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Glucocorticoids regulate plasma membrane potential during rat thymocyte apoptosis in vivo and in vitro
1Molecular Endocrinology Group, Laboratory of Signal Transduction, National Institute of Environmental Health Sciences, National Institutes of Health, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599, USA.
Abstract:
Glucocorticoids induce a series of profound biochemical changes in thymocytes that initiate apoptosis; however, the pathways beyond receptor transactivation that lead to this form of cell death are not fully understood. In this study, we report a novel site of action for glucocorticoids at the site of the plasma membrane. Specifically, we find that glucocorticoids induce the loss of plasma membrane potential both in vivo and in vitro. The glucocorticoid-induced loss of plasma membrane potential in cultured primary isolated rat thymocytes was both dose and time dependent. Other steroid hormones, including progesterone, estrogen, and testosterone, fail to alter the depolarization state of the thymocyte plasma membrane. Interestingly, other nonsteroid stimuli that also activate apoptosis in thymocytes also lead to cellular depolarization. In contrast, HeLa cells, which contain functional glucocorticoid receptors but do not die in response to hormone, do not alter their plasma membrane potential in response to glucocorticoids, indicating a strong association between depolarization and apoptosis. Furthermore, the ability of glucocorticoids to depolarize the plasma membrane of thymocytes required the interaction of glucocorticoids with their cognate receptor, because RU486 failed to depolarize thymocytes and antagonized the effect of glucocorticoids. Finally, experiments using inhibitors of transcription and translation indicated that the loss of plasma membrane potential in thymocytes following glucocorticoid treatment required de novo gene expression. The results of these studies establish that the loss of plasma membrane potential is an early important feature of glucocorticoid-induced apoptosis of thymocytes.
Insights
Glucocorticoids trigger thymocyte apoptosis by disrupting plasma membrane potential, a novel finding requiring gene expression. This depolarization is linked to cell death, unlike other steroid hormones.
Area of Science:
- Cell Biology
- Endocrinology
- Immunology
Background:
- Glucocorticoids induce apoptosis in thymocytes, but the precise molecular pathways are not fully elucidated.
- Receptor transactivation is a known mechanism, but other cellular targets remain to be identified.
Purpose of the Study:
- To investigate novel mechanisms of glucocorticoid-induced apoptosis in thymocytes.
- To identify the role of plasma membrane potential in glucocorticoid action.
Main Methods:
- Primary rat thymocytes were treated with glucocorticoids in vitro and in vivo.
- Plasma membrane potential was measured using established techniques.
- Experiments involved various steroid hormones, non-steroid apoptotic stimuli, and HeLa cells.
- RU486 (a glucocorticoid receptor antagonist) and inhibitors of transcription/translation were utilized.
Main Results:
- Glucocorticoids induce a dose- and time-dependent loss of plasma membrane potential in thymocytes.
- This effect was specific to glucocorticoids, as other steroid hormones did not cause depolarization.
- Apoptotic non-steroid stimuli also induced cellular depolarization.
- HeLa cells, despite having functional glucocorticoid receptors, did not depolarize, linking depolarization to apoptosis.
- Glucocorticoid-induced depolarization required receptor interaction and de novo gene expression.
Conclusions:
- Loss of plasma membrane potential is an early and critical event in glucocorticoid-induced thymocyte apoptosis.
- This finding reveals a novel, non-transcriptional or post-transcriptional site of action for glucocorticoids at the plasma membrane.
- The results highlight the importance of cellular bioenergetics and membrane integrity in programmed cell death.