| Summary: | 碩士 === 國立成功大學 === 細胞生物及解剖學研究所 === 92 === Abstract
The pathological hallmarks of Alzheimer’s disease (AD) are the deposition of neuritic plaques extracellularly and accumulation of neurofibrillary tangles intracellularly in the patient’s brain. Extensive studies have demonstrated that neuritic plaques are usually surrounded by activated microglia and astrocytes. The major component of neuritic plaques is the 40 to 42 amino acid long �-amyloid peptide (A�). The abnormal Aβ aggregations in brain could induce various pathologies, such as dystrophic neuritis, synaptic losses, and retraction of the neuritic arbor strongly, which all suggest a pivotal role of Aβ in the AD pathogenesis. Furthermore, A� may exert indirect neuron toxicity through the activation of microglia. Some studies suggest that activated microglia cells might be the link between A� deposition and neuronal degeneration. Therefore, the first objective of this study was to evaluate Aβ peptide-induced neuronal toxicity in microglia-like THP-1 cells. Human neuroblastoma cells, SH-SY5Y, were used to monitor the neurotoxicity of THP-1 secreted neurotoxins. Our results showed that A� was capable of inducing SH-SY5Y cell death in a dose-dependent manner with a LD50 = 0.4�M of A�, whereas the THP-1 cells were spared. When the THP-1 cells were treated with 200nM of Aβ, the THP-1 condition media (CM) demonstrated neurotoxicity with a maximal effect at 3h. To avoid the confounding effects in serum, the culture medium was switched from the serum-containing medium to serum-free medium. In the serum-free culture condition, A� induced SH-SY5Y cell death in a dose-dependent manner (LD50 = 0.15�M). Such effects were not observed in THP-1 cells. THP-1 cells were then treated with different concentrations of A� for 15 min and the THP-1 CM was transferred to SH-SY5Y cells for 24h. I found that the viabilities of SH-SY5Y cells decreased as the THP-1 cell treated Aβ concentrations increased. These results suggested that microglia can release neurotixins upon the stimulation by Aβ oligomers and prefibrillar aggregated were the proximal effectors of neurotoxicity. The second objective of my study was to identify the A� oligomer species that responsible for its neurotoxicity. A� peptides were separated into different molecular weight fractions using graded membrane filtration and the toxicity of each fraction was examined. My results showed that soluble form of A� ranging over 30 kDa was the major A� species causing the toxicity.
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