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Prohibitin在姜黄素诱导人成骨肉瘤MG-63细胞凋亡过程中的调控机制
石松林 赵振利 王国红 刘凡 路锟 杨玲 韩荣 李晓 李祺福
解剖学报 ›› 2015, Vol. 46 ›› Issue (1) : 72-80.
Prohibitin在姜黄素诱导人成骨肉瘤MG-63细胞凋亡过程中的调控机制
Regulatory mechanism of prohibitin in human osteosarcoma MG-63 cells during apoptosis induced by curcumin
目的 探讨姜黄素（curcumin）诱导成骨肉瘤细胞凋亡前后，Prohibitin（PHB）蛋白的定位与表达变化，及PHB与凋亡相关基因产物的共定位关系。
方法 选择性抽提经姜黄素诱导处理前后的人成骨肉瘤MG-63细胞核基质蛋白，以蛋白质组学、免疫印迹法和激光扫描共焦显微技术对PHB在核基质中存在、分布及其与凋亡相关基因产物在MG-63细胞中的共定位关系进行了研究。结果 双向凝胶电泳、质谱鉴定和免疫印迹法结果显示，PHB存在于MG-63细胞核基质蛋白组分中，细胞凋亡后在细胞核基质蛋白中表达下调。免疫荧光显微镜观察显示，PHB定位于MG-63细胞核基质纤维上，经姜黄素处理后出现分布位置与表达水平的变化。激光扫描共焦显微镜观察结果显示，PHB在MG-63细胞凋亡过程中与Bax、Bcl-2、Fas、P53、c-Myc和Rb等基因产物具有共定位关系，且共定位区域发生了变化。 结论 PHB是一种核基质结合蛋白；PHB在MG-63细胞凋亡过程中的表达与分布变化，及其与细胞凋亡相关基因的共定位关系，对MG-63细胞凋亡具有重要影响。
Objective To investigate the expression and distribution of prohibitin in cellular nuclear matrix and the co-localization between prohibitin and the products of apoptosis-related genes in human osteosarcoma MG-63 cell before and after curcumin treatment. Methods Nuclear matrix proteins were selectively extracted and subjected to two-dimensional gel electrophoresis analysis, immunoblotting, and microscopy. Results Two-dimensional polyacrylamide Gel electrophoresis and mass spectrum analysis showed that prohibitin existed in the nuclear matrix protein components of the MG-63 cell and its expression were down-regulated by curcumin. The change of prohibitin was further confirmed by immunoblotting. The immunofluorescence microscopy observation demonstrated that the prohibitin located in the filaments of the nuclear matrix of MG-63 cell and its distribution and expression were altered after curcumin treatment. Laser scanning confocal microscopy revealed that there were co-localization between prohibitin and the products of Bax, Bcl-2, Fas, P53, c-Myc, and Rb genes in MG-63 cell, and the site of co-localization was altered by curcumin treatment. Couclusion Current research identifiys that prohibitin is a nuclear matrix protein existing in the nuclear matrix. The alteration of expression and distribution of prohibitin, and the relationship between prohibitin and multiple apoptosis-related genes may play a pivotal role in the process of human osteosarcoma MG-63 cell apoptosis. These results contribute to the elucidation of the regulatory mechanisms of tumor cellular apoptosis.
Prohibitin / 核基质 / 人成骨肉瘤MG-63细胞 / 姜黄素 / 免疫印迹法
Prohibitin / Nuclear matrix / Human osteosarcoma MG-63 cell / Curcumin
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国家自然科学基金资助项目;福建省自然科学基金
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