姜黄色素活性单体成分抗血管生成的实验研究

Role of active components of curcumin in anti-angiogenesis in vitro

目的 探讨中药姜黄的活性单体成分姜黄素(姜黄素Ⅰ)、脱甲氧基姜黄素(姜黄素Ⅱ)和双脱甲氧基姜黄素(姜黄素Ⅲ)对血管生成的抑制作用。方法 ①以体外培养的人脐静脉内皮细胞和人肺微血管内皮细胞为对象,研究3种单体对内皮细胞增殖的抑制作用和诱导内皮细胞凋亡的效果;②建立裸鼠人肺腺癌细胞A5 49移植瘤模型,腹腔注射双脱甲氧基姜黄素,研究其对荷癌裸鼠肿瘤血管生成的影响。结果 ①MTT检测3种姜黄色素作用72h后对内皮细胞增殖抑制作用的IC50值分别为0 5 2 5、0 3 99、0 12 5 μg ml,在浓度为4μg ml时,3种姜黄色素作用48h后均导致内皮细胞出现明显的增殖抑制(P<0 0 5 ) ,作用72h时达最高值;②流式细胞仪分析浓度为4μg ml时姜黄素Ⅲ可将内皮细胞阻止于S期(P <0 0 5 ) ,浓度增至8μg ml以上时不但可引起S期细胞比例增加(P <0 0 1) ,同时又有明确的诱导凋亡作用;③免疫组化显示姜黄素Ⅲ实验组裸鼠瘤组织内新生血管的数量明显减少,与阴性对照组相比相差非常显著(P <0 0 1) ;姜黄素Ⅲ组肿瘤组织中VEGF表达水平,VEGF受体Flt 1、KDR水平与阴性对照组相比均相差非常显著(P <0 0 1)。结论 3种姜黄色素单体均具有确切的抗人内皮细胞和肺腺癌裸鼠移植瘤血管生成作用,其中姜黄素Ⅲ最强,机制可能与其抑制VEGF及其受体的表达有关。

Objective To explore the anti-angiogenesis effects of active components (curcumin-1, curcumin-2 and curcumin-3) of Curcuma Longa. Methods ①Human umbilical vein endothelial cell lines (HUVEC) and human pulmonary microvascular endothelial cell lines(HPMEC) were employed to analyze and compare the inhibitory effects on cell proliferation and apoptosis of the active components. ②Nude mice bearing with human lung adenocarcinoma cell line A549 were injected intraperitoneally with purified curcumin-3 to observe the inhibitory effects on angiogenesis. Results ①The IC 50 values of inhibitory effect of curcumin-1, curcumin-2 and curcumin-3 on HUVEC growth were 0.525 μg/ml, 0.399 μg/ml and 0.125 μg/ml, respectively. At the concentration of 4 μg/ml, curcumin-1, curcumin-2 and curcumin-3 were able to inhibit effectively the proliferation of endothelial cell at 48 h post-treatment (P<0.05), and reached the peak at 72 h post-treatment. ②At the concentration of 4 μg/ml, curcumin-3 could effectively retard the endothelial cell at S phase at 24 h post-treatment (P<0.05). But when the concentration was 8 μg/ml or more, curcumin-3 could also induce obvious apoptosis of endothelial cell. ③MVD (micro vein density) in curcumin-3 group was lower than that in control group, respectively (P<0 01). Curcumin-3 markedly suppressed the expression of vascular endothelium growth factor (VEGF) and its receptors, Flt-1 and KDR in xenografted tumor tissues, as compared with the control group (P<0.01). Conclusion Curcumin-1, curcumin-2 and curcumin-3 can significantly inhibit angiogenesis in cultured human endothelial cells and tumor tissues in nude mice bearing with human lung adenocarcinoma cell line A549. The bioactivity of curcumin-3 is significantly higher than that of curcumin-1 and curcumin-2, respectively. The possible mechanisms involved in the suppression of angiogenesis may be related to down-regulation of VEGF, Flt-1and KDR in tumor cells and endothelial cells.