Ginkgo biloba Extract and its Flavonol and Terpenelactone Fractions do not Affect β-Secretase mRNA and Enzyme Activity Levels in Cultured Neurons and in Mice

Abstract

Numerous clinical trials have reported beneficial effects of the Ginkgo biloba extract EGb761 in the prevention and therapy of cognitive disorders including Alzheimer’s disease (AD). Although neuroprotective properties of EGb761 have been consistently reported, the molecular mechanisms of EGb761 and the specific role of its major constituents, the flavonols and terpenlactones, are largely unknown. One major hallmark of AD is the deposition of amyloid-beta (Aβ) as amyloid plaques in the brain. Aβ is a cleavage product of amyloid precursor protein (APP). Certain proteases, called β-secretases (BACE), are crucial in the formation of Aβ. The purpose of the present study was to investigate the efficacy of EGb761 and its flavonol and terpenelactone fraction to modulate BACE-1 enzyme activity and mRNA levels in vitro and in vivo. Neither EGb761 nor its fractions affected BACE-1 activity in vitro. Furthermore, also in Neuro-2a cells and wild-type as well as transgenic (Tg2576) laboratory mice, no significant effect of EGb761 on BACE-1 enzyme activity and mRNA levels were observed. Current findings suggest that BACE-1 may not be a major molecular target of EGb761 and its flavonol and terpenelactone fraction.

Key words

Ginkgo biloba - Ginkgoaceae - EGb761 - β-secretase - Alzheimer’s disease - amyloid-beta - Tg2576

