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Human Egg Maturity Assessment and Its Clinical Application
Published on: August 19, 2019
Assessment of meiotic spindle configuration and post-warming bovine oocyte viability using polarized light microscopy
J N Caamaño1, C Díez, B Trigal
1Genética y Reproducción Animal, Centro de Biotecnología Animal, Servicio Regional de Investigación y Desarrollo Agroalimentario, Gijón, Principado de Asturias, Spain. jncaamano@serida.org
Abstract:
The objectives of this study were to assess the efficiency of polarized light microscopy (PLM) in detecting microtubule-polymerized protein in in vitro-matured bovine oocytes; to examine its effects on oocyte developmental competence; and to assess the meiotic spindle of in vitro-matured oocytes after vitrification/warming and further assessment of oocyte developmental competence. In the first experiment, the presence of microtubule-polymerized protein (MPP) was confirmed as a positive PLM signal detected in 99.1% of analysed oocytes (n = 115), which strongly correlated (r = 1; p < 0.0001) with the presence of MPP as confirmed by immunostaining. In the second experiment, oocytes (n = 651) were exposed or not (controls) to PLM for 10 min and then fertilized and cultured in vitro. Oocytes exposed to PLM did not significantly differ from controls with regard to cleavage, total blastocyst and expanded blastocyst rates and cell numbers. In the third experiment, meiotic spindles were detected in 145 of 182 oocytes (79.6%) following vitrification and warming. Interestingly, after parthenogenetic activation and in vitro culture, oocytes that displayed a positive PLM signal PLM(+) differed significantly from PLM(-) in cleavage and Day 8 blastocyst rates. These results suggest that polarized light microscopy is an efficient system to detect microtubule-polymerized protein in in vitro-matured bovine oocytes and does not exert detrimental effects on bovine oocyte developmental competence. Moreover, PLM could be used as a tool to assess post-warming viability in vitrified bovine oocytes.
Insights
Polarized light microscopy (PLM) effectively detects microtubule-polymerized protein in bovine oocytes without harming their development. This technique can also assess the viability of vitrified oocytes after warming.
Area of Science:
- Reproductive Biology
- Cell Biology
- Microscopy
Background:
- Assessing oocyte quality is crucial for successful in vitro fertilization and embryo development.
- Microtubule-polymerized protein (MPP) organization is vital for meiotic spindle function and oocyte competence.
- Polarized light microscopy (PLM) offers a non-invasive method for visualizing microtubule structures.
Purpose of the Study:
- To evaluate the efficacy of PLM in detecting MPP in in vitro-matured bovine oocytes.
- To determine the impact of PLM exposure on oocyte developmental competence.
- To assess the utility of PLM in evaluating meiotic spindle integrity after cryopreservation.
Main Methods:
- In vitro maturation of bovine oocytes.
- Detection of MPP using PLM and immunostaining for validation.
- Exposure of oocytes to PLM (10 min) or control conditions.
- Fertilization and in vitro culture of oocytes.
- Vitrification and warming of oocytes, followed by assessment of meiotic spindles.
- Parthenogenetic activation and in vitro culture of vitrified/warmed oocytes.
Main Results:
- PLM detected MPP in 99.1% of oocytes, correlating perfectly with immunostaining.
- PLM exposure (10 min) did not significantly affect oocyte cleavage, blastocyst rates, or cell numbers.
- Post-warming, PLM signal presence correlated with improved cleavage and blastocyst development rates after parthenogenetic activation.
- PLM identified meiotic spindles in 79.6% of vitrified-warmed oocytes.
Conclusions:
- PLM is an efficient and non-detrimental method for detecting MPP in bovine oocytes.
- PLM does not impair oocyte developmental competence.
- PLM shows potential as a tool for assessing the viability of vitrified-warmed bovine oocytes.
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