Selection of developmentally competent immature equine oocytes with brilliant cresyl blue stain prior to in vitro maturation with equine growth hormone.
Abstract: Immature oocytes synthesize a variety of proteins that include the enzyme glucose-6-phosphate dehydrogenase (G6PDH). Brilliant cresyl blue (BCB) is a vital blue dye that assesses intracellular activity of G6PDH, an indirect measure of oocyte maturation. The objective was to evaluate the BCB test as a criterion to assess developmental competence of equine oocytes and to determine if equine growth hormone (eGH) enhanced in vitro maturation (IVM) of equine oocyte. Cumulus-oocytes complexes (COCs) were recovered by aspirating follicles 0.05). Maturation was not affected significantly by BCB classification, but the maturation rate was higher for oocytes that had been exposed to exogenous eGH versus controls (16/28, 57.1% versus 8/26, 30.8%, P < 0.05). In the present study, the BCB test was not useful for predicting competent equine oocytes prior to IVM. However, eGH enhanced equine oocyte maturation in vitro.
Publication Date: 2013-02-01 PubMed ID: 23369728DOI: 10.1017/S096719941200072XGoogle Scholar: Lookup
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Summary
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This research studied the use of brilliant cresyl blue (BCB) to identify immature horse oocytes capable of maturation and development, and checked if equine growth hormone (eGH) aided in the in vitro maturation of these oocytes. The results showed that while BCB was not effective as a predictive tool, the use of eGH did improve in vitro maturation rates for equine oocytes.
Introduction to the Study and Methodology
- Researcher’s main purpose was to test BCB as a means to determine the developmental competence of horse oocytes (egg cells), and to examine the effect of equine Growth Hormone (eGH) on the in vitro maturation (outside the body) of these oocytes.
- The study retrieved Cumulus-oocytes complexes (COCs), structures that are made up of an oocyte surrounded by cumulus cells, by aspirating follicles of a certain size (less than 30 mm in diameter) from ovaries obtained from an abattoir. The COCs were then assessed based on their morphology.
- The COCs were then subjected to BCB, a dye that indicates glucose-6-phosphate dehydrogenase (G6PDH) activity. G6PDH is an enzyme whose intracellular activity can give an indirect measure of oocyte maturation. Based upon the color change, the COCs were classified as BCB positive (BCB+) or BCB negative (BCB-).
Dividing the Study Groups
- COCs were arranged into the following groups: (a) IVM medium (a culture medium for in vitro maturation); (b) eGH group (those treated with equine growth hormone); (c) BCB-/IVM (those that were BCB negative in the maturation medium); (d) BCB+/IVM (those that were BCB positive in the maturation medium); (e) BCB-/eGH (those that were BCB negative and treated with equine growth hormone); and (f) BCB+/eGH (those that were BCB positive and treated with equine growth hormone).
- Following this categorization, the COCs were cultured in vitro for 30 hours under specific environmental conditions. After the incubation period, the nuclear maturation (an important step in cell division and growth) was evaluated.
Research Findings
- After staining with BCB, 46.2% of the 39 COCs selected for the study were classified as BCB+, and 53.8% as BCB-. However, the researchers found that BCB classification did not significantly affect maturation rates.
- The research did reveal that the group exposed to exogenous eGH had higher oocyte maturation rates (57.1% versus 30.8% in controls).
- Though the BCB test was not effective in predicting which oocytes were competent prior to IVM, the study found that eGH did enhance in vitro maturation of equine oocytes.
Cite This Article
APA
Pereira GR, Lorenzo PL, Carneiro GF, Bilodeau-Goeseels S, Kastelic JP, Esteller-Vico A, Lopez-Bejar M, Liu IK.
(2013).
Selection of developmentally competent immature equine oocytes with brilliant cresyl blue stain prior to in vitro maturation with equine growth hormone.
Zygote, 22(4), 500-504.
https://doi.org/10.1017/S096719941200072X Publication
Researcher Affiliations
- Federal University of Pelotas - UFPEL,School of Veterinary Medicine,Campus Capão do Leão,s/n° - Mailbox 354,Zip 96010-900,Pelotas,RS,Brasil.
- Animal Physiology Department,Veterinary School,Universidad Complutense de Madrid,Madrid 28040,Spain.
- Animal Reproduction Department,Federal Rural University of Pernambuco,Garanhuns,PE 52296-901,Brazil.
- Lethbridge Research Centre,P.O. Box 3000,5403 1Avenue S.,Lethbridge,AB T1J4B1,Canada.
- Lethbridge Research Centre,P.O. Box 3000,5403 1Avenue S.,Lethbridge,AB T1J4B1,Canada.
- Department of Population Health and Reproduction,School of Veterinary Medicine,University of California,Davis,CA 95616,USA.
- Departament de Sanitat i d'Anatomia Animals,Autonomous University of Barcelona,Edifici V 08193 Bellaterra,Barcelona,Spain.
- Department of Population Health and Reproduction,School of Veterinary Medicine,University of California,Davis,CA 95616,USA.
MeSH Terms
- Animals
- Cells, Cultured
- Female
- Growth Hormone / pharmacology
- Horses
- In Vitro Oocyte Maturation Techniques / methods
- Oocytes / cytology
- Oocytes / drug effects
- Oocytes / physiology
- Oogenesis
- Oxazines / analysis
- Oxazines / metabolism
- Staining and Labeling / methods
Citations
This article has been cited 7 times.- Bittner-Schwerda L, Herrera C, Wyck S, Malama E, Wrenzycki C, Bollwein H. Brilliant Cresyl Blue Negative Oocytes Show a Reduced Competence for Embryo Development after In Vitro Fertilisation with Sperm Exposed to Oxidative Stress.. Animals (Basel) 2023 Aug 14;13(16).
- Pan P, Huang X. The Clinical Application of Growth Hormone and Its Biological and Molecular Mechanisms in Assisted Reproduction.. Int J Mol Sci 2022 Sep 15;23(18).
- Lin Y, Xie B, Li X, Li R, Ma C, Zhu J, Qiao J. Supplementation of the In Vitro Maturation Culture Medium of Mouse Oocytes with Growth Hormone Improves Pregnancy Outcomes.. Reprod Sci 2021 Sep;28(9):2540-2549.
- Dutta R, Mandal S, Lin HA, Raz T, Kind A, Schnieke A, Razansky D. Brilliant cresyl blue enhanced optoacoustic imaging enables non-destructive imaging of mammalian ovarian follicles for artificial reproduction.. J R Soc Interface 2020 Nov;17(172):20200776.
- Li Y, Liu H, Yu Q, Liu H, Huang T, Zhao S, Ma J, Zhao H. Growth Hormone Promotes in vitro Maturation of Human Oocytes.. Front Endocrinol (Lausanne) 2019;10:485.
- Fu B, Ren L, Liu D, Ma JZ, An TZ, Yang XQ, Ma H, Zhang DJ, Guo ZH, Guo YY, Zhu M, Bai J. Subcellular Characterization of Porcine Oocytes with Different Glucose-6-phosphate Dehydrogenase Activities.. Asian-Australas J Anim Sci 2015 Dec;28(12):1703-12.
- Ashry M, Lee K, Mondal M, Datta TK, Folger JK, Rajput SK, Zhang K, Hemeida NA, Smith GW. Expression of TGFβ superfamily components and other markers of oocyte quality in oocytes selected by brilliant cresyl blue staining: relevance to early embryonic development.. Mol Reprod Dev 2015 Mar;82(3):251-64.
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