干扰HMGB1表达影响视网膜母细胞瘤细胞恶性生物学行为的实验研究

Effects of interfering with HMGB1 expression on malignant biological behavior of retinoblastoma cells

目的探讨高迁移率族蛋白1(HMGB1)对视网膜母细胞瘤(RB)细胞增殖、凋亡等行为的调控作用及机制。方法检测比较RB细胞株Y79细胞和正常视网膜上皮细胞株ARPE-19细胞HMGB1表达情况。通过转染慢病毒HMGB1-siRNA干扰Y79细胞HMGB1表达,分为对照组、siRNA NC组和siRNA HMGB1组,分析对细胞增殖、凋亡及细胞周期的影响。利用裸鼠成瘤模型研究干扰HMGB1表达对RB肿瘤生长的影响,并分析Toll样受体4(TLR4)、髓样分化因子88(MyD88)、核转录因子-κB(NF-κB)表达情况,探讨其作用机制。结果Y79细胞中HMGB1表达明显高于ARPE-19细胞(P<0.05),干扰HMGB1表达明显抑制Y79细胞增殖。siRNA HMGB1组细胞凋亡率明显高于对照组和siRNA NC组(P<0.05)。与对照组和siRNA NC组比较,siRNA HMGB1组细胞G_(0)/G_(1)期比例明显增高,S期和G_(2)/M期明显减少(P<0.05)。裸鼠成瘤模型显示siRNA HMGB1组肿瘤体积和质量均低于对照组(P<0.05)。与ARPE-19细胞比较,Y79细胞TLR-4、MyD88及NF-κB蛋白和mRNA水平均明显增高(P<0.05)。与对照组和siRNA NC组比较,siRNA HMGB1组细胞TLR-4、MyD88及NF-κB蛋白和mRNA水平均明显降低(P<0.05)。结论干扰RB细胞HMGB1表达可通过TLR-4/MyD88/NF-κB信号通路抑制细胞增殖、促进细胞凋亡。

Objective To investigate the regulation effect and mechanism of high mobility group box-1 protein(HMGB1)on the proliferation and apoptosis of Retinoblastoma(RB)cells.Methods The expression of HMGB1 in RB cell line Y79 cells and normal retinal epithelial cell line ARPE-19 cells,was detected and compared.The expression of HMGB1 was interfered by transfection of lentivirus HMGB1-siRNA in Y79 cells,divided into the control group,the siRNA NC group and the siRNA HMGB1 group,and the effects on cell proliferation,apoptosis and cell cycle were analyzed.The effect of interfering HMGB1 expression on RB growth was studied in nude mouse tumorigenic model,and the expression of toll-like receptor 4(TLR4),myeloid differentiation factor 88(MyD88)and nuclear transcription factor-κB(NF-κB)was analyzed to explore the mechanism.Results The expression of HMGB1 in Y79 cells was significantly higher than that in ARPE-19 cells(P<0.05).The interference with HMGB1 expression significantly inhibited the proliferation of Y79 cells.The apoptosis rate in siRNA HMGB1 group was significantly higher than that in the control group and siRNA NC group(P<0.05).Compared with the control group and siRNA NC group,the proportion of G_(0)/G_(1) phase cells in siRNA HMGB1 group was significantly increased,while S phase and G_(2)/M phase cells were significantly decreased(P<0.05).Nude mouse tumor model showed that the tumor volume and weight in siRNA HMGB1 group were lower than those in the control group(P<0.05).Compared with ARPE-19 cells,TLR-4,MyD88,NF-κB,and mRNA levels in Y79 cells were significantly increased(P<0.05).Compared with the control group and the siRNA NC group,TLR-4,MyD88,NF-κB,and mRNA levels in the siRNA HMGB1 group were significantly decreased(P<0.05).Conclusion Interfering with HMGB1 expression can inhibit RB cells can inhibit cell proliferation and promote apoptosis through TLR-4/MyD88/NF-κB signal pathway.