NOS-Based Biopolymers: Towards Novel Thromboresistant No-Release Materials

Nitric Oxide releasing biopolymers have the potential to prolong vascular graft and stent potency without adverse systemic vasodilation. It was reported in literature that eNOS-overexpressing endothelial cell seeding of synthetic small diameter vascular grafts decreased human platelet aggregation by 46 and bovine aortic smooth muscle cell proliferation by 67.2 in vitro. We hypothesized that incorporating the enzyme nitric oxide synthase (NOS) in biocompatible polymeric matrix will provide a source of NO that utilizes endogenous compounds to maintain an unlimited supply of NO. To test this hypothesis, we have incorporated the enzyme nitric oxide synthase into a polyethyleneimine film using a layer-by-layer electrostatic deposition. This approach will provide a source of NO that utilizes endogenous compounds available in the blood matrix to maintain a constant supply of NO at the blood/device interface. When coated onto the surface of various blood-contacting implantable medical devices, it will provide NO fluxes at levels equal or greater than the normal endothelial cells, and for extended time periods. This configuration will help solve the issues of both thrombosis and stenosis that occur as side effects for several types of biomedical implants. Our results indicate a proof of principle of a new approach for making antithrombotic coatings for medical devices and implants based on NO release. We have demonstrated that NOS-based polymetric films successfully generate NO under physiologic conditions at small levels equal to and higher than those observed for endothelial cells. The level of NO release can be fine-tuned through varying the number of NOS layers in the film buildup. We have shown that NO fluxes from our NOS-based PEI films are sustained for prolonged periods of time, which has the potential of producing efficient, short and long-term, antithrombotic coatings for medical devices and blood-contacting tools such as stents and catheters. We also show that NO release from these coatings successfully dec

NOS-Based Biopolymers: Towards Novel Thromboresistant No-Release Materials

Nitric Oxide releasing biopolymers have the potential to prolong vascular graft and stent potency without adverse systemic vasodilation. It was reported in literature that eNOS-overexpressing endothelial cell seeding of synthetic small diameter vascular grafts decreased human platelet aggregation by 46 and bovine aortic smooth muscle cell proliferation by 67.2 in vitro. We hypothesized that incorporating the enzyme nitric oxide synthase (NOS) in biocompatible polymeric matrix will provide a source of NO that utilizes endogenous compounds to maintain an unlimited supply of NO. To test this hypothesis, we have incorporated the enzyme nitric oxide synthase into a polyethyleneimine film using a layer-by-layer electrostatic deposition. This approach will provide a source of NO that utilizes endogenous compounds available in the blood matrix to maintain a constant supply of NO at the blood/device interface. When coated onto the surface of various blood-contacting implantable medical devices, it will provide NO fluxes at levels equal or greater than the normal endothelial cells, and for extended time periods. This configuration will help solve the issues of both thrombosis and stenosis that occur as side effects for several types of biomedical implants. Our results indicate a proof of principle of a new approach for making antithrombotic coatings for medical devices and implants based on NO release. We have demonstrated that NOS-based polymetric films successfully generate NO under physiologic conditions at small levels equal to and higher than those observed for endothelial cells. The level of NO release can be fine-tuned through varying the number of NOS layers in the film buildup. We have shown that NO fluxes from our NOS-based PEI films are sustained for prolonged periods of time, which has the potential of producing efficient, short and long-term, antithrombotic coatings for medical devices and blood-contacting tools such as stents and catheters. We also show that NO release from these coatings successfully dec

NOS-based Biopolymers; Towards Novel Thromboresistant No-Release Materials

Nitric Oxide releasing biopolymers have the potential to prolong vascular graft and stent potency without adverse systemic vasodilation. It was reported in literature that eNOS-overexpressing endothelial cell seeding of synthetic small diameter vascular grafts decreased human platelet aggregation by 46 and bovine aortic smooth muscle cell proliferation by 67.2 in vitro. We hypothesized that incorporating the enzyme nitric oxide synthase (NOS) in biocompatible polymeric matrix will provide a source of NO that utilizes endogenous compounds to maintain an unlimited supply of NO. To test this hypothesis, we have incorporated the enzyme nitric oxide synthase into a polyethyleneimine film using a layer-by-layer electrostatic deposition. This approach will provide a source of NO that utilizes endogenous compounds available in the blood matrix to maintain a constant supply of NO at the blood/device interface. When coated onto the surface of various blood-contacting implantable medical devices, it will provide NO fluxes at levels equal or greater than the normal endothelial cells, and for extended time periods. This configuration will help solve the issues of both thrombosis and stenosis that occur as side effects for several types of biomedical implants. Our results indicate a proof of principle of a new approach for making antithrombotic coatings for medical devices and implants based on NO release. We have demonstrated that NOS-based polymetric films successfully generate NO under physiologic conditions at small levels equal to and higher than those observed for endothelial cells. The level of NO release can be fine-tuned through varying the number of NOS layers in the film buildup. We have shown that NO fluxes from our NOS-based PEI films are sustained for prolonged periods of time, which has the potential of producing efficient, short and long-term, antithrombotic coatings for medical devices and blood-contacting tools such as stents and catheters. We also show that NO release from these coatings successfully dec

Functional Layer-by-Layer Thin Films of Inducible Nitric Oxide (NO) Synthase Oxygenase and Polyethylenimine: Modulation of Enzyme Loading and NO-Release Activity

Nitric oxide (NO) release counteracts platelet aggregation and prevents the thrombosis cascade in the inner walls of blood vessels. NO-release coatings also prevent thrombus formation on the surface of blood-contacting medical devices. Our previous work has shown that inducible nitric oxide synthase (iNOS) films release NO fluxes upon enzymatic conversion of the substrate L-arginine. In this work, we report on the modulation of enzyme loading in layer-bylayer (LbL) thin films of inducible nitric oxide synthase oxygenase (iNOSoxy) on polyethylenimine (PEI). The layer of iNOSoxy is electrostatically adsorbed onto the PEI layer. The pH of the iNOSoxy solution affects the amount of enzyme adsorbed. The overall negative surface charge of iNOSoxy in solution depends on the pH and hence determines the density of adsorbed protein on the positively charged PEI layer. We used buffered iNOSoxy solutions adjusted to pHs 8.6 and 7.0, while saline PEI solution was used at pH 7.0. Atomic force microscopy imaging of the outermost layer shows higher protein adsorption with iNOSoxy at pH 8.6 than with a solution of iNOSoxy at pH 7.0. Graphite electrodes with PEI/iNOSoxy films show higher catalytic currents for nitric oxide reduction mediated by iNOSoxy. The higher enzyme loading translates into higher NO flux when the enzyme-modified surface is exposed to a solution containing the substrate and a source of electrons. Spectrophotometric assays showed higher NO fluxes with iNOSoxy/PEI films built at pH 8.6 than with films built at pH 7.0. Fourier transform infrared analysis of iNOSoxy adsorbed on PEI at pH 8.6 and 7.0 shows structural differences of iNOSoxy in films, which explains the observed changes in enzymatic activity. Our findings show that pH provides a strategy to optimize the NOS loading and enzyme activity in NOS-based LbL thin films, which enables improved NO release with minimum layers of PEI/NOS