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本研究旨在建立一套血液恶性肿瘤的体外细胞培养模型。构建几种带有GFP(绿色荧光蛋白)表达并能够编码不同白血病/淋巴瘤相关融合蛋白的逆转录病毒载体(如:TEL-PDGFR,Rabaptin5-PDGFR,p210BCR-ABL,AML1-ETO,NPM-ALK);将病毒载体导入小鼠的原始骨髓细胞,转导的骨髓细胞随后培养在含有10%FCS的IMDM中。将细胞分为:无生长因子组和含有不同生长因子的各种组合组:①c-kit联合flt3(KL+FL)组,②IL-3+TPO+G-CSF+Hyper-IL-6(3/T/G/H6)组,③KL+FL+3/T/G/H6组。用流式细胞术检测肿瘤融合蛋白联合各种生长因子支持骨髓转导细胞的自我更新、扩增生长的能力。结果表明,转导的细胞能够持续对数级的生长扩增。KL联合FL能够支持转导编码TEL-PDGFR、Rabaptin5-PDGFR、AML1-ETO、NPM-ALK病毒载体的骨髓细胞自我更新、生长扩增;而转导p210 BCR-ABL病毒载体的骨髓细胞除需要KL和FL外,还需要IL-3的参与。扩增培养的细胞形态与所对应的肿瘤基因相关的血液肿瘤细胞相一致。结论:本研究建立了一套体外模型培养体系,提供了一种研究血液恶性肿瘤的方法,可用于筛选血液肿瘤的治疗性药物。
This study aimed to establish a set of in vitro cell culture models of hematological malignancies. Several retroviral vectors (eg TEL-PDGFR, Rabaptin5-PDGFR, p210BCR-ABL, AML1-ETO, NPM-ALK with GFP (green fluorescent protein) expression and encoding different leukemia / lymphoma associated fusion proteins ); The viral vector was introduced into the mouse primary bone marrow cells and the transduced bone marrow cells were subsequently cultured in IMDM containing 10% FCS. The cells were divided into: non-growth factor group and various combination groups containing different growth factors: ①c-kit combined with flt3 (KL + FL) group, ②IL-3 + TPO + G-CSF + Hyper-IL-6 T / G / H6) group, ③KL + FL + 3 / T / G / H6 group. Flow cytometry was used to detect the ability of tumor fusion protein and various growth factors to support self-renewal and expansion of bone marrow transduced cells. The results show that the transduced cells can continue logarithmic growth expansion. KL combined with FL can support the self-renewal and growth expansion of bone marrow cells transduced with TEL-PDGFR, Rabaptin5-PDGFR, AML1-ETO and NPM-ALK viral vectors, while the bone marrow cells transduced with p210 BCR- And FL, but also the need for IL-3 participation. Expansion of the cultured cell morphology and the corresponding tumor gene-associated blood tumor cells are consistent. Conclusion: This study established a set of in vitro model culture system and provided a method for studying hematological malignancies, which could be used to screen hematological tumors for therapeutic drugs.