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Pharmacopsychiatry 2003; 36: 8-14
DOI: 10.1055/s-2003-40454
Original Paper
© Georg Thieme Verlag Stuttgart · New York

Pharmacological Studies Supporting the Therapeutic Use of Ginkgo biloba Extract for Alzheimer’s Disease

B. Ahlemeyer1 , J. Krieglstein1
  • 1Institute for Pharmacology und Toxicology, Department of Pharmacy, Philipps University of Marburg, Germany
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Publication History

Publication Date:
07 July 2003 (online)

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The standardized Ginkgo biloba extract EGb 761® (definition see editorial) has been shown to produce neuroprotective effects in different in vivo and in vitro models. Since EGb 761® is a complex mixture containing flavonoid glycosides, terpene lactones (non-flavone fraction) and various other constituents, the question arises as to which of these compounds mediates the protective activity of EGb 761®. Previous studies have demonstrated that the non-flavone fraction was responsible for the antihypoxic activity of EGb 761®. Thus, we examined the neuroprotective and anti-apoptotic ability of the main constituents of the non-flavone fraction, the ginkgolides A, B, C, J and bilobalide. In focal cerebral ischemia models, the administration of bilobalide (5 - 20 mg/kg, s. c.) 60 min before ischemia dose-dependently reduced the infarct area in mouse brain and the infarct volume in rat brain 2 days after the onset of the injury. 30 minutes of pretreatment with ginkgolide A (50 mg/kg, s. c.) and ginkgolide B (100 mg/kg, s. c.) reduced the infarct area in the mouse model of focal ischemia. In primary cultures of hippocampal neurons and astrocytes from neonatal rats, ginkgolide B (1 μM) and bilobalide (10 μM) protected the neurons against damage caused by glutamate (1 mM, 1 h) as evaluated by trypan blue staining. In addition, bilobalide (0.1 μM) was able to increase the viability of cultured neurons from chick embryo telencepalon when exposed to cyanide (1 mM, 1h). Furthermore, we attempted to find out whether ginkgolides A, B, and J and bilobalide were also able to inhibit neuronal apoptosis (determined by nuclear staining with Hoechst 33 258 and TUNEL-staining). Ginkgolide B (10 μM), ginkgolide J (100 μM) and bilobalide (1 μM) reduced the apoptotic damage induced by serum deprivation (24h) or treatment with staurosporine (200 nM, 24h) in cultured chick embryonic neurons. Bilobalide (100 μM) rescued cultured rat hippocampal neurons from apoptosis caused by serum deprivation (24h), whereas ginkgolide B (100 μM) and bilobalide (100 μM) reduced apoptotic damage induced by staurosporine (300 nM, 24h). Ginkgolide A failed to affect apoptotic damage neither in serum-deprived nor in staurosporine-treated neurons. The results suggest that some of the constituents of the non-flavone fraction of EGb 761® possess neuroprotective and anti-apoptotic capacity, and that bilobalide is the most potent one. In contrast, ginkgolic acids (100 - 500 μM) induced neuronal death, which showed features of apoptosis as well as of necrosis, but these constituents were removed from EGb 761® below an amount of 0.0005 %. Taking together, there is experimental evidence for a neuroprotective effect of EGb 761® that agrees with clinical studies showing the efficacy of an oral treatment in patients with mild and moderate dementia.

