EGFP标记的A20细胞构建小鼠B细胞播散型淋巴瘤模型

Establishment of a Disseminated Model of Mouse B-Cell Lymphoma with A20 Cell Lines Expression Enhanced Green Fluorescent Protein

目的:A20细胞是来源于同系Balb/c小鼠大B细胞淋巴瘤的细胞系,能通过尾静脉接种建立小鼠B细胞播散型淋巴瘤的模型,但由于该细胞系缺乏细胞表面特异性标志而难于监测。本研究使用增强型绿色荧光蛋白(EGFP)标记A20细胞,试图建立易于监测的小鼠B细胞播散型淋巴瘤模型。方法:用含增强型绿色荧光蛋白(EGFP)的慢病毒载体将标记基因EGFP转入A20细胞,通过流式分选出EGFP+的A20细胞,体外培养后通过尾静脉注射接种于同系Balb/c小鼠体内,用流式细胞仪监测其外周血EGFP+细胞的百分率。当小鼠出现消瘦、毛发竖立、嗜睡等体征时,将小鼠处以安乐死;取动物脏器行石蜡包埋、病理切片、HE染色。结果:尾静脉注射1×106细胞于6只Balb/c小鼠体内,接种后15天可在外周血中检测到EGFP+细胞,平均生存时间为29.6±0.8天;在肝脏、脾脏、脊椎和淋巴结等多脏器成瘤,流式检测瘤细胞EGFP表达阳性。结论:经尾静脉注射接种A20细胞可建立小鼠B细胞播散型淋巴瘤模型,A20细胞经含EGFP的慢病毒标记后易于通过流式进行监测,为通过动物体内试验评价靶向治疗的疗效提供了保证。

Objective： The A20 cells are B lymphocytes from a syngeneic Balb/c mouse with a large B-cell lymphoma, and a disseminated model of mouse B-cell lymphoma can be established by using A20 cell line via tail vein injection. The model was hard to monitor because of no specific cell surface markers. Our study is to present a disseminated model of mouse B-cell lymphoma easy to be monitored by using A20 cells labled with EGFP. Methods： A20 cells were transfected with a lentivirus vector containing a gene encoding enhanced green fluorescent protein （EGFP）, EGFP＋A20 cells were sorted by FACS and intravenously injected into syngeneic Balb/c mouse, and the percentage of EGFP± cells were evaluated by flow cytometry. The mice were euthanized at the first indication of morbidity （weight loss, ruffed fur, lethargy）. Organs were paragon-imbedded, sectioned and analyzed by microscopy. Results： 1× 10^6 cells were inoculated into 6 Balb/c mouse via tail vein injection, EGFP± cells in peripheral blood were detected 15 days post inoculation. The average time of survival was 29.6 ± 0.8 days. Organs such as liver, spleen, spine and lymph nodes were involved, while flow cytometry showed the tumor cells were positive for EGFP. Conclusion： A disseminated model of mouse B-cell lymphoma can be established by using A20 cell line via tail vein injection, which was easy to be traced after transfection of A20 cells with a lentivirus vector containing a gene encoding enhanced green fluorescent protein （EGFP）, providing a useful experimental platform for further in vivo target therapy researches.