烟曲霉素抑制小鼠结直肠癌生长及转移的体内外研究

Fumagillin inhibits colorectal cancer growth and metastasis in mice:an in vivo and in vitro study of antiangiogenesis

背景与目的:血管新生与肿瘤生长密切相关,烟曲霉素(Fumagillin)可特异性抑制血管内皮细胞的增殖,但其对结直肠癌的治疗作用尚不十分清楚。本研究旨在探讨Fumagillin对结直肠癌细胞系生长的抑制作用及其作用机制。方法:将WiDr或HT-29细胞以5×105/L的量分别接种于20只SCID小鼠背部皮下,接种4周后每2日腹腔注射Fumagillin(0.1mg/kg)或Cyclo(1mg/kg)(Arg-Gly-Asp-D-Phe-Val)持续4周,然后处死动物并测量原发瘤质量及原发瘤内微血管密度。在体外培养脐带静脉内皮细胞(HUVECs),向培养基内加入Fumagillin(0.01mg/kg)或Cyclo(0.1mg/kg),观察其对HUVECs增殖及微管形成的抑制作用。以基因芯片技术筛选Fumagillin处理后HUVECs的基因表达变化,并以定量PCR和免疫印迹技术对基因芯片的结果做进一步验证。结果:Fumagillin处理组小鼠的结直肠癌原发瘤质量及肿瘤内CD105阳性的微血管数量显著小于对照组小鼠(P<0.05),体外实验显示,Fumagillin处理后HUVECs的增殖和微管形成受到明显抑制,基因芯片检测显示Fumagillin处理后HUVECs有71个基因表达上调和143个基因表达下调,表达变化的基因多与细胞黏附、移动、增殖和基因转录有关。定量PCR和免疫印迹技术发现Fumagillin抑制HUVECs表达cyclinE2,白细胞活化黏附因子(activated leukocyte cell adhesion molecule,ALCAM)和细胞间黏附分子-1(intercellular adhesion molecule-1,ICAM-1)。结论:Fumagillin通过抑制新生血管的形成来抑制结直肠癌的生长。

Background and purpose：Fumagillin is an inhibitor of type 2 methionine aminopeptidase that can block blood vessel formation. However, its molecular mechanism and therapeutic value in colon cancer still remain to be elucidated.In this study, the effect of Fumagillin on the growth of colon cancer was examined. Methods：Twenty mice were divided into 4 groups and injected subcutaneously with 5×105/L WiDr or HT-29 cells in 200 μL phosphate-buffered saline （PBS） respectively. After 4 weeks, intraperitoneal injections of Fumagillin （0.1 mg/kg）, Cyclo （1 mg/kg）, or both were given every 2 days for 4 weeks. The tumor weight and microvessel density （MVD） were examined. Geneexpression profiles were examined by microarray analysis of human umbilical endothelial cells （HUVECs）. Results： The Fumagillin-treated mice showed smaller tumor mass and lower MVD-CD105 levels than control ones. In vitro proliferation and tube formation of HUVEC was also significantly decreased by Fumagillin. Microarray analysis of Fumagillin-treated HUVECs showed up-regulation of 71 genes and down-regulation of 143 genes. Expression changes were involved in cell proliferation, migration, adhesion and gene transcription. Quantitative real time-polymerase chain reaction and Western blot revealed decreased expression of cyclin E2, activated leukocyte cell adhesion molecule （ALCAM）, and intercellular adhesion molecule-1 （ICAM-1） genes in the presence of Fumagillin. Conclusion： Fumagillin was found to suppress colorectal cancer growth by suppressing angiogenesis. The down-regulation of cyclin E2, ALCAM and ICAM-1 by fumagillin may be involved in the anti-angiogenesis.