【作者】 汤静；

【导师】 孟力；

【作者基本信息】 华中科技大学 ， 血液病学， 2012， 硕士

【摘要】 【目的】收集骨髓标本，检测其中的Fn表达情况；以急性淋巴系白血病细胞系Jurkat为研究对象，部分模拟急性淋巴细胞白血病的骨髓微环境，观察Jurkat在与骨髓基质细胞MSCs作用后，对化疗药物吡柔比星药物敏感性的改变，研究白血病骨髓微环境介导化疗耐药的部分机制。【方法】收集骨髓标本，检测不同病例骨髓液标本上清中Fn水平的高低；使用CCK-8的方法检测单独培养的Jurkat细胞对化疗药物吡柔比星的药物敏感性；体外建立急性淋巴系白血病细胞系Jurkat与骨髓基质细胞MSCs共培养接触耐药模型；体外建立Jurkat细胞与Fn作用的粘附耐药模型；使用CCK-8和流式细胞仪的方法分别检测Jurkat细胞在MSCs共培养接触耐药模型、Fn粘附耐药模型、Jurkat细胞单独培养模型中的Jurkat细胞对化疗药物吡柔比星在相同药物浓度作用下细胞凋亡情况的改变；使用流式细胞仪检测Jurkat细胞在MSCs接触耐药模型和Fn粘附耐药模型中细胞周期阻滞的改变；使用ELISA检测在MSCs共培养接触耐药模型中Fn表达水平的高低。【结果】与正常人相比较，血液肿瘤患者的骨髓标本上清中所分泌的Fn水平明显的升高，其结果有统计学意义；ELISA检测在Jurkat细胞与MSCs共培养的接触耐药模型中收集的上清中存在Fn；在与骨髓基质细胞MSCs共培养接触耐药模型中，Jurkat细胞在相同浓度化疗药物吡柔比星作用下，在同一时间点上，细胞凋亡数目较单独培养组有明显下降，结果具有统计学意义；在Fn作用的粘附模型中，Jurkat细胞在同一时间点上，对化疗药物吡柔比星在相同药物浓度作用下细胞凋亡数目较单独培养组也有明显下降，结果具有统计学意义；Jurkat细胞在MSCs和Fn作用下，细胞出现明显的周期阻滞，有统计学意义。【结论】血液肿瘤病人与正常人相比，骨髓中可分泌更高水平的Fn；骨髓基质细胞MSCs能分泌Fn，并可能通过Fn的作用，使细胞凋亡减少，并诱导发生G1期阻滞。更多还原

【Abstract】 [Objective] Collect bone marrow specimens to detect the expression of Fn. Then partly simulate the ALL bone marrow microenvironment to demonstrate drug sensitivity changes of acute lymphoid leukemia cell line Jurkat to pirarubicin, when Jurkat cells exposed to bone marrow stramal cells(MSCs) and fibronectin(Fn).And discuss drug resistance in acute leukemia mediated by bone marrow microenvironment and certain mechanism.[Methods] collect some bone marrow specimens of different patients, detect the expression of Fn in bone marrow samples; use the CCK-8method to detecte the drug sensitivity of singly cultured Jurkat cells to pirarubicin in vitro; establish a co-culture model of Jurkat cells and bone marrow stromal cells(MSCs)/fibronectin(Fn) in vitro; use CCK-8and flow cytometry to detect changes of Jurkat cell apoptosis, then analyse the drug sensitivity changes of pirarubicin at the same concentration; use ELISA to detect the differences of Fn expression of level Jurkat cells in MSCs co-culture model compared with single cultured Jurkat cells model at different concentration of pirarubicin; using flow cytometry to demonstrate changes of the cycle arrest of Jurkat cells in the MSCs and Fn co-culture model compared with single cultured Jurkat cells model.[results] compared to samples of normal persons, patients with haemal tumor diseases demonstrated significantly high expression of Fn, the results were statistically significant; detections of ELISA in MSCs co-culture model showed expression of Fn in bone marrow stromal cells; In MSCs/Fn co-culture model, cell apoptosis changed obviously compared to single Jurkat cells cultured model at the same concentration of pirarubicin at the same time, with statistical significance; there were statistical significance of cell cycle arrest induced by MSCs/Fn co-culture model compared to single Jurkat cells model at the same concentration of pirarubicin at the same time.[Conclusion]bone marrow of patients with haemal tumor diseases can secrete higher levels of Fn compared with nomal persons; bone marrow stromal cells can secrete Fn MSCs, and probobly induce cell apoptosis, and G1cycle arrest in this way.更多还原