人肝肿瘤细胞的裸小鼠原位癌建模条件优化及评价  

Optimization and Evaluation of Conditions for Orthotopic Nude Mouse Models of Human Liver Tumor Cells

作　　者：孟雨 梁冬丽 郑琳琳 周园园[2] 王朝霞[2] MENG Yu;LIANG Dongli;ZHENG Linlin;ZHOU Yuanyuan;WANG Zhaoxia(School of Life Sciences and Biotechnology,Shanghai Jiao Tong University,Shanghai 200240,China;Laboratory Animal Center,Shanghai Jiao Tong University,Shanghai 200240,China)

机构地区：[1]上海交通大学生命科学技术学院,上海200240 [2]上海交通大学实验动物中心,上海200240

出　　处：《实验动物与比较医学》2024年第5期511-522,共12页Laboratory Animal and Comparative Medicine

摘　　要：目的优化通过注射人肝肿瘤细胞株构建原位癌裸小鼠模型的条件,并探索适宜的给药治疗时间。方法选用稳定表达萤光素酶报告基因(LUC)的人肝细胞癌Hep3B与肝母细胞瘤HepG2细胞株,使用小动物活体成像系统分析萤光素酶发光强度与肝肿瘤细胞数量之间的线性相关性,验证人源肝肿瘤细胞的发光效率。在5周龄雌性BALB/c裸小鼠的肝叶原位接种不同浓度(8×10^(6)、2.4×10^(7)、7.2×10^(7)个/mL)、不同重悬介质(PBS、Matrigel)的人肝肿瘤细胞悬液HepG2-LUC和Hep3B-LUC(共12组,每组7只),分别构建人肝肿瘤裸小鼠原位癌模型。每7 d为1个周期记录各组小鼠体重,用小动物活体成像系统定期监测原位肿瘤的生长过程,观察肿瘤生长趋势。接种肿瘤细胞后第35天剖取小鼠肝脏,制备病理切片,进行苏木精-伊红(hematoxylin and eosin,HE)染色观察组织病理学变化。结果两种人肝肿瘤细胞株的发光强度均与细胞数量呈正相关(R^(2)=0.9831,R^(2)=0.9705),适宜用于原位癌模型的构建。HepG2-LUC高浓度组,HepG2-LUC+Matrigel低、中、高浓度组,Hep3B-LUC中、高浓度组与Hep3B-LUC+Matrigel低、中、高浓度组均成功造模。HepG2-LUC+Matrigel高浓度组较低浓度与中浓度组小鼠的体重显著下降(P<0.05),Hep3B-LUC+Matrigel高浓度组较低浓度与中浓度组小鼠的体重也显著下降(P<0.05)。成功造模组小鼠的荧光发光强度随时间呈指数型增长(R^(2)>0.9500),且在移植后14 d发光强度至少可达到1.0×10^(7) p/(s·cm^(2)·sr)。HepG2-LUC低、中浓度组和Hep3B-LUC低浓度组小鼠肝脏未见明显的病理学变化,其余组肝脏肿瘤和肝细胞病变明显。结论对于HepG2-LUC细胞株,推荐肝叶原位注射2.4×10^(7)个/mL(50μL)且与Matrigel重悬的混合细胞液体造模,并于造模后第7天给药或采取预后措施;而对于Hep3B-LUC细胞株,推荐肝叶原位注射7.2×10^(7)个/mL(50μL)(不与Matrigel重悬混合)造模,并于造模后�Objective The study aims to optimize the conditions for constructing orthotopic nude mouse models of liver cancer by injecting human liver tumor cell lines and to explore appropriate timings for drug administration.Methods Human hepatocellular carcinoma Hep3B and hepatoblastoma HepG2 cell lines,which stably expressing the luciferase reporter gene(LUC),were selected.The linear correlation between the luciferase luminescence intensity and the number of liver tumor cells was analyzed using a Small Animal In Vivo Imaging system to verify the luminescent efficiency of the human liver tumor cells.Different concentrations(8×10^(6),2.4×10^(7),7.2×10^(7) cells/mL)and resuspension media(PBS,Matrigel)of human liver tumor cell suspensions HepG2-LUC and Hep3B-LUC were orthotopically inoculated into the liver lobes of 5-week-old female BALB/c nude mice(12 groups,7 mice each)to construct human liver tumor nude mouse orthotopic cancer models.Every 7 days,the weights of mice were recorded,and the growth of orthotopic tumors was monitored using the Small Animal In Vivo Imaging system.On day 35 post-cell inoculation,mouse livers were dissected,and pathological slices were prepared for HE staining to observe histopathological changes in liver tissues.Results The luminescence intensity of human liver tumor cell lines was positively correlated with the number of cells(R^(2)=0.9831,R^(2)=0.9705),indicating their suitability for orthotopic model construction.Successful modeling was achieved in the high-concentration groups of HepG2-LUC,the low-,medium-,and high-concentration groups of HepG2-LUC+Matrigel,the medium-and high-concentration groups of Hep3B-LUC,and the low-,medium-,and high-concentration groups of Hep3BLUC+Matrigel.For both HepG2-LUC+Matrigel and Hep3B-LUC+Matrigel groups,mice in the highconcentration groups exhibited significantly reduced body weight compared to the low-and mediumconcentration groups(both with P<0.05).The luminescence intensity of successfully modeled mice increased exponentially over time(R^(2)>0.9500),

关 键 词：肝癌 原位移植 HepG2-LUC Hep3B-LUC BALB/C裸小鼠 

分 类 号：R73-3[医药卫生—肿瘤] Q95-33[医药卫生—临床医学]