摘要
目的：Rb94基因转染裸鼠肺腺癌皮下移植瘤，观察Rb94基因对人肺腺癌裸鼠移植瘤的抑制作用及对鞘氨醇激酶（sphingosine kinase，SphK）表达的影响。方法：收集一定数量的对数生长期A549细胞，生理盐水重悬后每只裸鼠右后肢皮下接种5×10^6／0．1mL A549细胞悬液，待瘤体生长至一定体积后随机分为4组，分别为对照组、空载体组、脂质体组、转染Rb94基因组；每组裸鼠瘤体内分别注射PBS、质粒pIRES、脂质体（lipofectamine）、质粒pIRES～Rb94，观察肿瘤生长情况并检测抑瘤率；裸鼠脱颈处死后，采用Western bolt法检测瘤组织中Rb94基因表达情况，并采用实时RT—PCR和免疫荧光法（immunonuorescence，IF）观察其对裸鼠肺腺癌皮下移植瘤中SphK表达的影响。结果：对照组及转染组裸鼠在接种部位均有移植瘤生成；与空白对照组、脂质体组、空载体组相比，转染Rb94基因组的裸鼠移植瘤的生长速度明显减慢，肿瘤的体积较小，重量较轻，有显著性差异（P〈0．05），而3个对照组之间，无显著性差异（P〉0．05）；空白对照组、空载体组、脂质体纽抑瘤率分别为0、11．78％、7．86％，而转染Rb94基因组平均抑瘤率高达51．99％，肿瘤生长明显抑制：SohK1在mRNA和蛋白水平表达下调而SohK2表达上调。结论：Rb94基因成功转染裸鼠肺腺癌皮下移植瘤；Rb94基因抑制裸鼠皮下肿瘤生长；Rb94基因抑制SphK1的表达，增强SphK2表达。
Objective： To construct an animal model of plRES-Rb94 transfection of the A549 adenocarcinoma cells to, investigate the inhibitory action and effect on the expression of SphK in a mouse xenograft model. Methods： Cell suspensions at 5×10^6/0.1mL were subcutaneously injected into flanks of every nude mouse. Then the inoculated mice were divided into 4 groups as follows： with PBS; with lipofectamine; with plRES empty vector; and with plRES-Rb94. The inhibitory rate was observed and the expression of the Rb94 gene was detected by Western blot. Then its effect on SphK expression was determined by RT-PCR and immunofluorescence. Results： The growth of tumor graft was obviously inhibited in the treatment group. The inhibitory ratio of the transplanted tumor was 51.99% in the pIRES-Rb94 group, higher than that in the control group （0）, lipofectamine group （7.86%） and plRES group （11.78%）, with significant differences between plRES-Rb94 group and the other three groups （P〈0.01）, The expression of SphK1 was down-regulated and the expression of SphK2 was up-regulated. Conclusion： Mouse xenograft model of lung adenocaroinoma was successfully constructed using eukaryotic expression plasmid containing plRES-Rb94 gene. Rb94 can effectively inhibit the growth of lung adenocarcinoma and the expression of SphK1 but can enhance SphK2 expression.
出处
《中国肿瘤临床》
CAS
CSCD
北大核心
2010年第9期481-484,共4页
Chinese Journal of Clinical Oncology
基金
山东省科技攻关项目资助(编号:2006GG3202008)~~
