氨磷汀在体外减轻足叶乙甙所致的骨髓间充质干细胞损伤

Amifostine alleviates the etoposide-induced damage to bone marrow-derived mesenchymal stem cells in vitro

目的研究氨磷汀在体外对化疗药物所致的骨髓间充质干细胞(MSCs)损伤的影响,为其应用于慢性粒细胞白血病(CML)患者的造血干细胞移植提供实验依据。方法采集CML患者的骨髓细胞,应用极限稀释法获取单克隆来源的MSCs,并在低血清培养液中培养和扩增。按实验分组分别加入氨磷汀(氨磷汀组)、足叶乙甙(VP-16组)、氨磷汀和足叶乙甙(联合处理组),并设没有处理的对照组。在药物作用一定时间后,计数MSCs,绘制生长曲线;流式细胞仪检测MSCs的免疫表型及凋亡率;在甲基纤维素半固体培养中检测MSCs作为滋养层支持造血的能力。结果MSCs生长曲线显示,与对照组相比,氨磷汀组的MSCs生长未受明显影响,联合处理组MSCs的生长显著好于VP-16组。培养48 h后,对照组、氨磷汀组、联合处理组和VP-16组的MSCs凋亡率分别为(4.9±0.8)%、(5.6±1.2)%、(18.7±3.4)%和(33.8±5.1)%,氨磷汀组与对照组相比,差异无统计学意义;联合处理组高于对照组,但明显低于VP-16组(P<0.05)。形成粒-单核系集落、红系集落和多系集落的产率,氨磷汀组与对照组间的差异无统计学意义,联合处理组低于对照组(P<0.05),但显著高于VP-16组(P<0.05)。氨磷汀作用前后未见MSCs免疫表型发生明显改变。结论氨磷汀在体外对CML患者骨髓来源MSCs的增殖、凋亡、造血支持能力以及免疫表型无明显影响,但能显著改善足叶乙甙对MSCs所致的生长抑制及功能损伤,提示在预处理中加用氨磷汀可能成为促进CML患者造血干细胞移植后造血恢复的一种新策略。

Objective To provide the experimental basis for the applied value of amifostine in the mesenchymal stem cells （MSCs） transplantation of patients with chronic myelogenous leukemia （CML） by studying the effects of amifostine on the etoposide （VP-16）-induced damage to bone marrow-derived MSCs in votro. Methods MSCs were obtained from CML patients, cultured in vitro, and then divided into 4 groups （control group, amifostine group, amifostine ＋ VP-16 group and VP-16 group）. The immunophenotype, growth and apoptosis of the MSCs were investigated and hematopoiesis support of MSCs in vitro was detected by long-term bone marrow culture and the methylcellulose progenitor assay. Results The growth curve showed that amifostine had no significant effects on the proliferation of MSCs, while growth of MSCs in amifostine ＋ VP-16 group was more vigorous significantly than in VP-16 group. The apoptosis rate of MSCs in control group, amifostine group, amifostine＋ VP-16 group and VP-16 group was 4. 9 % ±0. 8 %, 5.6 % ± 1.2 %, 18.7 0.% ± 3.4 % and 33.8 % ± 5.1%, respectively. There was significant difference between amifostine ＋ VP-16 group and control group or VP-16 group （both P〈0.05）, but not between amifostine group and control group. There was no significant difference in the ability of hematopoiesis support between amifostine group and control group （P〉0. 05）, but that in amifostine ＋ VP-16 group was significantly lower than in control group, and significantly higher than in VP-16 group （both P〈0. 05）. There was no significant changes in the phenotype of MSCs before and after the treatment with amifostine. Conclasion Amifostine alone has no obvious effects on proliferation, apoptosis, immuophenotype or ability of hematopoiesis support of the MSCs in vitro alone, but can protect MSCs from injury by VP-16, suggesting that amifostine can be a new way to improve the recovery of hematopoiesis ability of CML patients subject to stem cells transplantation.