目的 研究去甲斑蝥素聯(lián)合馬法蘭對多發(fā)性骨髓瘤細胞增殖與凋亡的影響及其作用機制。方法 去甲斑蝥素單獨或聯(lián)合馬法蘭給藥后，MTT法觀察對人多發(fā)性骨髓瘤細胞U266株生長的抑制作用，流式細胞術(shù)檢測細胞凋亡率，Western blotting檢測核因子-κB P65（NF-κB P65）、NF-κB抑制因子IκBα、磷酸化IκBα（p-IκBα）、survivin、Bcl-2和Bax蛋白表達的影響。制備多發(fā)性骨髓瘤小鼠模型，觀察去甲斑蝥素及其與馬法蘭聯(lián)合給藥對腫瘤生長的抑制作用。結(jié)果 去甲斑蝥素能增強馬法蘭的細胞毒和誘導(dǎo)細胞凋亡作用。與馬法蘭單獨給藥組比較，聯(lián)合給藥組使U266細胞核NF-κB P65和胞漿p-IκBα的表達量分別由1.13±0.08、0.83±0.08降至0.26±0.02、0.090±0.002；馬法蘭對IκBα的表達無影響，但去甲斑蝥素能促進IκBα的表達，二者合用能顯著促進胞漿IκBα的表達；與馬法蘭單獨給藥組比較，聯(lián)合給藥組survivin和Bcl-2的表達量分別由1.03±0.10、0.72±0.05降至0.52±0.04、0.06±0.01，而Bax由0.29±0.03升至0.75±0.06。體內(nèi)實驗也證實去甲斑蝥素能增強馬法蘭的抗腫瘤作用。結(jié)論 去甲斑蝥素通過抑制NF-κB/p-IκBα途徑，調(diào)節(jié)下游信號分子survivin、Bcl-2和Bax的表達，增強馬法蘭的抗骨髓瘤作用。

Abstract: Objective To investigate the effects and mechanisms of norcantharidin (NCTD) combined with melphalan (Mel) on proliferation and apoptosis of multiple myeloma (MM) cells. Methods Human MM cell line U266 cells were treated with NCTD alone or in combination with Mel. MTT and Annexin V/PI flow cytometry were used to determine the rates of cell viability and apoptosis. The protein expression of nuclear factor-κB P65 (NF-κB P65), NF-κB P65 inhibitor IkBα, phosphorylated IκBα (p-IκBα), survivin, Bcl-2, and Bax was determined by Western blotting. The model of mice with MM was established, and the inhibition of combination of NCTD and Mel on tumor growth was observed. Results NCTD potentiated the cytotoxicity and pro-apoptotic effects induced by Mel. Compared with the group treated with Mel alone, the expression levels of NF-κB P65 in U266 nucleus and p-IκBα in cytoplasm in NCTD and Mel combination group decreased from 1.13 ± 0.08 and 0.83 ± 0.08 to 0.26 ± 0.02 and 0.090 ± 0.002, respectively. Mel showed no effect on IκBα expression, but NCTD could enhance its expression, and the combination of NCTD and Mel could obviously enhance the expression. The expression of survivin and Bcl-2 decreased from 1.03 ± 0.10 and 0.72 ± 0.05 to 0.52 ± 0.04 and 0.06 ± 0.01, while the expression of Bax increased from 0.29 ± 0.03 to 0.75 ± 0.06 respectively. In vivo results also demonstrated that NCTD could increase the antitumor effects of Mel. Conclusion The results suggest that NCTD could potentialize the anti-MM effects through inhibiting NF-κB/p-IκBα signal pathway and regulating the espression of survivin, Bcl-2, and Bax.

河北省中醫(yī)藥管理局資助項目（2007136）