References

  • 1 Ahlemeyer B, Selke D, Schaper C, Klumpp S, Krieglstein J. Ginkgolic acids induce neuronal death and activate protein phosphatase type-2C.  Eur J Pharmacol. 2001;  430 1-7
    CrossrefPubMedSearch in Google Scholar
  • 2 Ahlemeyer B, Möwes A, Krieglstein J. Inhibition of serum deprivation- and staurosporine-induced neuronal apoptosis by Ginkgo biloba extract and some of its constituents.  Eur J Pharmacol. 1999;  367 423-430
    CrossrefPubMedSearch in Google Scholar
  • 3 Ahlemeyer B, Krieglstein J. Nenroprotective effects of the Ginkgo biloba extract. In: Lawson LD, Bauer R. (Eds.) Phytomedicine of Europe: Chemistry and Biological activity. ACS Symposium Series 691, American Chemical Society Washington DC; 1998: pp. 210-220
    Search in Google Scholar
  • 4 Backhauß C, Karkoutly C, Welsch M, Krieglstein J. A mouse model of focal cerebral ischemia for screening neuroprotective drug effects.  J Pharmacol Meth. 1992;  27 23-27
    PubMedSearch in Google Scholar
  • 5 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 β-amyloid.  Eur J Neurosci. 2000;  12 1882-1890
    CrossrefPubMedSearch in Google Scholar
  • 6 Bazan N G, Squinto S P, Braquet P, Panetta T, Marcheselli V L. Platelet activating factor and polyunsaturated fatty acids in cerebral ischemia or convulsions: intracellular PAF-binding sites and activation of a Fos/Jun/AP-1 transcriptional signalling system.  Lipids. 1991;  26 1236-1242
    PubMedSearch in Google Scholar
  • 7 Bielenberg G W, Sauer D, Nuglisch J, Beck T, Roßberg C, Mennel H D, Krieglstein J. Effects of emopamil on postischemic blood flow and neuronal damage in rat brain.  Naunyn Schmiedeberg’s Arch Pharmacol. 1989;  339 230-235
    PubMedSearch in Google Scholar
  • 8 Chatterjee S S, Gabard B. Protective effect of an extract of Ginkgo biloba and other hydroxyl radical scavengers against hypoxia. VIII International Congress on Pharmacology Tokyo; 1981: pp. 483-486
    Search in Google Scholar
  • 9 Chen C, Wei T T, Gao Z H, Zhao B L, Hou J W, Xu H B, Xin W J, Packer L. Different effects of the constituents of EGb 761® on apoptosis in rat cerebellar granule cells induced by hydroxyl radicals.  Biochem Mol Biol Int. 1999;  47 397-405
    PubMedSearch in Google Scholar
  • 10 DeFeudis F V. Ginkgo biloba extract (EGb 761®). From chemistry to the clinic. Ullstein Medical Wiesbaden; 1998
    Search in Google Scholar
  • 11 Droy-Lefaix M T. Effect of the antioxidant action of Ginkgo biloba extract (EGb 761®) on aging and oxidative stress.  Age. 1997;  20 141-149
    PubMedSearch in Google Scholar
  • 12 Ernst E, Pittler M H. Ginkgo biloba for dementia: A systematic review of double-blind, placebo-controlled trials.  Clin Drug Invest. 1999;  17 301-308
    CrossrefPubMedSearch in Google Scholar
  • 13 Ginsberg M D, Busto R. Rodent models of cerebral ischemia.  Stroke. 1989;  20 1627-1642
    PubMedSearch in Google Scholar
  • 14 Guidetti C, Paracchini S, Lucchini S, Cambieri M, Marzatico F. Prevention of neuronal cell damage induced by oxidative stress in- vitro: effect of different Ginkgo biloba extracts.  J Pharm Pharmacol. 2001;  53 387-392
    CrossrefPubMedSearch in Google Scholar
  • 15 Husain S R, Cillard J, Cillard P. Hydroxyl radical scavenging activity of flavonoids.  Phytochemistry. 1987;  26 2489-2491
    CrossrefPubMedSearch in Google Scholar
  • 16 Janssens D, Remacle J, Drieu K, Michiels C. Protection of mitochondrial respiration activity by bilobalide.  Biochem Pharmacol. 1999;  58 109-119
    CrossrefPubMedSearch in Google Scholar
  • 17 Kanowski S, Herrmann W M, Stephan K, Wierich W, Hörr 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 of multi-infarct dementia.  Pharmacopsychiatry. 1996;  29 47-56
    Thieme ConnectPubMedSearch in Google Scholar