References

• 1 Selkoe D J. Amyloid protein and Alzheimer’s disease.  Sci Am. 1991;  265 68-78
• 2 Tong Y, Zhou W, Fung V, Christensen M A, Qing H, Sun X. et al . Oxidative stress potentiates BACE1 gene expression and Abeta generation.  J Neural Transm. 2005;  112 455-69
• 3 Dewachter I, Van Leuven F. Secretases as targets for the treatment of Alzheimer’s disease: the prospects.  Lancet Neurol. 2002;  1 09-16.
• 4 DeFeudis F V, Drieu K. Ginkgo biloba extract (EGb761) and CNS functions: basic studies and clinical applications.  Curr Drug Targets. 2000;  1 25-58
• 5 Le Bars P L, Velasco F M, Ferguson J M, Dessain E C, Kieser M, Hoerr R. Influence of the severity of cognitive impairment on the effect of the Ginkgo biloba extract EGb 761 in Alzheimer’s disease.  Neuropsychobiology. 2002;  45 19-26
• 6 Kanowski S, Herrmann W M, Stephan K, Wierich W, Horr R. Proof of efficacy of the Ginkgo biloba special extract EGb 761 in outpatients suffering from mild to moderate primary degenerative dementia of the Alzheimer type or multi-infarct dementia.  Pharmacopsychiatry. 1996;  29 47-56
• 7 Kanowski S, Hoerr R. Ginkgo biloba extract EGb 761 in dementia: intent-to-treat analyses of a 24-week, multi-center, double-blind, placebo-controlled, randomized trial.  Pharmacopsychiatry. 2003;  36 297-303
• 8 Dongen M V, van Rossum E, Kessels A, Sielhorst H, Knipschild P. Ginkgo for elderly people with dementia and age-associated memory impairment: a randomized clinical trial.  J Clin Epidemiol. 2003;  56 367-76
• 9 Williams B, Watanabe C MH, Schultz P G, Rimbach G, Krucker T. Age-related effects of Ginkgo biloba extract on synaptic plasticity and excitability.  Neurobiol Aging. 2004;  25 955-62
• 10 Smith J V, Luo Y. Studies on molecular mechanisms of Ginkgo biloba extract.  Appl Microbiol Biotechnol. 2004;  64 465-72
• 11 Song W, Guan H J, Zhu X Z, Chen Z L, Yin M L, Cheng X F. Protective effect of bilobalide against nitric oxide-induced neurotoxicity in PC12 cells.  Acta Pharmacol Sin. 2000;  21 415-20
• 12 Watanabe C MH, Wolffram S, Ader P, Rimbach G, Packer L, Maguire J J. et al . The in vivo neuromodulatory effects of the herbal medicine Ginkgo biloba.  PNAS. 2001;  98 6577-80
• 13 Farzan M, Schnitzler C E, Vasilieva N, Leung D, Choe H. BACE2, a β-secretase homolog, cleaves at the beta site and within the amyloid-beta region of the amyloid-beta precursor protein.  PNAS. 2000;  97 9712-7
• 14 Cai H, Wang Y, McCarthy D, Wen H, Borchelt D R, Price D L. et al . BACE1 is the major β-secretase for generation of Aβ peptides by neurons.  Nat Neurosci. 2001;  4 233-4
• 15 Luo Y, Bolon B, Kahn S, Bennett B D, Babu-Khan S, Denis P. et al . Mice deficient in BACE1, the Alzheimer's beta-secretase, have normal phenotype and abolished beta-amyloid generation.  Nat Neurosci. 2001;  4 231-2
• 16 Vassar R, Bennett B D, Babu-Khan S, Kahn S, Mendiaz E A, Denis P. et al . β-Secretase cleavage of Alzheimer's amyloid precursor protein by the transmembrane aspartic protease BACE.  Science. 1999;  286 735-41
• 17 Yan R, Bienkowski M J, Shuck M E, Miao H, Tory M C, Pauley A M. et al . Membrane-anchored aspartyl protease with Alzheimer's disease β-secretase activity.  Nature. 1999;  402 533-7
• 18 Sinha S, Anderson J P, Barbour R, Basi G S, Caccavello R, Davis D. et al . Purification and cloning of amyloid precursor protein β-secretase from human brain.  Nature. 1999;  402 37-40
• 19 Ohnishi S, Takano K. Amyloid fibrils from the viewpoint of protein folding.  Cell Mol Life Sci. 2004;  61 511-24
• 20 Sinha S, Lieberburg I. Cellular mechanisms of beta-amyloid production and secretion.  PNAS. 1999;  96 11 049-53
• 21 Citron M, Oltersdorf T, Haass C, McConlogue L, Hung A Y, Seubert P. et al . Mutation of the β-amyloid precursor protein in familial Alzheimer's disease increases β-protein production.  Nature. 1992;  360 672-4
• 22 Bodendorf U, Danner S, Fischer F, Stefani M, Sturchler-Pierrat C, Wiederhold K H. et al . Expression of human beta-secretase in the mouse brain increases the steady-state level of beta-amyloid.  J Neurochem. 2002;  80 799-806
• 23 Hsiao K, Chapman P, Nilsen S, Eckman C, Harigaya Y, Younkin S. et al . Correlative memory deficits, Abeta elevation, and amyloid plaques in transgenic mice.  Science. 1996;  274 99-102
• 24 Hsiao K K, Borchelt D R, Olson K, Johannsdottir R, Kitt C, Yunis W. et al . Age-related CNS disorder and early death in transgenic FVB/N mice overexpressing Alzheimer amyloid precursor proteins.  Neuron. 1995;  15 1203-18
• 25 Monografie: Trockenextrakt (35 - 67 : 1) aus Ginkgo biloba Blättern, extrahiert mit Aceton-Wasser. Bundesanzeiger 46 1994 Nr. 133: 7361-66
• 26 Ader P, Wessmann A, Wolffram S. Bioavailability and metabolism of the flavonol quercetin in the pig.  Free Radic Biol Med. 2000;  28 1056-67.
• 27 Biber A, Koch E. Bioavailability of ginkgolides and bilobalide from extracts of Ginkgo biloba using GC/MS.  Planta Med. 1999;  65 192-93
• 28 Borenfreund E, Puerner J A. Toxicity determined in vitro by morphological alterations and neutral red absorption.  Toxicol Lett. 1985;  24 119-24
• 29 de Boer V CJ, Dihal A A, van der Woude H, Arts I CW, Wolffram S, Alink G M. et al . Tissue distribution of quercetin in rats and pigs.  J Nutr. 2005;  135 1718-25
• 30 Bastianetto S, Ramassamy C, Dore S, Christen Y, Poirier J, Quirion R. The Ginkgo biloba extract (EGb 761) protects hippocampal neurons against cell death induced by beta-amyloid.  Eur J Neurosci. 2000;  12 1882-90
• 31 Christensen M A, Zhou W, Qing H, Lehman A, Philipsen S, Song W. Transcriptional regulation of BACE1, the β-amyloid precursor protein β-Secretase, by Sp1.  Mol Cell Biol. 2004;  24 865-74
• 32 Marcinkiewicz M, Seidah N G. Coordinated expression of beta-amyloid precursor protein and the putative beta-secretase BACE and alpha-secretase ADAM10 in mouse and human brain.  J Neurochem. 2000;  75 2133-43.
• 33 Luo Y, Smith J V, Paramasivam V, Burdick A, Curry K J, Buford J P. et al . Inhibition of amyloid-beta aggregation and caspase-3 activation by the Ginkgo biloba extract EGb761.  PNAS. 2002;  99 2197-202
• 34 Colciaghi F, Borroni B, Zimmermann M, Bellone C, Longhi A, Padovani A. et al . Amyloid precursor protein metabolism is regulated toward alpha-secretase pathway by Ginkgo biloba extracts.  Neurobiol Dis. 2004;  16 454-60