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Updated: Jul 27, 2025
Evaluation of the Spindle Assembly Checkpoint Integrity in Mouse Oocytes
Published on: September 13, 2022
Thiol Protease Inhibitor, E-64-d, Prevents Spindle Formation during Mouse Oocyte Maturation
Naohiro Hashimoto1, Shintaro Iwashita2, Yoko Shoji-Kasai2
1Biological Laboratory, Faculty of Science, Tokyo Institute of Technology, Meguro, Tokyo 152, Japan.
Abstract:
E-64-d, a membrane permeant derivative of E-64, the thiol protease inhibitor, was found to prevent meiotic maturation of mouse oocytes in a dose dependent manner. When immature mouse oocytes were incubated with E-64-d for up to 14 hr, first polar body emission was blocked to 36% at 200 μg/ml and 6% at 400 μg/ml, but germinal vesicle breakdown occurred normally. Cytological analysis revealed that meiotic spindles were not formed, while chromosome condensation occurred. Thus, E-64-d prevents oocytes from progressing to the first meiotic metaphase. When exposed to E-64-d after 8 hr of incubation without E-64-d, one-fourth of oocytes completed the first meiotic division but never progressed to the second metaphase. In three-fourth of the oocytes inhibited to emit the first polar body, spindles disappeared after incubation with E-64-d. The results suggest that E-64-d promotes disassembly of meiotic spindles resulting in inhibition of meiotic maturation. We propose that thiol protease is involved in spindle formation in mouse meiotic maturation.
Insights
E-64-d, a thiol protease inhibitor, blocks mouse oocyte maturation by preventing meiotic spindle formation. This suggests thiol proteases are crucial for spindle assembly during oocyte meiosis.
Area of Science:
- Developmental Biology
- Cell Biology
- Reproductive Biology
Background:
- Meiotic maturation is essential for oocyte developmental competence.
- Thiol proteases play diverse roles in cellular processes.
- The specific role of thiol proteases in oocyte meiotic spindle formation remains unclear.
Purpose of the Study:
- To investigate the effect of E-64-d, a thiol protease inhibitor, on mouse oocyte meiotic maturation.
- To elucidate the role of thiol proteases in the formation of meiotic spindles during oocyte maturation.
Main Methods:
- Incubation of immature mouse oocytes with varying concentrations of E-64-d.
- Assessment of meiotic maturation markers, including germinal vesicle breakdown and first polar body emission.
- Cytological analysis of meiotic spindle formation and chromosome condensation.
Main Results:
- E-64-d inhibited first polar body emission in a dose-dependent manner.
- Germinal vesicle breakdown occurred normally, but meiotic spindle formation was prevented.
- E-64-d exposure post-maturation also inhibited progression to the second metaphase and induced spindle disassembly.
Conclusions:
- E-64-d inhibits mouse oocyte meiotic maturation by promoting meiotic spindle disassembly.
- Thiol proteases are implicated in the process of meiotic spindle formation during mouse oocyte maturation.
- These findings highlight a novel role for thiol proteases in regulating key events of female meiosis.
