流式细胞仪检测急性婴幼儿白血病干细胞

Determination of leukemia stem cells in childhood acute myeloid leukemia by flow cytometry

背景:有关儿童急性髓细胞性白血病干细胞含量的测定及急性髓细胞性白血病患儿缓解后白血病干细胞含量与急性白血病微小残留病之间关系的研究国内外未见报道。目的:通过测定急性髓细胞性白血病干细胞或急性髓细胞性白血病干细胞-IPIC在儿童急性白血病患儿骨髓单个核细胞中的含量,研究急性髓细胞性白血病患儿缓解后白血病干细胞含量与急性白血病微小残留病水平之间的关联。方法:收集白血病患儿113例次。采用骨髓单个核细胞分离及单细胞悬液制成单细胞悬液,进行单个核细胞染色、急性髓细胞性白血病干细胞分析及根据初诊免疫表型获得白血病相关表型,并采用该白血病相关免疫表型进行单抗组合和流式细胞术测定分析。结果与结论:①初诊急性髓细胞性白血病组骨髓单个核细胞中急性髓细胞性白血病干细胞含量明显高于初诊急性淋巴细胞性白血病组和非肿瘤对照组(P均<0.017),初诊急性淋巴细胞性白血病组骨髓单个核细胞中急性髓细胞性白血病干细胞-IPIC含量显著高于非肿瘤对照组(P<0.017)。②对33例次缓解急性髓细胞性白血病患儿急性髓细胞性白血病干细胞和急性白血病微小残留病相关性分析发现,两者存在显著负相关性。结果提示,①急性髓细胞性白血病干细胞-IPIC也存在于初诊急性淋巴细胞性白血病患儿骨髓细胞中,且当急性淋巴细胞性白血病获完全缓解时急性髓细胞性白血病干细胞-IPIC含量却没有下降,但非肿瘤对照组标本中急性髓细胞性白血病干细胞-IPIC的含量极微。②急性髓细胞性白血病患儿缓解后骨髓中急性髓细胞性白血病干细胞含量和急性白血病微小残留病水平之间存在着明显的负相关。

BACKGROUND： Few studies are reported at home and abroad regarding detection of leukemia stem cell concentration in childhood acute myeloid leukemia and the correlation between leukemia stem cell level in childhood acute myeloid leukemia after remission and minimal residual disease level. OBJECTIVE： To determine leukemia stem cell level and leukemia stem cell-IPIC level in childhood acute myeloid leukemia both at initial diagnosis and at remission and correlate them to minimal residual disease level in childhood acute myeloid leukemia. METHODS： A total of 113 samples from patients with childhood acute myeloid leukemia were collected. All heparinized bone marrow mononuclear cells were separated by cell density gradient centrifugation on ficoll-hypaque solution. After washes with PBS containing 0.1% fetal calf serum, mononuclear cell suspension was prepared. Mononuclear cells were stained with fluochrome labeled monoclonal antibodies. Leukemia stem cell level was determined and leukemia-related immunophenotypes were acquired. Immunophenotype determination and flow cytometry analysis were performed. RESULTS AND CONCLUSION： Leukemia stem cell level in childhood acute myeloid leukemia group at initial diagnosis was significantly higher than that in acute lymphocytic leukaemia and non-malignancy control groups （both P＆lt;0.017）. The leukemia stem cell-IPIC level in childhood acute lymphocytic leukemia at initial diagnosis was significantly higher than that in the non-malignancy control group （P＆lt;0.017）. There was a significant negative correlation between leukemia stem cell level and minimal residual disease level in childhood acute myeloid leukemia. The results indicate that （1） the phenotypically same leukemia stem cell populations were also found in the bone marrow of patients with acute lymphocytic leukaemia at initial diagnosis, but the levels were not significantly different when complete remission was achieved. These phenotypically same populations were hardly found in the non-malignancy control group. （2） There was a significant negative correlation between leukemia stem cell level in childhood acute myeloid leukemia and minimal residual disease level in childhood acute myeloid leukemia patients after remission.