Local Release Of Nitric Oxide Research Articles

Pillar[5]arene stabilized gold nanoparticles for the enhanced light-triggered nitric oxide release with antibacterial and antibiofilm activities

The reactivity of guests confined in well-defined host cavities act significantly different from that in bulk systems. In this work, pillar[5]arene not only works as a stabilizer for the fabrication of gold nanoparticles, but also act as an accelerator for the enhanced photo-triggered release of gaseous nitric oxide (NO) from encapsulated NO donor. The host–guest complex can dramatically improve the photoactivity of loaded guest molecule to generate antibacterial NO molecules through the confinement effect of pillar[5]arene cavity. The synergistic photo-triggered localized NO release and photo-thermal effect of host–guest complex demonstrated impressive antibacterial behaviors against Gram-negative bacterial strain Escherichia coli, Gram-positive bacterial strain Staphylococcus aureus and methicillin-resistant Staphylococcus aureus. Moreover, this biocompatible photo-responsive complex can also inhibit bacterial biofilm formation and disrupt the pre-existing biofilms.

(366) Potential Effectiveness of Focused Vibration Treatment for Psychogenic Erectile Dysfunction: A Case Series

Abstract Introduction The current non-pharmacological treatment of psychogenic erectile dysfunction (pED) is based on cognitive-behavioral interventions that often involve the participation of the partner, such as desensitization procedures, sensory focusing, and couples counseling. However, it is important to note that in many cases, patients may not have a stable partner or their partner cannot participate in the treatment. The application of local mechanical vibration has shown promising results in increasing skin blood flow and nitric oxide production in healthy young adults. The relaxation of smooth muscle in the penis initiates and maintains an erection. The local release of nitric oxide from autonomic dilator nerves and vascular endothelium is crucial in initiating and sustaining this process. Technological advances have allowed the design of vibrating devices that men can use during sexual encounters. Objective To determine if the on-demand use of a focused vibration device can increase blood flow and promote relaxation of erectile tissue to improve erectile function in patients with pED. Methods Seven sexually active men diagnosed with pED based on criteria for Erectile Disorder from the Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition (DSM-5) and without a stable partner, were instructed to use a vibrating device called TENUTO II during their sexual encounters for duration of 8 weeks. This device generates focal vibrations on the penis and the perineal area in intervals of 10 seconds of vibration followed by 10 seconds of pause, with a frequency of 50Hz and an amplitude of 2mm. The five-item Spanish version of the International Index of Erectile Function (IIEF-5) questionnaire was used to assess symptoms before and after the procedure and the Erectile Dysfunction Inventory of Treatment Satisfaction (EDITS) questionnaire was used for evaluating satisfaction with treatment. Results After the using the device, all patients enrolled with a mean age of 36.1 years (range 28-45) experienced an improvement in erectile function, mean IIEF-5 score pretreatment was 11(range 5–13) mean posttreatment IIEF score was 20 (range 18-27). No adverse effects were reported during the treatment. Conclusions This case series suggests that use of an on-demand focused vibration device could be a potentially effective and safe therapeutic option for men with pED. However, further studies with a larger number of participants and a more rigorous research design are needed to confirm these preliminary findings. Disclosure Yes, this is sponsored by industry/sponsor: MysteryVibe. Clarification: Industry funding only - investigator initiated and executed study. Any of the authors act as a consultant, employee or shareholder of an industry for: Mysteryvibe.

In Situ Photo-crosslinkable Hyaluronic Acid/Gelatin Hydrogel for Local Nitric Oxide Delivery.

