Preventing Staphylococci Surgical Site Infections with a Nitric Oxide-Releasing Poly(lactic acid-co-glycolic acid) Suture Material.

Of the 27 million surgeries performed in the United States each year, a reported 2.6% result in a surgical site infection (SSI), and Staphylococci species are commonly the culprit. Alternative therapies, such as nitric oxide (NO)-releasing biomaterials, are being developed to address this issue. NO is a potent antimicrobial agent with several modes of action, including oxidative and nitrosative damage, disruption of bacterial membranes, and dispersion of biofilms. For targeted antibacterial effects, NO is delivered by exogenous donor molecules, like S-nitroso-N-acetylpenicillamine (SNAP). Herein, the impregnation of SNAP into poly(lactic-co-glycolic acid) (PLGA) for SSI prevention is reported for the first time. The NO-releasing PLGA copolymer is fabricated and characterized by donor molecule loading, leaching, and the amount remaining after ethylene oxide sterilization. The swelling ratio, water uptake, static water contact angle, and tensile strength are also investigated. Furthermore, its cytocompatibility is tested against 3T3 mouse fibroblast cells, and its antimicrobial efficacy is assessed against multiple Staphylococci strains. Overall, the NO-releasing PLGA copolymer holds promise as a suture material for eradicating surgical site infections caused by Staphylococci strains. SNAP impregnation affords robust antibacterial properties while maintaining the cytocompatibility and mechanical integrity.

Bioinspired superhydrophobic surfaces with silver and nitric oxide-releasing capabilities to prevent device-associated infections and thrombosis.

Bacteria-associated infections and thrombus formation are the two major complications plaguing the application of blood-contacting medical devices. Therefore, functionalized surfaces and drug delivery for passive and active antifouling strategies have been employed. Herein, we report the novel integration of bio-inspired superhydrophobicity with nitric oxide release to obtain a functional polymeric material with anti-thrombogenic and antimicrobial characteristics. The nitric oxide release acts as an antimicrobial agent and platelet inhibitor, while the superhydrophobic components prevent non-specific biofouling. Widely used medical-grade silicone rubber (SR) substrates that are known to be susceptible to biofilm and thrombus formation were dip-coated with fluorinated silicon dioxide (SiO2) and silver (Ag) nanoparticles (NPs) using an adhesive polymer as a binder. Thereafter, the resulting superhydrophobic (SH) SR substrates were impregnated with S-nitroso-N-acetylpenicillamine (SNAP, an NO donor) to obtain a superhydrophobic, Ag-bound, NO-releasing (SH-SiAgNO) surface. The SH-SiAgNO surfaces had the lowest amount of viable adhered E. coli (> 99.9 % reduction), S. aureus (> 99.8 % reduction), and platelets (> 96.1 % reduction) as compared to controls while demonstrating no cytotoxic effects on fibroblast cells. Thus, this innovative approach is the first to combine SNAP with an antifouling SH polymer surface that possesses the immense potential to minimize medical device-associated complications without using conventional systemic anticoagulation and antibiotic treatments.

Catalytic effect of transition metal-doped medical grade polymer on S-nitrosothiol decomposition and its biological response.

