Attachment of Fibrinogen on Ion Beam Treated Polyurethane.

Protein-stable coverage of the artificial implant is a key problem for biocompatibility. In the present study, a protein layer was attached covalently to a polyurethane surface treated by an ion beam. A plasma system consisting of a vacuum chamber (0.8 Pa pressure) with a high voltage electrode powered by a short pulse (20 µS pulse duration and 200 Hz pulse repetition) generator was designed. Polyurethane with a formulation certified as a material for medical implants was treated by nitrogen ions with an energy of 20 keV and 5 × 1014-1016 ions/cm2 fluence range. Wettability measurements, X-ray photoelectron, Raman, Fourier transform infrared attenuated total reflection, and ellipsometry spectra showed a significant change in the structure of the surface layer of the treated polyurethane. The surface of the treated polyurethane contained a carbonised layer containing condensed aromatic clusters with terminal free radicals. The surface energy of polyurethane surface increased from 33 to 65 mJ/m2. The treated polyurethane surface became capable of adsorbing and chemically binding protein (fibrinogen). The designed system for ion beam treatment can be used for surface activation of biomedical polymer devices, where a total protein coverage is required.

Foreign Body Reaction (Immune Response) for Artificial Implants Can Be Avoided: An Example of Polyurethane in Mice for 1 Week.

Despite great success with artificial implants for the human body, modern implants cannot solve major health problems. The reason is an immune reaction of organisms to artificial implants, known as the foreign body reaction. We have found a way to avoid or decrease the foreign body reaction. The surface of an artificial implant is modified with condensed aromatic structures containing free radicals, which provide a covalent attachment of host proteins in a native conformation. The total protein coverage prevents the direct contact of immune cells with the implant surface, and the immune cells are not activated. As a result, the immune response of the organism is not generated, and the artificial implant is not isolated from the tissue; there is no collagen capsule, low activity of macrophages, low cell proliferation, and low inflammatory activity.

Extracellular-Vesicle-Based Coatings Enhance Bioactivity of Titanium Implants-SurfEV.

Extracellular vesicles (EVs) are nanoparticles released by cells that contain a multitude of biomolecules, which act synergistically to signal multiple cell types. EVs are ideal candidates for promoting tissue growth and regeneration. The tissue regenerative potential of EVs raises the tantalizing possibility that immobilizing EVs on implant surfaces could potentially generate highly bioactive and cell-instructive surfaces that would enhance implant integration into the body. Such surfaces could address a critical limitation of current implants, which do not promote bone tissue formation or bond bone. Here, we developed bioactive titanium surface coatings (SurfEV) using two types of EVs: secreted by decidual mesenchymal stem cells (DEVs) and isolated from fermented papaya fluid (PEVs). For each EV type, we determined the size, morphology, and molecular composition. High concentrations of DEVs enhanced cell proliferation, wound closure, and migration distance of osteoblasts. In contrast, the cell proliferation and wound closure decreased with increasing concentration of PEVs. DEVs enhanced Ca/P deposition on the titanium surface, which suggests improvement in bone bonding ability of the implant (i.e., osteointegration). EVs also increased production of Ca and P by osteoblasts and promoted the deposition of mineral phase, which suggests EVs play key roles in cell mineralization. We also found that DEVs stimulated the secretion of secondary EVs observed by the presence of protruding structures on the cell membrane. We concluded that, by functionalizing implant surfaces with specialized EVs, we will be able to enhance implant osteointegration by improving hydroxyapatite formation directly at the surface and potentially circumvent aseptic loosening of implants.

Covalent Biofunctionalization of the Inner Surfaces of a Hollow-Fiber Capillary Bundle Using Packed-Bed Plasma Ion Implantation.

