热应激对慢性粒细胞白血病细胞系K562中热休克蛋白gp96表达的影响及其意义

Effect of Heat Stress on Expression of gp96 in K562 Cell Line of the Chronic Myeloid Leukemia and Its Significance

本研究查明不同热应激条件对慢性粒细胞白血病细胞系K562细胞中热休克蛋白gp96表达量的影响,为寻找增加K562细胞中gp96的提取量的最佳热应激条件提供实验依据。用免疫组织化学方法检测经38、40、42、44、46及48℃水浴热休克处理30分钟后的K562细胞中gp96表达量的变化;以平均阳性细胞百分率和平均荧光强度为指标,用流式细胞术研究40、44、48及52℃各水浴热休克处理30分钟对K562细胞中gp96表达量的影响;利用免疫印记原理,用Westernblot技术研究K562细胞经40、44、48与52℃水浴各休克处理30分钟后gp96表达量的变化。结果表明免疫组织化学方法显示gp96主要在胞浆内表达,在48℃水浴热休克处理30分钟时,强阳性的细胞数最多,积分也最高;流式细胞术显示平均阳性细胞百分率和平均荧光强度均在48℃水浴热休克处理30分钟组达高峰,各温度间gp96表达量的差异有统计学意义;Westernblot显示48℃水浴热休克处理30分钟时gp96的表达量达到高峰,各温度间gp96表达量的差异有统计学意义。结论48℃水浴热休克处理30分钟,gp96在K562细胞中的表达量达到高峰,该条件可作为提取K562细胞gp96的一个有效预处理条件。

This study was purposed to investigate the effect of different heat stress conditions on expression level of heat shock protein gp96 in K562 cell line of chronic myeloid leukemia in order to provide experiment basis for seeking optimal heat stress condition increasing extraction amount of gp96 from K562 cells. The expression changes of gp96 in K562 cell line was detected by immunocytochemistry under 38,40,42,44,46 and 48℃ for 30 minutes in water, by flow cytometry under 40,44,48 and 52℃ for 30 minutes in water, by Western blot under 40,44,48 and 52 ℃ for 30 minutes in water. Immunocytochemistry assay showed that gp96 existed mainly in cytoplasm. The peak of gp96 expression was at 30 minutes after 48℃ in water. The result of flow cytometry was consistent to immunocytochemitry detection results under temperatures 40,44,48 and 52~C （ P 〈 0. 01 ）. Western blot showed that detection result was the same as the immunocytochemistry and flow cytometry detections. In conclusion, the expression of gp96 in K562 cell line reached peak at 30 minutes after 48℃ in water. This condition may be an effective preparative condition for extraction of gp96 from K562 cells.