| Summary: | 碩士 === 國立陽明大學 === 神經科學研究所 === 94 === Histopathological features in AD brain include the extracellular senile plaques, intracellular neurofibrillary tangles, synapses loss, neuronal cell loss and reactive glial cell around the plaques. Senile plaques are extracellular protein aggregation mainly composed of full-length of b-amyloid peptides containing 40 to 42 amino acids (Ab1-40 and Ab1-42). Activated astrocytes have been found in the vicinity of senile plaques in AD brain and suggested to be correlated with the formation of senile plaques. Apoptotic astrocytes are found in white matter lesions in AD postmortem brain.
Serum deprivation is a common pretreatment used for astrocyte studies. Serum deprivation sensitized astrocytes to oxidative stresses. In our study, primary astrocytes were serum deprived. Serum deprivation alone changed primary astrocytes from flattened and polygonal to process-bearing stellate cells, which look like activated astrocytes distributed around the senile plaques. More process-bearing were occurred by Ab25-35 under serum deprivation. The morphological observation suggested that Ab25-35 augmented the activation of primary astrocytes under serum deprivation. The purpose in our study was to investigate whether Ab25-35 augments cell death in primary astrocytes under serum deprivation. Necrosis was observed by PI staining and quantified by the release of LDH. Serum deprivation alone induced cell death, and Ab25-35 significantly increased necrosis under serum deprivation. Apoptosis was triggered by serum deprivation and Ab25-35 augmented DNA fragmentation. However, the cell number revealed by trypan blue and WST1 assay was not decreased by Ab25-35. The number of condensed nuclei were significantly increased by Ab25-35 for 36 h. Double staining in Hoechst 33258 with PI revealed that all necrotic astrocytes (PI positive) were colocalized with condensed nuclei, but the condensed nuclei could also under proliferation as shown by staining of Hoechst 33258 with Ki67. Consistently, DNA synthesis detected by [3H] thymidine incorporation was also increased by Ab25-35. Our observations suggested that cell death and DNA synthesis occurred simultaneously. However, there was no difference in mitochondrial membrane potential by Ab25-35. Although serum deprivation alone induced obvious effects on morphological changes, cell death, and aberrant cell cycle, Ab25-35 augmented cell death in primary astrocytes under serum deprivation.
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