Novel Nitric Oxide (NO)-Releasing Polymers and their Biomedical Applications.

Two common factors that can cause complications with indwelling biomedical devices are thrombus and infection. Nitric oxide (NO) is known to be a potent inhibitor of platelet activation and an antimicrobial agent. In this dissertation work, two novel approaches to achieving long-term NO release from polymers were studied and evaluated for their potential biomedical applications. First, S-nitroso-N-acetypenicillamine (SNAP)-doped Elast-eon E2As creates an inexpensive polymer that can locally deliver physiologically relevant levels of NO. SNAP was found to be stable in E2As during shelf-life stability and ethylene oxide sterilization studies. The SNAP/E2As polymer was coated on the inner walls of extracorporeal circulation (ECC) circuits and was found to preserve the platelet count at ~100% of baseline and reduce thrombus area after 4 h blood flow in a rabbit model. The SNAP/E2As polymer was also used to fabricate NO-releasing catheters that were implanted in sheep veins for 7 d. The SNAP/E2As catheters significantly reduced the amount of thrombus and bacterial adhesion. In the second approach, the NO release from diazeniumdiolated dibutylhexanediamine (DBHD/N2O2)-doped polymers was significantly improved using poly(lactic-co-glycolic acid) (PLGA) additives. Acid-capped PLGA additives were found to cause high initial bursts of NO, while ester-capped PLGA additives extended the NO release for up to 14 d. Poly(vinyl chloride)- and Elast-eon E2As were used as the base polymers for combined DBHD/N2O2 and PLGA coatings on the inner walls of ECC circuits. After 4 h of blood flow, the E2As-based NOrel circuits preserved platelets at a higher level than PVC-based NOrel circuits (97% and 80% of baseline, respectively). This demonstrates that the inherent hemocompatibility properties of the base polymer can also influence the efficiency of the NO release coatings. A DBHD/N2O2-doped SG-80A polymer material was also studied and used to fabricate patches that were applied to scald burn wounds infected with Acinetobacter baumannii. The NO released from these patches applied to the wounds is shown to significantly reduce the A. baumannii infection after 24 h. The results for both of types of NO-releasing polymers studied here demonstrated greatly enhanced hemocompatibility properties that can be applied to a wide variety of blood-contacting medical devices.PHDChemistryUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/109037/1/brisbel_1.pd

In Vitro and in Vivo Study of Sustained Nitric Oxide Release Coating Using Diazeniumdiolate-doped Poly(vinyl chloride) Matrix with Poly(lactide-co-glycolide) Additive

Nitric oxide (NO) is an endogenous vasodilator as well as natural inhibitor of platelet adhesion and activation that can be released from a NO donor species, such as diazeniumdiolated dibutylhexanediamine (DBHD/N2O2) within a polymer coating. In this study, various Food and Drug Administration approved poly(lactic-co-glycolic acid) (PLGA) species were evaluated as additives to promote a prolonged NO release from DBHD/N2O2 within a plasticized poly(vinyl chloride) (PVC) matrix. When using an ester-capped PLGA additive with a slow hydrolysis time, the resulting coatings continuously release between 7 and 18 × 10−10 mol cm−2 min−1 NO for 14 days at 37 °C in PBS buffer. The corresponding pH changes within the polymer films were visualized using pH sensitive indicators and are shown to correlate with the extended NO release pattern. The optimal combined diazeniumdiolate/PLGA-doped NO release (NOrel) PVC coating was evaluated in vitro and its effect on the hemodynamics was also studied within a 4 h in vivo extracorporeal circulation (ECC) rabbit model of thrombogenicity. Four out of 7 control circuits clotted within 3 h, whereas all the NOrel coated circuits were patent after 4 h. Platelet counts on the NOrel ECC were preserved (79 ± 11% compared to 54 ± 6% controls). The NOrel coatings showed a significant decrease in the thrombus area as compared to the controls. Results suggest that by using ester-capped PLGAs as additives to a conventional plasticized PVC material containing lipophilic diazeniumdiolates, the NO release can be prolonged for up to 2 weeks by controlling the pH within the organic phase of the coating

Attenuation Of Thrombin-Mediated Fibrin Formation: Via Changes In Fibrinogen Conformation Induced By Reaction With S -Nitroso- N -Acetylpenicillamine, But Not S -Nitrosoglutathione

