Abstract
Importance of germinal vesicle breakdown (GVBD) in the development chronology of oocyte development can be gauged from the fact that it has been the target of several studies attempting to improve oocyte competence. Manipulation of this event rests on its precise and explicit documentation. Pertinently, we decided to compare three different methods for their efficacy and precision in assessing GVBD. Immature buffalo oocytes were subjected to GVBD inhibition using Roscovitine (Ros) and Cilostamide (Cil) under different concentrations of each as well as their combination, along with control. After 24 h of inhibition, chromatin assessment was done using aceto-orcein staining, hoechst staining and anti-lamin staining. Hoechst staining and anti-lamin staining proved to be easier and less time consuming when compared to aceto-orcein, which was cumbersome and lengthy. Aceto-orcein and hoechst staining were equally ambiguous, while antilamin staining was most precise, accurate and clear in characterizing an oocyte as germinal vesicle (GV) or GVBD. We conclude by stating that anti-lamin staining is an easy and specific technique for assessing GVBD in buffalo oocytes.
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Terasaki, M., Redistribution of cytoplasmic components during germinal vesicle breakdown in starfish oocytes, J. Cell Sci., 1994, vol. 107, no. 7, pp. 1797–1805.
Metwally, M., Cutting, R., Tipton, A., Skull, J., Ledger, W.L., and Li, T.C., Effect of increased body mass index on oocyte and embryo quality in IVF patients, Reprod. Biomed. Online, 2007, vol. 15, no. 5, pp. 532–538.
Leibfried, L. and First, N.L., Characterization of bovine follicular oocytes and their ability to mature in vitro, J. Anim. Sci., 1979, vol. 48, no. 1, pp. 76–86.
Lonergan, P., Dinnyes, A., Fair, T., Yang, X., and Boland, M., Bovine oocyte and embryo development following meiotic inhibition with butyrolactone I, Mol. Reprod. Dev., 2000, vol. 57, no. 2, pp. 204–209.
Hashimoto, S., Minami, N., Takakura, R., and Imai, H., Bovine immature oocytes acquire developmental competence during meiotic arrest in vitro, Biol. Reprod., 2002, vol. 66, no. 6, pp. 1696–1701.
Prentice-Biensch, J.R., Singh, J., Alfoteisy, B., and Anzar, M., A simple and high-throughput method to assess maturation status of bovine oocytes: Comparison of anti-lamin A/C-DAPI with an aceto-orcein staining technique, Theriogenology, 2012, vol. 78, no. 7, pp. 1633–1638.
Bézard, J., Bøgh, I.B., Duchamp, G., Hyttel, P., and Greve, T., Comparative evaluation of nuclear morphology of equine oocytes aspirated in vivo and stained with Hoechst and orcein, Cells Tissues Organs, 2002, vol. 170, no. 4, pp. 228–236.
Bézard, J., Mekarska, A., Goudet, G., Duchamp, G., and Palmer, E., Timing of in vivo maturation of equine preovulatory oocytes and competence for in vitro maturation of immature oocytes collected simultaneously, Equine Vet. J., 1997, vol. 29, no. S25, pp. 33–37.
Goudet, G., Leclercq, L., Bezard, J., Duchamp, G., Guillaume, D., and Palmer, E., Chorionic gonadotropin secretion is associated with an inhibition of follicular growth and an improvement in oocyte competence for in vitro maturation in the mare, Biol. Reprod., 1998, vol. 58, no. 3, pp. 760–768.
Gerace, L. and Burke, B., Functional organization of the nuclear envelope, Annu. Rev. Cell Biol., 1988, vol. 4, pp. 335–374.
Hall, V.J., Cooney, M.A., Shanahan, P., Tecirlioglu, R.T., Ruddock, N.T., and French, A.J., Nuclear lamin antigen and messenger RNA expression in bovine in vitro produced and nuclear transfer embryos, Mol. Reprod. Dev., 2005, vol. 72, no. 4, pp. 471–482.
Lénárt, P., Rabut, G., Daigle, N., Hand, A.R., Terasaki, M., and Ellenberg, J., Nuclear envelope breakdown in starfish oocytes proceeds by partial NPC disassembly followed by a rapidly spreading fenestration of nuclear membranes, J. Cell Biol., 2003, vol. 160, no. 7, pp. 1055–1068.
Ogushi, S., Fulka, J., Jr., and Miyano, T., Germinal vesicle materials are requisite for male pronucleus formation but not for change in the activities of CDK1 and MAP kinase during maturation and fertilization of pig oocytes, Dev. Biol., 2005, vol. 286, no. 1, pp. 287–298.
Arnault, E., Doussau, M., Pesty, A., Lefevre, B., and Courtot, A.M., Lamin A/C, caspase-6, and chromatin configuration during meiosis resumption in the mouse oocyte, Reprod. Sci., 2010, vol. 17, no. 2, pp. 102–115.
Kalous, J., Solc, P., Baran, V., Kubelka, M., Schultz, R.M., and Motlik, J., PKB/AKT is involved in resumption of meiosis in mouse oocytes, Biol. Cell., 2006, vol. 98, no. 2, pp. 111–123.
Keefer, C.L., Stice, S.L., and Matihews, D.L., Bovine inner cell mass cells as donor nuclei in the production of nuclear transfer embryos and calves, Biol. Reprod., 1994, vol. 50, no. 4, pp. 935–939.
Datta, T.K. and Goswami, S.L., Feasibility of harvesting oocytes from buffalo (Bubalus bubalis) ovaries by different methods, Buffalo J., 1998, vol. 14, pp. 277–284.
Nagai, T., Ebihara, M., Onishi, A., and Kubo, M., Germinal vesicle stages in pig follicular oocytes collected by different methods, J. Reprod. Dev., 1997, vol. 43, no. 4, pp. 339–343.
Motlik, I., Koefoed-Johnson, H.H., and Fulka, J., Breakdown of the germinal vesicle in bovine oocytes cultivated in vitro, J. Exp. Zool., 1978, vol. 205, no. 2, pp. 377–384.
Sun, X.S., Liu, Y., Yue, K.Z., Ma, S.F., and Tan, J.H., Changes in germinal vesicle (GV) chromatin configurations during growth and maturation of porcine oocytes, Mol. Reprod. Dev., 2004, vol. 69, no. 2, pp. 228–234.
Hinrichs, K., Choi, Y.H., Love, L.B., Varner, D.D., Love, C.C., and Walckenaer, B.E., Chromatin configuration within the germinal vesicle of horse oocytes: changes post mortem and relationship to meiotic and developmental competence, Biol. Reprod., 2005, vol. 72, no. 5, pp. 1142–1150.
Viuff, D., Madison, V., Hyttel, P., Avery, B., and Greve, T., Fluorescent intravital staining of bovine oocytes and zygotes, Theriogenology, 1991, vol. 35, no. 1, pp. 291–291.
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Kumar, S., Dholpuria, S., Chaubey, G.K. et al. Assessment of nuclear membrane dynamics using anti-lamin staining offers a clear cut evidence of germinal vesicle breakdown in buffalo oocytes. Cytol. Genet. 52, 80–85 (2018). https://doi.org/10.3103/S0095452718010061
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DOI: https://doi.org/10.3103/S0095452718010061