Ginkgolic acids induce neuronal death and activate protein phosphatase type-2C.
The standardized extract from Ginkgo biloba (EGb 761) is used for the treatment of dementia. Because of allergenic and genotoxic effects, ginkgolic acids are restricted in EGb 761 to 5 ppm. The question arises whether ginkgolic acids also have neurotoxic effects. In the present study, ginkgolic acids caused death of cultured chick embryonic neurons in a concentration-dependent manner, in the presence and in the absence of serum. Ginkgolic acids-induced death showed features of apoptosis as we observed chromatin condensation, shrinkage of the nucleus and reduction of the damage by the protein synthesis inhibitor cycloheximide, demonstrating an active type of cell death. However, DNA fragmentation detected by the terminal-transferase-mediated ddUTP-digoxigenin nick-end labeling (TUNEL) assay and caspase-3 activation, which are also considered as hallmarks of apoptosis, were not seen after treatment with 150 microM ginkgolic acids in serum-free medium, a dose which increased the percentage of neurons with chromatin condensation and shrunken nuclei to 88% compared with 25% in serum-deprived, vehicle-treated controls. This suggests that ginkgolic acid-induced death showed signs of apoptosis as well as of necrosis. Ginkgolic acids specifically increased the activity of protein phosphatase type-2C, whereas other protein phosphatases such as protein phosphatases 1A, 2A and 2B, tyrosine phosphatase, and unspecific acid- and alkaline phosphatases were inhibited or remained unchanged, suggesting protein phosphatase 2C to play a role in the neurotoxic effect mediated by ginkgolic acids.
Ahlemeyer
B
B
Institut für Pharmakologie und Toxikologie, Fachbereich Pharmazie der Philipps-Universität Marburg, Ketzerbach 63, D-35032, Marburg, Germany.
Selke
D
D
Schaper
C
C
Klumpp
S
S
Krieglstein
J
J
D000818:Animals; D014151:Anti-Anxiety Agents / Q000494:pharmacology / Q000633:toxicity; D017209:Apoptosis; D001921:Brain / Q000187:drug effects / Q000196:embryology; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002642:Chick Embryo; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D020441:Ginkgo biloba; D020287:In Situ Nick-End Labeling; D007527:Isoenzymes / Q000378:metabolism; D009474:Neurons / Q000187:drug effects* / Q000502:physiology; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D010936:Plant Extracts / Q000737:chemistry; D054648:Protein Phosphatase 2; D000071636:Protein Phosphatase 2C; D029701:Saccharomyces cerevisiae Proteins; D012459:Salicylates / Q000737:chemistry / Q000494:pharmacology* / Q000633:toxicity; D013194:Staining and Labeling; D013997:Time Factors; D014343:Trypan Blue; D000096983:Ginkgo Extract
D014151:Anti-Anxiety Agents; D053148:Caspase 3; D020169:Caspases; D007527:Isoenzymes; D010749:Phosphoprotein Phosphatases; D010936:Plant Extracts; D054648:Protein Phosphatase 2; D000071636:Protein Phosphatase 2C; D029701:Saccharomyces cerevisiae Proteins; D012459:Salicylates; D014343:Trypan Blue; C063170:Ginkgo biloba extract; D000096983:Ginkgo Extract; C112485:ginkgolic acid; C087358:PTC1 protein, S cerevisiae