Surface Modification Of Polyethylene Terephthalate For Improvement Of Anticoagulation And Endothelialization

In recent years, synthetic polymers are widely used in biomedical materials for artificial organs including cardiovascular artificial organs. However, once blood contact with polymers, complicated interaction will happen on the interface so as to form coagulating reaction and thrombus. In that case, the farther application of synthetic polymers in blood-contacting devices was restricted. It is believed that the endothelialization of the blood contacting device surface may overcome the problem. Therefore, a surface that enhances cell-material interaction and promotes the endothelialization at a shorter time is necessary.In our research, natural proteins like gelatin and collagen molecules were immobilized onto the biomedical poly(ethylene terephthalate) (PET, dacron) surfaces. However, the immobilization of gelatin or collagen on the surface of materials may accelerate the aggregation of the platelets before the formation of endothelial layer. Hence, surface anticoagulative modification before endothelialization is necessary. Our study focuses on the co-immobilization of heparin (or albumin) and collagen (or gelatin) molecules on the PET surface which can construct the anticoagulant surface before the endothelium formation and also accelerate the endothelialization of polymer surface.The surface constitution of the modified PET is determined by X-ray photoelectron spectroscopy (XPS). According to the resurts of XPS, The detection of S2p, S2s might be in virtue of the sulfonic groups of the heparin chemically attached to the membrane surfaces, which demonstrates that of the molecules of heparin and collagen or gelatin co-immobilized on the PET surfaces. Comparing to the only collagen and gelatin immobilized surface, the nitrogen contents increase 4.74% and 5.64% on the surface of albumin-collagen (PET-CL-BSA) and albumin-gelatin (PET-Gel-BSA) co-immobilized PET film surface. This proves that albumin combined with collagen or gelatin molecules are effectively co-immobilized onto the PET film surfaces. The results of static contact angle indicate the hydrophilicity of the modified PET surface is more remarkably improved than that of the unmodified PET surface.The determination of carboxyl group on the acrylic acid grafted PET surface is 5.29×10^-9mol/cm². According to the protein measurement, the concentration of gelatin immobilized on only gelatin-PET (PET-Gel) is 2.02μg/cm², while the concertration of collagen immobilized on only collagen-PET (PET-CL) is 3.14μg/cm². The quantification of heparin on heparin-collagen co-immobilized surface (PET-CL-Hep) is 1.73μg/cm², and heparin on heparin-gelatin co-immobilized surface(PET-CL-Hep) is 1.91μg/cm².The results of lactate dehydrogenase(LDH) and the platelet adhesion test in vitro all indicat that the co-immobilization of heparin or albumin with collagen or gelatin molecules could improve the anticoagulation. The percentage of platelet adhered on the surface of PET-CL-Hep is 12.83%, PET-Gel-Hep 17.24%, PET-CL-BSA 10.53%, and PET-Gel-BSA 12.19%, thus the percentages of adhered platelet decreased significantly compared to those of untreated PET (26.21%) and the PET-Gel surface (40.38%) , the PET-CL surface (47.21%). The results of coagulant factor tests reveal that the activated partially thromboplastion time (APTT) of PET-CL-Hep is 68.9s and PET-Gel-Hep is 65.5s, which is longer than those of the virgin PET (59.2s) and the plasma (54s). The change of APTT indicates that the modified PET films can suppress intrinsic coagulant system. The number of denatured fibrinogen on different modified surfaces is also determined by enzyme linked immunosorbent assay (ELISA). The resulte shows that the proption of denatured fibrinogen on PET-CL-Hep is 25.32%, on PET-Gel-Hep is 22.48%, on PET-CL-BSA is 19.20%, and on PET-Gel-Hep is 20.59%, which is decreased comparing the unmodified PET (35.57%). All show that after the co-immobilization of heparin (albumin) and collagen (gelatin), the anticoagulation of the film surfaces has improved.The results of endothelial cells culture in vitro shows significant difference in the behavior of cell adhesion and proliferation among different materials. PET-CL-Hep and PET-Gel-Hep films support cell attachment and proliferation more than unmodified film surfaces. The ECs on the PET-CL-BSA and PET-Gel-BSA can spread and proliferate. Quantitative analysis of cell adhesion is performed for these different modified and untreated PET surfaces. For the 120 hour cultured samples, the number of cells on PET-CL-Hep, PET-Gel-Hep, PET-CL-BSA, PET-Gel-BSA are 9.37, 9.73, 10.66,9.07(×10⁴cells/cm²), and the activity of cell is 202.3%, 205.9%, 214.8%, 199.4%. This is in accordance with the optical microscopy images, which suggests that cell adhesion onto PET is favoured by surface co-immobilization of heparin (albumin) with collagen (gelatin).