An increasing number of studies have shown that the local release of nitric oxide (NO) from hydrogels stimulates tissue regeneration by modulating cell proliferation, angiogenesis, and inflammation. The potential biomedical uses of NO-releasing hydrogels can be expanded by enabling their application in a fluid state, followed by controlled gelation triggered by an external factor. In this study, we engineered a hydrogel composed of methacrylated hyaluronic acid (HAGMA) and thiolated gelatin (GELSH) with the capacity for in situ photo-cross-linking, coupled with localized NO release. To ensure a gradual and sustained NO release, we charged the hydrogels with poly(l-lactic-co-glycolic acid) (PLGA) nanoparticles functionalized with S-nitrosoglutathione (GSNO), safeguarding SNO group integrity during photo-cross-linking. The formation of thiol-ene bonds via the reaction between GELSH's thiol groups and HAGMA's vinyl groups substantially accelerated gelation (by a factor of 6) and increased the elastic modulus of hydrated hydrogels (by 1.9-2.4 times). HAGMA/GELSH hydrogels consistently released NO over a 14 day duration, with the release of NO depending on the hydrogels' equilibrium swelling degree, determined by the GELSH-to-HAGMA ratio. Biocompatibility assessments confirmed the suitability of these hydrogels for biological applications as they display low cytotoxicity and stimulated fibroblast adhesion and proliferation. In conclusion, in situ photo-cross-linkable HAGMA/GELSH hydrogels, loaded with PLGA-GSNO nanoparticles, present a promising avenue for achieving localized and sustained NO delivery in tissue regeneration applications.

Photochemical Nitric Oxide Release from S‐Nitrosothiol‐Functionalized Chitosan

AbstractS‐nitrosothiols (RSNOs) are nitric oxide (NO) donor molecules with potential biomedical applications since they mimic NO biological functions, such as vasodilation and inflammatory process regulation. In this work, chitosan (CS), a natural biopolymer widely used as a biomaterial, is functionalized with thioglycolic acid (TGA) or mercaptosuccinic acid (MSA) in reactions mediated by a carbodiimide, yielding CS with thiol contents of 88 µmol g–1 (CS‐TGA) and 126 µmol g–1 (CS‐MSA). The subsequent S‐nitrosation of CS‐TGA and CS‐MSA generates the corresponding S‐nitroso derivatives, CS‐TGA(SNO) and CS‐MSA(SNO). The UV–Vis spectral changes associated with the S‐NO bond cleavage of CS‐TGA(SNO) and CS‐MSA(SNO), monitored in solution over 18 days of storage in a common refrigerator, reveal that these S‐nitroso CS derivatives display enhanced thermal stability in the dark, with half‐life values of 36.1 for CS‐TGA(SNO) and 16.1 days for CS‐MSA(SNO). Conversely, CS‐TGA(SNO) and CS‐MSA(SNO) can release NO photochemically on‐demand under visible light irradiation in a first‐order process triggered by photon absorption that leads to a half‐life of 33 min for both S‐nitroso CS. These results show that CS‐TGA(SNO) and CS‐MSA(SNO) may emerge as new CS‐based biomaterials for providing light‐triggered local NO release in biomedical applications.

Enhancement of Nitric Oxide Bioavailability by Modulation of Cutaneous Nitric Oxide Stores.

The generation of nitric oxide (NO) in the skin plays a critical role in wound healing and the response to several stimuli, such as UV exposure, heat, infection, and inflammation. Furthermore, in the human body, NO is involved in vascular homeostasis and the regulation of blood pressure. Physiologically, a family of enzymes termed nitric oxide synthases (NOS) generates NO. In addition, there are many methods of non-enzymatic/NOS-independent NO generation, e.g., the reduction of NO derivates (NODs) such as nitrite, nitrate, and nitrosylated proteins under certain conditions. The skin is the largest and heaviest human organ and contains a comparatively high concentration of these NODs; therefore, it represents a promising target for many therapeutic strategies for NO-dependent pathological conditions. In this review, we give an overview of how the cutaneous NOD stores can be targeted and modulated, leading to a further accumulation of NO-related compounds and/or the local and systemic release of bioactive NO, and eventually, NO-related physiological effects with a potential therapeutical use for diseases such as hypertension, disturbed microcirculation, impaired wound healing, and skin infections.

