小白菊内酯对不同环氧合酶-2表达的鼻咽癌细胞株中干性侧群细胞转归的影响

Effects of parthenolide on the cellular outcomes of stem-like side population cells in nasopharyngeal carcinoma cell lines with different level of COX-2

目的:探讨倍半萜烯内酯(sesquiterpene Iactones,SLs)类化合物对不同环氧合酶-2(cyclooxygenase-2,COX-2)表达水平的人鼻咽癌(nasopharyngeal carcinoma,NPC)细胞中癌干细胞(cancer stem cells,CSCs)转归的影响。方法:分别采用实时定量PCR(qRT-PCR)和Western blot检测人鼻咽癌高分化CNE1L和低分化CNE2L细胞株中COX-2 mRNA和蛋白本底表达水平。给予不同剂量SLs活性成分小白菊内酯(parthenolide,PN)处理2种细胞株后,再分别采用噻唑蓝(MTT)法测定细胞增殖活性;qRT-PCR检测细胞中干性相关基因COX-2、ABCG2、BMI1、OCT4 mRNA水平;流式细胞术分析干性侧群(side population,SP)细胞的相对含量;平板克隆集落形成实验检测干性细胞的自我更新和分化增殖能力。结果:CNE1L和CNE2L细胞中COX-2 mRNA和蛋白本底水平显著不同,在CNE1L细胞中明显高于CNE2L细胞(P<0.05);与对照组比较,PN处理后,细胞增殖抑制率随PN浓度(10～100μmol/L)增加呈剂量依赖性增高(P<0.05),细胞中COX-2 mRNA在CNE1L中升高、在CNE2L中降低,干性相关基因ABCG2、BMI1 mRNA在CNE1L细胞中升高、而OCT4 mRNA在两株细胞中均降低(P均<0.05);SP细胞抑制率亦随PN浓度(2.5～40μmol/L作用24 h)增加呈剂量依赖性增高(P<0.05);而细胞的集落形成能力随PN浓度(0.25～1.5μmol/L作用72 h)增加显著降低(P<0.05);上述结果在CNE1L和CNE2L细胞株间差异均有统计学意义(P<0.05),在CNE1L细胞中变化作用更强。结论:PN对不同NPC细胞株中的干性细胞功能具有抑制作用且存在明显差别,可能与不同细胞间COX-2表达的差异性有关。

OBJECTIVE：To investigate effects of the sesquiterpene lactones（SLs） compound on the induced outcomes of cancer stem cells（CSCs） in human nasopharyngeal carcinoma（NPC） cells with different levels of cyclooxygenase-2（COX-2）.METHODS：The background levels of COX-2 mRNA and COX-2 protein were determined by the real time PCR（qRT-PCR） and Western blot in CNEIL and CNE2L cell lines.After treated with different doses of parthenolide（PN）,proliferation of cells was determined by MTT assay,the transcription levels of COX-2,ABCG2, BMI1,OCT4 mRNA were determined by qRT-PCR,the relative content of side population（SP） cells was examined by flow cytometry,and the self-renewal and differentiation proliferation of stem-like cells were measured by colony formation assay.RESULTS：The background levels of COX-2 mRNA and protein were significantly higher in CNE1L compared with CNE2L cells（P〈0.05）.Compared with control group,in PN-treated group the proliferation inhibition rate was increased in dose-dependent（10-100μmol/L for 24 h） manner（P〈0.05）.The mRNA level of stemness relative genes,including COX-2,ABCG2,BMI1 mRNA was increased in CNEIL cells,but COX-2 mRNA was decreased in CNE2L cells,while the level of OCT4 mRNA was reduced in both cell lines.The SP inhibition rate（%） was also increased with dose-dependent pattern（2.5-40μmol/L for 24 h）（P〈0.05）,and the colony formation was significantly lower（0.25- 1.5μmol/L for 72 h）（P〈0.05）.All the results above were statistically different between CNE1L and CNE2L （P〈0.05）,with effects in CNE1L cell more remarkable.CONCLUSION：There were significantly different effects of PN on the inhibition of CSCs function in different NPC cells.The varied expression of COX-2 between cell lines may be related to the susceptibility.