Loss of cyclin A and G1-cell cycle arrest are a prerequisite of ceramide-induced toxicity in human arterial endothelial cells
| Autor: | Ioakim Spyridopoulos, Karl R. Karsch, Kerida Shook, Dorothea I. Axel, R. Viebahn, Petra Mayer |
|---|---|
| Rok vydání: | 2001 |
| Předmět: |
Ceramide
Programmed cell death Physiology Cell Cyclin A Cell Culture Techniques Apoptosis Cell Count Biology chemistry.chemical_compound Sphingosine Physiology (medical) Phosphoprotein Phosphatases medicine Humans Enzyme Inhibitors Protein Phosphatase Inhibitor Dose-Response Relationship Drug G1 Phase Arteries Cell cycle Cell biology Endothelial stem cell medicine.anatomical_structure chemistry biology.protein Endothelium Vascular Cardiology and Cardiovascular Medicine Cell Division |
| Zdroj: | Cardiovascular Research. 50:97-107 |
| ISSN: | 0008-6363 |
| Popis: | Background: Ceramide is an important messenger of TNF- and lipid-induced apoptosis. We previously demonstrated the adverse effect of TNF in the process of reendothelialization as well as the dependence of its effect on cell-cycle regulation. The current study was designed to investigate the linkage between ceramide induced toxicity and growth arrest in human endothelial cells. Methods and results: Cultured human arterial endothelial cells (HAEC) served as an in-vitro model to test the cellular effects of C2-ceramide (C2). C2-induced cell death in HAECs occurred time- and dose-dependently. The LD50 in subconfluent cells was three times lower than in confluent cell layers (25 vs. 75 μM). C2 caused up to 70% inhibition of BrdU and [3H]thymidine incorporation at non-toxic concentrations as a result of G1 cell-cycle arrest. Downregulation of cyclin A and p21Cip1/Waf1 protein expression was observed independently of C2-toxicity, while expression of other cell-cycle regulatory genes was not affected. Inhibition of cyclin A protein expression by sequence-specific antisense-oligonucleotides was paralleled by significant growth-inhibition. The protein phosphatase inhibitor okadaic acid induced endothelial cell proliferation, which was completely abrogated by C2. In contrast, aphidicolin-synchronized endothelial cells demonstrated elevated cyclin A levels along with 30% higher BrdU-incorporation and 70% less C2-toxicity. G1-arrested cells, however, showed significantly enhanced C2-toxicity, lack of cyclin A expression and induction of uncleaved caspase-3 (CPP32). Conclusions: Ceramide abrogates endothelial cell proliferation independently of apoptosis or necrosis at low concentrations (≤10 μM) through loss of cyclin A expression with subsequent G1 cell-cycle arrest. Synchronization of HAECs in S-phase with aphidicolin overcomes C2-induced G1-arrest and partially blocks ceramide toxicity. These findings demonstrate the dependence of ceramide toxicity on cell cycle regulation, suggesting a strong bidirectional relationship between cell-cycle control and cell death in vessel biology. |
| Databáze: | OpenAIRE |
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