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目的探讨白血病成骨细胞对急性髓性白血病(AML)细胞屏蔽化疗杀伤的作用,及其与促血小板生成素/血小板生成素受体(TPO/c-MPL)信号通路的关系。方法分离培养白血病骨髓间充质干细胞并行成骨诱导,ELISA法检测成骨细胞分泌血小板生成素(TPO)水平,流式细胞仪检测人红白血病细胞株(HEL)c-MPL表达。成骨细胞与HEL细胞共培养,观察在化疗药物柔红霉素干预下HEL细胞存活率。结果白血病骨髓成骨细胞培养d7、d14 TPO表达分别为(176.84±15.32)ng/mL和(198.24±13.49)ng/mL,明显高于正常对照的TPO水平,分别为d7(140.13±20.12)和d14(157.66±18.82)ng/mL。应用柔红霉素干预后,AML成骨细胞与HEL细胞共培养组半抑制浓度(IC50)为(3.320±0.218)μg/mL,高于HEL+正常来源成骨细胞组(2.236±0.167)μg/mL和HEL+hFOB1.19组(2.254±0.111)μg/mL。生长曲线研究发现,在不同浓度DNR干预下,AML成骨细胞与HEL细胞共培养组中HEL细胞的细胞存活率最大,均高于其他HEL共培养组。结论白血病成骨细胞能有效屏蔽化疗药物对白血病细胞的杀伤作用,这种作用可能与白血病骨髓微环境内TPO/c-MPL信号通路相关。
Objective To investigate the effect of leukemic osteoblasts on cytotoxicity of acute myelogenous leukemia (AML) cells and its relationship with TPO / c-MPL signaling pathway. Methods Bone marrow-derived mesenchymal stem cells were isolated and cultured for osteogenic induction. The levels of TPO secreted by osteoblasts were detected by ELISA. The expression of c-MPL in human erythroleukemia cell line (HEL) was detected by flow cytometry. Osteoblasts were co-cultured with HEL cells to observe the survival rate of HEL cells under the intervention of daunorubicin. Results The expressions of d7 and d14 TPO in leukemia osteoblasts were (176.84 ± 15.32) ng / mL and (198.24 ± 13.49) ng / mL, respectively, which were significantly higher than those of the normal controls (d7 = 140.13 ± 20.12 and d14 (157.66 ± 18.82) ng / mL. Compared with HEL + normal osteoblast group (2.236 ± 0.167) μg / mL, the half inhibitory concentration (IC50) of AML osteoblast and HEL cell co-culture group was (3.320 ± 0.218) μg / mL and HEL + hFOB1.19 group (2.254 ± 0.111) μg / mL. Growth curve study found that under the different concentrations of DNR intervention, the cell survival rate of HEL cells co-cultured with AML osteoblasts and HEL cells was the highest, which were higher than other HEL co-culture groups. Conclusion Leukemic osteoblasts can effectively block the killing effect of chemotherapeutic drugs on leukemia cells, which may be related to the TPO / c-MPL signaling pathway in leukemic bone marrow microenvironment.