://doi.org/10.1071/RD13004
Submitted: 4 January 2013 Accepted: 23 March 2013 Published: 9 May 2013
Abstract
In the present study, the effects of replacing glucose with pyruvate–lactate and supplementing these in vitro culture (IVC) media with hyaluronic acid (HA) on porcine embryo development and sex ratio were examined. The in vitro-produced (IVP) porcine embryos were cultured in NCSU-23 medium with 0.0, 0.5 or 1.0 mg mL–1 HA, and with either 5.55 mM glucose (IVC-Glu) or pyruvate (0.17 mM)–lactate (2.73 mM) from 0 to 48 h post insemination (h.p.i.) and then with glucose from 48 to 168 h.p.i. (IVC-PL). Those embryos cultured with IVC-PL had significantly higher blastocyst rates (23.7 ± 1.5%) than those cultured with IVC-Glu (14.27 ± 2.75%). At 1.0 mg mL–1, HA tended to skew the sex ratio of blastocysts towards males in those embryos cultured in IVC-PL, and led to a significant decrease in the blastocyst rate compared with embryos cultured in the presence of 0.5 and 0.0 mg mL–1 HA and IVC-Glu (4.28 ± 0.28% vs 11.01 ± 1.42% and 10.14 ± 2.77%, respectively) and IVC-PL (14.37 ± 1.35% vs 20.96 ± 2.85% and 22.99 ± 1.39%, respectively). In contrast, there were no significant differences in the total cell number per blastocyst or in apoptosis rates. In conclusion, pyruvate and lactate were the preferred energy substrates in the early stages of IVP porcine embryos. Moreover, 1.0 mg mL–1 HA significantly decreased the percentage of blastocyst rates in both the IVC-Glu and IVC-PL groups, but only by a preferential loss of female embryos for those cultured in IVC-PL.
Additional keywords: blastocyst, glucose, hyaluronan, pig, pyruvate–lactate.
References
Bermejo-Alvarez, P., Rizos, D., Lonergan, P., and Gutierrez-Adan, A. (2011). Transcriptional sexual dimorphism during preimplantation embryo development and its consequences for developmental competence and adult health and disease. Reproduction 141, 563–570.| Transcriptional sexual dimorphism during preimplantation embryo development and its consequences for developmental competence and adult health and disease.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3MXmvFCju7Y%3D&md5=bfaa79dbaea49627c7ed846e11bf15ffCAS | 21339284PubMed |
Block, J., Bonilla, L., and Hansen, P. J. (2009). Effect of addition of hyaluronan to embryo culture medium on survival of bovine embryos in vitro following vitrification and establishment of pregnancy after transfer to recipients. Theriogenology 71, 1063–1071.
| Effect of addition of hyaluronan to embryo culture medium on survival of bovine embryos in vitro following vitrification and establishment of pregnancy after transfer to recipients.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1MXjsV2qsro%3D&md5=366550580e59c2de893687884f563881CAS | 19157530PubMed |
Brison, D. R., and Schultz, R. M. (1997). Apoptosis during mouse blastocysts formation: evidence for a role for survival factors including transforming growth factor alpha. Biol. Reprod. 56, 1088–1096.
| Apoptosis during mouse blastocysts formation: evidence for a role for survival factors including transforming growth factor alpha.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK2sXisl2mu7Y%3D&md5=a77daa022476d3bbf3d634a66def175fCAS | 9160705PubMed |
Casas, I., Sancho, S., Briz, M., Pinart, E., Bussalleu, E., Yeste, M., and Bonet, S. (2009). Freezability prediction of boar ejaculates assessed by functional sperm parameters and sperm proteins. Theriogenology 72, 930–948.
| Freezability prediction of boar ejaculates assessed by functional sperm parameters and sperm proteins.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1MXhtFGlsr3E&md5=ec4c79d985e859b9058624eef80df13aCAS | 19651432PubMed |
Coy, P., Grullon, L., Canovas, S., Romar, R., Matas, C., and Aviles, M. (2008). Hardening of the zona pellucida of unfertilized eggs can reduce polyspermic fertilization in the pig and cow. Reproduction 135, 19–27.
