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利用人血管内皮细胞的细胞核而建立的体外转录分析体系可灵敏地定量测定血管生成素促进的RNA转录。反应体系中血管内皮细胞核先在缓冲液中与血管生成素混合 ,然后加四种核糖核苷三磷酸混合物启动反应 ,并以 [α 3 2 P]CTP作为示踪剂确定新生RNA的产量。结果显示 ,该体外分析体系的最适反应温度为 30℃ ,最佳反应时间是 30min ,在此反应条件下血管生成素可定量促进RNA转录 ,并存在着剂量效应。研究发现 ,体系中过高浓度的血管生成素降解RNA产物 ,提示细胞具有控制该因子在细胞核内积聚的生物学机制 ,以保证其在细胞内恰当地发挥作用。抑制血管生成素诱导的RNA转录可能可以抑制血管新生 ,因而可能是治疗肿瘤的一个新的分子靶。
An in vitro transcription assay system that utilizes the nucleus of human vascular endothelial cells enables sensitive quantitative determination of angiogenin-promoted RNA transcription. In the reaction system, the vascular endothelial cell nucleus was first mixed with angiogenin in buffer, and then four kinds of ribonucleoside triphosphates were added to start the reaction. [Α 3 2 P] CTP was used as tracer to determine the yield of newborn RNA. The results showed that the optimum reaction temperature was 30 ℃ and the optimal reaction time was 30 min. Under the reaction conditions, angiogenin could promote RNA transcription quantitatively, and there was dose effect. The study found that high concentrations of angiogenin in the system degraded RNA products, suggesting that cells have a biological mechanism that controls the accumulation of this factor in the nucleus to ensure that it functions properly within the cell. Inhibition of angiogenin-induced RNA transcription may inhibit angiogenesis and may therefore be a new molecular target for the treatment of tumors.