  • 18 Karcher L, Zagermann P, Krieglstein J. Effect of an extract of Ginkgo biloba on rat brain energy metabolism in hypoxia.  Naunyn-Schmiedeberg’s Arch Pharmacol. 1984;  327 31-35
    PubMedSearch in Google Scholar
  • 19 Kleijnen J, Knipschild P. Ginkgo biloba.  Lancet. 1992;  340 1136-1139
    CrossrefPubMedSearch in Google Scholar
  • 20 Krieglstein J, Beck T, Seibert A. Influence of an extract of Ginkgo biloba on cerebral blood flow and metabolism.  Life Sci. 1986;  39 2327-2334
    CrossrefPubMedSearch in Google Scholar
  • 21 Krieglstein J, Ausmeier F, El-Abhar H, Lippert K, Welsch M, Rupalla K, Henrich-Noack P. Neuroprotective effects of Ginkgo biloba constituents.  Eur J Pharm Sci. 1995;  3 39-48
    PubMedSearch in Google Scholar
  • 22 Le Bars P, Katz M M, Herman N, Itil T M, Freedman A M, Schatzberg A F. A placebo-controlled, double-blind, randomized trial of an extract of Ginkgo biloba for dementia.  J Am Med Assoc. 1997;  278 1327-1332
    CrossrefPubMedSearch in Google Scholar
  • 23 Le Bars P L, Velasco P M, Ferguson J M, Dessain E G, Kieser M, Hörr 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
    CrossrefPubMedSearch in Google Scholar
  • 24 Lugasi A, Horhavovich P, Dworschak E. Additional information to the in vitro antioxidant activity of Ginkgo biloba.  L Phytother Res. 1999;  13 160-162
    PubMedSearch in Google Scholar
  • 25 Marcocci L, Packer L, Droy-Lefaix M T, Sekaki A, Gardes-Albert M. Antioxidant action of Ginkgo biloba extract EGb 761®.  Methods Enzymol. 1994;  234 462-475
    PubMedSearch in Google Scholar
  • 26 Ni Y, Zhao B, Hou J, Xin W. Preventive effect of Ginkgo biloba extract on apoptosis in rat cerebellar neuronal cells induced by hydroxyl radicals.  Neurosci Lett. 1996;  214 115-118
    CrossrefPubMedSearch in Google Scholar
  • 27 Nuglisch J, Karkoutly C, Mennel H D, Roßberg C, Krieglstein J. Protective effect of nimodipine against ischemic neuronal damage in rat hippocampus without changing postischemic cerebral blood flow.  J Cereb Blood Flow Metab. 1990;  10 654-659
    PubMedSearch in Google Scholar
  • 28 Nunez D, Chignard M, Korth R, Le Couedic J P, Norel X, Spinnewyn B, Braquet P, Benveniste J. Specific inhibition of paf-acether-induced platelet activition by BN 52 021 and comparison with the paf-acether inhibitors Kadsurenone and CV 3988.  Eur J Pharmacol. 1986;  123 197-205
    CrossrefPubMedSearch in Google Scholar
  • 29 Oberpichler H, Beck T, Abdel-Rahman M M, Bielenberg G W, Krieglstein J. Effects of Ginkgo biloba constituents related to protection against brain damage caused by hypoxia.  Pharmacol Res Commun. 1988;  20 349-368
    PubMedSearch in Google Scholar
  • 30 Oberpichler H, Sauer D, Roßberg C, Mennel H D, Krieglstein J. PAF-antagonist ginkgolide B reduced postischemic neuronal damage in rat brain hippocampus.  J Cereb Blood Flow Metab. 1990;  10 133-135
    PubMedSearch in Google Scholar
  • 31 Oppenheim R W. Cell death during development of the nervous system.  Ann Rev Neurosci. 1991;  14 453-501
    PubMedSearch in Google Scholar
  • 32 Oyama Y, Fuchs P A, Katayam N, Noda K. Myricetin and quercetin, the flavonoid constituents of Ginkgo biloba extract, greatly reduce oxidative metabolism in both resting and Ca2+-loaded brain neurons.  Brain Res. 1994;  28 125-129
    PubMedSearch in Google Scholar
  • 33 Oyama Y, Chikashia L, Ueha T, Kanemaru K, Noda K. Ginkgo biloba extract protects brain neurons against oxidative stress induced by hydrogen peroxide.  Brain Res. 1996;  18 349-352
    PubMedSearch in Google Scholar
  • 34 Panetta T, Marcheselli V L, Braquet P, Spinnewyn B, Bazan N G. Effect of a platelet activating factor antagonist (BN 52 021) on free fatty acids, diacylglycerols, polyphosphoinositides and blood flow in the gerbil brain: inhibition of ischemia-reperfusion induced cerebral injury.  Biochem Biophys Res Commun. 1987;  149 580-587
    CrossrefPubMedSearch in Google Scholar