Hollow-fiber capillary bundles are widely used in the production of medical devices for blood oxygenation and purification purposes such as in cardiopulmonary bypass, hemodialysis, and hemofiltration, but the blood interfacing inner surfaces of these capillaries provide poor hemocompatibility. Here, we present a novel method of packed-bed plasma ion implantation (PBPII) for the modification of the inner surfaces of polymeric hollow-fiber bundles enclosed in a cassette. The method is simple and can be performed on an intact hollow-fiber bundle cassette by the placement of a hollow cylindrical electrode, connected to a negative high-voltage pulse generator, around the cassette. The method does not require the insertion of electrodes inside the capillaries or the cassette. Nitrogen gas is fed into the capillaries inside the cassette by connecting the inlet of the cassette to a gas source. Upon the application of negative high-voltage bias pulses to the electrode, plasma is ignited inside the cassette, achieving the surface modification of both the internal and external surfaces of the capillaries. Fourier transform infrared-attenuated total reflectance spectroscopy of the PBPII-treated capillaries revealed the formation of aromatic C═C bonds, indicating the progressive carbonization of the capillary surfaces. The PBPII treatment was found to be uniform along the capillaries and independent of the radial position in the cassette. Atomic force microscopy of cross sections through the capillaries revealed that the increased stiffness associated with the carbonized layer on the inner surface of the PBPII-treated capillary has a depth (∼40 nm) consistent with that expected for ions accelerated by the applied bias voltage. The modified internal surfaces of the capillary bundle showed a greatly increased wettability and could be biofunctionalized by covalently immobilizing protein directly from the buffer solution. The direct, reagent-free protein immobilization was demonstrated using tropoelastin as an example protein. Covalent binding of the protein was confirmed by its resistance to removal by hot sodium dodecyl sulfate detergent washing, which is known to disrupt physical binding.

The protein corona determines the cytotoxicity of nanodiamonds: implications of corona formation and its remodelling on nanodiamond applications in biomedical imaging and drug delivery.

The use of nanodiamonds for biomedical and consumer applications is growing rapidly. As their use becomes more widespread, so too do concerns around their cytotoxicity. The cytotoxicity of nanodiamonds correlates with their cellular internalisation and circulation time in the body. Both internalisation and circulation time are influenced by the formation of a protein corona on the nanodiamond surface. However, a precise understanding of both how the corona forms and evolves and its influence on cytotoxicity is lacking. Here, we investigated protein corona formation and evolution in response to two classes of nanodiamonds, pristine and aminated, and two types of proteins, bovine serum albumin and fibronectin. Specifically, we found that a corona made of bovine serum albumin (BSA), which represents the most abundant protein in blood plasma, reduced nanodiamond agglomeration. Fibronectin (FN9-10), the second most abundant protein found in the plasma, exhibited a significantly higher nanodiamond binding affinity than BSA, irrespective of the nanodiamond surface charge. Finally, nanodiamonds with a BSA corona displayed less cytotoxicity towards nonphagocytic liver cells. However, regardless of the type of corona (FN9-10 or BSA), both classes of nanodiamonds induced substantial phagocytic cell death. Our results emphasise that a precise understanding of the corona composition is fundamental to determining the fate of nanoparticles in the body.

Biological impact of nanodiamond particles - label free, high-resolution methods for nanotoxicity assessment.

Current methods for the assessment of nanoparticle safety that are based on 2D cell culture models and fluorescence-based assays show limited sensitivity and they lack biomimicry. Consequently, the health risks associated with the use of many nanoparticles have not yet been established. There is a need to develop in vitro models that mimic physiology more accurately and enable high throughput assessment. There is also a need to set up new assays that offer high sensitivity and are label-free. Here we developed 'mini-liver' models using scaffold-free bioprinting and used these models together with label-free nanoscale techniques for the assessment of toxicity of nanodiamond produced by laser-assisted technology. Results showed that NDs induced cytotoxicity in a concentration and exposure-time dependent manner. The loss of cell function was confirmed by increased cell stiffness, decreased cell membrane barrier integrity and reduced cells mobility. We further showed that NDs elevated the production of reactive oxygen species and reduced cell viability. Our approach that combined mini-liver models with label-free high-resolution techniques showed improved sensitivity in toxicity assessment. Notably, this approach allowed for label-free semi-high throughput measurements of nanoparticle-cell interactions, thus could be considered as a complementary approach to currently used methods.

Enhanced biocompatibility of polyurethane-type shape memory polymers modified by plasma immersion ion implantation treatment and collagen coating: An in vivo study.

