摘要
目的:探讨DNA损伤激活的长链非编码RNA(NORAD)对多发性骨髓瘤(MM)细胞增殖及对硼替佐米(BTZ)化疗耐药性的影响。方法:应用逆转录聚合酶链反应(RT-PCR)检测健康人和骨髓瘤患者、不同MM细胞系中NORAD表达差异,并分析其与MM患者生存率关系;运用慢病毒转染技术,干扰细胞系中NORAD的表达,检测其表达差异;定量PCR(qPCR)检测骨髓瘤细胞系中人类非编码RNA-144a-3p(hsa-miR-144a-3p)的表达;采用双荧光酶素报告验证NORAD和hsa-miR-144a-3p的结合;通过RNA结合蛋白免疫沉淀(RIP)实验进一步验证LncRNA和miRNA的结合;细胞计数试剂8(CCK8)法检测各细胞处理后细胞活性。细胞克隆试验检测细胞增殖能力。结果:MM患者细胞系中NORAD常高表达,提示预后差,生存期短;敲除NORAD基因后,MM耐药细胞系对BTZ药物敏感性增加。MM患者hsa-miR-144a-3p高表达,在骨髓瘤细胞系中加用miR-144a-3p类似物时细胞增殖能力下降;NORAD低表达可显著抑制细胞增殖,细胞对药物敏感性增强。细胞克隆形成实验也证实NORAD低表达可抑制癌细胞增殖。结论:NORAD高表达可增强MM对BTZ化疗耐药性,NORAD可直接与hsa-miR-144a-3p结合,上调其在MM细胞中的表达,进一步调控癌细胞增殖,同时调控患者对BTZ药物敏感性。
Objective:To explore the effects of LncRNA NORAD on proliferation and BTZ chemotherapy resistance of multiple myeloma(MM)cells.Method:RT-PCR was used to detect the difference of NORAD expression in healthy people,MM,and different MM cell lines.The relationship between NORAD expression and MM patients'production was analyzed.The lentivirus transfection technique was used to interfere with the expression of NORAD in cell lines.The expression of hsa-miR-144a-3q was detected by qPCR.The binding of NORAD and hsa-miR-144a-3p was verified by double fluorescence report.The binding of LncRNA and miRNA was further verified by RIP experiment.The CCK8 assay method was used to detect the cell activity after treatment.Cell cloning test was used to detect cell proliferation.Results:High expression of NORAD in MM patients and cell lines suggests poor prognosis and short survival.After NORAD gene was knocked out,the sensitivity of MM drug-resistant cell line to BTZ increased.The high expression of hsa-miR-144a-3p in myeloma patients was detected by qPCR,and the cell proliferation ability decreased when miR-144a-3p analogues were added to myeloma cell lines.Low expression of NORAD could significantly inhibit cell proliferation and enhance drug sensitivity of cells.Cell clonogenesis experiment also confirmed that low expression of NORAD could inhibit the proliferation of cancer cells.Conclusions:High expression of NORAD can enhance the resistance of MM to BTZ.NORAD can directly bind to hsa-miR-144a-3p and upregulate its expression in MM cells.hsa-miR-144a-3p can further regulate the proliferation of cancer cells and regulate the drug sensitivity of patients to BTZ.
作者
许超平
常伟
刘晓倩
刘仕明
肖颖
XU Chao-ping;CHANG Wei;LIU Xiao-qian;LIU Shi-ming;XIAO Yin(Department of Hematology,Puren Hospital Affiliated to Wuhan University of Science and Technology,Hubei Wuhan 430081,China)
出处
《内科急危重症杂志》
2023年第6期490-494,共5页
Journal of Critical Care In Internal Medicine
基金
武汉市卫生计生委科研计划资助项目(No:WX18Z35)。
