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I.R.F. / Aging news / Diseases / 02111801

Alterations in Content and Phosphorylation State of Cytoskeletal Proteins in the Sciatic Nerve During Ageing and Alzheimer's Disease
Posted on: November 18, 2002

Alzheimer's disease is a chronic degenerative dementing disorder that is neuropalhologically defined by the occurrence of senile plaques, neurofibrillary tangles and neuropil threads in the central nervous system. The neurofibrillar degeneration is characterized by the appearance of paired helical filaments, which are found in neurofibrillary tangles, neurophil threads and dystrophic neurites surrounding senile plaques. These paired helical filaments contain TAU protein in an abnormally high-phosphorylated state. The function of TAU protein in neurons is to regulate the stabilization of microtubules that are determinants of the cellular morphology and serve as a track for microtubule based transport mechanisms. The ability of TAU protein to stabilize microtubules depends on the TAU isoform and the degree of phosphorylation. The highly phosphorylated TAU species have a much-reduced ability to bind to microtubules, which are likely to result in destabilization of the axonal cytoskeleton, disruption of axonal transport and impairment of neuronal viability.

Neurofibrillary degeneration, however, is not universal to all brain regions and cell types. It is well documented for neurons of the entorhinal cortex, hippocampus and neocortex and for a number of subcortical neurons, which all have in common that they are mainly long-axon neurons. Investigations of neurofibrillary degeneration in the peripheral nervous system are, however, quite limited, although a large number of these ganglion cells are long axonal neurons, too. Perturbations of intracellular microtubule-based transport mechanisms may have an important impact on the function of such long-axonal neurons. Indeed, there are studies reporting neurological symptoms in more than 85% of patients with AD have recently described the occurrence of neurofibrillary tangles in the peripheral autonomic ganglion cells supporting earlier reports.

The microtubule-associated protein TAU represents a family of different isoforms generated by alternative splicing. In the human CNS, six different isoforms referred to as low molecular weight TAU (LMW) with an apparent molecular weight ranging from 48,000 to 65.000 are expressed from a mRNA with a size of 6RB. However, in the peripheral nervous system, TAU protein is derived from a larger mRNA of 8kB translated into different TAU isoforms with an apparent molecular weight of 100,000 to 120,000 commonly known as high molecular weight TAU (HMW). This HMW-TAU can preferentially be found in the spinal cord and optical nerve and is described as the only TAU species found in the adult rat sciatic nerve, dorsal and ventral roots and trigeminal nerve.

The expression of this HMW-TAU species may confer increased stabilization and spacing of the axonal microtubules. Together with the neurofilaments it determines the caliber of the axon. In contrast to the expression of HMW-TAU that has been described as the only TAU species in the rat peripheral nervous system, in the human sciatic nerve it was found three groups of TAU proteins that react with the polyclonal TAU antibody BR134. Besides LMW-TAU and HMW-TAU isoforms, furthermore, a third class of TAU proteins with a molecular size between 66 and 84kDa was detected. These latter bands were reactive for BR134 but not for TAU-1. This might suggest that these proteins harboring the BR134 epitope might represent the so-called "middle molecular weight" (MMW)-TAU isoforms. This MMW-TAU might either arise from proteolytically processed HMW-TAU, where the TAU-1 epitope is masked by phosphorylation or from dimerization of LMW-TAU. The main difference in TAU protein expression between the human nervous system and the nervous system of rodents resides in the down regulation of the juvenile TAU isoforms in rodents, while it persists in human brain throughout life. This difference might explain the abundant LMW-TAU expression in the sciatic nerve of adult human.

TAU isoforms, furthermore, are differentially expressed between sensory fibers and motor fibers. Scientists reported a higher percentage of HMW-TAU in the sensory parts of the spinal cord compared to non-sensory parts. A potentially different composition of sciatic nerve fibers might, thus, also explain this discrepancy between rodents and humans.

Both during ageing and in AD, the elevated phosphorylation of TAU protein in the sciatic nerve at Seri 99/202 leads to a loss of TAU-1 reactivity. This increased phosphorylation may be accompanied by phosphorylation of other sites found on PHF-TAU and may potentially lead to the formation of PHF in the peripheral nervous system. In order to address this question it was used the B5-2 sandwich ELISA previously developed to map neurofibrillary degeneration in the central system. The specificity of the monoclonal antibody B5-2 for PHF-TAU is based on the recognition of a conformational epitope on PHF-TAU, which involves a phosphorylation site. No increase in immunoreactivity could be observed in any of the AD cases using the B5-2 sandwich ELISA or the PHF-preparation in combination with western blotting against TAU protein. These results argue against widespread neurofibrillary degeneration within the peripheral nervous system in AD. There are reports, however, describing the occurrence of neurofibrillary structures in autonomic ganglion cells, the celiac ganglia and the upper cervical ganglia. Scientists observed the occurrence of neurofibrillary tangles and neurophil threads in autonomic ganglion cells in 5 cases out of 120 non-selective autopsy cases. Three of those five NFT-positive cases had clinical symptoms of senile dementia and neuropathological signs of AD. These reports might present rare events of neurofibrillary pathology in the peripheral nervous system.

The lack of detection of PHF-TAU in the sciatic, however, could possibly arise from the fact that neurofibrillary pathology may be confined to cell bodies and is not detectable in the axonal fibers. In our previous investigation of neurofibrillary degeneration in the CNS, however, we were able to demonstrate that the main PHF-TAU load arises from neuropil threads localized in neurites and not from NFTs restricted to cell bodies.

In summary, we can conclude that neurofibrillary degeneration is not a common denominator in the sciatic nerve of AD patients. Nevertheless, the reduction of neurofilament and TAU protein indicates a degenerative process. These pathological alterations may have a serious impact on the function of the peripheral nervous system and might be related to the neurological symptoms associated with AD. This loss of neurofilament and TAU protein does not correlate with the stage of the disease rated according to GDS. The alterations of cytoskeletal proteins in the sciatic nerve are, therefore, likely to represent a more general phenomenon associated with AD than a result of the terminal immobility of the patients. The present observation on the lack of a paired helical filament-like aggregation of TAU despite an increased state of phosphorylation indicates that at least in peripheral neurons increased TAU-phosphorylation is not necessarily accompanied by the formation of paired helical filaments. Analyzing principal differences in the expression, posttranslational modification and metabolism of TAU between central and peripheral neurons might, therefore, help to get a better insight into the mechanism of paired helical filament formation.

Source: Johannes M. Holzer and colleagues, Journal of Neural Transmission (1999).
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