Mesoporous calcium peroxide-ignited NO generation for amplifying photothermal immunotherapy of breast cancer

L-arginine (LA) as a semiessential amino acid within human body has shown great promise in cancer therapy since it can continuously generate nitric oxide (NO) in the presence of inducible NO synthase (iNOS) or reactive oxygen species (ROS). However, NO production efficiency in tumor tissue is severely restricted by the hypoxic and H2O2-deficient tumor microenvironment (TME). Herein, mesoporous calcium peroxide (CaO2) with the ability of supplying O2/H2O2 has been prepared to encapsulate and oxidize LA to achieve controllable local release of NO in the acidic TME. Interestingly, the generated NO can aggravate the abnormal retention of Ca2+ in cells, thus resulting in amplified oxidative stress and immunogenic cell death. To improve the stability of CaO2 in the body fluid, polydopamine (PDA) is further introduced to coat on the surface of LA-CaO2 and its high photothermal conversion property will reinforce the immunogenic dying of tumor cells. Such a NO-enhanced phototherapeutic nanoplatform, LA-CaO2@PDA, exhibit the most effective treatment in inhibiting primary and distant tumor growth and suppressing lung metastasis of 4T1 tumor by provoking a strong antitumor immune response.

3D printed nitric oxide-releasing poly(acrylic acid)/F127/cellulose nanocrystal hydrogels.

Hydrogels have been used as matrices for the topical delivery of nitric oxide (NO) for achieving vasodilation, wound healing and analgesic actions. More recently, supramolecular hydrogels comprised of poly(acrylic acid) (PAA) and micellar Pluronic F127 (F127), prepared by thermal reaction, emerged as a suitable matrix for the incorporation of hydrophilic NO donors, such as S-nitrosoglutathione (GSNO). Herein, we describe an innovative method for the three-dimensional (3D) printing of cellulose nanocrystal (CNC)-containing and semi-interpenetrating PAA/F127 hydrogels by PAA photopolymerization via digital light processing (DLP), in the absence of organic solvents. Scanning electron microscopy showed that, differently from typical porous PAA-based hydrogels, the 3D printed PAA/F127/CNC hydrogels have dense morphology. By using transmission electron microscopy we confirmed for the first time the presence of F127 micelles in the printable resin, and their preservation after the photopolymerization process. The F127 micelles conferred compressive recoverability to the 3D printed PAA/F127/CNC hydrogels, widening their potential applications as soft biomaterials. PAA/F127/CNC hydrogels charged with GSNO are shown to release NO spontaneously upon hydration at initial rates that depend on the GSNO charge and are higher in the presence of CNC. As local NO release may exert cell proliferation action, 3D printed PAA/F127/CNC/GSNO hydrogels may serve as a versatile soft biomaterial for local NO delivery in regenerative medicine and other biomedical applications.

Atherogenic diet-diminished endothelial glycocalyx contributes to impaired vasomotor properties in rat.

Hypercholesterolemia- and atherosclerosis-caused vasomotor property dysfunction may be involved in many clinic manifestations of atherosclerosis, including angina, acute myocardial infarction, and sudden cardiac death. However, its underlying mechanism is not clear. The endothelial glycocalyx is a protective surface layer on the endothelial cells, serving as a molecular sieve, cell adhesion modulator, and mechanosensor for blood flow. In the present study, we demonstrated by confocal microscopy in Sprague-Dawley (SD) male rats fed a 12-wk high-cholesterol diet (HC) compared with the normal diet (NC) that the dimension of the endothelial glycocalyx reduced significantly in both the common carotid artery (2.89 ± 0.41 µm and 3.25 ± 0.44 μm, respectively) and the internal sinus region (2.35 ± 0.07 µm and 3.46 ± 0.86 μm, respectively). Furthermore, we showed by real-time PCR that this dimension modification of endothelial glycocalyx may be attributed to a significant downregulation of heparan sulfate proteoglycan (HSPG)-related genes, including syndecan-3, glypican-1, and EXT1, not resulting from an enhanced shedding of sulfated glycosaminoglycans (sGAGs) from the vessel wall to the plasma. Meanwhile, the mean contraction and relaxation forces of the common carotid artery with responses to norepinephrine (NE) and acetylcholine (ACh) decreased ~0.34- and 0.13-fold, respectively, accompanied by a lower level of nitric oxide (NO) release. These findings suggest that the atherogenic high cholesterol diet diminished endothelial glycocalyx and disturbed the local NO release, thus contributing to the impaired vasomotor properties of the vessel.NEW & NOTEWORTHY Twelve-week high-cholesterol (HC) diet reduces the thickness of the endothelial glycocalyx in Sprague-Dawley (SD) male rats, which is mainly attributed to a downregulation of heparan sulfate proteoglycan-related genes (syndecan-3, glypican-1, EXT1), not resulting from an enhanced shedding of sulfated glycosaminoglycans (sGAGs) into the plasma. HC-diminished glycocalyx may disturb its mechanotransduction of local shear stress, lower nitric oxide (NO) release, and impair vasomotor responses to norepinephrine (NE) and acetylcholine (ACh).

