川芎嗪抑制人视网膜母细胞瘤WERI-Rb1细胞增殖的机制研究

Effects of tetramethylpyrazine on human retinoblastoma cell line WERI-Rb1 and its mechanism

目的探索川芎嗪(TMP)对WERI-Rb1细胞增殖的影响及TMP下调人WERI-Rb1细胞中趋化因子受体4(CXCR4)的转录的分子机制。方法 (1)TMP及CXCR4的拮抗剂AMD3100药物分别处理WERIRb1细胞,相同浓度PBS及DMSO作为对照,活细胞成像仪观察WERI-Rb1细胞形态及密度变化。(2)WERIRb1细胞经TMP处理后,采用荧光定量逆转录PCR、免疫荧光及蛋白免疫印迹法分别从mRNA和蛋白质水平检测TMP对WERI-Rb1细胞CXCR4及NRF1表达的影响。结果 (1)活细胞成像仪观察发现TMP处理组与AMD3100处理组的细胞状态较正常组差,增殖明显受抑制。(2)荧光定量逆转录PCR结果显示:与对照组比较,TMP处理组WERI-Rb1细胞中CXCR4和NRF1mRNA表达均明显下降(P<0.05);免疫荧光及蛋白免疫印迹实验结果显示,与对照组比较,TMP处理组WERI-Rb1细胞中CXCR4和NRF1蛋白表达均明显下降(P<0.05)。结论TMP对WERI-Rb1细胞的增殖有明显抑制作用,且能下调WERI-Rb1细胞中CXCR4及NRF1的表达水平。

Objective To characterize the bioactivity of tetramethylpyrazine( TMP) on human WERI- Rb1 cells and its transcription regulatory mechanism of TMP- mediated down- regulation of CXCR4. Methods( 1) WERI- Rb1 cells were treated with TMP and AMD3100. The control cells were treated with DMSO or PBS at the same concentration.The morphology and cell density of WERI- Rb1 cells was observed and recorded by the living cells imager.( 2) WERI-Rb1 cells were treated with TMP; then real- time RT- PCR,immunofluorescence and western blot were performed to detect the expression levels of CXCR4 and NRF1 in WERI- Rb1 cells treated with TMP. Results( 1) TMP and AMD3100 notably inhibited the proliferation of WERI- Rb1 cells,as evidenced by cell morphology and density analysis.( 2) TMP down- regulated CXCR4 and NRF1 expression in WERI- Rb1 cells at both mRNA and protein levels( P < 0. 05),as evidenced by real- time RT- PCR,immunofluorescence and western blot assays. Conclusion TMP significantly inhibits the proliferation of WERI- Rb1 cells and down- regulates its expression of CXCR4 and NRF1.