SZ-21拮抗血管生成的初步研究

本研究的目的是从SZ-1，-2，-21，-51，-65及262(GPⅡb/Ⅲa McAb)中筛选出与ECV304（人脐静脉内皮细胞株）起反应的单抗，研究其抗血管生成的能力。用流式细胞仪筛选与ECV304起反应的单抗，用酶标法检测反应的单抗对ECV304、肺癌细胞株A549粘附、迁移的作用，采用鸡绒毛尿囊膜（CAM）血管生成分析法观察其体外抗血管生成作用，以Annexin Ⅴ TITC盒检测单抗对A549细胞的凋亡作用。结果表明：筛选出了与ECV304起反应的SZ-21，其对ECV304与基质纤连蛋白（Fn）、胶原蛋白（Col）Ⅳ间粘附、迁移均有抑制作用，鸡绒毛尿囊膜经TNF-α诱导血管生成后SZ-21可抑制其新生。进一步研究发现SZ-21对肿瘤细胞A549（肺癌细胞株）与基质Fn，ColⅣ间粘附、迁移亦有拮抗作用，对A549细胞可诱导其凋亡。结论：SZ-21通过对整合素（integrin）β3的抑制和诱导凋亡的作用表现了抗血管生成的能力，整合素与Fn，ColⅣ的结合部位也不限于RGD序列，而具有不同的识别信号。

Inhibition of Angiogenesis Properties by SZ-21

The aim of this study is to screen out the monoclonal antibody reactive to a kind of endothelial cell line (ECV304) from SZ-1, -2, -21, -22, -51, -65, -262 and explore its function of anti-angiogenesis. The inhibitory effects of monoclonal antibody reactive to ECV304 and human lung carcinom cells (A549) adhesion and migration to extracellular matrix proteins (i.e, fibronectin and collagen IV) were studied by ELISA, the inhibition of angiogenesis in vivo was analyzed by chick chorioallantoic membrane (CAM) assays, the percentage of apoptotic cells in A549 cells was assayed by flow cytometric Annexin-V-FITC/PI dual labeling technique. The results showed that SZ-21 exhibited inhibitory effects on human umbilical vein endothelial cell line (ECV304) and pulmonary cancer cell line (A549) adhesion and migration to extracellular matrix proteins (i.e, fibronectin and collagen IV). In addition, it disrupted ongoing angiogenesis on the chick chorioallantoic membrane (CAM) model. SZ-21 also induced apoptosis of the A549 cells. In conclusion, SZ-21 inhibits angiogenesis in vivo and in vitro by blocking integrin beta(3) and inducing apoptosis. SZ-21 recognized the sequence beta(3) 28 - 35 which is far away from the RGD ligation site on GPIIIa. Integrin may interact the extracellular matrix via recognition sites other than RGD sequence.