As one of the promising smart materials, polyurethane-type shape memory polymers (SMPU) have been extensively investigated as potential biomedical implant materials. However, the hydrophobicity and bio-inertness of SMPU are major problems for biomedical applications. We applied plasma immersion ion implantation (PIII) to increase surface wettability and enable one-step covalent, functionalisation of SMPU with biological molecules to create a tuneable, biocompatible surface. The changes of surface properties due to PIII treatment in nitrogen plasma were determined by measurements of morphology, contact angle, surface energy, and nanoindentation. Collagen attachment on SMPU with and without PIII treatment was measured by Attenuated total reflectance-Fourier transform infrared (ATR-FTIR). To investigate in vivo biocompatibility, SMPU with/without PIII and with/without collagen were subcutaneously implanted in mice. SMPU implants with surrounding tissue were collected at days 1, 3, 7, 14 and 28 to study acute/subacute inflammatory responses at histopathological and immunohistochemical levels. The results show that PIII treatment improves wettability and releases residual stress in the SMPU surfaces substantially. Covalent attachment of collagen on PIII treated SMPU in a single step incubation was demonstrated by its resistance to removal by rigorous Sodium Dodecyl Sulfonate (SDS) washing. The in-vivo results showed significantly lower acute/subacute inflammation in response to SMPU with PIII treatment + collagen coating compared to untreated SMPU, collagen coated untreated SMPU, and PIII treated SMPU, characterised by lower total cell numbers, macrophages, neovascularisation, cellular proliferation, cytokine production, and matrix metalloproteinase production. This comprehensive in vivo study of PIII treatment with protein coating demonstrates that the combination of PIII treatment and collagen coating is a promising approach to enhance the biocompatibility of SMPU, facilitating its application as an implantable biomaterial.

Plasma-Activated Substrate with a Tropoelastin Anchor for the Maintenance and Delivery of Multipotent Adult Progenitor Cells.

Conventional wound therapy utilizes wound coverage to prevent infection, trauma, and fluid and thermal loss. However, this approach is often inadequate for large and/or chronic wounds, which require active intervention via therapeutic cells to promote healing. To address this need, a patch which delivers multipotent adult progenitor cells (MAPCs) is developed. Medical-grade polyurethane (PU) films are modified using plasma immersion ion implantation (PIII), which creates a radical-rich layer capable of rapidly and covalently attaching biomolecules. It is demonstrated that a short treatment duration of 400 s maximizes surface activation and wettability, minimizes reduction in gas permeability, and preserves the hydrolytic resistance of the PU film. The reactivity of PIII-treated PU is utilized to immobilize the extracellular matrix protein tropoelastin in a functional conformation that stably withstands medical-grade ethylene oxide sterilization. The PIII-treated tropoelastin-functionalized patch significantly promotes MAPC adhesion and proliferation over standard PU, while fully maintaining cell phenotype. Topical application of the MAPC-seeded patch transfers cells to a human skin model, while undelivered MAPCs repopulate the patch surface for subsequent cell transfer. The potential of this new wound patch as a reservoir for the sustained delivery of therapeutic MAPCs and cell-secreted factors for large and/or non-healing wounds is indicated in the findings.

Plasma processing of PDMS based spinal implants for covalent protein immobilization, cell attachment and spreading.

PDMS is widely used for prosthetic device manufacture. Conventional ion implantation is not a suitable treatment to enhance the biocompatibility of poly dimethyl siloxane (PDMS) due to its propensity to generate a brittle silicon oxide surface layer which cracks and delaminates. To overcome this limitation, we have developed new plasma based processes to balance the etching of carbon with implantation of carbon from the plasma source. When this carbon was implanted from the plasma phase it resulted in a surface that was structurally similar and intermixed with the underlying PDMS material and not susceptible to delamination. The enrichment in surface carbon allowed the formation of carbon based radicals that are not present in conventional plasma ion immersion implantation (PIII) treated PDMS. This imparts the PDMS surfaces with covalent protein binding capacity that is not observed on PIII treated PDMS. The change in surface energy preserved the function of bound biomolecules and enhanced the attachment of MG63 osteosarcoma cells compared to the native surface. The attached cells, an osteoblast interaction model, showed increased spreading on the treated over untreated surfaces. The carbon-dependency for these beneficial covalent protein and cell linkage properties was tested by incorporating carbon from a different source. To this end, a second surface was produced where carbon etching was balanced against implantation from a thin carbon-based polymer coating. This had similar protein and cell-binding properties to the surfaces generated with carbon inclusion in the plasma phase, thus highlighting the importance of balancing carbon etching and deposition. Additionally, the two effects of protein linkage and bioactivity could be combined where the cell response was further enhanced by covalently tethering a biomolecule coating, as exemplified here with the cell adhesive protein tropoelastin. Providing a balanced carbon source in the plasma phase is applicable to prosthetic device fabrication as illustrated using a 3-dimensional PDMS balloon prosthesis for spinal implant applications. Consequently, this study lays the groundwork for effective treatments of PDMS to selectively recruit cells to implantable PDMS fabricated biodevices.

