白血病骨髓基质细胞黏附对DNR在Jurkat细胞内蓄积及分布的影响

Effects of adhesion mediated by BMSCs from leukemia patients on DNR accumulation and distribution in the Jurkat cells

目的初步探讨细胞黏附介导耐药模型中DNR在Jurkat细胞内的蓄积浓度及分布特征。方法分离、培养骨髓基质细胞,Jurkat细胞与60Co照射的基质细胞层黏附培养,构建细胞黏附介导耐药模型,0.5μmol/L DNR作用24h,FITC-AnnexinV/PI标记后流式细胞仪定量Jurkat细胞的凋亡率及DNR在Jurkat细胞内蓄积浓度,荧光显微镜观察其荧光信号在Jurkat细胞内的分布。结果白血病基质细胞黏附组、正常基质细胞黏附组Jurkat细胞凋亡率分别为(6.05±0.54)%、(8.48±0.86)%,与悬浮对照组(25.74±6.15)%比较均相差非常显著(P<0.01),且白血病基质细胞组与正常基质细胞组间相差显著(P<0.05)。基质细胞黏附并未降低Jurkat细胞内DNR的蓄积浓度,其红色荧光信号在Jurkat细胞内仍呈均匀、弥漫性分布。结论白血病骨髓基质细胞黏附能介导Jurkat细胞耐药,但其耐药机制可能独立于药物泵出增多致细胞内药物蓄积浓度降低这一经典耐药机制。

Objective To investigate DNR accumulation and distribution in Jurkat cells adhered to the cultured BMSCs from leukemia patients. Methods To construct the CAM-DR model,we co-cultured Jurkat cells with the irradiated BMSCs layer by ^60Co. The apoptosis percentage of Jurkat cells exposured to 0.5μmol/L DNR for 24h was quantitated by FACScan with FITC-Annexin V/PI stain. The DNR accumulation in Jurkat cell was detected by FACScan machine and the distribution of DNR cells was observed under fluorescence microscope. Results, We identified the function of the model and found that the Jurkat cells in the model showed a decreased sensitivity to DNR, the apoptosis percentages for normal BMSCs,leukemic BMSCs and noadhered cotrol were of（8.48±0.86）%, （6.05±0.54）% and （25.74±6.15）%. As compared with suspension contol, the apoptosis percentages of Jurkat cells adhered to normal BMSCs and leukemic BMSCs had significant difference in apoptosis percentages （ P〈0.01 ）. Moreover, the apoptosis percentages of Jurkat cells adhered to leukemic BMSCs were lower than those of Jurkat cells adhered to normal BMSCs. Interestingly, intracellular drug accumulation in adhered Jurkat cells was not significantly different from the suspended cells（P〉0.05）. Similar distribution that fluorescene signals from DNR were distributed diffusely in both adhered Jurkat cells and suspended cells were observed in adhered and suspended Jurkat cells under fluorescent microscope. Conclusion The bone marrow stromal cells from leukemia patients have stronger ability in protecting Jurkat cells exposured chemotherapeutics than normal bone marrow stromal cells. But the mechanisms associated with cell adhesion mediated drug resistance may be independent the classic mechanism.