热休克诱导体外梅克尔软骨细胞死亡的研究

M. Kubo, K. Ishizeki
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引用次数: 1

摘要

为了明确热应激诱导的体外Meckel软骨细胞死亡是否涉及凋亡或坏死,我们使用免疫组织化学进行形态学检查,包括brdu结合、TUNEL方法、光镜和电镜。将Meckel软骨细胞从17天的胚胎小鼠中酶分离出来,暴露在40-60℃的温度下诱导细胞死亡。暴露于40-45℃亚致死温度下,BrdU的掺入促进,tunel阳性凋亡细胞出现频率高。在超微结构水平上,凋亡细胞的死亡表现为染色质凝聚、核分节和凋亡小体的形成。相比之下,致死温度(50-60°C)诱导的细胞死亡在tunel阳性细胞和含有BrdU的细胞中有所减少。高温处理的细胞细胞器膜明显破坏。热休克蛋白(HSP) -27和70的免疫染色显示,这些蛋白在较低温度下持续分泌,但在致死温度下有减少的趋势。我们的研究结果表明,Meckel软骨细胞在亚致死温度下表现出凋亡,但在致死温度下发生坏死。
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Analysis of Cell Death in Meckel's Chondrocytes In Vitro Induced by Heat-Shock Treatment
To clarify whether in vitro cell death in Meckel's chondrocytes induced by heat-stress treatment involves apoptosis or necrosis, we performed morphological examinations using immunohistochemistry including BrdU-incorporation, the TUNEL method, and light and electron microscopy. Cell death was induced in Meckel's chondrocytes isolated enzymatically from 17-day embryonic mice, by exposure to temperatures 40-60°C. On exposure t0 sublethal temperatures of 40-45°C, the incorporation of BrdU was facilitated, and TUNEL-positive apoptotic cells appeared with high frequency. At the ultrastructural level, apoptotic cell death was characterized by chromatin condensation, nuclear segmentation, and the formation of apoptotic bodies. In contrast, cell death induced by lethal temperature (50-60°C) presented decreases in TUNEL-positive cells and cell incorporating BrdU. High temperature-treated cells showed apparent disruption of the membrane of cell organelles. Immunostaining for heat shock proteins (HSP) -27 and-70 revealed that these proteins are secreted continuously at lower temperatures, but have a tendency toward decrease on exposure to lethal temperatures. Our results suggest that Meckel's chondrocytes demonstrate apoptosis at sublethal temperatures, but undergo necrosis at lethal temperatures.
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