Nitric oxide (NO)-release from polymer metal composites is achieved through the incorporation of NO donors such as S-nitrosothiols (RSNO). Several studies have shown that metal nanoparticles catalytically decompose RSNO to release NO. In polymer composites, the NO surface flux from the surface can be modulated by the application of metal nanoparticles with a varying degree of catalytic activity. In this study, we compare the NO-releasing polymer composite design strategy - demonstrating how different ways of incorporating RSNO and metal nanoparticles can affect NO flux, donor leaching, or biological activity of the films. The first approach included blending both the RSNO and metal nanoparticle in the matrix (non-layered), while the second approach involved dip-coating metal nanoparticle/polymer layer on the RSNO-containing polymer composite (layered). Secondly, we compare both designs with respect to metal nanoparticles, including iron (Fe), copper (Cu), nickel (Ni), zinc (Zn), and silver (Ag). Differential NO surface flux is observed for each metal nanoparticle, with the Cu-containing polymer composites showing the highest flux for layered composites, whereas Fe demonstrated the highest NO flux for non-layered composites in 24 h. Additionally, a comparative study on NO flux modulation via the choice of metal nanoparticles is shown. Furthermore, mouse fibroblast cell viability when exposed to leachates from the polymer metal composites was dependent on (1) the design of the polymer composite where the layered approach performed better than non-layered composites (2) diffusion of metal nanoparticles from the composites plays a key role. Antibacterial activity on methicillin-resistant Staphylococcus aureus was also dependent on individual metal nanoparticles and flux levels in a 24 h in vitro CDC bioreactor study. Therefore, the study establishes the need for a layered polymer metal composite strategy that synergizes NO flux without negatively affecting biocompatibility.

Nanoarchitectonics of nitric oxide releasing supramolecular structures for enhanced antibacterial efficacy under visible light irradiation.

Light-controlled therapies offer a promising strategy to prevent and suppress infections caused by numerous bacterial pathogens. Excitation of exogenously supplied photosensitizers (PS) at specific wavelengths elicits levels of reactive oxygen intermediates toxic to bacteria. Porphyrin-based supramolecular nanostructure frameworks (SNF) are effective PS with unique physicochemical properties that have led to their widespread use in photomedicine. Herein, we developed a nitric oxide (NO) releasing, biocompatible, and stable porphyrin-based SNF (SNF-NO), which was achieved through a confined noncovalent self-assembly process based on π-π stacking. Characterization of the SNFs via scanning electron microscopy (SEM) and transmission electron microscopy (TEM) analysis showed the formation of three-dimensional, well-defined octahedral structures. These SNF-NO were shown to exhibit a red shift due to the noncovalent self-assembly of porphyrins, which also show extended light absorption to broadly cover the entire visible light spectrum to enhance photodynamic therapy (PDT). Under visible light irradiation (46 J cm-2), the SNF generates high yields of singlet oxygen (1O2) radicals, hydroxyl radicals (HO), superoxide radicals (O2), and peroxynitrite (ONOO-) radicals that have shown potential to enhance antimicrobial photodynamic therapy (APDT) against Gram-positive methicillin-resistant Staphylococcus aureus (MRSA) and Gram-negative Escherichia coli (E. coli). The resulting SNFs also exhibit significant biofilm dispersion and a decrease in biomass production. The combination of robust photosensitizer SNFs with nitric oxide-releasing capabilities is dynamic in its ability to target pathogenic infections while remaining nontoxic to mammalian cells. The engineered SNFs have enormous potential for treating and managing microbial infections.

Nitric Oxide-Releasing Nanofibrous Scaffolds Based on Silk Fibroin and Zein with Enhanced Biodegradability and Antibacterial Properties.

Clinical applications of scaffolds and implants have been associated with bacterial infection resulting in impaired tissue regeneration. Nanofibers provide a versatile structure for both antimicrobial molecule delivery and tissue engineering. In this study, the nitric oxide (NO) donor molecule S-nitrosoglutathione (GSNO) and the natural biodegradable polymer zein (ZN) were combined with silk fibroin (SF) to develop antibacterial and biodegradable nanofibrous scaffolds for tissue engineering applications. The compatibility and intermolecular interactions of SF and ZN were studied using differential scanning calorimetry and Fourier transform infrared spectroscopy. The incorporation of ZN increased the hydrophobicity of the fibers and resulted in a more controlled and prolonged NO release profile lasting for 48 h. Moreover, the degradation kinetics of the fibers was significantly improved after blending with ZN. The results of tensile testing indicated that the addition of ZN and GSNO had a positive effect on the strength and stretchability of SF fibers and did not adversely affect their mechanical properties. Finally, due to the antibacterial properties of both NO and ZN, the SF-ZN-GSNO fibers showed a synergistically high antibacterial efficacy with 91.6 ± 2.5% and 77.5 ± 3.1% reduction in viability of adhered Staphylococcus aureus and Escherichia coli after 24 h exposure, respectively. The developed NO-releasing fibers were not only antibacterial but also non-cytotoxic and successfully enhanced the proliferation and growth of fibroblast cells, which was quantitatively studied by a CCK-8 assay and visually observed through fluorescent staining. Overall, SF-ZN-GSNO fibers developed in this study were biodegradable and highly antibacterial and showed great cytocompatibility with fibroblasts, indicating their promising potential for a range of tissue engineering and medical device applications.