| Hardening of the zona pellucida of unfertilized eggs can reduce polyspermic fertilization in the pig and cow.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1cXhs1CksLw%3D&md5=9cb6b444a593807c436a3a00a7c3f877CAS | 18159080PubMed |
Dang-Nguyen, T. Q., Somfai, T., Haraguchi, S., Kikuchi, K., Tajima, A., Kanai, Y., and Nagai, T. (2011). In vitro production of porcine embryos: current status, future perspectives and alternative applications. Anim. Sci. J. 82, 374–382.
| In vitro production of porcine embryos: current status, future perspectives and alternative applications.Crossref | GoogleScholarGoogle Scholar | 21615829PubMed |
Dattena, M., Mara, L., Bin, T. A., and Cappai, P. (2007). Lambing rate using vitrified blastocysts is improved by culture with BSA and hyaluronan. Mol. Reprod. Dev. 74, 42–47.
| Lambing rate using vitrified blastocysts is improved by culture with BSA and hyaluronan.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD28Xht1yjsb%2FE&md5=4802bf3e89bbfda9d7024cb8239c6e7fCAS | 16929524PubMed |
Dittmann, J., Keller, C., and Harisch, G. (1998). Inhibition of adenylate cyclise of bat hepatic membranes by glycosaminoglycans. Life Sci. 63, 2199–2208.
| Inhibition of adenylate cyclise of bat hepatic membranes by glycosaminoglycans.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK1cXnslymsL8%3D&md5=8a7cd6c1e0e912671a688c20d5fa8ac9CAS | 9851312PubMed |
Gandhi, A. P., Lane, M., Gardner, D. K., and Krisher, R. L. (2001). Substrate utilization in porcine embryos cultured in NCSU23 and G1.2/G2.2 sequential culture media. Mol. Reprod. Dev. 58, 269–275.
| Substrate utilization in porcine embryos cultured in NCSU23 and G1.2/G2.2 sequential culture media.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3MXhtVSiu7o%3D&md5=f53be782b02f290eb3e36cf0a021f90eCAS | 11170267PubMed |
Gardner, D. K., Rodriegez-Martinez, H., and Lane, M. (1999). Fetal development after transfer is increased by replacing protein with the glycosaminoglycan hyaluronan for mouse embryo culture and transfer. Hum. Reprod. 14, 2575–2580.
| Fetal development after transfer is increased by replacing protein with the glycosaminoglycan hyaluronan for mouse embryo culture and transfer.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK1MXntFWmsLs%3D&md5=c1069621e7d908f8978ab053eaa57d14CAS | 10527990PubMed |
Gil, M. A., Cuello, C., Parrilla, I., Vazquez, J. M., Roca, J., and Martinez, E. A. (2010). Advances in swine in vitro embryo production technologies. Reprod. Domest. Anim. 45, 40–48.
| Advances in swine in vitro embryo production technologies.Crossref | GoogleScholarGoogle Scholar | 20591064PubMed |
Kano, K., Miyano, T., and Kato, S. (1998). Effects of glycosaminoglycans on the development of in vitro-matured and -fertilized porcine oocytes to the blastocyst stage in vitro. Biol. Reprod. 58, 1226–1232.
| Effects of glycosaminoglycans on the development of in vitro-matured and -fertilized porcine oocytes to the blastocyst stage in vitro.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK1cXivFCltLk%3D&md5=a696a8dcf28f0515762534c65209cbb9CAS | 9603257PubMed |
Karja, N. W., Kikuchi, K., Fahrudin, M., Ozawa, M., Somfai, T., Ohnuma, K., Noguchi, J., Kaneko, H., and Nagai, T. (2006). Development to the blastocyst stage, the oxidative state, and the quality of early developmental stage of porcine embryos cultured in alteration of glucose concentrations in vitro under different oxygen tensions. Reprod. Biol. Endocrinol. 4, 54.
| Development to the blastocyst stage, the oxidative state, and the quality of early developmental stage of porcine embryos cultured in alteration of glucose concentrations in vitro under different oxygen tensions.Crossref | GoogleScholarGoogle Scholar | 17087833PubMed |
Kikuchi, K., Onishi, A., Kashiwazaki, N., Iwamoto, M., Noguchi, J., Kaneko, H., Akita, T., and Nagai, T. (2002). Successful piglet production after transfer of blastocysts produced by a modified in vitro system. Biol. Reprod. 66, 1033–1041.
