CAJ | 학술논문

目的通过慢病毒转染C6胶质瘤细胞建立稳定表达绿色荧光蛋白(EGFP)的C6细胞株.方法用含2个启动子延长因子-1α(EF-1α)和CMV的慢病毒转染C6胶质瘤细胞,荧光显微镜下观察表达EGFP的C6细胞,噻唑蓝(MTT)法比较C6和EGFP-C6胶质瘤细胞生长曲线;建立裸鼠皮下和大鼠颅内胶质瘤模型评价其致瘤性,测量肿瘤体积、苏木素-伊红(HE)染色、免疫组织化学法检测胶质纤维酸性蛋白(GFAP)染色、磁共振成像(MRI)监测肿瘤体积.结果在倒置和荧光显微镜下观察EGFP-C6细胞形态与C6细胞相似,MTF方法检测其细胞增殖能力(0.53±0.02)与C6细胞(0.51±0.01)相似,裸鼠皮下模型结果表明C6细胞和EGFP-C6细胞的肿瘤体积[(2.20±0.91) cm3比(2.10 ±0.84) cm3]、HE染色、GFAP的表达未见明显区别,并且颅内模型结果也表明,C6细胞和EGFP-C6细胞的肿瘤体积[(19.5±2.3)cm3比(20.5±2.3) cm3]、荷瘤鼠生存期、成瘤率及颅外转移率未见明显区别.结论成功建立以EGFP为生物标记的C6细胞株,其生物学特性与C6细胞相似.
목적 통과만병독전염C6효질류세포건립은정표체록색형광단백(EGFP)적C6세포주.방법 용함2개계동자연장인자-1α(EF-1α)화CMV적만병독전염C6효질류세포,형광현미경하관찰표체EGFP적C6세포,새서람(MTT)법비교C6화EGFP-C6효질류세포생장곡선;건립라서피하화대서로내효질류모형평개기치류성,측량종류체적、소목소-이홍(HE)염색、면역조직화학법검측효질섬유산성단백(GFAP)염색、자공진성상(MRI)감측종류체적.결과 재도치화형광현미경하관찰EGFP-C6세포형태여C6세포상사,MTF방법검측기세포증식능력(0.53±0.02)여C6세포(0.51±0.01)상사,라서피하모형결과표명C6세포화EGFP-C6세포적종류체적[(2.20±0.91) cm3비(2.10 ±0.84) cm3]、HE염색、GFAP적표체미견명현구별,병차로내모형결과야표명,C6세포화EGFP-C6세포적종류체적[(19.5±2.3)cm3비(20.5±2.3) cm3]、하류서생존기、성류솔급로외전이솔미견명현구별.결론 성공건립이EGFP위생물표기적C6세포주,기생물학특성여C6세포상사.
Objective To establish a stable C6 glioma line expressing enhanced green fluorescent protein (EGFP).Methods C6 glioma cells were transfected with the lentivirus vector enhancer promoter CMV and elongation factor-1α (EF-1α).C6 cells expressing EGFP were observed under the fluorescence microscopy and screened by flow cytometry.Tumorigenicity was evaluated by establishment of mouse skin and intracranial glioma model.Tumor volume was measured,and hematoxylin and eosin (HE) staining,immunohistochemical glial fibrillary acidic protein (GFAP) staining and MRI were used to monitor the tumor volume.Results Morphology observed under a fluorescence microscope and proliferation detected by methyl thiazol tetrazolium (MTT) of EGFP-C6 cells were similar to those of C6 cells (0.53 ±0.02 vs.0.51 ± 0.01).Subcutaneous model results showed there were no significant differences in tumor volume [C6:(2.20 ±0.91) cm3 and EGFP-C6:(2.10 ±0.84) cm3],HE staining and GFAP staining between EGFP-C6 and C6 cells.Intracranial model results showed there were no significant differences in tumor volume calculated by MRI [C6:(20.5±2.3) cm3 and EGFP-C6:(19.5±2.3) cm3],the survival of tumor-bearing rats,tumor formation rate,and the extracranial metastasis rate.Conclusion EGFP-C6 cell line expressing EGFP as a marker was established.Biological characteristics of EGFP-C6 cells were similar to those of C6 cells.