Homocysteine-thiolactone induces caspase-independent vascular endothelial cell death with apoptotic features.
OBJECTIVE
Cell death is generally classified into two large categories: apoptosis, which represents active, physiological programmed cell death, and necrosis, which represents passive cell death without underlying regulatory mechanisms. Apoptosis plays an important role in tissue homeostasis and its role in endothelium integrity can be influenced by the functional status of endothelial cells. Homocysteine, a sulfated amino-acid product of methionine demethylation, is an independent risk factor for vascular disease (arterial and venous thombosis). Our goal was to investigate the thiol-derivatives effect on the endothelial cell apoptosis.


METHODS
Three parameters were measured: mitochondrial membrane potential using DiOC6(3) as the probe, DEVDase activation, and phosphatidylserine exposure on the cell surface with fluorosceinated annexin V labeling which allows apoptosis to be distinguished from necrosis.


RESULTS
Homocysteine-thiolactone induced endothelial cell apoptosis in a concentration-dependent manner (range: 50-200 microM), independently of the caspase pathway. Only homocysteine-thiolactone, among the thiol derivatives tested, induced apoptosis. Apoptosis was not influenced by the serum concentration in culture medium, suggesting that the observed apoptotic process could occur in vivo. None of the inhibitors used (e.g., leupeptin, fumosinin B1, catalase, or z-VAD-fmk) was able to prevent homocysteine-induced apoptosis of vascular endothelial cells.


CONCLUSIONS
The apoptosis of vascular endothelial cells induced by high concentration of homocysteine-thiolactone might be one step atherosclerotic cardiovascular disease, and contribute to its complication.
Mercié
P
P
Laboratoire d'Hématologie, Université Victor Ségalen Bordeaux 2, France. patrick.mercie@chu-bordeaux.fr
Garnier
O
O
Lascoste
L
L
Renard
M
M
Closse
C
C
Durrieu
F
F
Marit
G
G
Boisseau
R M
RM
Belloc
F
F
D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D002232:Carbocyanines; D020169:Caspases / Q000502:physiology*; D016923:Cell Death / Q000187:drug effects / Q000502:physiology; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D003470:Culture Media; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000166:cytology* / Q000187:drug effects* / Q000201:enzymology; D005456:Fluorescent Dyes; D006710:Homocysteine / Q000031:analogs & derivatives* / Q000494:pharmacology*; D006801:Humans; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008928:Mitochondria / Q000187:drug effects / Q000502:physiology; D010447:Peptide Hydrolases / Q000378:metabolism; D010718:Phosphatidylserines / Q000378:metabolism; D011506:Proteins / Q000378:metabolism / Q000494:pharmacology
D002232:Carbocyanines; D003470:Culture Media; D005456:Fluorescent Dyes; D010718:Phosphatidylserines; D011506:Proteins; D006710:Homocysteine; C007392:3,3'-dihexyl-2,2'-oxacarbocyanine; C007957:homocysteine thiolactone; D010447:Peptide Hydrolases; D020169:Caspases; C108711:DEVDase