| Successful piglet production after transfer of blastocysts produced by a modified in vitro system.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD38XitlClur8%3D&md5=0da6918593cb618cc40294d92c3d461fCAS | 11906923PubMed |
Kim, H. S., Lee, G. S., Hyun, S. H., Lee, S. H., Nam, D. H., Jeong, Y. W., Kim, S., Kang, S. K., Lee, B. C., and Hwang, W. S. (2004). Improved in vitro development of porcine embryos with different energy substrates and serum. Theriogenology 61, 1381–1393.
| Improved in vitro development of porcine embryos with different energy substrates and serum.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2cXhsVOmu74%3D&md5=3eb65f624eb9c57c3f8b5a50bb40b621CAS | 15036970PubMed |
Kim, H. S., Lee, G. S., Hyun, S. H., Nam, D. H., Lee, S. H., Jeong, Y. W., Kim, S., Kim, J. H., Kang, S. K., Lee, B. C., and Hwang, W. S. (2005). Embryotropic effect of glycosaminoglycans and receptors in development of porcine pre-implantation embryos. Theriogenology 63, 1167–1180.
| Embryotropic effect of glycosaminoglycans and receptors in development of porcine pre-implantation embryos.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2MXhtlSltb4%3D&md5=6b3c5a0ede2beeb4c2cf3fefc374cb1cCAS | 15710201PubMed |
Kimura, K., Iwata, H., and Thompson, J. G. (2008). The effect of glucosamine concentration on the development and sex ratio of bovine embryos. Anim. Reprod. Sci. 103, 228–238.
| The effect of glucosamine concentration on the development and sex ratio of bovine embryos.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2sXhsVSmtLfL&md5=a605557f5c543c4d62395224262e3bd5CAS | 17198747PubMed |
King, W. A., Yadav, B. R., Xu, K. P., Picard, L., Sirard, M. A., Verini Supplizi, A., and Betteridge, K. J. (1991). The sex ratios of bovine embryos produced in vivo and in vitro. Theriogenology 36, 779–788.
| The sex ratios of bovine embryos produced in vivo and in vitro.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BD283pvFyjuw%3D%3D&md5=ec262078d17a869580ce0cc551204241CAS | 16727046PubMed |
Kurebayashi, S., Miyake, M., Katayama, M., Miyano, T., and Kato, S. (1995). Improvement of developmental abilty to the blastocyst stage by addition of hyaluronic acid to chemically defined medium in diploid porcine eggs matured in-vitro and subsequently electro-activated. J. Mamm. Ova. Res 12, 119–125.
| Improvement of developmental abilty to the blastocyst stage by addition of hyaluronic acid to chemically defined medium in diploid porcine eggs matured in-vitro and subsequently electro-activated.Crossref | GoogleScholarGoogle Scholar |
Lane, M., and Gardner, D. K. (2000). Lactate regulates pyruvate uptake and metabolism in the preimplantation mouse embryo. Biol. Reprod. 62, 16–22.
| Lactate regulates pyruvate uptake and metabolism in the preimplantation mouse embryo.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3cXhslKmtA%3D%3D&md5=05d985e15b94625715243a333c9e15e1CAS | 10611062PubMed |
Lane, M., and Gardner, D. K. (2007). Embryo culture medium: which is the best? Best Pract. Res. Clin. Obstet. Gynaecol. 21, 83–100.
| Embryo culture medium: which is the best?Crossref | GoogleScholarGoogle Scholar | 17090393PubMed |
Lopes, A. S., Wrenzycki, C., Ramsing, N. B., Herrmann, D., Niemann, H., Lovendahl, P., Greve, T., and Callesen, H. (2007). Respiration rates correlate with mRNA expression of G6PD and GLUT1 genes in individual bovine in vitro-produced blastocysts. Theriogenology 68, 223–236.
| Respiration rates correlate with mRNA expression of G6PD and GLUT1 genes in individual bovine in vitro-produced blastocysts.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2sXmsFKgsLk%3D&md5=1257456bf95981928cb93f3f404411baCAS | 17559924PubMed |
Marei, W. F., Ghafari, F., and Fouladi-Nashta, A. A. (2012). Role of hyaluronic acid in maturation and further early embryo development of bovine oocytes. Theriogenology 78, 670–677.
