人促血管生成素2单链抗体对裸鼠人肝癌血管生成及肿瘤生长的作用

Effects of angiopoietin 2 single-chain antibody on human hepatocellular carcinoma in nude mice

目的观察促血管生成素2（An萨）单链抗体（scFv-An舀）对人肝癌血管生成及肿瘤生长的作用。方法体外培养人脐静脉内皮细胞（HUVEC），各组分别添加血管内皮生长因子（VEGF）、VEGF＋Ang2、VEGF＋Ang2＋ScFv-Ang2，通过HUVEC增殖测定、迁移实验、小管形成实验观察ScFv-Ang2在体外对HUVEC生物学行为的影响；建立荷人肝癌细胞株MHCC97的肝原位移植瘤模型裸小鼠24只，随机分为实验组和对照组，实验组瘤内注射ScFv-Ang2（4×10^11 pfu／m1），对照组予以生理盐水，测量肿瘤大小、肺转移灶数目、癌组织CD31蛋白表达及微血管密度（MVD）计数，观察ScFv-Ang2对肝细胞癌（HCC）血管生成及肿瘤生长的作用。结果体外实验中观察到ScFv-Antg2可抑制VEGF和ArtS2诱导的HUVEC增殖、迁移及小管形成；ScFv-Ang2瘤内注射组肿瘤体积、重量、肺转移灶数目、CD31表达水平以及MVD计数均显著低于对照组（P〈0．05）。结论ScFv-Ang2在体外实验中具有抑制HUVEC形成血管的作用，体内实验中对裸鼠人肝癌具有抑制血管生成和肿瘤生长的作用。

Objective To observe the effects of single-chain antibody against human angiopoietin 2 （Ang2） on the angiogenesis and growth of hepatocellular carcinoma in nude mice. Methods Human umbilical vein endothelial cells （HUVECs） were cultured and grouped in vitro. Vascular endothelial growth factor （VEGF）, VEGF and Ang2, or VEGF ＋ Ang2 ＋ single-chain antibody （ScFv）-Ang2 was added into each group. Methyl thiazol tetrazolium （MTr） assay, Transwell assay and Matrigel assay were used to de- tect the effects of ScFv-Ang2 on angiogenie ability in vitro. A total of 24 nude mice bearing human hepato- cellular carcinoma of MHCC97 cell line with a high metastatic potential were established and randomly di- vided into 2 groups. Experimental group accepted intratumoral injection of ScFv-Ang2, and saline injection group was set as placebo-control. Mice were sacrificed on the 21 st day, tumor size and weight and metasta- ses in lungs were measured, the expression of CD31 in tumor tissues was detected by using immunohisto- chemistry, and microvessel density （MVD） was counted. Results In vitro experiment concerning HUVECs, ScFv-Ang2 inhibited VEGF- and Ang2-induced proliferation, migration and tube formation of HUVECs. Statistical indexes including tumor weight and volume, metastases to lungs, CD31 expression and MVD counting in ScFv-Ang2 injection group were all significantly lower than in placebo-control group （P 〈 0. 05）. Conclusion In addition to the inhibition of blood vessel formation of HUVECs in vitro, ScFv-Ang2 exerts significant inhibitory effect on tumor growth, anglogenesis and metastasis of human hepa- tocellular carcinoma in nude mice.