Biochemical and Structural Studies of Paired Helical Filaments and Senile Plaque Amyloid in Alzheimer’s Disease
During aging of the human brain and particularly in Alzheimer’s disease (AD), abnormal fibers accumulate both inside neurons (in neurofibrillary tangles and the neurites of senile plaques) and extracellularly (as amyloid in senile plaque cores and in vessels). Both types of fibers show birefringence with Congo red, but their ultrastructures are distinct. The principal intraneuronal fibers are paired helical filaments (PHF) with maximal diameters of 20–24 nm in situ. However, we have recently observed in several cases of AD that both neuronal cell bodies and neurites can display a complex mixture of abnormal fibers including, in addition to an abundance of PHF, straight filaments varying from 10 to 20 nm in diameter, most commonly ∿15 nm. The extracellular fibers in the amyloid cores of senile plaque are generally approximately 10 nm in diameter. We have found that both PHF and amyloid core fibers are highly insoluble in a variety of detergents and denaturants including sodium dodecyl sulfate, reducing agents, urea, and guanidine HCl. The PHF are also resistant to a variety of specific and non-specific proteases; we have not yet fully determined the extent of resistance of senile plaque amyloid to these proteases. We have raised highly specific antibodies to PHF which decorate neurofibrillary tangles both in AD brain tissue sections and following the isolation and partial purification of the tangles. We have also raised a polyconal antiserum which immunolabels SDS-isolated senile plaque cores. Taking advantage of the dense spherical nature of the senile plaque core and its intactness following SDS-extraction, we have developed a novel method for purification of cores from postmortem brain using a combination of sucrose density centrifugation and fluorescence activated cell sorting. This flow cytometry technique produces highly purified, intact cores in quantities suitable for further biochemical and structural analyses. Results of initial analyses of the purified cores are provided. The evidence available to date suggests that although PHF and extracellular amyloid core fibers are morphologically distinct, they have similar amino acid compositions and share major antigenic determinants.
KeywordsSucrose Urea Lithium Bromide Glycine
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