HSP27高表达与人白血病多药耐药细胞K562/VCR的关系

背景与目的:白血病细胞多药耐药(multidrug resistance,MDR)是白血病化疗失败的常见原因,虽然已有研究揭示了一些肿瘤MDR机制,但目前仍然不能完全解释MDR现象。本文旨在应用蛋白质组学方法筛选白血病耐药相关蛋白,并研究其与白血病MDR的关系,为进一步阐明白血病MDR发生的分子机制提供理论依据。方法:二维聚丙烯酰胺凝胶电泳(two-dimensional electrophoresis,2-DE)技术分离白血病细胞K562和人白血病耐药细胞K562/VCR的总蛋白,用基质辅助激光解吸电离飞行时间质谱(matrix assisted laser desorption/ionization-time offlight-mass spectrometry,MALDI-TOF-MS)对差异表达的蛋白质点进行鉴定。应用反义核酸技术将分离鉴定差异表达蛋白的反义核酸转染至耐药细胞,Western blot检测转染后差异蛋白的表达情况,MTT法检测转染后细胞存活率,流式细胞仪检测转染后细胞凋亡率。结果:在K562/VCR细胞与K562细胞中鉴定出一差异表达蛋白点,并用质谱分析证实其为热休克蛋白27(heat shock protein27,HSP27)。用HSP27反义核酸转染K562/VCR细胞,在不同浓度长春新碱作用下,HSP27反义核酸转染的K562/VCR细胞的存活率较对照错义核酸转染组明显降低(P<0.05),流式细胞仪显示细胞凋亡率为16.37%,明显高于对照组(P<0.05)。结论:HSP27在K562/VCR细胞中高表达,其表达抑制后,K562/VCR细胞对长春新碱的敏感性增强。

Correlation of high expression of HSP27 to multidrug resistance of leukemia cell line K562/VCR

BACKGROUND & OBJECTIVE: Multidrug resistance (MDR) is a major obstacle preventing effective treatment of leukemia. The mechanisms of MDR in leukemic cells have been broadly explored, but they are still unclear. We used proteomic tools to screen MDR-related proteins in vincristine-resistant leukemia cell line K562/VCR, and analyzed the mechanism of MDR in leukemia. METHODS: Two-dimensional electrophoresis (2-DE) was used to extract total proteins from K562/VCR and K562 cells. The proteins expressed differentially between the two cell lines were identified by matrix-assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS). The antisense oligonucleotide (ASO) of the protein was transfected into K562/VCR cells; mis-sense oligonucleotide (MSO) of the protein was also transfected as control. The expression of the protein was detected by Western blot. Cell survival was detected by MTT assay. Cell apoptosis was detected by flow cytometry (FCM). RESULTS: Heat shock protein 27 (HSP27), a differential expression protein between K562/VCR and K562 cells, was identified. When treated with vincristine, the survival rate of K562/VCR cells was significantly lower in HSP27 ASO group than in HSP27 MSO group (P<0.05). The apoptosis rate of K562/VCR cells was significantly higher in HSP27 ASO group than in HSP27 MSO group (16.37% vs. 3.08%, P<0.05). CONCLUSION: HSP27 is highly expressed in K562/VCR cells, and the suppression of its expression by HSP27 ASO could enhance chemosensitivity of K562/VCR cells to vincristine.