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目的研究2-甲氧基雌二醇(2ME2)诱导骨髓瘤细胞系分化的分子机制。方法应用细胞形态学方法、流式细胞术分析、ELISA法和RT-PCR方法,观察2ME2对于骨髓瘤细胞系LP-1、CZ-1、NCI-H929细胞分化的影响,同时检测细胞分化时X盒结合蛋白1(XBP-1)mRNA的改变情况。继而采用XBP-1、Blimp-1、pax-5硫代反义寡核苷酸(ASODN)作为干预手段,观察其对于2ME2诱导骨髓瘤细胞系分化作用的影响及mRNA水平的变化。结果2ME2可诱导骨髓瘤细胞系LP-1、CZ-1、NCI-H929细胞向成熟阶段分化,伴随着细胞分化的过程XBP-1mRNA的表达水平上调。XBP-1ASODN和Blimp-1ASODN可以部分抑制2ME2促骨髓瘤细胞系分化作用,而pax-5ASODN可以加强2ME2促骨髓瘤细胞系分化作用。经pax-5ASODN作用72h,经形态学观察,细胞向成熟阶段分化,表现为细胞胞核缩小,胞浆丰富,核浆比例下降,核仁减少或消失,核染色质变粗、变密;细胞表面标志CD49e表达率增高;同时细胞分泌轻链蛋白升高。经Blimp-1ASODN作用72h后,pax-5mRNA水平显著升高;而XBP-1mRNA水平下降。结论2ME2可诱导骨髓瘤细胞系LP-1、CZ-1、NCI-H929细胞向成熟阶段分化,伴随着细胞的分化XBP-1mRNA的表达水平上调。Blimp-1可通过抑制pax-5mRNA的表达上调XBP-1mRNA的表达水平,促进骨髓瘤细胞分化。
Objective To investigate the molecular mechanism of 2-methoxyestradiol (2ME2) -induced myeloma cell line differentiation. Methods Cell morphology, flow cytometry, ELISA and RT-PCR methods were used to observe the effect of 2ME2 on the differentiation of myeloma cell line LP-1, CZ-1 and NCI-H929 cells. Box binding protein 1 (XBP-1) mRNA changes. Subsequently, XBP-1, Blimp-1, pax-5 antisense oligodeoxynucleotides (ASODN) were used as an intervention to observe the effect of ASBPN on the differentiation of 2ME2-induced myeloma cell lines and the changes of mRNA levels. Results 2ME2 induced the differentiation of myeloma cell lines LP-1, CZ-1 and NCI-H929 into mature stage, and the expression of XBP-1 mRNA was up-regulated during cell differentiation. XBP-1 ASODN and Blimp-1 ASODN can partially inhibit 2ME2-promoting myeloma cell line differentiation, while pax-5ASODN can enhance the differentiation of 2ME2-promoting myeloma cell line. The morphological observation showed that the cells differentiated into mature stage after being treated with pax-5ASODN for a long time. The cell nucleus was reduced, the cytoplasm was rich, the ratio of nuclear plasma reduced, the nucleolus was reduced or disappeared, and the chromatin became thicker and denser. Marked CD49e expression increased; at the same time the cells secreted light chain protein increased. After 72h treatment with Blimp-1 ASODN, the level of pax-5 mRNA increased significantly, while the level of XBP-1 mRNA decreased. Conclusion 2ME2 can induce myeloma cell line LP-1, CZ-1 and NCI-H929 to differentiate into mature stage with the up-regulated expression of XBP-1 mRNA. Blimp-1 can promote the differentiation of myeloma cells by up-regulating the expression of XBP-1 mRNA by inhibiting the expression of pax-5 mRNA.