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目的:构建骨髓基质细胞-白血病共培养模型模拟白血病微环境,探讨肿瘤坏死因子相关的凋亡诱导配体(TRAIL)逆转黏附介导白血病耐药的有效性。方法:构建Jurkat细胞共培养模型、扫描电镜观察。以0.5μmol/LDNR作用6和12h,流式细胞仪检测Jurkat细胞TRAIL-R1、TRAIL-R2的表达。分为TRAIL组、DNR组和TRAIL协同DNR组,AnnexinⅤ-FITC/PI标记后流式细胞仪定量Jurkat细胞凋亡率,并检测细胞内DNR蓄积浓度。MTT法测定IC50,计算200ng/mLTRAIL作用18h的耐药逆转倍数。结果:0.5μmol/LDNR能上调TRAIL-R2的表达,作用6h即可达高峰,但不能上调TRAIL-R1的表达,骨髓基质细胞黏附均不影响Jurkat细胞TRAIL-R1和TRAIL-R2的表达。骨髓基质细胞黏附明显抑制DNR的凋亡诱导效应,但对TRAIL的凋亡诱导效应无明显影响,且与DNR间有协同效应。200ng/mLTRAIL能使Jurkat细胞对DNR的IC50由2.30μmol/L降至0.049μmol/L,逆转倍数FR为46.94倍,但未观察到TRAIL上调Jurkat细胞内DNR的蓄积浓度。结论:TRAIL协同DNR能有效逆转黏附介导Jurkat细胞耐药,其机制不依赖于白血病细胞内DNR蓄积浓度的变化。
OBJECTIVE: To construct a model of bone marrow stromal cells-leukemia co-culture model to simulate leukemia microenvironment, and to explore the effectiveness of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) reversing adhesion-mediated drug resistance in leukemia. Methods: Jurkat cells co-culture model was constructed and observed by scanning electron microscopy. The expression of TRAIL-R1 and TRAIL-R2 in Jurkat cells was detected by flow cytometry with 0.5μmol / LDNR for 6 and 12 h. The cells were divided into three groups: TRAIL group, DNR group and TRAIL group. The apoptosis rate of Jurkat cells was determined by flow cytometry with Annexin V-FITC / PI and the concentration of intracellular DNR accumulation was detected. IC50 was measured by MTT method, and the reversal multiple of drug resistance was calculated after 18h of 200ng / mL TRAIL. Results: 0.5μmol / L LDNR up-regulated TRAIL-R2 expression and peaked at 6h, but did not up-regulate TRAIL-R1 expression. Bone marrow stromal cell adhesion did not affect TRAIL-R1 and TRAIL-R2 expression in Jurkat cells. Adhesion of bone marrow stromal cells significantly inhibited the apoptosis-inducing effect of DNR, but had no obvious effect on the apoptosis-inducing effect of TRAIL, and had a synergistic effect with DNR. The IC50 of Jurkat cells to DNR decreased from 2.30μmol / L to 0.049μmol / L, and the reversal fold FR was 46.94 times at 200ng / mLTRAIL, but no accumulation of DNR in Jurkat cells was observed by TRAIL up-regulation. CONCLUSION: TRAIL combined with DNR can effectively reverse the adhesion-mediated drug resistance of Jurkat cells, and its mechanism does not depend on the concentration of DNR accumulated in leukemia cells.