Polydopamine-Modified Copper-Doped Titanium Dioxide Nanotube Arrays for Copper-Catalyzed Controlled Endogenous Nitric Oxide Release and Improved Re-Endothelialization.

The controllable release is necessary for ideal drug delivery technologies. Because of their high specific surface area and high porosity, titanium dioxide nanotubes (TNTs) have been widely used as drug carriers in medical devices. By loading copper as the catalyst, nitric oxide (NO) generation was facilitated by catalyzing the decomposition of renewable endogenous NO donors in vivo. Herein, the long-term controllable release profile of NO is highlighted owing to the multilayer polydopamine (PDA) cap structure. Different layers of PDA are used to adjust the NO release behavior, and the results show that three layers of PDA can not only effectively prevent the burst release of NO but also maintain long-term stable release of copper ion and NO. The bioactivity of the NO generated from three-layer PDA-modified copper-loaded TNTs (PDA-3L-NTCu2) and unmodified copper-loaded TNTs (NTCu2) are verified by our work, indicating effective inhibition of platelet activation, thrombosis, inflammation, and intimal hyperplasia. Importantly, the PDA-3L-NTCu2 show selectively promote the growth of endothelial cells in vitro and outstanding re-endothelialization for 4 weeks in vivo, as compared to NTCu2, TNTs, and 316L stain steel. This study suggests that copper-loaded with PDA modification helps us achieve controlled long-term stable local NO release with well-retained bioactivity and enhanced re-endothelialization.

Cannabinoid receptor type 1 in the bed nucleus of the stria terminalis modulates cardiovascular responses to stress via local N-methyl-D-aspartate receptor/neuronal nitric oxide synthase/soluble guanylate cyclase/protein kinase G signaling.

Endocannabinoid neurotransmission in the bed nucleus of the stria terminalis is involved in the control of cardiovascular responses to stress. However, the local mechanisms involved is this regulation are not known. The purpose of this study was to assess an interaction of bed nucleus of the stria terminalis endocannabinoid neurotransmission with local nitrergic signaling, as well as to investigate the involvement of local N-methyl-D-aspartate glutamate receptor and nitric oxide signaling in the control of cardiovascular responses to acute restraint stress by bed nucleus of the stria terminalis endocannabinoid neurotransmission in rats. The first protocol evaluated the effect of intra-bed nucleus of the stria terminalis microinjection of the selective cannabinoid receptor type 1 receptor antagonist AM251 in nitrite/nitrate content in the bed nucleus of the stria terminalis following restraint stress. The other protocols evaluated the impact of local pretreatment with the selective N-methyl-D-aspartate glutamate receptor antagonist LY235959, the selective neuronal nitric oxide synthase inhibitor Nω-propyl-L-arginine, the soluble guanylate cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one, or the protein kinase G inhibitor KT5823 in restraint-evoked cardiovascular changes following bed nucleus of the stria terminalis treatment with AM251. Bilateral microinjection of AM251 into the bed nucleus of the stria terminalis increased local nitric oxide release during restraint stress. Bed nucleus of the stria terminalis treatment with the cannabinoid receptor type 1 receptor antagonist also enhanced the tachycardia caused by restraint stress, but without affecting arterial pressure increase and sympathetic-mediated cutaneous vasoconstriction. The facilitation of restraint-evoked tachycardia following bed nucleus of the stria terminalis treatment with the cannabinoid receptor type 1 receptor antagonist was completely inhibited by local pretreatment with LY235959, Nω-propyl-L-arginine, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one, or KT5823. Our results provide evidence that bed nucleus of the stria terminalis endocannabinoid neurotransmission inhibits local N-methyl-D-aspartate/neuronal nitric oxide synthase/soluble guanylate cyclase/protein kinase G signaling, and this mechanism is involved in the control of the cardiovascular responses to stress.

