目的：探讨HT22细胞氧糖剥夺再灌注及Grasp65过表达干预后高尔基体的形态变化及其可能机制。方法利用小鼠海马神经元细胞系HT22为研究对象，HT22细胞经氧糖剥夺再灌注损伤及Grasp65过表达干预后，采用MTT法检测细胞存活率；Hoechest33258荧光染色法评估细胞凋亡；并应用细胞免疫荧光技术观察高尔基体的形态；应用Western blot技术检测GM130和GAAP蛋白的表达。结果氧糖剥夺再灌注可导致HT22细胞的活性显著降低（P＜0．05），凋亡率显著增高（P＜0．05）；并可导致高尔基体形态的异常，随着再灌注时间的延长，高尔基体逐渐发生碎裂，尤其以再灌注12 h和24 h最为明显；GM130、GAAP的表达水平在氧糖剥夺再灌注后出现下降，特别是在再灌注12 h、24 h后出现了显著下降（P＜0．05）。过表达Grasp65后，HT22细胞在氧糖剥夺再灌注所致高尔基体碎裂出现减少（P＜0．05），碎裂程度减轻，同时GM130和GAAP的表达均显著增加（P＜0．05），HT22细胞的存活率大大提高（P＜0．05），凋亡率显著降低（P＜0．05）。结论缺血再灌注损伤的细胞模型中，同样发生了高尔基体的碎裂；过表达Grasp65可以减轻氧糖剥夺再灌注损伤所致的高尔基体碎裂，并可以减少HT22细胞的凋亡，其机制可能与上调GM130和GAAP的表达有关。%Objecive To investigate Morphological alterations of Golgi apparatus after oxygen-glucose deprivation followed by reperfusion (OGD/R) and intervening by over-expressed Grasp65 in HT22 cells and their underlying mecha-nism.Methods Using mouse hippocampal neuronal cell line HT 22 as the research object .After OGD/R and intervening by over-expressed Grasp65,using MTT method to detect cell viability and hoechest 33258 fluorescence staining method to e-valuate cell apoptosis .Morphology of Golgi apparatus were observed with cytochemistry immunofluorescence .The protein ex-pressions of GM130 and GAAP were detected by Western blot .Results HT22 cells activity was significantly decreased ( P<0.05) via OGD/R induction and apoptosis rate increased significantly (P<0.05).OGD/R could also lead to abnormal Golgi morphology .Along with the reperfusion time extended ,the Golgi apparatus gradually fragmented ,it was the most obvi-ous at 12 h and 24 h of reperfusion .The protein expression levels of GM 130 and GAAP were decreased significantly at 12 h and 24 h time points after OGD/R (P<0.05).Performing OGD/R treatment after over-expressed Grasp65 in HT22 cells, Golgi fragmentation were significantly decreased (P<0.05) and the degree of fragmentation alleviated .The protein expres-sions of GM130 and GAAP were significantly increased (P<0.05).HT22 cells survival rate was greatly improved (P<0.05) and apoptosis rate decreased significantly (P<0.05).Conclusions In the cell model of ischemia reperfusion inju-ry,the same happened to fragmentation of the Golgi apparatus .Over-expression of Grasp65 could alleviate OGD/R-induced Golgi fragmentation and reduce HT 22 cells apoptosis ,possibly related to the up-regulation of the expression of GM 130 and GAAP.
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