Effect of Thymidine on $In$ $Vitro$ Maturation of Immature Porcine Follicular Oocytes

돼지 미성숙 난포란의 체외성숙에 미치는 Thymidine 처리 효과

  • Received : 2012.03.20
  • Accepted : 2012.03.26
  • Published : 2012.03.31

Abstract

The objective of this study was to examine the effect of thymidine treatment during $in$ $vitro$ maturation (IVM) of porcine follicular oocytes on blastocyst development. Porcine oocytes were treated with thymidine (10 mM, 20 mM and 30 mM) for 2 or 6 hr in the preiods of IVM I and/or II. The survival rates of the blastocysts in the 6 hr treatment groups of 10 mM and 20 mM during IVM I period were significantly higher than those of control group ($p$<0.05). However, the survival rate of the blastocysts in the 2 hr treatment group of 20 mM during IVM II period was significantly higher than control group ($p$<0.05). Furthermore, the survival rate of the blastocysts in the 6 hr treatment group of 30 mM during IVM II period was significantly lower than control group ($p$<0.05). Consistent with the previous result, blastocyst development of both IVM I and II treatment group was also showed as similar pattern. Total and apoptotic cell numbers of blastocysts derived from thymidine treated porcine oocytes were examined by using Tunel assay. The results showed that there was no significant differences in total cell number of blastocysts between thymidine treated and untreated groups. However, apoptosis-positive cells in the thymidine treated group (6 hr IVM I) were significantly lower than those of other groups ($p$<0.05). Taken together, these results indicate that high quality oocytes were selected by DNA synthesis mechanism according to high concentration thymidine treatment during porcine oocyte maturation. Therefore, we concluded that presumptive selected oocytes by thymidine treatment during maturation periods improved the further embryo development and embryonic quality of IVF embryos by decreasing the incidence of apoptosis in preimplantation porcine embryos.

