Massia S P, Hubbell J A
Department of Chemical Engineering, University of Texas, Austin 78712-1062.
J Biomed Mater Res. 1991 Feb;25(2):223-42. doi: 10.1002/jbm.820250209.
The attachment, spreading, spreading rate, focal contact formation, and cytoskeletal organization of human umbilical vein endothelial cells (HUVECs) were investigated on substrates that had been covalently grafted with the cell adhesion peptides Arg-Gly-Asp (RGD) and Tyr-Ile-Gly-Ser-Arg (YIGSR). This approach was used to provide substrates that were adhesive to cells even in the absence of serum proteins and with no prior pretreatment of the surface with proteins of the cell adhesion molecule (CAM) family. This approach was used to dramatically enhance the cell-adhesiveness of substrates that were otherwise cell-nonadhesive and to improve control of cellular interactions with cell-adhesive materials by providing stably bound adhesion ligands. Glycophase glass was examined as a model cell-nonadhesive substrate prior to modification, and polyethylene terephthalate (PET) and polytetrafluoroethylene (PTFE) were examined as representative materials for biomedical applications. The peptides were surface-coupled by their N-terminal amine to surface hydroxyl moieties using tresyl chloride chemistry. Prior to peptide grafting, the PET and PTFE were surface hydroxylated to yield PET-OH and PTFE-OH. The PET-OH was less cell-adhesive and the PTFE-OH was much more cell-adhesive than the native polymers. Radioiodination of a C-terminal tyrosine residue was used to quantify the amount of peptide coupled to the surface, and these amounts were 12.1 pmol/cm2 on glycophase glass, 139 fmol/cm2 on PET-OH, and 31 fmol/cm2 on PTFE-OH. Although the glycophase glass did not support adhesion or spreading even in the presence of serum, the RGD- and YIGSR-grafted glycophase glass did support adhesion and spreading, even when the only serum protein that was included was albumin. Although PET and PTFE-OH supported adhesion when incubated in serum-supplemented medium, neither of these materials supported adhesion with only albumin present, indicating that cell adhesion is mediated by adsorbed CAM proteins. When these materials were peptide-grafted, however, extensive adhesion and spreading did occur even when only albumin was present. Since the peptide grafting is quite easily controlled and is temporally stable, while protein adsorption is quite difficult to precisely control and is temporally dynamic, peptide grafting may be advantageous over other approaches employed to improve long-term cell adhesion to biomaterials.
在通过共价接枝细胞黏附肽精氨酸 - 甘氨酸 - 天冬氨酸（RGD）和酪氨酸 - 异亮氨酸 - 甘氨酸 - 丝氨酸 - 精氨酸（YIGSR）的底物上，研究了人脐静脉内皮细胞（HUVECs）的附着、铺展、铺展速率、焦点接触形成和细胞骨架组织。这种方法用于提供即使在没有血清蛋白且无需事先用细胞黏附分子（CAM）家族的蛋白质对表面进行预处理的情况下仍能黏附细胞的底物。这种方法用于显著增强原本不具有细胞黏附性的底物的细胞黏附性，并通过提供稳定结合的黏附配体来改善细胞与细胞黏附材料相互作用的控制。在改性之前，将糖相玻璃作为一种典型的非细胞黏附性底物进行研究，并将聚对苯二甲酸乙二酯（PET）和聚四氟乙烯（PTFE）作为生物医学应用的代表性材料进行研究。使用三氟甲磺酸氯化学方法，通过肽的N端胺将肽表面偶联到表面羟基部分。在肽接枝之前，将PET和PTFE进行表面羟基化处理，得到PET - OH和PTFE - OH。PET - OH的细胞黏附性比天然聚合物低，而PTFE - OH的细胞黏附性比天然聚合物高得多。通过对C端酪氨酸残基进行放射性碘化来定量偶联到表面的肽的量，这些量在糖相玻璃上为12.1 pmol/cm²，在PET - OH上为139 fmol/cm²，在PTFE - OH上为31 fmol/cm²。尽管即使在有血清的情况下糖相玻璃也不支持细胞黏附或铺展，但接枝了RGD和YIGSR的糖相玻璃即使在仅含白蛋白作为唯一血清蛋白时也确实支持细胞黏附与铺展。虽然PET和PTFE - OH在补充血清的培养基中孵育时支持细胞黏附，但当仅存在白蛋白时，这两种材料都不支持细胞黏附，这表明细胞黏附是由吸附的CAM蛋白介导的。然而，当这些材料进行肽接枝后，即使仅存在白蛋白时也会发生广泛的细胞黏附和铺展。由于肽接枝相当容易控制且在时间上是稳定的，而蛋白质吸附很难精确控制且在时间上是动态的，因此与用于改善细胞对生物材料长期黏附的其他方法相比，肽接枝可能具有优势。
