干扰VEGF表达联合放射线治疗食管癌细胞移植瘤的实验研究

【作者】 封巍； 郑晓；

【机构】 浙江省肿瘤医院放疗科十一病区；

【摘要】 目的分别研究两种干扰VEGF表达方法联合照射对裸鼠移植瘤的影响,并观察各组裸鼠肿瘤病理组织学和细胞生物学特征的变化,为VEGF基因治疗联合放射治疗食管癌提供理论依据。方法使用已转染反义VEGFcDNA肿瘤细胞,建立裸鼠爪垫建立移植瘤模型。使用未转染的TE-1肿瘤细胞建立裸鼠爪垫移植瘤模型,在裸鼠移植瘤照射结束后开始使用反义VEGF寡核苷酸加脂质体转染。将32只BALB/c/nu裸鼠随机分为两大组8小组,每小组4只裸鼠:反义VEGFcDNA组分为①对照组(TE-11组)②反义VEGFcDNA组(TE-1-A组)③单纯照射组(TE-1-R1组)④反义VEGFcDNA加照射组(TE-1-AR组)。反义组寡核苷酸组分为①对照组(TE-12组)②反义VEGF寡核苷酸组(TE-1-AO组)③单纯照射组(TE-1-R2组)④反义VEGF寡核苷酸加照射组(TE-1-AOR组)。观察各组裸鼠移植瘤照射后的生长情况,移植瘤体积变化情况,应用RT-PCR、western blotting检测移植瘤组织中VEGF的表达和透射电镜分析裸鼠肿瘤组织中细胞凋亡情况。结果照射前:已转染入反义VEGFcDNA的组与其它组相比较成瘤潜伏期延长、瘤体生长速度慢。照射后:在反义VEGFcDNA组:可以看到TE-1-A组与TE-1-AR组肿瘤平均体积差别较小,而此二组与对照、单纯照射组差别较明显。反义组寡核苷酸组:TE-1-AOR组与其它3组瘤体平均体积差别非常明显。两大组中:反义组与反义加照射组裸鼠体内TE-1细胞的VEGF mRNA与蛋白表达产物相对光密度比值明显小于对照组和单纯照射组,凋亡增加。结论抗VEGF治疗可有效地抑制裸鼠移植瘤的生长,对增加裸鼠移植瘤放射治疗效果有一定作用。更多还原

【Abstract】 Objective To investigate the effect of antisense VEGF on the radiation sensitivity of esophageal cancer ceils in vivo. Methods 32 male BLBA/C/nu nude mice were randomly divided into eight groups and eight kinds of prepared cells were subcutaneously injected on the paw pat of mice(2×10 6/100μl/mice):The volume of the subcutaneous tumors was measured before and after irradiated.The expression level of VEGF mRNA and protein were examined in different groups by RT-PCR,western blotting respectively.Apoptotic cell and cell cycle distribution were detected in transfected cells by FCM assay and electron micrographs. Results In the tumor formation experiment of nude mice.Latent period of the tumor formation of the antisense-cDNA group was lengthened than that of control group or irradiation group.After irradiation,TE- 1-A group and TE-1-AR groups’ average volume are similar,but these two groups’ tumor average volume were smaller than the other groups.After radiation,TE-1-AOR group has the smallest average volume.TE-1-R2 group average volume was smaller than TE-1-AO.The decreased expression level of endogenous VEGFmRNA and protein were found in antisene group by RT-PCR and western blotting analysis.Apoptotic cells were found in antisense group cells by FCM assay..Observing by TE-1 cells in antisense group showed the characteristics of apoptosis. Conclusions Antisense VEGF could inhibit the proliferation of esophageal cancer cell in the nude mice. Combining Antisense VEGF and radiotherapy could get better control for esophageal cancer,but this aspect need to explore farther.更多还原