HOXA4调控Wnt/β-catenin信号通路对裸鼠移植胶质瘤的抑制作用

Inhibitory effect of HOXA4 on transplantation glioma of nude mice through regulating Wnt/β-catenin signal pathway

目的:建立人胶质瘤荷瘤裸鼠模型,探讨HOXA4对胶质瘤U251细胞体内生长的影响以及对Wnt/β-catenin信号通路的调控作用及其机制。方法:采用慢病毒转染,建立胶质瘤U251细胞稳转同源异型盒基因A4 (HOXA4)siRNA细胞系(si-HOXA4)和稳转空载体阴性对照U251细胞系(si-NC)。取对数生长期U251、si-NC和si-HOXA4细胞接种于BALB/c裸鼠颈背部皮下,建立荷瘤裸鼠模型,命名为对照组、si-NC组和si-HOXA4组。观察各组裸鼠成瘤情况并绘制肿瘤生长曲线;培养21d处死裸鼠后剥离肿瘤组织,测量肿瘤体积和质量;qRT-PCR法检测各组裸鼠肿瘤组织中HOXA4、CTNNB1和Gsk3βmRNA相对表达量;免疫组织化学(IHC)法检测各组裸鼠肿瘤组织中HOXA4、β-catenin、Gsk3β、CyclinD1和P53蛋白表达水平。结果:si-HOXA4组裸鼠肿瘤体积和质量小于si-NC组和对照组(P<0.05);si-HOXA4组裸鼠肿瘤组织中HOXA4mRNA相对表达量和HOXA4蛋白表达水平低于其他2组(P<0.05);与si-NC组和对照组比较,si-HOXA4组裸鼠肿瘤组织中CTNNB1mRNA相对表达量降低(P<0.05),β-catenin和CyclinD1蛋白表达水平亦降低(P<0.05),但Gsk3β和P53蛋白表达水平明显升高(P<0.05)。结论:抑制人胶质瘤U251细胞HOXA4表达可在体内通过Wnt/β-catenin信号通路调控CyclinD1和P53蛋白的表达,从而抑制荷瘤裸鼠肿瘤的生长。

Objective:To establish the glioma-bearing nude mouse models,and to investigate the effect of HOXA4 on the growth of glioma U251 cells in vivo and its regulatory effect on the Wnt/β-catenin signal pathway and its mechanism.Methods:The glioma U251 cell line stably transfected with HOXA4 siRNA(si-HOXA4)and the U251 cell line stably transfected with blank vector(si-NC)were established by lentivirus transfection.The U251,si-NC,and si-HOXA4 cells were respectively inoculated under the skin of the neck and back of the BALB/c nude mice to establish the glioma-bearing nude mouse models named as control group,si-NC group,and si-HOXA4 group.The tumorigenesis of nude mice in various groups were observed and the tumor growth curve was drawn.The tumor tissue was stripped after the mice were sacrificed on the 21 th day,and the volume and weight of tumor were measured;the relative mRNA expression amounts of HOXA4,CTNNB1,and Gsk3βin tumor tissue of the nude mice in various groups were detected by qRT-PCR method;the expression levels of HOXA4,β-catenin,Gsk3β,CyclinD1,and P53 proteins in tumor tissue of the nude mice in various groups were detected by immunohistochemistry(IHC)method.Results:Compared with si-NC group and control group,the volume and weight of tumor of the nude mice in si-HOXA4 group were significantly decreased(P<0.05).The relative expression amount of HOXA4 mRNA and the expression level of HOXA4 protein in si-HOXA4 group were significantly lower than those in the other groups(P<0.05).Compared with si-NC group and control group,the relative expression amount of CTNNB1 mRNA and the expression levels ofβ-catenin and CyclinD1 proteins in si-HOXA4 group were significantly decreased(P<0.05),and the expression levels of Gsk3β and P53 proteins were significantly increased(P<0.05).Conclusion:Inhibition of HOXA4 expression in human glioma U251 cells can regulate the expressions of CyclinD1 and P53 through Wnt/-catenin signal pathway in vivo,thus inhibiting the tumor growth of glioma-bearing nude mice.