三氧化二砷对人多发性骨髓瘤KM3细胞凋亡和染色体区域稳定蛋白mRNA表达的影响
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摘要
目的:探讨中药"砒霜"的主要成分三氧化二砷(As_2O_3)对人多发性骨髓瘤KM3细胞株增殖与凋亡的影响及其可能的作用机制。方法:培养KM3细胞株,采用CCK-8法检测不同浓度As_2O_3对KM3细胞株增殖的影响,绘制增殖抑制曲线;根据CCK-8结果,选用10、20、30μmol/L浓度的As_2O_3作用于KM3细胞,然后以流式细胞术检测不同浓度As_2O_3诱导KM3细胞凋亡情况,以实时定量PCR方法检测干预后KM3细胞染色体区域稳定蛋白CRM1 mRNA表达情况。结果:As_2O_3可抑制KM3细胞增殖,且具有明显的时间和剂量效应;不同浓度梯度的As_2O_3均能诱导KM3细胞凋亡,与对照组相比均具有显著差异,其中30μmol/L浓度作用后细胞凋亡率明显高于其余两个浓度(P<0.05);CRM1 mRNA表达水平在各浓度梯度中均有所下降,与对照组相比均具有显著差异,而30μmol/L浓度组与其余两个浓度梯度组相比下调CRM1 mRNA表达的作用更强(P<0.05)。结论:As_2O_3可抑制KM3细胞增殖并诱导其凋亡,其作用机制可能与下调CRM1 mRNA的表达有关。
Objective: To explore the effect of arsenic trioxide(As_2O_3), a main ingredient of white arsenic, on the proliferation and apoptosis of the KM3 cell line of human multiple myeloma and its possible mechanism. Methods: KM3 cell lines were cultured and the effects of arsenic trioxide on the proliferation of KM3 cell lines were detected by CCK-8, and the proliferation inhibition curve was plotted. According to cck-8 results, the KM3 cells were intervened in 10, 20, 30μmol/L concentrations of arsenic trioxide, then, the cell apoptosis of KM3 cells was detected by flow cytometry, and the mRNA expression of CRM1 in KM3 cell chromosome area was detected by real-time quantitative PCR method. Results: Arsenic trioxide inhibited KM3 cell proliferation and had obvious time and dose effects. The apoptosis of KM3 cells was induced by arsenic trioxide of different concentration gradients and had statistically different from the control group. The cell apoptosis rate in 30μmol/L concentration was significantly higher than the other concentration gradient(P<0.05). The mRNA expression of CRM1, a stability protein in chromosome region, was decreased in each concentration gradient. Compared with the control group, there was a statistical difference(P<0.05). The effect of down-regulating CRM1 mRNA expression was stronger in 30μmol/L concentration than the other two concentration gradient groups(P<0.05). Conclusion: Arsenic trioxide could inhibit the proliferation of KM3 cells and induces apoptosis, and its mechanism may be related to the down-regulating CRM1 mRNA expression, a stability protein in chromosome region.
Objective: To explore the effect of arsenic trioxide(As_2O_3), a main ingredient of white arsenic, on the proliferation and apoptosis of the KM3 cell line of human multiple myeloma and its possible mechanism. Methods: KM3 cell lines were cultured and the effects of arsenic trioxide on the proliferation of KM3 cell lines were detected by CCK-8, and the proliferation inhibition curve was plotted. According to cck-8 results, the KM3 cells were intervened in 10, 20, 30μmol/L concentrations of arsenic trioxide, then, the cell apoptosis of KM3 cells was detected by flow cytometry, and the mRNA expression of CRM1 in KM3 cell chromosome area was detected by real-time quantitative PCR method. Results: Arsenic trioxide inhibited KM3 cell proliferation and had obvious time and dose effects. The apoptosis of KM3 cells was induced by arsenic trioxide of different concentration gradients and had statistically different from the control group. The cell apoptosis rate in 30μmol/L concentration was significantly higher than the other concentration gradient(P<0.05). The mRNA expression of CRM1, a stability protein in chromosome region, was decreased in each concentration gradient. Compared with the control group, there was a statistical difference(P<0.05). The effect of down-regulating CRM1 mRNA expression was stronger in 30μmol/L concentration than the other two concentration gradient groups(P<0.05). Conclusion: Arsenic trioxide could inhibit the proliferation of KM3 cells and induces apoptosis, and its mechanism may be related to the down-regulating CRM1 mRNA expression, a stability protein in chromosome region.
