The Signaling Pathway of Oxysterol Induced Apoptosis in Chinese Hamster Ovary (CHO)-K1 Cells.

Author

Lin Yang, East Tennessee State University

Degree Name

PhD (Doctor of Philosophy)

Program

Biomedical Sciences

Date of Award

8-2002

Committee Chair or Co-Chairs

Michael S. Sinensky

Committee Members

Antonio E. Rusinol, Sankhavaram Panini, Stephen Armstrong, William Stone

Abstract

Apoptosis, a form of genetically programmed cell death, plays a key role in regulation of cellularity of the arterial wall. During atherogenesis, improper apoptosis may cause abnormalities of arterial morphogenesis, wall structural stability, and metabolisms. It has been well established that vascular cells undergo apoptosis after uptake of oxidized low-density lipoprotein (oxLDL). Thus, an analysis of the signaling pathway of apoptotic induction by oxLDL is of value in understanding the development of atherosclerotic plaque. In order to elucidate the signaling pathway of apoptosis induced by oxLDL, we have used Chinese hamster ovary (CHO)-K1 cells treated with a potent oxysterol, 25-hydroxycholesterol (25-OHC). In the present study, we find that oxLDL can induce apoptosis in any cell types if cells present the specific receptors on their surface to take up oxLDL, and that apoptosis-inducing activity is associated with oxysterol components in oxLDL. Oxysterol-induced apoptosis does not involve regulation of sterol regulatory element-binding protein proteolysis pathway. 25-OHC stimulates calcium uptake by CHO-K1 cells within 2 min after addition. Treatment of CHO-K1 cells with the calcium channel blocker nifedipine prevents 25-OHC induction of apoptosis. One possible signal transduction pathway initiated by calcium ion fluxes is the activation of cytosolic phospholipase A₂ (cPLA₂). We demonstrate that activation of cPLA₂ does occur in CHO-K1 treated with 25-OHC. Activation is evidenced by 25-OHC-induced relocalization of cPLA₂ to the nuclear envelope and arachidonic acid (AA) release. Loss of cPLA₂ activity by treatment with a cPLA₂ inhibitor results in an attenuation of AA release as well as of the apoptotic response to 25-OHC in CHO-K1 cells. CPLA2ûmediated liberation of AA leads to the formation of a cyclooxygenase product, probably a prostaglandin, which activates the transcription factor PPARγ and induces apoptosis. We also examined the execution phase of the apoptotic pathway in CHO-K1 cell death induced by 25-OHC. Oxysterol-induced apoptosis in CHO-K1 is accompanied by caspase activation and is preceded by mitochondrial cytochrome C release. Furthermore, treatment with a cPLA₂ inhibitor results in an inhibition of caspase-3 activation in CHO-K1 cells. These data provide strong evidence indicating that 25-OHC induces caspase-3-mediated apoptosis via an activation of calcium-dependent cPLA₂.

Document Type

Dissertation - unrestricted

Recommended Citation

Yang, Lin, "The Signaling Pathway of Oxysterol Induced Apoptosis in Chinese Hamster Ovary (CHO)-K1 Cells." (2002). Electronic Theses and Dissertations. Paper 678. https://dc.etsu.edu/etd/678

Copyright

Copyright by the authors.