  • 35 Pierre S, Jamme I, Droy-Lefaix M T, Nouvelot A, Maixent J M. Ginkgo biloba extract (EGb 761®) protects Na,K-ATPase activity during cerebral ischemia.  NeuroReport. 1999;  10 47-51
    PubMedSearch in Google Scholar
  • 36 Pietri S, Maurelli E, Drieu K, Culcasi M. Cardioprotective and anti-oxidant effects of the terpenoid constituents of Ginkgo biloba extract (EGb 761®).  J Mol Cell Cardiol. 1997;  29 733-742
    CrossrefPubMedSearch in Google Scholar
  • 37 Pincemail J, Thirion A, Dupuis M, Braquet P, Drieu K, Deby C. Ginkgo biloba extract inhibits oxygen species production generated by phorbol myristate acetate stimulated human leukocytes.  Experientia. 1987;  43 181-184
    PubMedSearch in Google Scholar
  • 38 Prehn J HM, Krieglstein J. Platelet-activating factor antagonists reduce excitotoxic damage in cultured neurons from embryonic chick telencephalon and protect the rat hippocampus and neocortex from ischemic injury in vivo.  J Neurosci Res. 1993;  34 179-188
    CrossrefPubMedSearch in Google Scholar
  • 39 Robak J, Gryglewski R J. Flavonoids are scavengers of superoxide anions.  Biochem Pharmacol. 1998;  37 837-841
    PubMedSearch in Google Scholar
  • 40 Schindowski K, Leutner S, Kressmann S, Eckert A, Müller W E. Age-related increase of oxidative stress-induced apoptosis in mice. Prevention by Ginkgo biloba extract (EGb 761®).  J Neural Transm. 2001;  108 969-978
    CrossrefPubMedSearch in Google Scholar
  • 41 Scholtyssek H, Damerau W, Wessel R, Schminke I. Antioxidative activity of ginkgolides against superoxide in an aprotic environment.  Chemico-Biol Interactions. 1997;  106 183-190
    PubMedSearch in Google Scholar
  • 42 Shi H, Niki E. Stoichiometric and kinetic studies on Ginkgo biloba extract and related antioxidants.  Lipids. 1998;  33 365-370
    PubMedSearch in Google Scholar
  • 43 Smith M L, Bendek G, Dahlgren N, Rosen I, Wieloch T, Siesjö B K. Models for studying long-term recovery following forebrain ischemia in the rat 2. A 2-vessel occlusion model.  ActaNeurol Scand. 1984;  69 385-401
    PubMedSearch in Google Scholar
  • 44 Tamura A, Graha D J, McCulloch J, Teasdale G M. Focal ischemia in the rat: 1. Description of technique and early neuropathological consequences following middle cerebral artery occlusion.  J Cereb Blood Flow Metab. 1981;  1 53-60
    PubMedSearch in Google Scholar
  • 45 Thompson C B. Apoptosis in the pathogenesis and treatment of disease.  Science. 1995;  267 1456-1462
    CrossrefPubMedSearch in Google Scholar
  • 46 Touvay C, Vilain B, Taylor J E, Etienne A, Braquet P. Proof of the involvement of platelet-activating factor (PAF-acether) in pulmonary complex immune system using a specific PAF acether receptor antagonist: BN 52 021.  Prog Lipid Res. 1986;  25 277-288
    PubMedSearch in Google Scholar
  • 47 Welsch M, Nuglisch J, Krieglstein J. Neuroprotective effect of nimodipine is not mediated by increased cerebral blood flow after transient forebrain ischemia in rats.  Stroke. 1990;  21 96-105
    PubMedSearch in Google Scholar
  • 48 Wettstein A. Cholinesterase inhibitors and Ginkgo extracts - are they comparable in the treatment of dementia? Comparison of published placebo-controlled efficacy studies of at least six months’ duration.  Phytomedicine. 2000;  6 393-401
    PubMedSearch in Google Scholar
  • 49 Zhou L J, Song W, Zhu X Z, Chen Z L, Yin M L, Cheng X F. Protective effect of bilobalide on amyloid beta-peptide 25 - 35-induced PC12 cell cytotoxicity.  Acta Pharmacol Sin. 2000;  21 75-79
    PubMedSearch in Google Scholar

Prof. Dr. Dr. Josef Krieglstein

Institut für Pharmakologie und Toxikologie

Fachbereich Pharmazie der Philipps-Universität Marburg

Ketzerbach 63

35032 Marburg

Germany

Phone: (49) 6421-2821311 (secretariat)

Fax: (49) 6421-2828918

Email: krieglst@mailer.uni-marburg.de