人红白血病斑马鱼异种模型的建立

Establishment of a zebrafish xenogeneic model of human erythroleukemia

目的利用斑马鱼胚胎建立人红白血病(HEL)细胞异种移植瘤模型,提供研究血液肿瘤的体内行为和实用药物测试的模型。方法将HEL细胞分为空白组和CM-DiI组,CM-DiI组用活细胞示踪剂进行标记;取野生型斑马鱼培育繁殖,产胚后将胚胎置于28℃孵育48 h至破膜,将破膜后斑马鱼分为空白对照组、PBS实验组、HEL实验组,将CM-DiI组细胞,通过显微注射方式移植到HEL实验组斑马鱼卵黄囊中,PBS实验组注射PBS,再将三组斑马鱼置于34℃温箱培养,通过体内外增殖实验及体内荧光观察等鉴定模型是否建立成功;将鉴定成功的斑马鱼进行阿糖胞苷药物暴露,分为50、100、200 nmol/L组及模型对照组,观察各组浓度药物暴露后异种移植瘤模型中白血病细胞的增殖及迁移情况。结果 CM-Dil组与空白组HEL细胞增殖状态相近,差异无统计学意义(P>0.05);通过体内外增殖实验证实了HEL细胞在斑马鱼体内出现了增殖,而HEL实验组斑马鱼存活量相对于空白对照组和PBS实验组降低(P<0.05);三个浓度的阿糖胞苷药物暴露组相对于模型对照组细胞增殖降低(P<0.05)。结论人急性红白血病HEL细胞移植入斑马鱼胚胎后可进行增殖,且增殖活动可被阿糖胞苷干预,证实异种移植瘤模型建立成功。

Objective A human erythroleukemia(HEL) cell xenograft model was established using zebrafish embryos to provide a model for studying in vivo behavior and practical drug testing of hematological tumors.Methods Divide the HEL cells into a blank group and a CM-DiI group. The CM-DiI group was labeled with a live cell tracer.Wild-type zebrafish were cultivated and reproduced. After embryogenesis, the embryos were incubated at 28 ℃ for 48 h until the membrane was broken. The zebrafish were divided into blank control group, PBS experimental group,and HEL experimental group. Cells of CM-DiI group were transplanted into zebrafish yolk sac of HEL experimental group by microinjection,and PBS experimental group was injected with PBS. The three groups of zebrafish were cultured in an incubator at 34 ℃,and whether the model was established successfully was identified through in vivo and in vitro proliferation experiments and in vivo fluorescence observation,the successfully identified zebrafish were exposed to cytarabine drug,divided into 50,100,200 nmol/L group and model control group,the proliferation and migration of leukemia cells in xenograft tumor models after drug exposure in each group were observed. Results The proliferation status of HEL cells in the CM-Dil group and the blank group was similar,and the difference was not statistically significant(P>0.05). The proliferation of HEL cells in zebrafish was confirmed by in vitro and in vivo proliferation experiments;The number of zebrafish in the HEL experimental group was lower than that in the blank control group and PBS injection group(P<0.05). Compared with the model control group,the proliferation of the three concentrations of cytarabine drug exposure group was lower(P<0.05). Conclusion Human acute erythroleukemia HEL cells can be proliferated after transplantation into zebrafish embryos,and the proliferative activity can be interfered by cytarabine,confirming the successful establishment of the xenograft model.