Apoptosis by 6-O-palmitoyl-L-ascorbic acid coincides with JNK-phosphorylation and inhibition of Mg2+-dependent phosphatase activity.
6-O-Palmitoyl ascorbic acid (PAA) has recently been used as a substitute for ascorbic acid because of its greater potency as an antioxidant. In detailed concentration response studies distinct cytotoxic effects of PAA at concentrations exceeding 100 microM were reported. Here we examined and further characterized this cytotoxicity. While ascorbic acid was tolerated well up to millimolar concentrations, PAA revealed an LC50 between 125 and 150 microM in rat GH3 tumor cells. Morphological and biochemical observations suggested the induction of apoptosis at concentrations exceeding 125 microM with a prominent activation of caspase 3 at 250 microM after 4 hr. A subsequent pronounced fragmentation of DNA (DNA-ladder) was detected after 6 hr and was further enhanced after 12 hr. The activation of caspases and the cleavage of its substrate PARP was preceded by a distinct increase in the phosphorylation of stress activated JNK-kinases. This observation suggested that the agent affected signal transduction mechanisms regulating protein phosphorylation at serine/threonine residues in the cell. No effect of PAA on protein phosphatase 2A (PP2A)-like activity was observed while magnesium-dependent protein phosphatase activity, presumably PP2C, was inhibited concentration-dependently up to 75% at the respective concentrations. Thus, the cytotoxic, pro-apoptotic effect of PAA might be related to the inhibition of PP2C and the activation of JNK.
Kaap
S
S
Institute of Pharmacology & Toxicology, University of Göttingen, Göttingen, Germany.
Brechlin
P
P
Quentin
I
I
Eger
K
K
Steinfelder
H J
HJ
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis; D001205:Ascorbic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology*; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D054648:Protein Phosphatase 2; D000071636:Protein Phosphatase 2C; D051381:Rats; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2; D000071636:Protein Phosphatase 2C; D001205:Ascorbic Acid; C031226:6-O-palmitoylascorbic acid