Previous work using a 4 h rabbit thrombogenicity model has shown that a nitric oxide (NO)-generating polymer extracorporeal circuit (ECC) with infusion of S-nitroso-N-acetyl-penicillamine (SNAP) preserved platelets even though platelets were activated, as shown by an increase in the glycoprotein p-selectin. The platelet preservation mechanism was shown to be due to a changing fibrinogen structure leading to attenuation of platelet aggregation. To understand the effects that SNAP, another RSNO, S-nitroso-glutathione (GSNO), as well as the non-RSNO, sodium nitroprusside (SNP), may have on human fibrinogen polymerization, this in vitro study evaluated the effects of released NO on thrombin-mediated fibrin formation and fibrinogen structure. Thrombin-induced fibrin formation at 300 μM SNAP (50 + 11% of baseline) was significantly reduced compared to SNAP\u27s parent, N-acetyl-penicillamine (NAP), (95 ± 13%) after 1 h of RSNO exposure. GSNO, its parent, glutathione (GSH), and 1000 ppm NO gas did not attenuate the thrombin-mediated fibrin formation. SNAP, NAP and SNP exposure for 1 h, however, did not decrease thrombin activity by directly inhibiting thrombin itself. Changes in fibrinogen conformation as measured by intrinsic tryptophan fluorescence significantly decreased in the 300 μM SNAP (38:057 ± 1196 mean fluorescence intensity (MFI)) and SNP (368:617 ± 541 MFI) groups versus the NAP control (47:937 ± 1196 MFI). However, infused 1000 ppm NO gas had no direct effect on the ITF after 1 h of incubation at 37 °C. High performance liquid chromatography (HPLC) showed that fibrinogen degradation by 0.03 U ml-1 thrombin was concentration-dependently reduced after 1 h with SNAP but not with NAP or SNP. Western blotting analysis was done on the RSNOs, SNAP and NAP, and the non-RSNO, SNP, incubated fibrinogen solutions and the results showed that the percent level of the Aγ dimer with respect to total Aγ dimer + γ monomer was significantly reduced in the case of the SNAP group when compared to the SNP group. These results suggest that NO donors such as SNAP and SNP induce fibrinogen conformational changes by potentially nitrosating fibrinogen tyrosine residues. These NO-mediated fibrinogen changes induced via NO donors may provide another mechanism of NO for improving thromboresistance in ECCs

Reduction of Thrombosis and Bacterial Infection via Controlled Nitric Oxide (NO) Release from SNitrosoNacetylpenicillamine (SNAP) Impregnated CarboSil Intravascular Catheters

[Image: see text] Nitric oxide (NO) has many important physiological functions, including its ability to inhibit platelet activation and serve as potent antimicrobial agent. The multiple roles of NO in vivo have led to great interest in the development of biomaterials that can deliver NO for specific biomedical applications. Herein, we report a simple solvent impregnation technique to incorporate a nontoxic NO donor, S-nitroso-N-acetylpenicillamine (SNAP), into a more biocompatible biomedical grade polymer, CarboSil 20 80A. The resulting polymer-crystal composite material yields a very stable, long-term NO release biomaterial. The SNAP impregnation process is carefully characterized and optimized, and it is shown that SNAP crystal formation occurs in the bulk of the polymer after solvent evaporation. LC-MS results demonstrate that more than 70% of NO release from this new composite material originates from the SNAP embedded CarboSil phase, and not from the SNAP species leaching out into the soaking solution. Catheters prepared with CarboSil and then impregnated with 15 wt % SNAP provide a controlled NO release over a 14 d period at physiologically relevant fluxes and are shown to significantly reduce long-term (14 day) bacterial biofilm formation against Staphylococcus epidermidis and Pseudonomas aeruginosa in a CDC bioreactor model. After 7 h of catheter implantation in the jugular veins of rabbit, the SNAP CarboSil catheters exhibit a 96% reduction in thrombus area (0.03 ± 0.01 cm(2)/catheter) compared to the controls (0.84 ± 0.19 cm(2)/catheter) (n = 3). These results suggest that SNAP impregnated CarboSil can become an attractive new biomaterial for use in preparing intravascular catheters and other implanted medical devices

Reduction of Thrombosis and Bacterial Infection via Controlled Nitric Oxide (NO) Release from S‑Nitroso‑N‑acetylpenicillamine (SNAP) Impregnated CarboSil Intravascular Catheters

Nitric oxide (NO) has many important physiological functions, including its ability to inhibit platelet activation and serve as potent antimicrobial agent. The multiple roles of NO in vivo have led to great interest in the development of biomaterials that can deliver NO for specific biomedical applications. Herein, we report a simple solvent impregnation technique to incorporate a nontoxic NO donor, S-nitroso-N-acetylpenicillamine (SNAP), into a more biocompatible biomedical grade polymer, CarboSil 20 80A. The resulting polymer-crystal composite material yields a very stable, long-term NO release biomaterial. The SNAP impregnation process is carefully characterized and optimized, and it is shown that SNAP crystal formation occurs in the bulk of the polymer after solvent evaporation. LC-MS results demonstrate that more than 70% of NO release from this new composite material originates from the SNAP embedded CarboSil phase, and not from the SNAP species leaching out into the soaking solution. Catheters prepared with CarboSil and then impregnated with 15 wt % SNAP provide a controlled NO release over a 14 d period at physiologically relevant fluxes and are shown to significantly reduce long-term (14 day) bacterial biofilm formation against Staphylococcus epidermidis and Pseudonomas aeruginosa in a CDC bioreactor model. After 7 h of catheter implantation in the jugular veins of rabbit, the SNAP CarboSil catheters exhibit a 96% reduction in thrombus area (0.03 ± 0.01 cm2/catheter) compared to the controls (0.84 ± 0.19 cm2/catheter) (n = 3). These results suggest that SNAP impregnated CarboSil can become an attractive new biomaterial for use in preparing intravascular catheters and other implanted medical devices