Objective: To establish the ADAR1 (adenosine deaminase that act on RNA 1) knockout MLL-AF9 acute myeloid leukemia (AML) mouse model, and to preliminarily investigate the effects of ADAR1 deletion on the development of AML.
Methods: The lineage⁻ (Lin⁻) cells of ER-CreADAR1(lox/lox) mice and their ADAR1(lox/lox) counterparts were enriched by magnetic activated cell sorting (MACS) and then transduced with retrovirus carrying MSCV- MLL/AF9-IRES-GFP fusion gene. The efficiency of transduction was detected by flow cytometry, and equal number of GFP⁺ cells were transplanted into lethally irradiated recipient mice. The recipient mice were treated with tamoxifen at 48 hours after transplantation to induce ADAR1 knockout and divided into following groups: experimental group (ER-Cre;ADAR1(lox/lox)+tamoxifen), control groups ((1)ER-Cre;ADAR1(lox/lox)+vechile, (2)ADAR1(lox/lox)+tamoxifen, (3)ADAR1(lox/lox)+vechile). The percentage of GFP⁺ cells in peripheral blood was examined at 10, 15 and 20 days respectively after transplantation and the survival of the recipient mice was observed. In vitro study, ER-Cre;ADAR1(lox/lox) and ADAR1(lox/lox) AML cells were cultured and the apoptosis rates of these cells 48 hours after 4-hydroxytamoxifen treatment were examined.
Results: The ADAR1 deletion MLL-AF9 AML mouse model was successfully established. Deletion of ADAR1 could decrease the percentage of GFP⁺ cells in the peripheral blood and significantly prolong the survival rate of recipient mice（P<0.05）. In vitro study showed that the cultured total cell number, percentage of GFP⁺ cells decreased and the apoptosis rate of AML cells increased.
Conclusion: Ablation of ADAR1 could delay the progression of AML in recipient mice. ADAR1 plays a critical role in the development and maintenance of murine MLL-AF9 AML.
目的: 建立敲除RNA腺苷脱氨酶1(ADAR1)的小鼠MLL-AF9融合基因急性髓系白血病（AML）模型，初步探讨ADAR1对AML发病的影响。
方法: 采用免疫磁珠法富集介导雌激素受体-重组酶Cre (ER-Cre)的ADAR1lox/lox及其对照ADAR1lox/lox小鼠骨髓Lineage−（Lin−）细胞，用携带MSCVMLL/AF9-IRES-GFP的逆转录病毒感染上述Lin−细胞，流式细胞术检测感染效率，移植相同数量细胞至致死剂量和半致死剂量照射受体小鼠中，建立MLL-AF9诱导的AML模型。移植48 h后诱导ADAR1敲除，体内实验分为实验组（ER-Cre;ADAR1lox/lox＋他莫昔芬）和对照组(①ER-Cre;ADAR1lox/lox +空载体、②ADAR1lox/lox+他莫昔芬、③ADAR1lox/lox＋空载体)，第10、15、20天分别检测小鼠外周血GFP+细胞比例，观察各组小鼠的存活情况。体外实验分组同上，将他莫昔芬改为4-羟基他莫昔芬，观察各组AML细胞并检测其凋亡情况。
结果: 成功建立敲除ADAR1的MLL-AF9融合基因AML小鼠模型。与对照组比较，体内实验中实验组AML小鼠在各时间点外周血GFP+细胞比例均降低，存活时间明显延长，差异均有统计学意义(P值均<0.05)；体外实验中实验组细胞总数、GFP+细胞比例均降低，Annexin Ⅴ+7-AAD+和Annexin Ⅴ+细胞比例均升高，差异均有统计学意义(P值均<0.05)。
结论: 敲除ADAR1可减缓AML的发病，增加AML细胞凋亡。ADAR1在MLL-AF9诱导的AML发生和维持过程中起关键作用。