| Summary: | 碩士 === 中國醫藥大學 === 醫學研究所 === 93 === Recent studies indicate tumor necrosis factor-alpha（TNF-α）is associated with Parkinson's disease（PD）. However whether TNF-α plays a neuron protective or neuron destructive role remains to be elucidated. In addition, the mechanism is still not clear. To determine the role of TNF-α in PD, we studied neurotoxic effect on dopaminergic neurons（PC-12 cells）induced by 1-methyl-4-phenyl-pyridinium（MPP+ ions；MPP+）in the presence or absence of TNF-α. PC-12 cells were either treated with MPP+ alone（600, 300, 150, 75 and 0μM） or in combination with TNF-α（15 or 30 ng/ml） for 24 hours and then assayed for cell viability. MTT Assay showed MPP+ treatment increased cell death in a dose-dependent manner. However, death rates were statistically higher in MPP+ plus TNF-α groups than those treated with either MPP+ or TNF-α only. Morphologically, we also observed similar trends. In addition, results from DAPI staining indicated the cell death was apoptotic. We further investigated possible signal molecules involved. Data from western blot analysis showed that caspase-3 and JNK activation increased when treated with MPP+ alone; and the increases were even higher in MPP+ plus TNF-α treated cells. However, the amount of phosphorylated c-Jun was not elevated in all treated
5
groups. We then detected the activation of JNK. Data from kinase assay showed JNK activity increased clearly in MPP+ plus TNF-α treatment cells. We later used Z-VAD （a caspase inhibitor） and a JNK inhibitor （SP600125） to determine whether blockade of caspase-3 and JNK activation can inhibit apoptosis. Data from TUNEL assay demonstrated caspase inhibition prevented the cell death efficiently. However, JNK inhibition only slightly prevented the cell death slightly.
Our data suggest that MPP+ caused cell apoptosis through caspase-3 activation, and the enhanced toxicity induced by the addition of TNF-α is aiso related to caspase-3 activation. In addition, JNK activation was involved in the intricate signaling pathways. Nevertheless, JNK activation may partially involved in the induction of apoptosis. Further investigation will be needed to elucidate the role of JNK activation in this apoptosis.
|
|---|