Bound ("Glassy") Rubber as a Free Radical Cross-linked Rubber Layer on a Carbon Black.

A model of rubber with a cross-linked rubber layer on a carbon black filler has been proposed. The cross-links are the result of free radical reactions generated by carbon atoms with unpaired electrons at the edge of graphitic sheets in a carbon black filler. The experimental study of the cross-linking reactions in polyisoprene was done on a flat carbonized surface after ion beam implantation. The cross-linking process in the polyisoprene macromolecules between two particles was simulated. The model with a cross-linked rubber layer on a carbon filler as a "glassy layer" explains the mechanical properties of the rubber materials.

Plasma Ion Implantation of Silk Biomaterials Enabling Direct Covalent Immobilization of Bioactive Agents for Enhanced Cellular Responses.

Silk fibroin isolated from Bombyx mori cocoons is a promising material for a range of biomedical applications, but it has no inherent cell-interactive domains, necessitating functionalization with bioactive molecules. Here we demonstrate significantly enhanced cell interactions with silk fibroin biomaterials in the absence of biofunctionalization following surface modification using plasma immersion ion implantation (PIII). Further, PIII treated silk fibroin biomaterials supported direct covalent immobilization of proteins on the material surface in the absence of chemical cross-linkers. Surface analysis after nitrogen plasma and PIII treatment at 20 kV revealed that the silk macromolecules are significantly fragmented, and at the higher fluences of implanted ions, surface carbonization was observed to depths corresponding to that of the ion penetration. Consistent with the activity of radicals created in the treated surface layer, oxidation was observed on contact with atmospheric oxygen and the PIII treated surfaces were capable of direct covalent immobilization of bioactive macromolecules. Changes in thickness, amide and nitrile groups, refractive index, and extinction coefficient in the wavelength range 400-1000 nm as a function of ion fluence are presented. Reactions responsible for the restructuring of the silk surface under ion beam treatment that facilitate covalent binding of proteins and a significant improvement in cell interactions on the modified surface are proposed.

EPDM Rubber Modified by Nitrogen Plasma Immersion Ion Implantation.

Ethylene-propylene diene monomer rubber (EPDM) was treated by plasma immersion ion implantation (PIII) with nitrogen ions of 20 keV energy and fluence from 1013 to 1016 ions/cm². The Fourier-transform infrared attenuated total reflection spectra, atomic force microscopy and optical microscopy showed significant structure changes of the surface. The analysis of an interface of PIII treated EPDM rubber with polyurethane binder showed a cohesive character of the adhesion joint fracture at the presence of solvent and interpreted as covalent bond network formation between the PIII treated rubber and the adhesive.

Effect of plasma immersion ion implantation on polycaprolactone with various molecular weights and crystallinity.

Polycaprolactone with five different molecular weights was spin-coated on silicon wafers and plasma immersion ion implanted (PIII) with ion fluence in the range 5 × 1014-2 × 1016 ions/cm2. The effects of PIII treatment on the optical properties, chemical structure, crystallinity, morphology, gel fraction formation and wettability were investigated. As in the case of a number of previously studied polymers, oxidation and hydrophobic recovery of the PIII treated PCL follow second order kinetics. CAPA 6250, which has the lowest molecular weight and the highest degree of crystallinity of the untreated PCL films studied, has the highest carbonization of the modified layer after PIII treatment. Untreated medical grade PCL films, mPCL PC12 (Perstorp) and mPCL OsteoporeTM have similar chemical structures and crystallinity. Accordingly, the chemical and structural transformations caused by PIII treatment and post-treatment oxidation are almost identical for these two polymers. In general, PIII treatment destroys the nano-scale lamellar structure and results in a reduction of PCL crystallinity. Examination after washing PIII treated PCL films in toluene confirmed our hypothesis that cross-linking due to PIII treatment is significantly higher in semi-crystalline PCL as compared with amorphous polymers.

Plasma mediated protein immobilisation enhances the vascular compatibility of polyurethane with tissue matched mechanical properties.

