Korean J Obstet Gynecol.
2001 Nov;44(11):2091-2097.
Apoptosis of human luteinized granulosa cell via change of mitochondria
- Affiliations
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- 1Department of Obstetrics and Gynecology, Ajou University School of Medicine, Suwon, Korea.
- 2Department of Molecular Science and Technology, Graduate School, Ajou University, Suwon, Korea.
- 3Medical Research Center, Ajou University School of Medicine, Suwon, Korea.
- 4Department of Biological Science, College of Natural Science, Ajou University, Suwon, Korea.
Abstract
OBJECTIVE
To determine the effects of extracellular ATP on mitochondrial function and apoptosis during human luteinized granulosa cell cultures.
METHODS
The addition of various concentrations of ATP (0, 0.1, 0.25, 0.5, 0.75 mM) to luteinized granulosa cells obtained during In vitro fertilization ovum pickup procedures. After culture for 24 hours, purinoceptor activity and functional changes in mitochondria were measured by patch clamp, flow cytometry, and confocal microscopy.
RESULTS
Measurement by patch clamp of the granulosa cell membrane potential after ATP addition to cultured granulosa cells showed that both the inward and outward currents were expressed. After treatment of the granulosa cells with JC-1, measurement of the mitochondrial activity by confocal microscopy showed that the with increasing concentrations of ATP the relative ratio of undamaged mitochondria (red/green ratio) tended to decrease (P=0.027). After double staining of the cultured granulosa cells with Annexin V and Propidium Iodide, quantitative flow cytometry analysis showed that apoptosis increased with increasing concentrations of ATP (7.88%, 8.44%, 11.40%, 13.52%, 18.57%).
CONCLUSION
The results of this study shows that apoptosis of granulosa cells increases with increasing extracellular ATP concentrations in cultured human luteinized granulosa cells. This is observed to be a consequence of cell membrane purinoceptor activity and functional changes in the mitochondria. It is therefore thought that remodelling processes of the ovarian tissue is regulated by neuroendocrine factors of the extracellular ATP.
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