褐藻多糖硫酸酯对6-羟基多巴胺所致MN9D细胞损伤的保护作用及其机制  

Neuroprotective effect of fucoidan against 6-OHDA-induced damage and its mechanism on MNgD cell line

作　　者：罗鼎真[1,2] 张全斌[3] 崔艳秋[1] 王晓民[1] 

机构地区：[1]首都医科大学教育部神经变性病学重点实验室,北京100069 [2]山东大学附属省立医院老年神经科,济南250021 [3]中国科学院青岛海洋研究所,青岛266071

出　　处：《中华神经医学杂志》2010年第3期217-221,共5页Chinese Journal of Neuromedicine

基　　金：基金项目：国家973计划项目（2006CB500700）

摘　　要：目的以6一羟基多巴胺（6一OHDA）为工具药，建立帕金森病（PD）的细胞模型，观察褐藻多糖硫酸酯（Fuc）对细胞模型的保护作用并初步探讨其作用机制。方法用不同浓度（12．5、25、50、100、200μmol／L）的6-OHDA处理MN9D细胞24h，挑选出合适浓度50μmol／L。再以50μmolfL6-OHDA处理MN9D细胞不同时间（6、12、24及48h），建立细胞损伤模型，挑选出合适时间24h。用0．01、0．1、1．0mg／mLFuc预孵育MN9D细胞1h后加入50μmol／L6-OHDA共同作用24h，以探讨Fuc的保护作用。MTT法检测细胞存活率，生化法测定乳酸脱氢酶（LDH）释放量．二氯荧光素乙二酯（DCF—DA）染色法检测细胞内的氧化应激水平。结果随着6．OHDA浓度增加或作用时间延长，MN9D细胞MTT值逐渐降低。50μmol/L6．OHDA处理细胞24h，MTT值明显下降，LDH释放量增加。而0．1、1．0mg／mL的Fuc预孵育1h可明显减轻MN9D细胞的损伤，提高MTT值并降低LDH释放量，细胞形态学改变与生化实验结果一致。50μmol/L6-OHDA作用8h可明显升高MN9D细胞内的氧化应激水平，而1．0mg／mL的Fuc预处理1h可以拮抗6-OHDA引起的细胞内氧化应激水平增高。结论Fuc可以有效的拮抗6-OHDA对MN9D细胞的损伤作用，其作用机制可能与抗氧化活性有关。Objective To study the neuroprotective effect offucoidan against 6-OHDA-induced damage on MN9D cell line and preliminarily explore its mechanism. Methods The MN9D cells were treated with different concentrations of 6-OHDA （12.5, 25, 50, 100 and 200 μmol/L） for 24 h and with 50 μmol/L 6-OHDA for different time courses （6, 12, 24 and 48 h） as well. Based on the above experiment, the best concentration （50 μmol/L） of 6-OHDA and time course （24 h） were selected and the damaged model of MN9D cell line was induced accordingly. The MN9D cells were pretreated with 0.01, 0.1 and 1.0 mg/mL fucoidan for 1 h, and then co-treated with 50 μmol/L 6-OHDA for 24 h. The viability of MN9D cells was detected by MTT assay; the LDH leakage was detected by bioassay; the level of intracellular xidative stress was observed by 2＇,7＇-dichlorodihydrofluorescein diacetate assay. Results MTT metabolic value decreased following the increase of 6-OHDA concentration or the extension of processing time. Treatmem with 50 μmol/L 6-OHDA for 24 h induced a significant decrease of MTT metabolic rate and an increase of LDH leakage rate in the MN9D cells. Pretreatment with 0.1 and 1.0 mg/mL fucoidan inhibited the death of 6-OHDA-induced cells, increased the MTT metabolic value and decreased the LDH leakage. Cell morphological changes were consistent with the biochemicalexperiments. Treatment with 50 μmol/L 6-OHDA for 8 h could significantly increased the generation of reactive oxygen species induced by 6-OHDA, while pretreatment with 1.0 mg/mL fucoidan for 1 h showed antagonism on that. Conclusion Fucoidan can protect MN9D cells from the death induced by 6-OHDA and the underlying mechanism may be involved in the antioxidant capacity of fucoidan.

关 键 词：帕金森病 6-羟基多巴胺 褐藻多糖硫酸酯 氧化应激 

分 类 号：R742.5[医药卫生—神经病学与精神病学]