Title

Modulation of Stress Responses and Attenuation of Oxidative Free Radicals In Alzheimer's Disease Models by the Ginkgo Biloba Extract EGb 761

Date of Award

2004

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biological Sciences

First Advisor

Yuan Luo

Advisor Department

Biological Sciences

Abstract

The prevalence of Alzheimer's disease (AD) among the public has reached shocking levels, striking those around us with alarming frequency. This debilitating disease, capable of erasing certain types of memory, vitality, sense of self, and even basic daily functions of those we love, seems to be an unstoppable force among our aging population. Medical therapies aimed at preventing this disease are dependent upon a better understanding of the AD etiology. Given the prevalence of AD in Western society and the integration of theories involving diet, environment, and heredity as possible causes, the role of free radicals in aging and AD is an important area of research. In this investigation, I used standardized Ginkgo biloba extract EGb 761, a known antioxidant and herbal supplement taken for memory enhancement, in an investigation of two strongly held theories of AD etiology and the cellular mechanisms of the therapeutic benefits gained by its use. Both in vitro and in vivo models displaying AD-associated mutations were employed to study the neurotoxic effects of Amyloid β (Aβ) and the alleviation of these effects by EGb 761 using immunochemical, enzymatic, and fluorescence techniques. First, the involvement of apoptosis, programmed cell death, as well as intracellular free radical levels produced by Aβ was investigated in vitro using a neuroblastoma cell line stably expressing an AD-associated double mutation, then similar studies were expanded to the in vivo investigation of the organism model of transgenic Caenorhabditis elegans nematodes constitutively expressing human Aβ. I demonstrated activation of the apoptotic machinery, evident by reduced mitochondrial integrity, enhanced expression of the pro-apoptotic caspase-3, and compromised viability in the AD-associated cells. I proved that the deleterious effects of Aβ in neuronal culture may be minimized by treatment with EGb 761. In addition, elevated hydrogen peroxide (H 2 O2 ) was observed by the dichlorofluorescein diacetate (DCF) method in both in vitro and in vivo models associated with AD, with significant attenuation by EGb 761 treatment at both conditions. Furthermore, an age-dependent increase in reactive oxygen species (ROS) was observed in wild type C. elegans and accelerated in the AD-associated mutants. Consistently, heightened protein carbonyls were observed in oxidative sensitive organisms. These results support the hypothesized involvement of both Aβ and ROS in association with AD and provide substantial evidence for the therapeutic potential of EGb 761 in the prevention and treatment of AD.