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目的:探讨粒细胞集落刺激因子(granulocyte colony stimulating factor,G-CSF)和AMD3100增加白血病细胞对化疗敏感性的作用及其机制。方法:用G-CSF、AMD3100单药或联合处理HL-60细胞株后,与HS-5上清或HS-5细胞共培养,采用CCK-8法检测细胞活力,用AnnexinⅤ试剂盒检测细胞凋亡,流式细胞术和蛋白印迹法分别检测膜表面CXCR4蛋白、总CXCR4蛋白表达情况,实时荧光定量PCR检测mRNA表达情况。结果:HS-5上清或细胞能够保护HL-60细胞免于自发或药物诱导的凋亡,而G-CSF和AMD3100均能够部分逆转这种保护作用,两药联合时更明显;G-CSF能够降低细胞膜表面和细胞质内总CXCR4蛋白及CXCR4 mRNA的表达,同时上调micro RNA-146a(miR-146a)表达;AMD3100只能降低膜表面CXCR4表达,对细胞质内总CXCR4蛋白、CXCR4 mRNA和miR-146a表达无影响。结论:G-CSF和AMD3100通过不同机制降低白血病细胞的CXCR4表达而阻断CXCR4/CXCL12信号轴,从而打破骨髓微环境-白血病细胞间的相互作用,直接增强化疗药物杀伤作用。
Objective: To investigate the effects of granulocyte colony stimulating factor (G-CSF) and AMD3100 on chemosensitivity of leukemia cells and its mechanism. Methods: HL-60 cells were treated with G-CSF, AMD3100 alone or in combination with HS-5 supernatant or HS-5 cells. Cell viability was detected by CCK-8 assay. Cell apoptosis was detected by Annexin V kit The expressions of CXCR4 and CXCR4 on the membrane surface were detected by flow cytometry and Western blot respectively. The mRNA expression of CXCR4 and CXCR4 was detected by real-time fluorescence quantitative PCR. Results: HS-5 supernatant or cells could protect HL-60 cells from spontaneous or drug-induced apoptosis, while both G-CSF and AMD3100 could partially reverse this protective effect. The combination of G-CSF and G-CSF The expression of CXCR4 and CXCR4 mRNA in the cytoplasm and the expression of CXCR4 mRNA in the cytoplasm decreased, while miR-146a (miR-146a) expression was upregulated. AMD3100 decreased the expression of CXCR4, 146a expression no effect. CONCLUSION: G-CSF and AMD3100 can down-regulate the CXCR4 / CXCL12 signal axis through different mechanisms to reduce the CXCR4 / CXCL12 signaling axis, thereby breaking the bone marrow microenvironment-leukemia cell interactions and directly enhancing chemotherapeutic drug-killing effects.