骨髓基质细胞通过上调纤维连接蛋白介导白血病Jurkat细胞化疗耐药

Bone Marrow Stromal Cells Mediate Drug Resistance of Jurkat Cells by Upregulating Fibronectin Level

目的探讨骨髓基质细胞通过上调纤维连接蛋白介导白血病Jurkat细胞化疗耐药。方法使用CCK-8法检测白血病Jurkat细胞对化疗药物吡柔比星的敏感性,绘制敏感曲线;体外建立Jurkat细胞与骨髓基质细胞(BMSC)共培养接触耐药模型,检测该模型中在吡柔比星作用下Jurkat细胞的抑制率,并检测在该模型中纤维连接蛋白(FN)的表达水平;体外建立Jurkat细胞与FN作用的粘附耐药模型,检测该模型中在吡柔比星作用下Jurkat细胞的抑制率;使用流式细胞仪检测在BMSC/FN耐药模型及单独培养模型中Jurkat细胞在吡柔比星作用下的凋亡率;收集复发难治血液肿瘤患者及正常人骨髓标本,检测标本上清中FN的水平。结果在与BMSC共培养接触耐药模型中,在相同浓度吡柔比星作用下,在同一时间点,Jurkat细胞凋亡率较单独培养组明显下降。在Jurkat细胞与BMSC共培养的接触耐药模型中,上清中分泌的FN明显上升。在FN作用的粘附耐药模型中,在相同浓度吡柔比星作用下,在同一时间点,Jurkat细胞凋亡率较单独培养组明显下降。与正常人比较,复发难治血液肿瘤患者骨髓上清中检测的FN水平明显上升。结论复发难治血液肿瘤患者的骨髓中,FN分泌增加;通过体外共培养模型提示骨髓基质细胞可以分泌FN,介导白血病Jurkat细胞发生化疗耐药,推测在白血病患者的骨髓微环境中,基质细胞可以为白血病细胞在化疗中提供耐药支持,黏附分子FN可能参与其中,是介导白血病细胞化疗耐药的重要原因。

Objective To discuss whether bone marrow stromal cells（BMSC）can promote drug resistance of Jurkat cells by upregulating fibronectin（FN）level.Methods CCK-8method was used to detect sensitivity of Jurkat cells to THP,and sensitivity curve was drawed.A co-cultured model of Jurkat cells with BMSC was established in vitro,and inhibition rate of Jurkat cells to THP was detected.FN expression in model of Jurkat cells with BMSC was determined.A co-cultured model of Jurkat cells with FN was established in vitro.Inhibition rate of Jurkat cells to THP of co-cultured model of Jurkat cells with FN was detected.Flow cytometry was employed to detect apoptosis rate of Jurkat cells to THP in co-cultured model of Jurkat cells with BMSC/FN and model of Jurkat cells alone.Bone marrow specimens of relapsed/refractory hematological malignancies patients and normal persons were collected,and the expression of FN in supernatant of bone marrow samples was detected.Results In BMSC co-cultured model,apoptosis rate of Jurkat cells changed significantly compared to model of Jurkat cells cultured alone at the same concentration of THP at the same time.The result showed significantly high expression of FN in supernatant in BMSC cocultured model compared with model of Jurkat cells cultured alone.In FN co-cultured model,apoptosis rate of Jurkat cells changed obviously compared to model of Jurkat cells cultured alone at the same concentration of THP at the same time.Compared to the supernatant of bone marrow samples of normal persons,the supernatant of patients with relapsed/refractory hematological malignancies demonstrated significantly high expression of FN.Conclusion Bone marrow microenvironment in specimens of patients with relapsed/refractory hematological malignancies can secrete higher level of FN than that in normal persons.In vitroexperiment,BMSC can upregulate FN level,and then decrease apoptosis of Jurkat cells,leading to drug resistance to THP.It may conclude that in bone marrow microenvironment of leukemia,BMCS could protect leukemia cells from chemotherapy,FN may be involved.This mechanism may be the possible reason of drug resistance in hematological malignancies.