引文
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[16]王清,李娟,谷景立,等.Bortezomib增强三氧化二砷抑制KM3细胞生长和促进凋亡机制的研究.中国病理生理杂志,2010,26(2):297-301
[17]Kevin T Nguyen,Michael P Holloway,Rachel A Altura.The CRM1nuclear export protein in normal development and disease.Int J Biochem Mol Biol,2012,3(2):137-151
[18]Sun H,Lin D C,Cao Q, et al.CRM1 inhibition promotes cytotoxicity in Ewing sarcoma cells by repressing EWS-FLI1-dependent IGF-1signaling.Cancer Research,2016,76(9):2687
[19]Fabi F,Adam P,Vincent K,et al.Inhibition of CRM1 activity sensitizes endometrial and ovarian cell lines to TRAIL-induced cell death.Cell Communication&Signaling Ccs,2018,16(1):39
[20]Liu X,Chong Y,Tu Y,et al.CRM1/XPO1 is associated with clinical outcome in glioma and represents a therapeutic target by perturbing multiple core pathways.Journal of Hematology&Oncology,2016,9(1):108
[21]Liu X,Chong Y,Liu H,et al.CRM1 inhibitor S109 suppresses cell proliferation and induces cell cycle arrest in renal cancer cells.Korean Journal of Physiology&Pharmacology Official Journal of the Korean Physiological Society&the Korean Society of Pharmacology,2016,20(2):161-168
[22]Wen Y S,Yong L Y,Yan H,et al.Phenethyl isothiocyanate suppresses the metastasis of ovarian cancer associated with the inhibition of CRM1-mediated nuclear export and mTOR-STAT3pathway.Cancer Biology&Therapy,2016,18(1):26-35
[23]Puente X S,Pinyol M,Quesada V,et al.Whole-genome sequencing identifies recurrent mutations in chronic lymphocytic leukaemia.Nature,2011,475:101-105
[24]Balatti V,Bottoni A,Palamarchuk A,et al.NOTCH1 mutations in CLL associated with trisomy 12.Blood,2012,119:329-331
[2]Rousselot P,Labaume S,Marolliau J P,et al.Arsenic trioxide and melarsoprol induce apoptosis in plasma cells from myeloma patients.Cancer Res,1999,59:1041-1048
[3]傅玉军,侯健,王东星,等.氧化砷诱导骨髓瘤细胞株KM3凋亡的研究.中华血液学杂志,1998,19(1):591
[4]Turner J G,Dawson J,Sullivan D M.Nuclear export of proteins and drug resistance in cancer.Biochem Pharmacol,2012,83:1021-1032
[5]Y-T Tai,Y Landesman,C Acharya,et al.CRM1 inhibition induces tumor cell cytotoxicity and impairs osteoclastogenesis in multiple myeloma:molecular mechanisms and therapeutic implications.Leukemia,2014,28(1):155-165
[6]GE Qun-Fang,OUYANG Gui-Fang,CHEN Ying,et al.Effects of arsenic trioxide combined with bortezomib on apoptosis of multiple myeloma cell line KM3 and its mechanisms.Journal of Experimental Hematology,2012,20(1):112-115
[7]Keiichi Sakakibara,Naoya Saito,Takuji Sato,et al.CBS9106 is a novel reversible oral CRM1 inhibitor with CRM1 degrading activity.Blood,2011,118:3922-3931
[8]方坚.多发性骨髓瘤中医诊治思路探讨.广州中医药大学学报,2013,30(4):581-582
[9]李仝,黄玉燕.多发性骨髓瘤从肾虚毒癖论治.北京中医药大学学报,2008,31(6):427-428
[10]贺芳,曾耀英,王通,等.三氧化二砷诱导Jurkat细胞凋亡过程中线粒体的变化.中国病理生理杂志,2008,24(8):1600-1603
[11]杜恒飞,于路佳,张金巧.去甲斑蝥素抗骨髓瘤作用及其机制研究.中草药,2011,42(12):2479-2483
[12]Zi Ma,Ken-ichiro Otsuyama,Shangqin Liu,et al.Baicalein, a component of Scutellaria radix from Huang-Lian-Jie-Du-Tang(HLJDT), leads to suppression of proliferation and induction of apoptosis in human myeloma cells.Blood,2005,105:3312-3318
[13]林圣云,沈楚云,蒋剑平,等.白花蛇舌草多糖诱导多发性骨髓瘤细胞凋亡及其机制研究.中华血液学杂志,2013,34(4):337-340
[14]Haiwen Ni,Wanzhou Zhao,Xiangtu Kong,et al.NF-Kappa B modulation is involved in celastrol induced human multiple myeloma cell apoptosis.PLoS One,2014,9(4):e95846
[15]王鸣明,朱琦,任志宏,等.砷剂诱导多发性骨髓瘤细胞socs-1基因去甲基化作用的研究.中国实验血液学杂志,2008,16(5):1064-1068
[16]王清,李娟,谷景立,等.Bortezomib增强三氧化二砷抑制KM3细胞生长和促进凋亡机制的研究.中国病理生理杂志,2010,26(2):297-301
[17]Kevin T Nguyen,Michael P Holloway,Rachel A Altura.The CRM1nuclear export protein in normal development and disease.Int J Biochem Mol Biol,2012,3(2):137-151
[18]Sun H,Lin D C,Cao Q, et al.CRM1 inhibition promotes cytotoxicity in Ewing sarcoma cells by repressing EWS-FLI1-dependent IGF-1signaling.Cancer Research,2016,76(9):2687
[19]Fabi F,Adam P,Vincent K,et al.Inhibition of CRM1 activity sensitizes endometrial and ovarian cell lines to TRAIL-induced cell death.Cell Communication&Signaling Ccs,2018,16(1):39
[20]Liu X,Chong Y,Tu Y,et al.CRM1/XPO1 is associated with clinical outcome in glioma and represents a therapeutic target by perturbing multiple core pathways.Journal of Hematology&Oncology,2016,9(1):108
[21]Liu X,Chong Y,Liu H,et al.CRM1 inhibitor S109 suppresses cell proliferation and induces cell cycle arrest in renal cancer cells.Korean Journal of Physiology&Pharmacology Official Journal of the Korean Physiological Society&the Korean Society of Pharmacology,2016,20(2):161-168
[22]Wen Y S,Yong L Y,Yan H,et al.Phenethyl isothiocyanate suppresses the metastasis of ovarian cancer associated with the inhibition of CRM1-mediated nuclear export and mTOR-STAT3pathway.Cancer Biology&Therapy,2016,18(1):26-35
[23]Puente X S,Pinyol M,Quesada V,et al.Whole-genome sequencing identifies recurrent mutations in chronic lymphocytic leukaemia.Nature,2011,475:101-105
[24]Balatti V,Bottoni A,Palamarchuk A,et al.NOTCH1 mutations in CLL associated with trisomy 12.Blood,2012,119:329-331