ALD Mediated Heparin Grafting On Nitinol For Self- Expanded Carotid Stents

Carotid-artery atherosclerosis is a common cause of ischemic stroke. Carotid-artery stenting (CAS) is one of the most effective treatments. However, In-stent restenosis (ISR) and re-endothelialization delay are two major issues of intravascular stent which affect clinical safety and reduce effects.Heparin, one of the most important natural anticoagulant material, has been used clinically to minimize thrombus formation on artificial surfaces. In the previous reports, the non-covalent heparinization of the metal surface has been proved as a potentially helpful strategy to improve the hemocompatibility and prevent thrombus formation of blood-contacting materials. But the rapid elution of heparin was still a challenge. In recent years, more and more approaches of surface heparinization have been reported, among them, layer-by-layer self-assemble and covalent immobilization are the two most widely used procedures. On a typical metallic surface, the immobilization of heparin is challenging due to the lack of reactive functional groups on a metallic surface and some preprocessing steps are needed.In this study, atomic layer deposition (ALD) technology was applied to deposit a layer (10nm) of Al2O3 on Nitinol surface as an intermediate functional layer. By using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide (EDC/NHS) chemistry, heparin was subsequently immobilized on the modified surfaces. The physical, chemical and biological properties of pristine and modified surfaces were also characterized and reported.The chemical, physical and biological properties of pristine and modified Nitinol surface were accessed by X-ray photoelectron spectroscopy and atomic force microscope. It was found that the pristine Nitinol surface is fairly homogeneous and smooth with a root-mean-square surface roughness (RMS) of about 18.1 nm in the analyzed 100μm2 region. After the surface grafting of heparin, the RMS value for NiTi-Heparin surface increased to about 68.5 nm. The obviously increase of RMS indicated the efficient grafting of heparin on Nitinol surface.The surface density of grafted heparin on Nitinol was determined by toluidine blue method, the heparin density on the NiTi-Heparin are 2.27 ± 0.05μg/cm2 right after been grafted onto Nitinol sheets. After stored in PBS as long as 30 days, the heparin density still kept at 1.96±0.05μg/cm2, The stable existence of heparin on NiTi-Heparin further proved the tight chemical grafting of heparin molecules on Nitinol surface.The predicted improvement in the biocompatibilities of modified Nitinol was confirmed by water contact angle measurement, protein adsorption, platelet adhesion, and plasma recalcification time determination. After being grafted with heparin, the water contact angle of Nitinol decreased significantly from 83° to 49°, the surface hydrophilicity of the NiTi-Heparin increased. The protein adsorption on the surface decreases. After been modified, the number of adhered platelets and the extent of aggregation decreased on the surface of the NiTi-Heparin sample, and most adherent platelets remain spherical and separated without pseudopodium. The PRT of the NiTi-Heparin was the longest, which was 1257 s comparing 325 s of glass and 863 s of Nitinol.The results of hemolysis assay, cell proliferation and cytotoxicity tests revealed that the grafting of heparin on NiTi kept the original positive performance of nitinol material. The hemolysis ratios of NiTi and NiTi-Heparin are both lower than 2%, which is far below the accepted threshold value of 5%. In the test of HUVEC proliferations and cytotoxicity assay, HUVEC kept their typical spindle shape and the similar cell density, which indicated that neither Nitinol nor modified Nitinol has influence on the proliferation of HUVEC.All the results indicate that ALD technology is of great potential for the manufacture of medical devices, especially for surface modifications and functionalization. ALD technology can help with modifications of inert metallic surfaces and therefore benefit implantable medical devices, especially intravascular stents.