Potent, Broad-Spectrum Antimicrobial Effects of S-Nitroso-N-acetylpenicillamine-Impregnated Nitric Oxide-Releasing Latex Urinary Catheters.

Although numerous prevention and intervention techniques have been developed to counteract catheter-associated urinary tract infections (CAUTIs), urinary catheters remain one of the most common sources of hospital-acquired infections. Nitric oxide (NO), a gaseous free radical responsible for regulating many physiological functions in the body, has gained immense popularity due to its potent, broad-spectrum antimicrobial activity, which is capable of combating medical device-associated infections. In this work, a straightforward solvent-swelling method was used to load the NO donor S-nitroso-N-acetyl-penicillamine (SNAP) into commercial latex catheters (SNAP-UCs) for the first time. The effects of swelling catheters with different concentrations of SNAP solutions (25-125 mg/mL SNAP in tetrahydrofuran (THF)) were studied by measuring the NO release kinetics, SNAP loading, and SNAP leaching. SNAP-UCs impregnated with a 50 mg/mL SNAP-THF solution were found to maximize the amount of SNAP loaded into the latex (0.115 ± 0.009 mg SNAP/mg catheter) and showed physiological levels of NO release (>2 × 10-10 mol min-1 cm-2) over 7 days and minimal SNAP leaching (<2%). SNAP-UCs showed impressive in vitro contact-based and diffusible antimicrobial efficacy against three CAUTI-associated pathogens, reducing the viability of adhered and planktonic Escherichia coli, Proteus mirabilis, and Staphylococcus aureus by ∼98.0 to 99.1% (adhered) and 86.3-96.3% (planktonic) compared to control latex catheters. In vitro cytotoxicity against 3T3 mouse fibroblasts using a CCK-8 assay showed that SNAP-UCs were noncytotoxic (>90% viability). In summary, SNAP-UCs show stable, noncytotoxic NO release characteristics capable of potent, broad-spectrum antimicrobial activity, demonstrating great potential for reducing the devastating effects associated with CAUTIs.

A Review on Antibacterial Silk Fibroin-based Biomaterials: Current State and Prospects.

Bacterial contamination of biomaterials is a common problem and a serious threat to human health worldwide. Therefore, the development of multifunctional biomaterials that possess antibacterial properties and can resist infection is a continual goal for biomedical applications. Silk fibroin (SF), approved by U.S. Food and Drug Administration (FDA) as a biomaterial, is one of the most widely studied natural polymers for biomedical applications due to its unique mechanical properties, biocompatibility, tunable biodegradation, and versatile material formats. In the last decade, many methods have been employed for the development of antibacterial SF-based biomaterials (SFBs) such as physical loading or chemical functionalization of SFBs with different antibacterial agents and bio-inspired surface modifications. In this review, we first describe the current understanding of the composition and structure-properties relationship of SF as a leading-edge biomaterial. Then we demonstrate the different antibacterial agents and methods implemented for the development of bactericidal SFBs, their mechanisms of action, and different applications. We briefly address their fabrication methods, advantages, and limitations, and finally discuss the emerging technologies and future trends in this research area.

