In vitro effect of biologically active coating on endothelialisation of Nitinol coils designed for the closure of intracardiac shunt

摘要

Objective: To evaluate in vitro the effect of biologically active coating on endothelialisation of Nitinol coils designed for the closure of intracardiac shunt. Methods: Covalently binding procedure was performed with chemical vapor deposition (CVD).Heparin(200 IU/ml ), r-hirudin(21 nmol/ml ), or fibronectin( 15 μg/ml), respectively, were bound to the basic coating( poly( amino- p- xylylene-co-p-xylylen) ). In vitro tests were performed on five different coating groups( n = 2 each) : uncoated Nitinol, basic coating, heparin-,r-hirudin-,and fibronectin-coating. Human umbilical vein endothelial cells(HUVECs) were cultured in the presence of tested coils for 48 and 72 h. Adhesion of HUVECs were evaluated with light microscopy,and with confocal laser scanning microscopy after double-staining of vinculin,phalloidin, Ki67 and fibronectin. Results: After 48 and 72 h of incubation,except HUVECs around the basic coating showing abnormal morphology, HUVECs were able to grow next to all kinds of coils. By CLSM adhering cells were typically confined to maximal three secondary coil windings. HUVECs adhered best to the fibronectin coated coils,followed by the uncoated Nitinol. HUVECs barely adhered to the basic coating. The adhesion to heparin coating or r-hirudin coating was observed with various cell densities. Heparin coating seemed to support cell adhesion better than r-hirudin coating did. The HUVECs assembled fibronectin matrix on the fibronectin coating and uncoated Nitinol coil. Vinculin was expressed on both fibronectin coating and uncoated Nitinol as well, mostly diffused,but occasionally also in focal contacts. Ki67 expression was mainly noticed on the uncoated Nitinol coil and fibronectin coated coil, only occasionally on other materials. The densities and morphology of HUVECs, adhered to the correspohding well surfaces, were similar for all wells. The only exception was the well with the basic coating coil after three days of incubation. In this case, the cell density was diminished, and the cells became more elongated. The cells grown on the well surfaces exhibited a more profound matrix assembly and more pronounced focal adhesions than the cells adhered to the coil surfaces did. Conclusion: Except for basic coating,biologically active substance coated or uncoated coils show no acute cytotoxicity on HUVECs. Due to better cell proliferation,cell matrix assembling,and focal adhesion,only fibronectin coating improves the endothelialisation of Nitinol coils.