Ginkgolide B inhibits the neurotoxicity of prions or amyloid-beta1-42

Abstract

BACKGROUND: Neuronal loss in Alzheimer's or prion diseases is preceded by the accumulation of fibrillar aggregates of toxic proteins (amyloid-beta1-42 or the prion protein). Since some epidemiological studies have demonstrated that the EGb 761 extract, from the leaves of the Ginkgo biloba tree, has a beneficial effect on Alzheimer's disease, the effect of some of the major components of the EGb 761 extract on neuronal responses to amyloid-beta1-42, or to a synthetic miniprion (sPrP106), were investigated. METHODS: Components of the EGb 761 extract were tested in 2 models of neurodegeneration. SH-SY5Y neuroblastoma cells were pre-treated with ginkgolides A or B, quercetin or myricetin, and incubated with amyloid-beta1-42, sPrP106, or other neurotoxins. After 24 hours neuronal survival and the production of prostaglandin E2 that is closely associated with neuronal death was measured. In primary cortical neurons apoptosis (caspase-3) in response to amyloid-beta1-42 or sPrP106 was measured, and in co-cultures the effects of the ginkgolides on the killing of amyloid-beta1-42 or sPrP106 damaged neurons by microglia was tested. RESULTS: Neurons treated with ginkgolides A or B were resistant to amyloid-beta1-42 or sPrP106. Ginkgolide-treated cells were also resistant to platelet activating factor or arachidonic acid, but remained susceptible to hydrogen peroxide or staurosporine. The ginkgolides reduced the production of prostaglandin E2 in response to amyloid-beta1-42 or sPrP106. In primary cortical neurons, the ginkgolides reduced caspase-3 responses to amyloid-beta1-42 or sPrP106, and in co-culture studies the ginkgolides reduced the killing of amyloid-beta1-42 or sPrP106 damaged neurons by microglia. CONCLUSION: Nanomolar concentrations of the ginkgolides protect neurons against the otherwise toxic effects of amyloid-beta1-42 or sPrP106. The ginkgolides also prevented the neurotoxicity of platelet activating factor and reduced the production of prostaglandin E2 in response to platelet activating factor, amyloid-beta1-42 or sPrP106. These results are compatible with prior reports that ginkgolides inhibit platelet-activating factor, and that platelet-activating factor antagonists block the toxicity of amyloid-beta1-42 or sPrP106. The results presented here suggest that platelet-activating factor antagonists such as the ginkgolides may be relevant treatments for prion or Alzheimer's diseases.

Figure 1

Ginkgolides protect neurons against amyloid-β_1-42: The survival of SH-SY5Y cells pre-treated with different concentrations of ginkgolide A (shaded circles) or ginkgolide B (open circles) for 3 hours, and thereafter incubated with 20 μM amyloid-β_1-42. Cell survival was measured 24 hours later using the WST-1 method. Each value represents the mean percentage cell survival ± SD from triplicate experiments repeated four times (12 observations).

Figure 2

Ginkgolide B protects neurons against amyloidogenic peptides: The survival of SH-SY5Y cells pre-treated with different concentrations of ginkgolide B for 3 hours and thereafter incubated with 5 μM sPrP106 (open squares), 10 μM HuPrP82-146 (open circles) or 20 μM amyloid-β_1-42 (shaded circles). Cell survival was measured 24 hours later using the WST-1 method. Each value represents the mean percentage cell survival ± SD from triplicate experiments repeated four times (12 observations).

Figure 3

The protective effect of ginkgolide B is non-competitive: Untreated SH-SY5Y cells were incubated for 24 hours with different concentrations of amyloid-β_1-42 (open circles) or sPrP106 (open squares). Cells pre-treated for 3 hours with 1 μM ginkgolide B were subsequently incubated for 24 hours with different concentrations of amyloid-β_1-42 (closed circles) or sPrP106 (closed squares). Cell survival was measured 24 hours later using the WST-1 method. Each point represents the mean ± SD from triplicate experiments repeated 4 times (12 observations).

Figure 4

Ginkgolides protect sPrP106 or amyloid-β damaged neurons against microglia. Primary cortical neurons were pre-treated with control medium (Con), with 1 μM ginkgolide A, with 1 μM ginkgolide B, with 25 μM quercetin (Quer) or with 25 μM myrecetin (Myr) for 3 hours were then incubated with 5 μM sPrP106 (open squares) or 10 μM amyloid-β_1-42 (shaded bars). Neuronal survival was determined 4 days later after the removal of microglia using the WST-1 assay. Values shown represent the percentage survival of treated cells compared to untreated cells. Each point represents the mean ± SD from triplicate experiments repeated 4 times (12 observations).