How can nitric oxide help osteogenesis?

Nitric oxide (NO) is endogenously produced free radical that plays important biological roles, such as, the promotion of vasodilation, angiogenesis, tissue repair, wound healing process, antioxidant, antitumoral and antimicrobial actions. Although the regenerative effects of NO in soft tissues have been extensively reported, its role in bone tissue repair has not been completely addressed. Both constitutive and inducible forms of NO synthase (NOS) are expressed in bone-derived cells, and some important cytokines, such as IL-1 and TNF, are potent stimulators of NO production. The effects of NO on bone tissue are dependent on its concentration. NO has dichotomous biological effects, at low concentrations (pico-nano molar range), NO may promote proliferation, differentiation and survival of osteoblasts, whereas at high concentrations (micromolar range) NO may inhibit bone resorption and formation. Therefore, at a certain concentration range, NO can avoid osteoclast-mediated bone resorption and promote osteoblast growth. Due to the potential beneficial effects of NO in bone tissue regeneration, the exogenous administration of NO might find important biomedical applications. As NO is a free radical and a gas, the administration of NO donors/generators has been explored in tissue repair. The delivery of NO to the bone using macromolecular NO releasing scaffolds has been shown to increase osteogenesis with a relevant impact in dental and orthopedist areas. In this sense, this review presents and discusses the recent and important progresses in the effects of NO/NO donors in bone tissue, and highlights the promising approach in the design and use of NO donors allied to biomaterials in the sustained and localized NO release for bone tissue regeneration.

Poly-ε-caprolactone/polysulfhydrylated polyester blend: A platform for topical and degradable nitric oxide-releasing materials

Polymeric biomaterials for implantable medical devices are subject to demands that go beyond passive biocompatibility. Inhibition of thrombus formation, reduced inflammatory response and the eventual complete bioabsorption of the implant are central challenges for emerging polymeric biomaterials. Nitric oxide (NO) released at the biomaterial/tissue interface may inhibit platelet adhesion and improve tissue integration in blood-contacting/implanted devices. In addition, local NO release may increase vasodilation, thereby improving ischemic conditions in topical applications. In this study, we present a novel degradable NO-releasing polymeric platform based on blended poly-ε-caprolactone (PCL)/polysulfhydrylated polyester (PSPE). PCL formed miscible blends with PSPE up to 45 wt% of PSPE. S-nitrosation of the PCL/PSPE blends converted the surface PSPE component into a NO-releasing polynitrosated polyester (PNPE). Real-time chemiluminescence NO detection showed that blends with different PCL:PSPE ratios allow surface S-nitrosothiol concentrations in the range of 0.5–1.5 μmol/cm2. Topical application of blended PCL/PNPE films on the healthy skin led to a 6–10-fold increase in the skin blood flow due to local NO delivery, as shown by in vivo laser-Doppler flowmetry. Blended PCL/PSPE films displayed higher hydrophilicity and a 2–11-fold increase in the hydrolytic mass loss, compared to pure PCL films. Therefore, PCL/PSPE blend is a potential biomaterial platform for local NO delivery and more quickly degradable biomedical devices.

Control of cardiovascular responses to stress by CRF in the bed nucleus of stria terminalis is mediated by local NMDA/nNOS/sGC/PKG signaling