| Role of hyaluronic acid in maturation and further early embryo development of bovine oocytes.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC38Xmt1GqsL4%3D&md5=929efcfed481808a0b87932be68d17c6CAS | 22541325PubMed |
Martín, I., García, T., Fajardo, V., Rojas, M., Pegels, N., Hernández, P. E., and Martín, I. G. (2009). SYBR-green real-time PCR approach for the detection and quantification of pig DNA in feedstuffs. Meat Sci. 82, 252–259.
| SYBR-green real-time PCR approach for the detection and quantification of pig DNA in feedstuffs.Crossref | GoogleScholarGoogle Scholar | 20416744PubMed |
Medvedev, S., Onishi, A., Fuchimoto, D. I., Iwamoto, M., and Nagai, T. (2004). Advanced in vitro production of pig blastocysts obtained through determining the time for glucose supplementation. J. Reprod. Dev. 50, 71–76.
| Advanced in vitro production of pig blastocysts obtained through determining the time for glucose supplementation.Crossref | GoogleScholarGoogle Scholar | 15007204PubMed |
Miyano, T., Hino-oka, R. E., Kano, K., Miyake, M., Kusunokir, H., and Katoi, S. (1994). Effects of hyaluronic acid on the development of 1- and 2-cell porcine embryos to the blastocyst stage in vitro. Theriogenology 41, 1299–1305.
| Effects of hyaluronic acid on the development of 1- and 2-cell porcine embryos to the blastocyst stage in vitro.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK2cXlslCmsr0%3D&md5=efdc995954d9160ca18fb390eabfdc2cCAS | 16727484PubMed |
Miyoshi, K., Umezu, M., and Sato, E. (1999). Effect of hyaluronic acid on the development of porcine 1-cell embryos produced by a conventional or new in vitro maturation/fertilization system. Theriogenology 51, 777–784.
| Effect of hyaluronic acid on the development of porcine 1-cell embryos produced by a conventional or new in vitro maturation/fertilization system.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK1MXjsFegtbY%3D&md5=c0d3fb729f94fac35ab5f4173a9e7374CAS | 10729002PubMed |
Nagai, T., Funahashi, H., Yoshioka, K., and Kikuchi, K. (2006). Up date of in vitro production of porcine embryos. Front. Biosci. 11, 2565–2573.
| Up date of in vitro production of porcine embryos.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD28XlvVagtr0%3D&md5=84d879f349968183679abe645e7fd716CAS | 16720334PubMed |
Palasz, A. T., Rodriguez-Martinez, H., Beltran-Breña, P., Perez-Garnelo, S., Martinez, M. F., Gutiérrez-Adán, A., and De la Fuente, J. (2006). Effects of hyaluronan, BSA, and serum on bovine embryo in vitro development, ultrastructure, and gene expression patterms. Mol. Reprod. Dev. 73, 1503–1511.
| Effects of hyaluronan, BSA, and serum on bovine embryo in vitro development, ultrastructure, and gene expression patterms.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD28XhtFGhtr7F&md5=cb74b2b700a549ecbd42da72a534f373CAS | 16902955PubMed |
Petters, R. M., and Wells, K. D. (1993). Culture of pig embryos. J. Reprod. Fertil. Suppl. 48, 61–73.
| 1:STN:280:DyaK2c7psVCktQ%3D%3D&md5=d1c333412cc96909299bbec3222f40ceCAS | 8145215PubMed |
Pursel, V. G., and Johnson, L. A. (1975). Freezing of boar spermatozoa: fertilizing capacity with concentrated semen and a new thawing procedure. J. Anim. Sci. 40, 99–102.
| 1:STN:280:DyaE2M%2FnslCjtw%3D%3D&md5=64a08be65961983b91263161a1d1a8deCAS | 1110222PubMed |
Rath, D., Long, C. R., Dobrinsky, J. R., Welch, G. R., Schreier, L. L., and Johnson, L. A. (1999). In vitro production of sexed embryos for gender preselection: high-speed sorting of X-chromosome-bearing sperm to produce pigs after embryo transfer. J. Anim. Sci. 77, 3346–3352.
