Angiogenesis promotion by fixing biotinylated VFGF on endothelial cells

我们开发了一种血管内药物输送系统，在。通过导管注射的药物被固定在目标组织的脉管系统中。活的内皮细胞的细胞蛋白首先被生物素化试剂直接生物素化，然后被生物素化的药物结合，或者使用生物素作为连接物，生物素化的药物。我们假设，如果将促进内皮细胞增殖和血管生成的血管内皮生长因子（vascular endothelial growth factor， VEGF）生物素化，并通过上述方法给予内皮细胞，其作用将比细胞仅仅暴露在内皮细胞中几分钟更大。在第一个实验中，我们研究了用上述方法给予内皮细胞的分子在细胞内的位置。用β-半乳糖苷酶标记的链亲和蛋白固定在生物基化的内皮细胞上。分别于0、1、2、3、6、12小时后收获细胞。用组织化学方法（x-gal）对酶进行可视化观察。电镜观察表明，该酶位于细胞质内0小时到达核膜。很少在细胞膜上发现。在第二个实验中，制作生物素化的VEGF，并将其固定在生物素化的内皮细胞上。虽然生物素化后的VEGF的活性与完整的VEGF相同，但将生长因子固定在细胞上对培养的内皮细胞的增殖没有那么有效。我们推测，通过细胞膜受体影响的药物不适合我们的药物递送方法，因为生物素化的药物进入细胞的速度太快，而没有与它们的受体结合。

We have developed an intravascular drug delivery system, in. which a drug injected from a catheter is fixed to the vasculature of the targeted tissue. Cellular protein of viable endothelial cells is first biotinylated directly by biotinylation reagents, and then bound by an avidinated drug or, using avidin as a linker, a biotinylated drug. We hypothesized that if vascular endothelial growth factor (VEGF), which promotes endothelial proliferation and angiogenesis, is biotinylated and given to the endothelium by the above method, its effect will become greater than when the cells are simply exposed to it for very few minutes. In the first experiment, we investigated intracellular locations of the molecule given to the endothelium by the above method. Streptavidht labeled with β-galactosidase was fixed to biotmylated endothelium, and.cells were harvested 0,1,2,3,6,12 hours after. The enzyme was visualized byhistochemical method (x-gal). Electron microscopic study revealed that the enzyme was located at 0 hour in the cytoplasm and reached nuclear membrane. Few were found on the cellular membrane. In the second experiment, biotinylated VEGF was made, and fixed to the biotinylated endothelial cells. Though the activity of the biotinylated VEGF was the same as that of intact VEGF, the fixation of the growth factor on the cells was not so effective for the proliferarion of cultured endothelial cells. We speculate that drugs that influence via the cell membrane receptors are not good candidates for our drug delivery method because the biotinylated drugs enter too fast into the cells without binding to their receptors.