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在构建稳定表达荧光素酶的人胰腺癌PANC-1细胞的基础上,建立适合活体成像研究的胰腺癌动物模型,为深入研究胰腺癌发病机制打下基础。该研究运用Lonza NucleofectorTM核转染系统将携带有荧光素酶基因(firefly luciferase,luc)的质粒(GV258)稳定转染PANC-1细胞,利用嘌呤霉素筛选出能稳定表达荧光素酶的细胞株PANC-1-LUC,并应用活体成像系统对其体内外生物发光进行检测。进一步构建PANC-1-LUC裸鼠皮下移植瘤,同样方法接种慢病毒载体构建的PANC-1-luc细胞作为对照,观察成瘤率、瘤体积,检测生物发光强度。体内外生物发光检测结果均显示,光通量值与细胞数之间呈显著的直线相关(P<0.01)。PANC-1-LUC细胞皮下接种裸鼠,1周成瘤率即达80%以上,瘤体积可达平均100 mm3。观察6周,裸鼠皮下肿瘤生长符合Gompertzian曲线。每周的生物发光检测结果均显示,PANC-1-LUC移植瘤的光强度与瘤体积有显著的线性关系(P<0.05)。以上结果说明,该文所构建的PANC-1-LUC细胞株能够稳定、长期地表达荧光素酶,更适合构建活体成像研究的裸鼠胰腺癌模型。
Based on the construction of human pancreatic cancer PANC-1 cells stably expressing luciferase, an animal model of pancreatic cancer suitable for in vivo imaging studies was established to lay a foundation for further studies on the pathogenesis of pancreatic cancer. In this study, a Lonza NucleofectorTM nucleofection system was used to stably transfect PANC-1 cells with a plasmid (GV258) carrying a luciferase gene (luci), and a cell line that stably expressed luciferase was selected using puromycin. PANC-1-LUC, and its in vivo and in vitro bioluminescent assays were performed using in vivo imaging systems. Subcutaneously implanted tumors of PANC-1-LUC in nude mice were further constructed. The same method was used to inoculate PANC-1-luc cells constructed with lentiviral vectors as controls. The rate of tumor formation, tumor volume, and bioluminescence intensity were measured. Both in vitro and in vivo bioluminescent assays showed that there was a significant linear correlation between light flux and cell number (P<0.01). Nude mice were inoculated subcutaneously with PANC-1-LUC cells. The tumorigenesis rate reached 80% at 1 week, and the average tumor volume was 100 mm3. At 6 weeks, the growth of subcutaneous tumors in nude mice was consistent with the Gompertzian curve. Weekly bioluminescence assays showed a significant linear relationship between light intensity and tumor volume in PANC-1-LUC xenografts (P<0.05). The above results show that the PANC-1-LUC cell line constructed in this paper can stably and long-term express luciferase, and it is more suitable for constructing a pancreatic cancer model of nude mice in vivo imaging studies.