| 1:CAS:528:DC%2BD3cXktFegtQ%3D%3D&md5=ba38d743ff257f7f011c0c75e006bf3aCAS | 10641883PubMed |
Salustri, A., Yanagishita, M., and Hascall, V. C. (1989). Synthesis and accumulation of hyaluronic acid and proteoglycans in the mouse cumulus cell-oocyte complex during follicle-stimulating hormone-induced mucification. J. Biol. Chem. 264, 13 840–13 847.
| 1:CAS:528:DyaL1MXlt1OgsLg%3D&md5=d733ea8e7d8603bc1e1356e6634e99b1CAS |
Stojkovic, M., Kölle, S., Peinl, S., Stojkovic, P., Zakhartchenko, V., Thompson, J. G., Wenigerkind, H., Reichenbach, H. D., Sinowatz, F., and Wolf, E. (2002). Effects of high concentrations of hyaluronan in culture medium on development and survival rates of fresh and frozen-thawed bovine embryos produced in vitro. Reproduction 124, 141–153.
| Effects of high concentrations of hyaluronan in culture medium on development and survival rates of fresh and frozen-thawed bovine embryos produced in vitro.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD38XmtFaksLg%3D&md5=ee69c3ac64bc5377f6af785b7d5061e7CAS | 12090927PubMed |
Stojkovic, M., Krebs, O., Kolle, S., Prelle, K., Assmann, V., Zakhartchenko, V., Sinowatz, F., and Wolf, E. (2003). Developmental regulation of hyaluronan-binding protein (RHAMM/IHABP) expression in early bovine embryos. Biol. Reprod. 68, 60–66.
| Developmental regulation of hyaluronan-binding protein (RHAMM/IHABP) expression in early bovine embryos.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3sXjtVyk&md5=3d9d236074e2e50a181e3db2dd0fd321CAS | 12493696PubMed |
Swain, J. E., Bormann, C. L., Clark, S. G., Walters, E. M., Wheeler, M. B., and Krisher, R. L. (2002). Use of energy substrates by various stage preimplantation pig embryos produced in vivo and in vitro. Reproduction 123, 253–260.
| Use of energy substrates by various stage preimplantation pig embryos produced in vivo and in vitro.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD38XhsFChtr4%3D&md5=1852c7f6c120cd1b2c0b4707ec845b2cCAS | 11866692PubMed |
Torner, E., Bussalleu, E., Briz, M. D., Gutiérrez-Adán, A., and Bonet, S. (2013). Sex determination of porcine embryos using a new developed duplex polymerase chain reaction procedure based on the amplification of repetitive sequences. Reprod. Fertil. Dev. 25, 417–425.
| Sex determination of porcine embryos using a new developed duplex polymerase chain reaction procedure based on the amplification of repetitive sequences.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3sXhslart7o%3D&md5=2d33d03b800adce46686ffa8fa501afeCAS | 23445818PubMed |
Toyokawa, K., Harayama, H., and Miyake, M. (2005). Exogenous hyaluronic acid enhances porcine parthenogenetic embryo development in vitro possibly mediated by CD44. Theriogenology 64, 378–392.
| Exogenous hyaluronic acid enhances porcine parthenogenetic embryo development in vitro possibly mediated by CD44.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2MXlt1Ggu7Y%3D&md5=567082658c25caee454e0fbb87fe464eCAS | 15955360PubMed |
Yadav, B. R., King, W. A., and Betteridge, K. J. (1993). Relationships between the completion of first cleavage and chromosomal complement, sex, and developmental rates of bovine embryos generated in vitro. Mol. Reprod. Dev. 36, 434–439.
| Relationships between the completion of first cleavage and chromosomal complement, sex, and developmental rates of bovine embryos generated in vitro.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DyaK2c7jslCmtA%3D%3D&md5=7d0fc7b62194da633b1a20674b3d1ac1CAS | 8305205PubMed |
Yoshioka, K., Suzuki, C., Tanaka, A., Anas, I. M., and Iwamura, S. (2002). Birth of piglets derived from porcine zygotes cultured in a chemically defined medium. Biol. Reprod. 66, 112–119.
| Birth of piglets derived from porcine zygotes cultured in a chemically defined medium.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD38Xht1yksQ%3D%3D&md5=336db91a01f713c0bc52fe07782a3876CAS | 11751272PubMed |