Nitric Oxide-Releasing Gelatin Methacryloyl/Silk Fibroin Interpenetrating Polymer Network Hydrogels for Tissue Engineering Applications.

Bacterial infection is one of the principal reasons for the failure of tissue engineering scaffolds. Therefore, the development of multifunctional scaffolds that not only are able to guide tissue regeneration but also can inhibit bacterial colonization is of great importance for tissue engineering applications. In this study, a highly antibacterial, biocompatible, and biodegradable scaffold based on silk fibroin (SF) and gelatin methacryloyl (GelMA) was prepared. Sequential cross-linking of GelMA and SF under UV irradiation and methanol treatment, respectively, resulted in the formation of interpenetrating network (IPN) hydrogels with a porous structure. In addition, impregnation of the hydrogels with a nitric oxide (NO) donor molecule, S-nitroso-N-acetylpenicillamine (SNAP), led to the development of NO-releasing scaffolds with strong antibacterial properties. According to the obtained results, the addition of SF to GelMA hydrogels caused an enhancement in the mechanical properties and NO release kinetics and prevented their rapid enzymatic degradation in aqueous media. Furthermore, swelling the GelMA-SF scaffolds with SNAP resulted in a bacteria reduction efficiency of >99.9% against Gram-positive (Staphylococcus aureus) and Gram-negative (Escherichia coli) bacteria. The scaffolds also showed great cytocompatibility in vitro by increasing the proliferation and supporting the adhesion of 3T3 mouse fibroblast cells. Overall, GelMA-SF-SNAP showed great promise to be used as a scaffold for tissue engineering and wound healing applications.

Nitric oxide releasing halloysite nanotubes for biomedical applications.

Halloysite nanotubes (HNTs) are natural aluminosilicate clay that have been extensivelyexplored fordelivery of bioactive agents in biomedical applications because of their desirable features including unique hollow tubular structure, good biocompatibility, high mechanical strength, and extensive functionality. For the first time, in this work, functionalized HNTs are developed as a delivery platform for nitric oxide (NO), a gaseous molecule, known for its important roles in the regulation of various physiological processes. HNTs were first hydroxylated and modified with an aminosilane crosslinker, (3-aminopropyl) trimethoxysilane (APTMS), to enable the covalent attachment of a NO donor precursor, N-acetyl-d-penicillamine (NAP). HNT-NAP particles were then converted to NO-releasing S-nitroso-N-acetyl-penicillamine HNT-SNAP by nitrosation. The total NO loading on the resulting nanotubes was 0.10 ± 0.07 µmol/mg which could be released using different stimuli such as heat and light. Qualitative (Fourier-transform infrared spectroscopy and Nuclear magnetic resonance) and quantitative (Ninhydrin and Ellman) analyses were performed to confirm successful functionalization of HNTs at each step. Field emission scanning electron microscopy (FE-SEM) showed that the hollow tubular morphology of the HNTs was preserved after modification. HNT-SNAP showed concentration-dependent antibacterial effects against Gram-positive Staphylococcus aureus (S. aureus), resulting in up to 99.6% killing efficiency at a concentration of 10 mg/mL as compared to the control. Moreover, no significant cytotoxicity toward 3T3 mouse fibroblast cells was observed at concentrations equal or below 2 mg/mL of HNT-SNAP according to a WST-8-based cytotoxicity assay. The SNAP-functionalized HNTs represent a novel and efficient NO delivery system that holds the potential to be used, either alone or in combination with polymers for different biomedical applications.

Synergistic Approach to Develop Antibacterial Electrospun Scaffolds Using Honey and S-Nitroso-N-acetyl Penicillamine.

