急性淋巴细胞白血病患儿骨髓间充质干细胞对K562细胞株药物耐受性的影响

摘要

Objective To study the effect of acute lymphoblastic leukemia (ALL) children bone marrow mesenchymal stem cells (MSCs) on the resistance of K562cell atd mechanism in vitro.Method MSCs were obtained from AL children bone marrow after derivation, cultivation and identification.The coculture of MSCs and K562 and K562 suspension were established.Effects of MSCs on the growth of K562 cells were investigated in vivo.The two kinds of cells treated with different concentration of adriamycin (ADM) and the rate of apoptosis was evaluated by flow cytometry.Cell cycle was determined by flow cytometry.RT-PCR was used to detect Bcl-2 and Bax in K562 cells.Result Compared with the cell growth curve of K562 alone, the K562 cell co-cultured with MSCs grew slower and the exponential phase of growth was not obvious.The apoptosis index of the K562 cells co- clutured with MSCs was (9.19 ±0.53)% examined by flow cytometry, and that of the K562 cells alone was 4.00 ± 0.37% respectively( P ＜ 0.05 ).The percentage of cells at G0/G1 phase was (50.2 ± 2.26) % and that at S phase was (37.03 ± 3.50) % in the group of K562 alone, but those of the K562 cells co - cultured with MSCs were (80.95 ± 3.83) % and ( 17.40 ± 1.50)% respectively( P ＜0.05).The result of RT-PCR suggested expression of Bcl-2/Bax of the K562 cell co-cultured with MSCs was higher than K562 alone.Conclusion ALL children MSCs suppressed the growth of K562 cell in vitro.Adhesion made K562 depress sensitive to ADM.The mechanism was perhaps caused by adhesion with MSCs, K562 cell cycle was changed and related to Bcl-2 gene high level expression.%目的 研究急性淋巴细胞白血病患儿骨髓间充质干细胞(MSCs)对K562细胞药物耐受性的影响,并初步探讨其机制.方法 从白血病患儿骨髓中分离、培养并鉴定MSCs;建立K562细胞株与MSCs共培养的体系,观察MSCs对K562细胞生长的影响;AnnexinV-FITC检测一定浓度的阿霉素(ADM)对K562细胞凋亡的影响;流式细胞仪测定不同培养条件下的K562细胞周期;RT-PCR检测K562细胞的Bcl-2和Bax基因.结果 和单独培养的K562细胞相比,与MSCs黏附共培养的K562细胞生长较为缓慢,未见明显的对数生长期.单独培养组细胞早期凋亡率为(9.19±0.53)%,而黏附培养组为(4.00±0.37)%,差异具有统计学意义(P＜0.05).黏附共培养组K562细胞,G0-G1期细胞比例为(80.95±3.83)%,显著高于单独培养组(50.2±2.26)%(P＜0.05);而S期细胞比例为(17.40±1.50)%,显著低于单独培养组(37.03±3.50)%(P＜0.05).相对于单独悬浮培养的K562细胞,黏附共培养的K562细胞Bcl-2基因相对表达明显增强,Bcl-2/Bax显著增高.结论 白血病患儿MSCs能抑制白血病细胞株K562生长,改变细胞周期而逃避药物的促凋亡作用,K562对阿霉素产生耐药性可能与黏附共培养后Bcl-2基因表达增强有关.