The aims of the present study were to assess an interaction of corticotropin-releasing factor (CRF) neurotransmission within the bed nucleus of the stria terminalis (BNST) with local nitrergic signaling, as well as to investigate an involvement of activation of local NMDA glutamate receptor and nitric oxide (NO) signaling in control of cardiovascular responses to acute restraint stress by BNST CRF neurotransmission in rats. We observed that CRF microinjection into the BNST increased local NO release during restraint stress. Furthermore, bilateral microinjection of CRF into the BNST enhanced both the arterial pressure and heart rate increases evoked by restraint stress, but without affecting the sympathetically-mediated cutaneous vasoconstriction. The facilitation of both pressor and tachycardiac responses to restraint stress evoked by BNST treatment with CRF were completely inhibited by local pretreatment with either the selective NMDA glutamate receptor antagonist LY235959, the selective neuronal nitric oxide synthase (nNOS) inhibitor Nω-Propyl-l-arginine (NPLA), the soluble guanylate cyclase (sGC) inhibitor 1H-[1,2,4]Oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) or the protein kinase G (PKG) inhibitor KT5823. Taken together, these results provide evidence that BNST CRF neurotransmission facilitates local NMDA-mediated glutamatergic neurotransmission and activates nitrergic signaling, and this pathway is involved in control of cardiovascular responses to stress.

Response of Local Nitric Oxide Release to Manual Acupuncture and Electrical Heat in Humans: Effects of Reinforcement Methods.

This study was to examine the influences of manual acupuncture (MA) and electrical heat corresponding to reinforcing methods on nitric oxide (NO) release over the skin regions in humans. A device with collecting solution was taped to the skin surface along pericardium (PC) or lung (LU) meridian. Acupuncture needles were gently inserted into PC 4 with reinforcing stimulation (low force/rate) for 20 minutes in the MA group. LU11 on the finger was heated (43-44°C) by electrical heat for 20 minutes. Biocapture was consecutively conducted for two 20-minute intervals during and after each treatment. Total nitrite and nitrate (NOx−) in the collecting samples were quantified using chemiluminescence in blinded fashion. Baseline NOx− levels are higher and tended to be higher over PC and LU acupoints during the 1st biocapture. NOx− levels over PC regions were consistently increased by MA during both intervals. NOx− concentrations over LU acupoints were increased and tended to be increased by electrical heat in the 1st and 2nd biocapture. The results suggest that reinforcing MA and electrical heat induce NO released from the local skin regions with higher levels at acupoints, which improve local circulation and contribute to the beneficial effects of the therapies.

Controlled delivery of nitric oxide in cardiovascular tissue engineering

Event Abstract Back to Event Controlled delivery of nitric oxide in cardiovascular tissue engineering Qiang Zhao1, Zhihong Wang1*, Jimin Zhang1 and Deling Kong1 1 Nankai University, Key Laboratory of Bioactive Materials (Ministry of Education), College of Life Sciences, China Nitric oxide (NO) is an important signaling molecule in the cardiovascular system. NO secreted by the endothelium holds important physiological function in maintaining the vessel patency and promoting tissue regeneration. Our group has previously prepared polysaccharide- and peptide- hydrogel based NO-releasing biomaterials by conjugation of glycosylated NO donors, which are highly stable and only decompose to release NO under the catalysis of corresponding enzymes, glycosidases. The two biomaterials showed significant effect in treatment of mouse hindlimb ischemia and skin injury. They can also be used as carrier for the delivery of stem cells, and improved therapeutic efficacy was observed in mouse myocardial infarction model by increasing cell engraftment and angiogenic paracrine action. In addition, we put forward a new concept that the enzyme immobilized on the vascular graft can catalyze exogenously supplied prodrug to release NO locally and sustainedly. In vivo implantation by replacing rat abdominal aorta demonstrated that localized NO release improved tissue regeneration, remodeling, as well as physiological function, which are critical for the small-diameter vascular grafts to maintain long-term patency. The work was financially supported by NSFC projects (No. 81171478, 81371699, and 81522023).

Origin of Long-Term Storage Stability and Nitric Oxide Release Behavior of CarboSil Polymer Doped with S-Nitroso-N-acetyl-D-penicillamine.

