甲磺酸乐伐替尼对猪胸导管内皮细胞增殖和侵袭的影响

Effect of lenvatinib mesylate on proliferation and invasion of endothelial cells in pig thoracic ducts

目的 研究甲磺酸乐伐替尼(Levatinib mesylate)对取自猪胸导管的淋巴管内皮细胞(LECs)增殖和侵袭的影响,探讨甲磺酸乐伐替尼对淋巴管生成作用的相关调控机制。方法 取健康猪的胸导管消化法进行LECs原代培养,用VEGFR-3(血管内皮生长因子-3)进行鉴定;采用EdU检测法检测甲磺酸乐伐替尼药物作用后LECs的增殖活力;采用Transwell侵袭实验检测甲磺酸乐伐替尼是否对LECs的侵袭能力有影响;Western blot(蛋白质印迹)法分别测定甲磺酸乐伐替尼药物作用后LECs的VEGFR-3靶蛋白的含量。结果 EdU结果显示,甲磺酸乐伐替尼药物作用后LECs增殖能力降低,当药物浓度达到10 nmol/L时,实验组EdU标记率(25±6)%明显低于对照组(49±9)%,LECs的增殖能力被甲磺酸乐伐替尼显著抑制(P<0.05);Transwell侵袭实验结果可见,LECs的侵袭能力能够被甲磺酸乐伐替尼抑制,当药物浓度达到10 nmol/L时,实验组LECs侵袭数量(66.33±8.93)低于对照组(138.67±7.02),LECs透膜数量明显减少(P<0.05);Western blot结果可见,甲磺酸乐伐替尼可以下调LECs的VEGFR-3蛋白表达,且随着药物浓度的增加,蛋白含量下调更为显著(P<0.05)。结论 甲磺酸乐伐替尼可通过作用于VEGFR-3靶蛋白,从而抑制LECs增殖和侵袭。

Objective To study the effect of levatinib mesylate on the proliferation and invasion of lymphat-ic endothelial cells(LECs)taken from pig thoracic ducts,and explore the relevant regulatory mechanisms of leva-tinib mesylate on lymphangiogenesis.Methods The thoracic duct digestion method of healthy pigs was used for pri-mary culture of LECs,and VEGFR-3(vascular endothelial growth factor-3)was used for identification;Using EdU detection method to detect the proliferation activity of LECs after the action of levatinib mesylate;Transwell invasion assay was used to detect whether the invasive ability of LECs after the action of levatinib mesylate;The Western blot method was used to determine the content of VEGFR-3 target protein in LECs after the action of levatinib mesylate.Results The EdU results showed that the proliferation ability of LECs was reduced after the action of levatinib me-sylate.When the drug concentration reached 10 nmol/L,the EdU labeling rate in the experimental group(25%±6%)was significantly lower than that in the control group(49%±9%),and the proliferation ability of LECs was significantly inhibited(P<0.05);The results of the Transwell invasion experiment showed that the invasive ability of LECs cells could be inhibited by lovatinib mesylate.When the drug concentration reached 10 nmol/L,the number of LECs invading the experimental group(66.33±8.93)was lower than that of the control group(138.67±7.02),the number of LECs membranes significantly decreased(P<0.05);Western blot results showed that levatinib mesylate can downregulate the expression of VEGFR-3 protein in LECs,and the protein content was more significantly down-regulated with the increase of drug concentration(P<0.05).Conclusion Levatinib mesylate can inhibit the prolifer-ation and invasion of LECs by acting on the VEGFR-3 target protein.