| 摘要: |
| 目的探讨去甲斑蝥素（NCTD）对人真皮淋巴管内皮细胞（HDLEC）体外三维培养淋巴管生成的影响及机制。方法建立HDLEC体外培养淋巴管生成模型，随机分为NCTD组（加1/3IC50NCTD40nM）与空白对照组，作用24h，观察两组淋巴管生成的形态变化；并采用免疫细胞化学、Westernblot试验和qPCR技术测定比较两组淋巴管生成因子（VEGF-C、VEGF-D和VEGFR-3）蛋白和基因表达。结果NCTD组的HDLEC细胞形态发生改变，很少形成管状结构，时间延长后细胞出现变圆、离壁、浮起、死亡；且NTCD组淋巴管生成（10.9±2.3）个，明显少于空白对照组（68.4±5.2）个（P＜0.01）。免疫细胞化学染色、Westernblot试验结果显示，NCTD对HDLEC三维培养淋巴管生成中的VEGF-C和VEGF-D的蛋白表达下调（P＜0.01）。qPCR检测显示，NCTD对VEGF-C、VEGF-D和VEGFR-3的基因表达明显降低（P＜0.01）。结论NCTD对通过下调淋巴管生成因子VEGF-C、VEGF-D的蛋白和基因表达，产生抗淋巴管生成的作用。 |
| 关键词: 去甲斑蝥素 人真皮淋巴管内皮细胞 三维培养 抗淋巴管生成 |
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| Inhibitory effect of norcantharidin on human dermal lymphatic endothelial cell in three-dimensional culture |
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| Abstract: |
| Objective To study the effect of norcantharidin (NCTD) on human dermal lymphatic epithelial cell (HDLEC) in three-dimensional culture and related molecular mechanisms. Methods Three dimensional model of lymphangiogenesis with HDLEC was established to mimic lymphatic capillary-like structures in vitro. The three-dimensional cultured HDLEC were randomly assigned to control group(no NCTD) and NCTD group(cultured with 40 nm NCTD) for 24 h. The morphological changes were observed by inverted microscope , the expressions of VEGF-C, VEGF-D, VEGFR-3 protein and MicroRNA in three- dimensional cultured HDLEC were measured by immunohistochemistry, western-blot and qPCR. Results The lymphangio- genesis model in vitro was established successfully by three-dimensional culture of HDLEC. The inhibitory effects of NCTD on lymphangiogenesis in vitro were observed. Compared to control group, the expression of VEGF-C and VEGF-D proteins in NCTD group was significantly down-regulated as showed by immunohistochemistry and Western-blot (P＜0.01), The MicroRNA
expression of VEGF-C, VEGF-D and VEGFR-3 in NCTD group was significantly decreased as shown by qPCR (P＜0.01). Conclusion Norcantharidin inhibits lymphangiogenesis by down-regulating the expression of VEGF-C and VEGF-D, suggesting that it might be an effective agent for targeting anti-lymphangiogenesis. |
| Key words: Norcantharidin Lymphatic endothelial cell Three-dimensional Anti-lymphangiogenesis |
