R(+)-methanandamide elicits a cyclooxygenase-2-dependent mitochondrial apoptosis signaling pathway in human neuroglioma cells.
OBJECTIVE
Cannabinoids have been associated with tumor regression and apoptosis of cancer cells. Recently, we have shown that R(+)-methanandamide (R(+)-MA) induces apoptosis of H4 human neuroglioma cells via a mechanism involving de novo expression of the cyclooxygenase-2 (COX-2) enzyme. The present study investigated a possible involvement of a mitochondrial-driven pathway in this process.


METHODS
Cell death was determined by the WST-1 cell viability test, and changes in apoptotic parameters [i.e., release of mitochondrial cytochrome c, activation of caspases, cleavage of poly(ADP-ribose) polymerase (PARP)] were detected by Western blotting.


RESULTS
H4 cells treated with R(+)-MA showed typical signs of mitochondrial apoptosis, i.e., release of mitochondrial cytochrome c into the cytosol and activation of initiator caspase-9. Moreover, activation of the executor caspase-3 was observed following cannabinoid treatment. Cells were fully protected from apoptotic cell death by the caspase-3 inhibitor Ac-DEVD-CHO, indicating a crucial role for caspase-3 activation in R(+)-MA-elicited apoptosis. Furthermore, cleavage of the caspase-3 target protein PARP was registered. All of the aforementioned effects were substantially reduced by the selective COX-2 inhibitor celecoxib (1 muM) at a pharmacologically relevant, nonapoptotic concentration.


CONCLUSIONS
R(+)-MA-induced apoptosis is mediated via a mitochondrial-dependent pathway that becomes activated, at least in part, through up-regulation of the COX-2 enzyme.
Eichele
Karin
K
Department of Experimental and Clinical Pharmacology and Toxicology, Friedrich Alexander University Erlangen-Nürnberg, Fahrstrasse 17, D-91054, Erlangen, Germany.
Weinzierl
Ulrike
U
Ramer
Robert
R
Brune
Kay
K
Hinz
Burkhard
B
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D001095:Arachidonic Acids / Q000494:pharmacology*; D015153:Blotting, Western; D001932:Brain Neoplasms / Q000473:pathology*; D016466:CHO Cells; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D000068579:Celecoxib; D002470:Cell Survival / Q000187:drug effects; D006224:Cricetinae; D051546:Cyclooxygenase 2 / Q000378:metabolism*; D052246:Cyclooxygenase 2 Inhibitors / Q000494:pharmacology; D045304:Cytochromes c / Q000378:metabolism; D005910:Glioma / Q000473:pathology*; D006801:Humans; D008928:Mitochondria / Q000187:drug effects*; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D011720:Pyrazoles / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects*; D013449:Sulfonamides / Q000494:pharmacology
D001095:Arachidonic Acids; D052246:Cyclooxygenase 2 Inhibitors; D011720:Pyrazoles; D013449:Sulfonamides; C088155:methanandamide; D045304:Cytochromes c; D051546:Cyclooxygenase 2; D011065:Poly(ADP-ribose) Polymerases; C505407:CASP3 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases; D000068579:Celecoxib