Keywords

References

  1. Abeydeera LR, Day BN (1997): In vitro penetration of pig oocytes in a modified Tris-buffered medium: effect of BSA, caffeine and calcium. Theriogenology 48(4):537-544. https://doi.org/10.1016/S0093-691X(97)00270-7
  2. Alm H, Torner H, Lohrke B, Viergutz T, Ghoneim IM, Kanitz W (2005): Bovine blastocyst development rate in vitro is influenced by selection of oocytes by brillant cresyl blue staining before IVM as indicator for glucose-6-phosphate dehydrogenase activity. Theriogenology 63:2194-2205. https://doi.org/10.1016/j.theriogenology.2004.09.050
  3. Blumenreich MS, Woodcock TM, Andreeff M, Hiddemann W, Chou TC, Vale K (1984): Effect of very high-dose thymidine infusions on leukemia and lymphoma patients. Cancer Res 44(5):2203-2207.
  4. Byun TH, Lee SH (1992): Morphological and cellular criteria ovaries, follicles and oocytes for in vitro maturation in the pig. Kor J Emb Tran 7:97-110.
  5. De LF, Van MP, Van BT, Kruip T (1992): Structural aspects of bovine oocytes maturation in vitro. Mol Reprod Dev 31:208-214. https://doi.org/10.1002/mrd.1080310308
  6. Downs SM (1997): Involvement of purine nucleotide synthetic pathways in gonadotropin-induced meiotic maturation in mouse cumulus cell-enclosed oocytes. Mol Reprod Dev 46(2):155-167. https://doi.org/10.1002/(SICI)1098-2795(199702)46:2<155::AID-MRD6>3.0.CO;2-P
  7. Ebeling S, Schuon C, Meinecke B (2007): Mitogenactivated protein kinase phosphorylation patterns in pig oocytes and cumulus cells during gonadotrophin- induced resumption of meiosis in vitro. Zygote 15(2):139-147. https://doi.org/10.1017/S0967199406004011
  8. Ebeling S, Topfer D, Meinecke B (2011): Steroidogenesis and the influence of MAPK activity during in vitro maturation of porcine cumulus oocyte complexes. Reprod Domest Anim 46(3):513-519. https://doi.org/10.1111/j.1439-0531.2010.01699.x
  9. Ebner T, Moser M, Sommergruber M, Tews G (2003): Selection based on morphological assessment of oocytes and embryos at different stages of preimplantation development. Hum Repord Update. 9:251-262. https://doi.org/10.1093/humupd/dmg021
  10. Evans R, Booth CG (1976): Inhibition of 125IUdR incorporation by supernatants from macrophage and lymphocyte cultures: a cautionary note. Cell Immunol 26(1):120-126. https://doi.org/10.1016/0008-8749(76)90354-3
  11. Fan HY, Tong C, Lian L, Li SW, Gao WX, Cheng Y (2003): Characterization of ribosomal S6 protein kinase p90rsk during meiotic maturation and fertilization in pig oocytes: mitogen-activated protein kinase- associated activation and localization. Biol Reprod 68(3):968-977. https://doi.org/10.1095/biolreprod.102.008839
  12. Fan HY, Sun QY (2004): Involvement of mitogenactivated protein kinase cascade during oocyte maturation and fertilization in mammals. Biol Reprod 70(3):535-547. https://doi.org/10.1095/biolreprod.103.022830
  13. Fulka J, Motlik J, Jilek, F (1986): Effect of cycloheximide on nuclear maturation of pig and mouse oocytes. J Reprod Fertil 77(1):281-285. https://doi.org/10.1530/jrf.0.0770281
  14. Funahashi H, Day BN (1993): Effects of the duration of exposure to hormone supplements on cytoplasmic maturation of pig oocytes in vitro. J Reprod Fertil 98:179-185. https://doi.org/10.1530/jrf.0.0980179
  15. Gordon I (2003): Recovering the bovine oocytes In : Laboratory Production of Cattle Embryos (Biotechnology in Agriculture No. 27). 2nd Ed. Cambrige UK P:79-111.
  16. Han JC, Park KS, Park HD (2000): Effects of hypoxanthin, aminopterine and thymidine(HAT) Added to media at day 5 on hatching of blastocyst in korean native cattle ESHRE P:158.
  17. Hulinska P, Martecikova S, Jeseta M, Machatkova M (2011): Efficiency of in vitro fertilization is influenced by the meiotic competence of porcine oocytes and time of their maturation. Anim Reprod Sci 124:112-117. https://doi.org/10.1016/j.anireprosci.2011.02.004
  18. Ishizaki C, Watanabe H, Bhuiyan MM, Fukui Y (2009): Developmental competence of porcine oocytes selected by brilliant cresyl blue and matured individually in a chemically defined culture medium. Theriogenology 72(1):72-80. https://doi.org/10.1016/j.theriogenology.2009.02.015
