Vascular growth factor binding kinetics to the endothelial cell basement membrane, with a kinetics-based correction for substrate binding
Vascular growth factor binding kinetics to the endothelial cell basement membrane, with a kinetics-based correction for substrate binding
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DOI：
10.1007/s10616-009-9212-1
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发表时间：
2009-07-01
期刊：
影响因子：
2.2
通讯作者：
Edelman, Elazer R.
中科院分区：
文献类型：
--
作者：
Clyne, Alisa Morss;Edelman, Elazer R.
血管生长因子，包括血管内皮生长因子和成纤维细胞生长因子-2，与基底膜中的硫酸乙酰肝素蛋白聚糖结合。虽然这种结合、储存和释放系统为受控药物释放装置提供了关键模型，但基底膜生长因子结合动力学尚未完全建立。我们修改了基底膜的内皮细胞生长因子结合动力学方案。基底膜对成纤维细胞生长因子-2 (K (d) = 185.8 nM) 具有较低的亲和力，且解离速率较慢 (k (off) = 0.00338 min(-1))。然而，生长因子以与基底膜惊人相似的方式与组织培养聚苯乙烯结合，结果令人困惑。由于底物结合无法被阻断，因此开发了基于结合动力学的校正技术来解释聚苯乙烯生长因子的结合。通过在几乎没有生长因子结合的细菌平板上进行结合动力学实验来验证该方法。这种新方法将提高我们对健康和疾病中细胞和蛋白质与基底膜相互作用的理解。它们还可以进一步应用于开发仿生药物输送系统。
Vascular growth factors, including vascular endothelial growth factor and fibroblast growth factor-2, bind to heparan sulfate proteoglycans in the basement membrane. While this binding, storage, and release system provides a critical model for controlled drug release devices, basement membrane-growth factor binding kinetics have not been fully established. We modified endothelial cell-growth factor binding kinetics protocols for the basement membrane. The basement membrane showed low affinity for fibroblast growth factor-2 (K (d) = 185.8 nM), with a slow off rate (k (off) = 0.00338 min(-1)). However, results were confounded by growth factor binding to tissue culture polystyrene in a manner strikingly similar to basement membrane. Since substrate binding could not be blocked, a binding kinetics based correction technique was developed to account for polystyrene growth factor binding. This method was validated by conducting binding kinetics experiments on bacteriologic plates that exhibit little growth factor binding. This novel method will improve our understanding of cell and protein interaction with the basement membrane in health and disease. They can also further be applied to develop biomimetic drug delivery systems.