The prolonged and localized delivery of nitric oxide (NO), a potent antithrombotic and antimicrobial agent, has many potential biomedical applications. In this work, the origin of the long-term storage stability and sustained NO release mechanism of S-nitroso-N-acetyl-d-penicillamine (SNAP)-doped CarboSil 20 80A polymer, a biomedical thermoplastic silicone-polycarbonate-urethane, is explored. Long-term (22 days) localized NO release is achieved by utilizing a cross-linked silicone rubber as topcoats, which can greatly reduce the amount of SNAP, NAP, and NAP disulfide leaching from the SNAP-doped CarboSil films, as measured by LC–MS. Raman spectroscopy and powder X-ray diffraction characterization of SNAP-doped CarboSil films demonstrate that a polymer–crystal composite is formed during the solvent evaporation process when SNAP exceeds its solubility in CarboSil (ca. 3.4–4.0 wt %). Further, when exceeding this solubility threshold, SNAP exists in an orthorhombic crystal form within the bulk of the polymer. The proposed mechanism of sustained NO release in SNAP-doped CarboSil is that the solubilized SNAP in the polymer matrix decomposes and releases NO, primarily in the water-rich regions near the polymer/solution interface, and the dissolved SNAP in the bulk polymeric phase becomes unsaturated, resulting in the dissolution of crystalline SNAP within the bulk of the polymer. This is a very slow process that ultimately leads to NO release at the physiological flux levels for >3 weeks. The increased stability of SNAP within CarboSil is attributed to the intermolecular hydrogen bonds between the SNAP molecules that crystallize. This crystallization also plays a key role in maintaining RSNO stability within the CarboSil polymer for >8 months at 37 °C (88.5% remains). Further, intravascular catheters fabricated with this new material are demonstrated to significantly decrease the formation of Staphylococcus aureus biofilm (a leading cause of nosocomial bloodstream infections) (in vitro) over a 7 day period, with 5 log units reduction of viable cell count on catheter surfaces. It is also shown that the NO release catheters can greatly reduce thrombus formation on the catheter surfaces during 7 h implantation in rabbit veins, when compared to the control catheters fabricated without SNAP. These results suggest that the SNAP-doped CarboSil system is a very attractive new composite material for creating long-term NO release medical devices with increased stability and biocompatibility.

Optimized polymeric film-based nitric oxide delivery inhibits bacterial growth in a mouse burn wound model

Nitric oxide (NO) has many biological roles (e.g. antimicrobial agent, promoter of angiogenesis, prevention of platelet activation) that make NO releasing materials desirable for a variety of biomedical applications. Localized NO release can be achieved from biomedical grade polymers doped with diazeniumdiolated dibutylhexanediamine (DBHD/N2O2) and poly(lactic-co-glycolic acid) (PLGA). In this study, the optimization of this chemistry to create film/patches that can be used to decrease microbial infection at wound sites is examined. Two polyurethanes with different water uptakes (Tecoflex SG-80A (6.2±0.7wt.%) and Tecophilic SP-60D-20 (22.5±1.1wt.%)) were doped with 25wt.% DBHD/N2O2 and 10wt.% of PLGA with various hydrolysis rates. Films prepared with the polymer that has the higher water uptake (SP-60D-20) were found to have higher NO release and for a longer duration than the polyurethane with the lower water uptake (SG-80A). The more hydrophilic polymer enhances the hydrolysis rate of the PLGA additive, thereby providing a more acidic environment that increases the rate of NO release from the NO donor. The optimal NO releasing and control SG-80A patches were then applied to scald burn wounds that were infected with Acinetobacter baumannii. The NO released from these patches applied to the wounds is shown to significantly reduce the A. baumannii infection after 24h (∼4 log reduction). The NO release patches are also able to reduce the level of transforming growth factor-β in comparison to controls, which can enhance re-epithelialization, decrease scarring and reduce migration of bacteria. The combined DBHD/N2O2 and PLGA-doped polymer patches, which could be replaced periodically throughout the wound healing process, demonstrate the potential to reduce risk of bacterial infection and promote the overall wound healing process.