  19. Khurana NK, Niemann H (2000): Energy metabolism in preimplantation bovine embryos derived in vitro or in vivo. Biol Reprod 62:847-856. https://doi.org/10.1095/biolreprod62.4.847
  20. Kidson A, Rubio-Pomar FJ, Van Knegsel A, Van Tol HT, Hazeleger W, Ducro-Steverink DW, Colenbrander B, Dieleman Sj, Bevers MM (2004): Qaulity of porcine blastocysts produced in vitro in the presence or absence of GH. Reproduction 127(2):165-177. https://doi.org/10.1530/rep.1.00086
  21. Kitagawa Y, Suzuki K, Yoneda A, Watanabe T (2004): Effects of oxygen concentration and antioxidants on the in vitro developmental ability, production of reactive oxygen species (ROS), and DNA fragmentation in porcine embryos. Theriogenology 62(7):1186-1197. https://doi.org/10.1016/j.theriogenology.2004.01.011
  22. Kwong WY, Adamiak SJ, Gwynn A, Singh R, Sinclair KD (2010): Endogenous folates and single-carbon metabolism in the ovarian follicle, oocyte and pre-implantation embryo. Reproduction 139(4):705-715. https://doi.org/10.1530/REP-09-0517
  23. Lee SL, Kumar BM, Kim JG, Ock SA, Jeon BG, Balasubramanian S, Choe SY, Rho GJ (2007): Cellular composition and viability of cloned bovine embryos using exogene-transfected somatic cells. Repord Domest Anim 42(1):44-52. https://doi.org/10.1111/j.1439-0531.2006.00723.x
  24. Liang CG, Huo LJ, Zhong ZS, Chen DY, Schatten H, Sun QY (2005): Cyclic adenosine 3',5'-monophosphate- dependent activation of mitogen-activated protein kinase in cumulus cells is essential for germinal vesicle breakdown of porcine cumulus-enclosed oocytes. Endocrinology 146(10):4437-4444. https://doi.org/10.1210/en.2005-0309
  25. Matwee C, Betts DH, King WA (2000): Apoptosis in the early bovine embryo. Zygote 8(1):57-68. https://doi.org/10.1017/S0967199400000836
  26. Mori T, Amano T, Shimizu H (2000): Roles of gap junctional communication of cumulus cells in cytoplasmic maturation of porcine oocytes cultured in vitro. Biol Reprod 62(4):913-919. https://doi.org/10.1095/biolreprod62.4.913
  27. Niwa K (1993): Effectiveness of in vitro maturation and in vitro fertilization techniques in pigs. J Reprod Fertil Suppl 48:49-59.
  28. Ock SA, Lee SL, Kim JG, Kumar BM, Balasubramanian S, Choe SY (2007): Development and quality of porcine embryos in different culture system and embryo-producing methods. Zygote 15(1):1-8. https://doi.org/10.1017/S0967199406003911
  29. Opitz HG, Niethammer D, Jackson RC, Lemke H, Huget R, Flad HD (1975): Biochemical characterization of a factor released by macrophages. Cell Immunol 18(1):70-75. https://doi.org/10.1016/0008-8749(75)90037-4
  30. Petters RM, Wells, KD (1993): Culture of pig embryos. J Reprod Fertil Suppl 48:61-73.
  31. Prather RS, Boice ML, Gibson J, Hoffman KE, Parry TW (1995): In vitro development of embryos from sinclair miniature pigs: A preliminary report. Theriogenology 43:1001-1007. https://doi.org/10.1016/0093-691X(95)00064-F
  32. Shimada M, Maeda T, Terada T (2001): Dynamic changes of connexin-43, gap junctional protein, in outer layers of cumulus cells are regulated by PKC and PI 3-kinase during meiotic resumption in porcine oocytes. Biol Reprod 64(4):1255-1263. https://doi.org/10.1095/biolreprod64.4.1255
  33. Snow MHL (1973): Abnormal development of preimplantation mouse embryos grown in vitro with thymidine. J Embryo Exp Morphol. 29:601-615.
  34. Sun QY, Nagai T (2003): Molecular mechanisms underlying pig oocyte maturation and fertilization. J Reprod Dev 49(5):347-359. https://doi.org/10.1262/jrd.49.347
  35. Suzuki H, Jeong BS, Yang X (2000): Dynamic changes of cumulus-oocyte cell communication during in vitro maturation of porcine oocytes. Biol Reprod 63(3):723-729. https://doi.org/10.1095/biolreprod63.3.723
  36. Toyama S, Ando T (1987): 單クローン抗體實驗マニュアル. pp 64-65.
  37. Yoshioka K, Suzuki C, Tanaka A, Anas IM, Iwamura S (2002): Birth of piglets derived from porcine zygotes cultured in a chemically defined medium. Biol Reprod 66(1):112-119. https://doi.org/10.1095/biolreprod66.1.112