MSCs分泌的肝细胞生长因子对人髓系白血病细胞系K562及其耐药株K562/G01增殖及药物敏感性的影响

Effects of hepatocyte growth factor secreted by bone marrow mesenchymal stem cells on proliferation and drug sensitivity of cell line K562 and resistant strain K562/G01

目的研究骨髓造血微环境中间充质干细胞(MSCs)分泌的肝细胞生长因子(HGF)对人白血病细胞K562及其耐药珠K562/G01细胞增殖及药物敏感性的影响。方法采用细胞贴壁法获取MSCs,选用人慢性髓系白血病急性红白血病变细胞系K562及其耐药系K562/G01。K562、K562/G01细胞分别与白血病患儿MSCs黏附共培养,实验组为加HGF抗体组,对照组加入等量培养液,通过显微镜观察白血病细胞在MSCs层上的黏附情况并计算黏附率,MTT法检测白血病细胞增殖情况,流式细胞仪测定白血病细胞周期。在共培养各组中加入伊马替尼孵育48 h后,流式细胞仪检测白血病细胞凋亡率。结果实验组中MSCs对K562和K562/G01细胞的黏附率分别为32.4%±3.2%和38.1%±4.8%,均分别低于对照组的51.6%±3.6%和62.2%±4.9%(P<0.05)。实验组中K652和K562/G01细胞的A值分别为(0.3102±0.0390)和(0.3246±0.0358),均分别低于对照组的(0.5216±0.0467)和(0.5908±0.0034)(P<0.05)。实验组中K652及K562/G01细胞处于G 0/G 1期的细胞比例均高于对照组,而处于S期的细胞比例低于对照组(P<0.05)。加入伊马替尼孵育48 h后白血病细胞凋亡情况:实验组中存活细胞比例低于对照组中存活细胞比例(P<0.05)。结论HGF可通过影响细胞黏附及细胞周期促进K562及K562/G01细胞存活与增殖,并降低伊马替尼对白血病细胞的杀伤作用。

Objective To study the effect of hepatocyte growth factor(HGF)secrete by mesenchymal stem cell in bone marrow hematopoietic microenvironmen on proliferation and drug sensitivity of leukemia cell line K562 and its drug-resistant variant K562/G01.Methods The MSCs were obtained from low serum medium.K562 cells and its drug resistant K562/G01 were used.The co-culture of MSCs with leukemia cells was prepared.Leukemia cells cultured with MSCs.According to existence of anti-HGF antibody or not,all groups were divided into control group and experiment group.The adhesion of leukemia cells on MSCs cell layer was observed under inverted fluorescence microscope and the adhesion rates were calculated.The pro1iferation of leukemia cells in the co-culture system was observed by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT).Cell cycle of leukemia cells in the co-culture system was investigated by flow cytometry.The survival rate of K562 cells and K562/G01 cells in the co-culture system was measured by flow cytometry after was added and incubated for 48 hours.Results After incu-bated with anti-HGF antibody for 24 hours,adhesion rates in experiment groups were significantly lower than those in control group(P<0.05).The proiferations of leukemia cells in experiment groups were all significantly lower than those in control groups in the co-culture system(P<0.05).The percentage of leukemia cells in G 0/G 1 phase in experiment group was higher than those in control group(P<0.05).On the contrary,the proportion of leukemia cells in S phase in experiment group was significantly lower than those in control group(P<0.05).Survival rate that of K562 cells and K562/G01 cells in the co-culture system after added imatinib incubation for 48 hours in experiment groups were respectively significantly lower than those in control groups(P<0.05).Conclusions The HGF promotes the cell survival and proliferation of K562 and K562/G01 cells by affecting cell adhesion and cell cycle,and reduces the cytotoxicity of imatinib on leukemia cells.