Bacterial infections have been increasingly recognized as the major reason for the failure of tissue engineering scaffolds. Therefore, there is a need for novel and multifunctional biomaterials that not only enhance tissue regeneration but also can combat infections. An antibacterial and bioactive scaffold was fabricated in this study by incorporation of honey and a nitric oxide (NO) donor, S-nitroso-N-acetyl-penicillamine (SNAP), into polylactic acid (PLA) nanofibers using a single-jet electrospinning method. The morphology of the prepared nanofibers was observed using a scanning electron microscope. PLA/honey/SNAP (PLA/HN/SNAP) nanofibers had an average diameter of 624.92 ± 137.69 nm and showed a sustained release of NO for 48 h. The scaffolds were characterized for their chemical composition via Fourier-transform infrared spectroscopy. Moreover, the tensile properties of nanofibers along with their wettability, water retention ability, and water vapor transmission rate were evaluated. The results of antibacterial studies revealed that the synergistic combination of honey and SNAP significantly reduced the viability of Gram positive Staphylococcus aureus and Gram negative Escherichia coli. In addition, qualitative and quantitative 3T3 fibroblast cell culturing experiments proved that the PLA/HN/SNAP scaffolds supported better cell attachment and proliferation compared to PLA. The promising results obtained in this study indicate that PLA/HN/SNAP nanofibrous scaffolds have great potential for tissue engineering applications.

Silk Nanoparticles: A Natural Polymeric Platform for Nitric Oxide Delivery in Biomedical Applications.

In this study, the preparation and characterization of nitric oxide (NO) releasing silk fibroin nanoparticles (SF NPs) are described for the first time. S-Nitroso-N-acetylpenicillamine (SNAP)-loaded SF NPs (SNAP-SF NPs) were prepared via an antisolvent/self-assembling method by adding a SNAP/ethanol solution to an aqueous SF solution and freeze-thawing. The prepared SNAP-SF NPs had a diameter ranging from 300 to 400 nm and an overall negative charge of -28.76 ± 0.73 mV. Among the different SNAP/SF ratios tested, the highest encapsulation efficiency (18.3 ± 1.3%) and loading capacity (9.1 ± 0.6%) values were attributed to the 1:1 ratio. The deconvolution of the amide I band in the FTIR spectra of SF NPs and SNAP-SF NPs showed an increase in the ß-sheet content for SNAP-SF NPs, confirming the hydrophobic interactions between SNAP and silk macromolecules. SNAP-SF NPs released up to 1.31 ± 0.02 × 10-10 mol min-1 mg-1 NO over a 24 h period. Moreover, SNAP-SF NPs showed concentration-dependent antibacterial effects against methicillin-resistant Staphylococcus aureus and Escherichia coli. Furthermore, they did not elicit any marked cytotoxicity against 3T3 mouse fibroblast cells at concentrations equal to or below 2 mg/mL. Overall, these results demonstrated that SNAP-SF NPs have great potential to be used as a NO delivery platform for biomedical applications such as tissue engineering and wound healing, where synergistic properties of SF and NO are desired.

Enhanced cellular response elicited by addition of amniotic fluid to alginate hydrogel-electrospun silk fibroin fibers for potential wound dressing application.

This study aimed to evaluate a novel bioactive wound dressing from alginate hydrogel-electrospun silk fibroin (SF) fibers with the ability to deliver amniotic fluid (AF) to the wound site. AF is highly enriched with multiple therapeutic agents that can promote cellular response and wound healing. In this study, electrospun SF fibers were first fabricated by electrospinning method and then combined with the alginate hydrogel (ALG) containing AF. Different dressings were made by changing the alginate to AF ratio. The scanning electron microscopy images revealed that SF fibers were fully covered with alginate hydrogel. In addition, the modulus of the dressing decreased by lowering ALG/AF ratios, while a reverse trend was observed for the elongation-at-break. In vitro AF release profiles indicated that an increase in the concentration of ALG leads to a reduction in the AF amount. Fibroblast culturing on the fabricated dressings demonstrated that cellular proliferation, spreading, and secretion of collagen enhanced with increasing AF. Taken together, our results provide a novel bioactive dressing with great potentials for speeding up the healing process in severe wounds.