Polyurethanes are a diverse class of polymers, with independently tunable mechanical and biodegradation properties making them a versatile platform material for biomedical implants. Previous iterations have failed to adequately embody appropriate mechanical and biological properties, particularly for vascular medicine where strength, compliance and multifaceted biocompatibility are required. We have synthesized a new polyurethane formulation with finely tuned mechanical properties, combining high strength and extensibility with a low Young's modulus. Additional cross-linking during synthesis enhanced stability and limits leaching. Under cyclic testing, hysteresis was minimal following completion of the initial cycles, indicating the robustness of the material. Building on this platform, we used plasma immersion ion implantation to activate the polymer surface and functionalized it with recombinant human tropoelastin. With tropoelastin covalently bound to the surface, human coronary endothelial cells showed improved attachment and proliferation. In the presence of heparinized whole blood, tropoelastin-coated polyurethane showed very low thrombogenicity in both static and flow conditions. Using this formulation, we synthesized robust, elastic prototype conduits which easily retained multiple sutures and were successfully implanted in a pilot rat aortic interposition model. We have thus created an elastic, strong biomaterial platform, functionalized with an important regulator of vascular biology, with the potential for further evaluation as a new synthetic graft material.

A sterilizable, biocompatible, tropoelastin surface coating immobilized by energetic ion activation.

Biomimetic materials which integrate with surrounding tissues and regulate new tissue formation are attractive for tissue engineering and regenerative medicine. Plasma immersion ion-implanted (PIII) polyethersulfone (PES) provides an excellent platform for the irreversible immobilization of bioactive proteins and peptides. PIII treatment significantly improves PES wettability and results in the formation of acidic groups on the PES surface, with the highest concentration observed at 40-80 s of PIII treatment. The elastomeric protein tropoelastin can be stably adhered to PIII-treated PES in a cell-interactive conformation by tailoring the pH and salt levels of the protein-surface association conditions. Tropoelastin-coated PIII-treated PES surfaces are resistant to molecular fouling, and actively promote high levels of fibroblast adhesion and proliferation while maintaining cell morphology. Tropoelastin, unlike other extracellular matrix proteins such as fibronectin, uniquely retains full bioactivity even after medical-grade ethylene oxide sterilization. This dual approach of PIII treatment and tropoelastin cloaking allows for the stable, robust functionalization of clinically used polymer materials for directed cellular interactions.

Structural Analysis and Protein Functionalization of Electroconductive Polypyrrole Films Modified by Plasma Immersion Ion Implantation.

Conducting polymers are good candidates for electronic biomedical devices such as biosensors, artificial nerves, and electrodes for brain tissue. Functionalizing the conducting polymer surface with bioactive molecules can limit adverse immune reactions to the foreign body and direct tissue integration. In this work, we demonstrate a simple one-step method to attach biomolecules covalently to a conductive polymer. Electrochemically synthesized polypyrrole was activated using plasma immersion ion implantation (PIII) in nitrogen. A short treatment with relatively low ion fluence (20 s) was found to enable direct covalent immobilization of protein upon incubation in a protein solution, while the protein is easily removed from untreated polypyrrole by washing in buffer. The covalent nature of the protein immobilization was demonstrated by its resistance to elution when repeatedly washed with SDS detergent. Changes in the surface properties and their evolution with time after PIII activation were studied by a combination of attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR), Raman spectroscopy, X-ray photoelectron spectroscopy (XPS), atomic force microscopy (AFM), cyclic voltammetry, and water contact angle measurements. Notable changes in the chemistry of the modified layer in polypyrrole include the appearance of nitrile groups that gradually disappear with time and oxidation of the surface that increases over time in air. The kinetics of surface energy are consistent with the generation of radicals in the modified layer that are lost predominantly through oxidation. The conductivity of the modified surface layer (64 nm in thickness) decreases for low fluence treatments and is partially restored after high fluence treatment. This simple surface modification process opens up the possibility of creating biologically active interfaces for electro-stimulating biomedical devices and electrical sensing of neurological processes.

Biospectroscopy of Nanodiamond-Induced Alterations in Conformation of Intra- and Extracellular Proteins: A Nanoscale IR Study.

The toxicity of nanomaterials raises major concerns because of the impact that nanomaterials may have on health, which remains poorly understood. We need to explore the fate of individual nanoparticles in cells at nano and molecular levels to establish their safety. Conformational changes in secondary protein structures are one of the main indicators of impaired biological function, and hence, the ability to identify these changes at a nanoscale level offers unique insights into the nanotoxicity of materials. Here, we used nanoscale infrared spectroscopy and demonstrated for the first time that nanodiamond-induced alterations in both extra- and intracellular secondary protein structures lead to the formation of antiparallel ß-sheet, ß-turns, intermolecular ß-sheet, and aggregation of proteins. These conformational changes of the protein structure may result in the loss of functionality of proteins and in turn lead to adverse effects.

