Affiliated Hospital of Nanjing University of TCM, Nanjing, China.
Acta Haematol. 2014;131(2):102-11. doi: 10.1159/000354770. Epub 2013 Oct 18.
Celastrol is an active compound extracted from the root bark of the traditional Chinese medicine Tripterygium wilfordii Hook F. In this study, we investigated the effect of celastrol on lipopolysaccharide (LPS)-activated LP-1 human multiple myeloma cell-induced angiogenesis, and identified its molecular mechanism of action. Migration of human umbilical vein endothelial cells (HUVECs) was tested using a wound-healing assay. HUVEC invasion was assayed using a Transwell chamber. Cell surface expression of Toll-like receptor 4 (TLR4) was analyzed by flow cytometry. Angiogenic factor vascular endothelial growth factor (VEGF) level was quantified by LUMINEX and protein expression was analyzed by Western blot. Translocation of nuclear factor-kappa B (NF-κB) was observed by fluorescence microscopy. Celastrol inhibited LPS-stimulated LP-1 human multiple myeloma-induced HUVEC migration and invasion in a concentration-dependent manner. Wound diameters increased by 72.9, 165.4 and 246.2% at 0.025, 0.05 and 0.1 μM, respectively, compared to LPS alone. A 45-74% inhibition of LPS-dependent cell invasion was achieved in the presence of 0.025-0.1 μM celastrol. Celastrol significantly downregulated LPS-induced TLR4 expression and inhibited LPS-induced VEGF secretion in LP-1 cells. VEGF levels decreased by 64.8, 84.4 and 92.9% after coexposure to celastrol at 0.025, 0.05 and 0.1 μM, respectively, compared to LPS alone. Celastrol also inhibited the IκB kinase (IKK)/NF-κB pathway induced by LPS. Protein levels of NF-κB p65, IKKα and IκB-α decreased in a dose-dependent manner after coexposure to celastrol. Celastrol also blocked nuclear translocation of the p65 subunit. These results suggest that celastrol inhibits LPS-induced angiogenesis by suppressing TLR4-triggered NF-κB activation.
三萜烯二醇是从雷公藤的根皮中提取的一种活性化合物。本研究探讨了三萜烯二醇对脂多糖（LPS）激活的 LP-1 人多发性骨髓瘤细胞诱导的血管生成的影响，并确定了其作用的分子机制。通过划痕愈合试验检测人脐静脉内皮细胞（HUVEC）的迁移。通过 Transwell 室测定 HUVEC 侵袭。通过流式细胞术分析 Toll 样受体 4（TLR4）的细胞表面表达。通过 LUMINEX 定量血管内皮生长因子（VEGF）水平，并通过 Western blot 分析蛋白表达。通过荧光显微镜观察核因子-κB（NF-κB）的易位。三萜烯二醇呈浓度依赖性抑制 LPS 刺激的 LP-1 人多发性骨髓瘤诱导的 HUVEC 迁移和侵袭。与 LPS 单独作用相比，在 0.025、0.05 和 0.1μM 时，划痕直径分别增加了 72.9%、165.4%和 246.2%。在 0.025-0.1μM 三萜烯二醇存在的情况下，LPS 依赖性细胞侵袭的抑制率为 45-74%。三萜烯二醇显著下调 LPS 诱导的 LP-1 细胞中 TLR4 的表达，并抑制 LPS 诱导的 VEGF 分泌。与 LPS 单独作用相比，在 0.025、0.05 和 0.1μM 时，共暴露于三萜烯二醇后 VEGF 水平分别降低了 64.8%、84.4%和 92.9%。三萜烯二醇还抑制 LPS 诱导的 IκB 激酶（IKK）/NF-κB 途径。在共暴露于三萜烯二醇后，NF-κB p65、IKKα 和 IκB-α 的蛋白水平呈剂量依赖性降低。三萜烯二醇还阻断了 p65 亚基的核易位。这些结果表明，三萜烯二醇通过抑制 TLR4 触发的 NF-κB 激活来抑制 LPS 诱导的血管生成。
