Prevention of cycloheximide-induced apoptosis in hepatocytes by adenosine and by caspase inhibitors

• Published in: Biochemical pharmacology Vol. 58; no. 12; pp. 1891 - 1898
• Main Authors: Blom, W.Marty, de Bont, Hans J.G.M, Meijerman, Irma, Mulder, Gerard J, Nagelkerke, J.Fred
• Format: Journal Article
• Language: English
• Published: New York, NY Elsevier Inc 15-12-1999; Elsevier Science
• Subjects: adenosine; Adenosine - pharmacology; Ageing, cell death; Animals; apoptosis; Apoptosis - drug effects; Biological and medical sciences; Caspase Inhibitors; caspases; Cell physiology; cycloheximide; Cycloheximide - antagonists & inhibitors; Cycloheximide - pharmacology; Drug Interactions; Enzyme Inhibitors - pharmacology; Fundamental and applied biological sciences. Psychology; In Vitro Techniques; liver; Liver - cytology; Liver - drug effects; Male; Membrane Potentials - drug effects; mitochondria; Mitochondria, Liver - drug effects; Mitochondria, Liver - physiology; Molecular and cellular biology; Protective Agents - pharmacology; Protein Synthesis Inhibitors - pharmacology; Rats; Rats, Wistar; cycloheximide; adenosine; apoptosis; liver; caspases; mitochondria; Adenosine; Rat; Rodentia; Enzyme inhibitor; In vitro; Biological activity; Cycloheximide; Vertebrata; Mitochondria; Mammalia; Hepatocyte; Animal; Mechanism of action; Apoptosis
• ISSN: 0006-2952; 1873-2968
• DOI: 10.1016/S0006-2952(99)00268-3

The mechanism by which cycloheximide induces apoptosis in isolated rat hepatocytes was studied. Cycloheximide (1–300 μM) induced apoptosis within 3–4 hr in the hepatocytes. Specific apoptotic characteristics such as blebbing, phosphatidyl serine (PS) exposure, chromatin condensation, and nuclear fragmentation were induced. Cycloheximide (CHX) dose dependently activated the caspase-3-like proteases, but not the caspase-1-like proteases. Pretreatment of the hepatocytes with 100 μM of the caspase inhibitors z-Val-Ala-DL-Asp-fluoromethylketone or Ac-Asp-Glu-Val-Asp-aldehyde completely abrogated the caspase activation and the apoptosis. Addition of adenosine (100 μM) reduced phosphatidyl serine exposure and other morphological characteristics of apoptosis by 50%; however, it did not prevent the activation of the caspases, suggesting that adenosine inhibited downstream of caspase activation. The adenosine receptor antagonist 8-[4-[[[(2-aminoethyl)amino]-carbonyl]methyl]oxy]phenyl]-1,3-dipropylxanthine abolished the capacity of adenosine to prevent apoptosis, indicating that prevention was receptor-mediated. During apoptosis, the mitochondrial membrane potential in apoptotic cells (cells with PS exposition) was decreased to 50–60% of the control value; in the population viable cells, however, the mitochondrial membrane potential remained stable. Prevention of apoptosis by the caspase inhibitor z-Val-Ala-DL-Asp-fluoromethylketone or adenosine prevented the decrease in mitochondrial membrane potential. In conclusion, CHX rapidly induces apoptosis in isolated rat hepatocytes, which is inhibited by adenosine at a relatively late step.