The effect of intermittent pneumatic compression of legs on the levels of nitric oxide related species in blood and on arterial function in the arm

BackgroundIntermittent pneumatic compression (IPC) of legs exerts beneficial local vascular effects, possibly through local release of nitric oxide (NO). However, studies demonstrating systemic transport of nitrogen oxide species and release of NO prompt the question of whether IPC could also exert nonlocal effects. We tested whether IPC (1) affects systemic levels of nitrite, S-nitrosothiols and red blood cell (RBC) NO, and (2) exerts vasoactive effects in the brachial artery (BA), although this hypothesis-generating pilot study did not investigate cause and effect relationship between (1) and (2). MethodsIn 10 healthy subjects, ages 24–39years, we measured plasma nitrite, plasma S-nitrosothiols and RBC-NO from venous blood samples drawn before and after IPC treatment. We also measured BA responses to 5min of upper arm occlusion at rest and during1h of leg IPC. ResultsThere was a significant decrease in plasma nitrite (112±26nM to 90±15nM, p=0.0008) and RBC-NO (129±72nM to 102±41nM, p=0.02). Plasma S-nitrosothiols were unchanged (5.79±4.81nM to 6.27±5.79nM, p=0.3). BA occlusion-mediated constriction (OMC) was significantly attenuated with IPC treatment (−43±13% to −33±12%, p=0.003). High-flow mediated BA dilation was unchanged (13.3±9.4% to 11.5±7.2%, p=0.2). ConclusionPlasma nitrite, RBC-NO, and BA OMC decreased with leg IPC. We hypothesize that this decrease in circulatory pool of plasma nitrite and RBC-NO may result from the transfer of their NO-bioactivity from blood to the hypoxic arm tissue, to be stored and released under hypoxic stress and oppose OMC. Future studies should investigate whether IPC-induced decreases in brachial OMC are caused by the changes in systemic NO activity, and whether leg IPC could benefit distant arterial function in systemic cardiovascular disease.

Effects of local nitric oxide release on human mesenchymal stem cell attachment and proliferation on gelatin hydrogel surface

Nitric oxide plays important roles in cardiovascular homeostasis, immune responses and wound repair. Therefore, polymers that release nitric oxide locally at the surface exhibit improved biocompatibility for biomedical implants through reducing neointimal hyperplasia and thrombosis caused by blood vessel wall damage. The objective of this article was to fabricate a nitric oxide–releasing gelatin hydrogel that can continuously generate nitric oxide at a physiologically relevant level and inhibit cell attachment and proliferation. The nitric oxide donor, S-nitroso-N-acetylpenicillamine (SNAP), was successfully conjugated to the gelatin hydrogel, which showed a rapid nitric oxide release in the first 2 h and then a slower but sustained release in the next 70-h period. Human mesenchymal stem cells (hMSCs), as a model cell line with wide biomedical applications, were used to examine the cell attachment and proliferation of the nitric oxide–releasing gelatin hydrogel. Compared with the control gelatin, the nitric oxide–releasing gelatin hydrogel demonstrated a 0·35 times lower hMSCs attachment at 6 h and a 3·15 times lower hMSCs proliferation after 72-h incubation. Moreover, hMSCs on nitric oxide–releasing gelatin exhibited a rounder cell shape and covered less cellular area than their counterparts on the control gelatin hydrogel. This gelatin hydrogel with local nitric oxide release at physiological level provides a promising therapeutic approach in enhancing the performance of biomedical implants.

Strategies for profiling native S-nitrosylation.

Cysteine is a uniquely reactive amino acid, capable of undergoing both nucleophlilic and oxidative post-translational modifications. One such oxidation reaction involves the covalent modification of cysteine via the gaseous second messenger nitric oxide (NO), termed S-nitrosylation (SNO). This dynamic post-translational modification is involved in the redox regulation of proteins across all phylogenic kingdoms. In mammals, calcium-dependent activation of NO synthase triggers the local release of NO, which activates nearby guanylyl cyclases and cGMP-dependent pathways. In parallel, diffusible NO can locally modify redox active cellular thiols, functionally modulating many redox sensitive enzymes. Aberrant SNO is implicated in the pathology of many diseases, including neurodegeneration, inflammation, and stroke. In this review, we discuss current methods to label sites of SNO for biochemical analysis. The most popular method involves a series of biochemical steps to mask free thiols followed by selective nitrosothiol reduction and capture. Other emerging methods include mechanism-based phosphine probes and mercury enrichment chemistry. By bridging new enrichment approaches with high-resolution mass spectrometry, large-scale analysis of protein nitrosylation has highlighted new pathways of oxidative regulation.