| Summary: | 碩士 === 國立陽明大學 === 生物化學研究所 === 88 === It is generally thought that the death of dopaminergic neurons induced by dopamine is due to the oxidative stress produced by dopamine. However, the mechanism involved is not clear. It has been shown that dopamine induced the death of various types of cells. In this study, PC12 cell, a dopamine-containing cell line derived from rat pheochromocytoma, was used as a model system to study the action mechanism of dopamine. Among the various types of cells examined, C6 glioma cells, 3T3 fibroblasts and PC12 cells, PC12 cells are most sensitive to 250 microM dopamine. The sensitivity of PC12 cells to dopamine decreased as the cell density increased. NO donor, such as hydroxylamine and sodium nitroprusside, enhanced the effect of dopamine. In contrast, the dopamine effect was partially reversed by pre-loading PC12 cells with 15microM BAPTA, a Ca2+ chelator. The glyceraldehyde-3-phosphate dehydrogenase was increased when PC12 cells were treated with dopamine.This study provide evidence to show that dopamine-induced cell death was enhanced by activation of Rac protein. Results indicated that Zn2+ enhanced dopamine toxicity and Zn2+accumulated in the vesicles upon entering the cell.
By co-culturing with a cell line derived from human microglial cells, C13NJ, the PC12 cell death induced by 250 microM dopamine was completely inhibited. The protective effect of co-culture with C13NJ cells did not require the contact between the two types of cells because the conditioned medium (C.M.) exhibited similar protective effects. In addition to the dopamine-induced cell death, the C.M.partially protected the PC12 cell death induced by ceramide and A23187, however the C.M. could not protect PC12 cell from staurosporine attact. Moreover I suggest that the protection from C.M. was not due to reduce the intracellular free-radical level. The C.M. obtained from murine J774 macrophages, C6 cells, 3T3 cells and PC12 cells showed similar protective effect, however, it required higher cell number than C13NJ cells to obtain the C.M. I have partially characterized the molecular identity of the factor; it was very polar, carried negative charge, heat-stable (70°C, 30 min), and its molecular weight was smaller than 500 Da. It appears that cells may secrete some factor(s) that protects PC12 cells from being damaged by dopamine. My results provide a further understanding of the mechanisms involved in the dopamine-induced cell death and may help to find ways to prevent the cells from unwanted death.
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參考文獻………………………..…………… 72
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