Plasma Ion Activated Expanded Polytetrafluoroethylene Vascular Grafts with a Covalently Immobilized Recombinant Human Tropoelastin Coating Reducing Neointimal Hyperplasia.

Expanded polytetrafluoroethylene (ePTFE) vascular conduits with less than or equal to 6 mm internal diameter typically occlude due to a combination of thrombus formation and neointimal hyperplasia. We hypothesized that by layering the polymerized elastin precursor, human tropoelastin, in the synthetic vessel lumen we could mimic the internal elastic lamina and so maintain low thrombogenicity while significantly reducing smooth muscle cell proliferation. The luminal surfaces of ePTFE conduits were activated with plasma immersion ion implantation (PIII) treatment to facilitate covalent attachment of tropoelastin. Multilayered tropoelastin vessels (2TE) enhanced endothelial cell attachment and proliferation in vitro and were superior to materials lacking the protein. In an ovine carotid interposition model of graft compatibility, partially tropoelastin coated vessels (1TE) thrombosed at a greater rate than control ePTFE, but 2TE maintained the same patency as controls. 2TE showed a significant reduction in neointimal area down to 9.7 ± 5.2% (p < 0.05) in contrast to 32.3 ± 3.9% for ePTFE alone. This reduction was due to a halving of the number of smooth muscle cells present and a corresponding reduction in their proliferation. 2TE, but not 1TE, enhanced the vascular compatibility of these materials: while both tropoelastin presentations increased in vitro endothelialization, only 2TE displayed the dual benefits of maintained hemocompatibility and simultaneously suppressed neointimal hyperplasia in vivo. We conclude that 2TE surface modification provides a significant improvement over ePTFE vascular conduits in a pilot large animal model study and presents an attractive path toward clinical applications for reduced diameter vessels.

Plasma-Activated Tropoelastin Functionalization of Zirconium for Improved Bone Cell Response.

The mechanical strength, durability, corrosion resistance, and biocompatibility of metal alloys based on zirconium (Zr) and titanium (Ti) make them desirable materials for orthopedic implants. However, as bioinert metals, they do not actively promote bone formation and integration. Here we report a plasma coating process for improving integration of such metal implants with local bone tissue. The coating is a stable carbon-based plasma polymer layer that increased surface wettability by 28%, improved surface elasticity to the range exhibited by natural bone, and additionally covalently bound the extracellular matrix protein, tropoelastin, in an active conformation. The thus biofunctionalized material was significantly more resistant to medical-grade sterilization by steam, autoclaving or gamma-ray irradiation, retaining >60% of the adhered tropoelastin molecules and preserving full bioactivity. The interface of the coating and metal was robust so as to resist delamination during surgical insertion and in vivo deployment, and the plasma process employed was utilized to also coat the complex 3D geometries typical of orthopedic implants. Osteoblast-like osteosarcoma cells cultured on the biofunctionalized Zr surface exhibited a significant 30% increase in adhesion and up to 70% improvement in proliferation. Cells on these materials also showed significant early stage up-regulation of bone marker expression (alkaline phosphatase, 1.8 fold; osteocalcin, 1.4 fold), and sustained up-regulation of these genes (alkaline phosphatase, 1.3 fold; osteocalcin, 1.2 fold) in osteogenic conditions. In addition, alkaline phosphatase production significantly increased (2-fold) on the functionalized surfaces, whereas bone mineral deposition increased by 30% above background levels compared to bare Zr. These findings have the potential to be readily translated to the development of improved Zr and Ti-based implants for accelerated bone repair.

Immobilization of bioactive plasmin reduces the thrombogenicity of metal surfaces.

Components of many vascular prostheses including endovascular stents, heart valves and ventricular assist devices are made using metal alloys. In these blood contacting applications, metallic devices promote blood clotting, which is managed clinically by profound platelet suppression and/or anticoagulation. Here it is proposed that the localized immobilization of bioactive plasmin, a critical mediator of blood clot stability, may attenuate metallic prosthesis-induced thrombus formation. Previously described approaches to covalently immobilize biomolecules on implantable materials have relied on complex chemical linker chemistry, increasing the possibility of toxic side effects and reducing bioactivity. We utilize a plasma deposited thin film platform to covalently immobilize biologically active plasmin on stainless steel substrates, including stents. A range of in vitro whole blood assays demonstrate striking reductions in thrombus formation. This approach has profound potential to improve the efficacy of a wide range of metallic vascular implants.