Chitosan sponges loaded with metformin and microalgae as dressing for wound healing: A study in diabetic bio-models.

Among the conditions caused by diabetes, the diabetic foot is a significant public health problem due to its delayed healing process. That makes it essential to design, manufacture, and apply auxiliary dressings during healing. In this work, chitosan sponges were developed and evaluated as wound dressings. Metformin, fucoidan, and exopolysaccharide from Porphyridium purpureum algae were loaded into the sponges and studied as healing promoters. The composite sponges were physicochemically, morphologically, and thermally characterized, allowing us to determine the chemical mechanisms involved in the sponge formation. The mechanical analysis demonstrated that sponge composites have shape memory and good mechanical performance under compression stress, showing a compressive strength above 30 kPa. These results correlated with the materials' porosity, influencing the swelling capacity that reached a maximum of 70 %. The morphology of materials was observed by SEM, resulting in folded films with surface porosity. The results of the biocompatibility tests confirmed that the materials are not cytotoxic or hemolytic and have good antibacterial activity. In vivo wound healing evaluation showed that metformin-loaded chitosan sponges regenerated skin tissue after 21 days of treatment, highlighting the rate of healing provided when exopolysaccharide was added to promote tissue regeneration, which can be corroborated by histological analysis. These results make chitosan sponge compounds promising dressings for diabetic foot wound treatment.

Use of Trehalose as an Additive to Bacteriophage Vb_Pd_PDCC-1: Long-Term Preservation Analysis and Its Biocontrol Against Vibrio diabolicus Infection.

Phage therapy is a promising alternative to control bacterial diseases and the increasing problem of antibiotic resistance. In this sense, this research evaluates the viability of lyophilized vibrio phage vB_Pd_PDCC-1 using trehalose as a preservative excipient at different concentrations (4, 2, 1, and 0.5% w/v) and its potential for phage therapy application against a pathogenic bacteria Vibrio diabolicus in brine shrimp nauplii (Artemia franciscana). The lyophilized phages were stored at 4 and 23 °C and rehydrated using biological sterile saline solution to test their viability at days 1, 15, and 60 post-lyophilization. The results showed that trehalose is beneficial in maintaining the viability of post-lyophilization phages (without titer losses) at 4 °C and even at room temperature (23 °C). When lyophilized phages with 4% w/v trehalose concentration were stored at 23 °C, they had not titer losses among the trials; viability and titer concentration were maintained up to 60 days at log 7. The use of lyophilized phage PDCC-1 increased brine shrimp survival and reduced Vibrio concentrations. The present study has identified trehalose as a promising lyophilization excipient to effectively preserve lyophilized bacteriophages for biotechnological applications and long-term storage.

Core-shell chitosan/Porphyridium-exopolysaccharide microgels: Synthesis, properties, and biological evaluation.

Advanced materials used in the biomedicine field comprises a diverse group of organic molecules, including polymers, polysaccharides, and proteins. A significant trend in this area is the design of new micro/nano gels whose small size, physical stability, biocompatibility, and bioactivity could lead to new applications. Herein a new synthesis route is described to obtain core-shell microgels based on chitosan and Porphyridium exopolysaccharides (EPS) crosslinked with sodium tripolyphosphate (TPP). First, the synthesis of EPS-chitosan gels through ionic interactions was explored, leading to the formation of unstable gels. Alternatively, the use of TTP as crosslinker agent led to stable core-shell structures. The influence of reaction temperature, sonication time, and exopolysaccharide concentration, pH and TPP concentration were determined as a function of particle size and polydispersity index (PDI). The obtained EPS-chitosan gels were characterized by TEM, TGA, and FTIR; followed by the assessment of protein load capacity, stability upon freezing, cytotoxicity, and mucoadhesivity. Experimentation revealed that the core-shell particles size ranges 100-300 nm, have a 52 % loading capacity for BSA and a < 90 % mucoadhesivity, and no toxic effects in mammalian cell cultures. The potential application of the obtained microgels in the biomedical field is discussed.

Two Methods of AuNPs Synthesis Induce Differential Vascular Effects. The Role of the Endothelial Glycocalyx.

AuNPs are synthesized through several methods to tune their physicochemical properties. Although AuNPs are considered biocompatible, a change in morphology or properties can modify their biological impact. In this work, AuNPs (~12 to 16 nm) capping with either sodium citrate (CA) or gallic acid (GA) were evaluated in a rat aorta ex vivo model, which endothelial inner layer surface is formed by glycocalyx (hyaluronic acid, HA, as the main component), promoting vascular processes, most of them dependent on nitric oxide (NO) production. Results showed that contractile effects were more evident with AuNPsCA, while dilator effects predominated with AuNPsGA. Furthermore, treatments with AuNPsCA and AuNPsGA in the presence or absence of glycocalyx changed the NO levels, differently. This work contributes to understanding the biological effects of AuNPs with different capping agents, as well as the key role that of HA in the vascular effects induced by AuNPs in potential biomedical applications.

In-Vitro silanization of dental enamel to prevent demineralization / Silanización in vitro sobre esmalte dental para prevenir desmineralización

Abstract Caries is a multifactorial disease that can negatively affect dental tissues through the demineralization process, which produces acids deriving from the metabolism of carbohydrates. Some strategies to prevent this process have been proposed, such as topical fluoride application, resin-based restorations, pit and fissures sealers, infiltrated resins, vaccines, mouthwashes, and several brushing techniques. Objective. To evaluate in vitro enamel hydrophobic modification as a method of prevention against demineralization. A descriptive and comparative study was carried out. Thirty premolars extracted for orthodontic reasons were obtained, encapsulated in epoxy resin, sectioned, and sanded to obtain specimens 3mm in thickness. The samples were pretreated with NaOCl and EDTA, incubated with 1 and 4% octadeyltrichlorosilane (OTS) or with 3 and 6% octadecyltriethoxysilane (TEOS) for 5min and for 8h. Subsequently, the samples were immersed in citric acid for 2 months. The samples were analyzed by their contact angle, infrared spectroscopy, scanning electron microscopy, atomic and confocal force, before and after treatment in citric acid. The samples coated with 1 and 4% OTS for 5min and 8h kept the silanizing agent on their surface after 2 months in citric acid. The treatment with TEOS was only effective at 6% with a reaction time of 5min. The modification with 1 and 4% OTS protects the surface of the tooth enamel from demineralization in acidic medium. The results indicate that treatment with 4% OTS is effective from 5min, which makes it appropriate in clinical practice.

Resumen Introducción. Caries es una enfermedad multifactorial que destruye en tejido dental por la desmineralización de ácidos generados en el metabolismo de carbohidratos. Algunos métodos preventivos, como fluoruro, resinas, selladores de fosetas y fisuras, resinas infiltradas, vacunas, enjuagues bucales, y un sinfín de técnicas de cepillado, han sido empleadas. Objetivo. Evaluar in vitro la modificación hidrofoba del esmalte como método preventivo en contra de la desmineralización. Materiales y Métodos. Un estudio decriptivo y comparativo fue empleado. Se obtuvieron treinta premolares sanos extraidos por razones ortodónticas y encapsulados en resina epóxica, seccionados y pulidos hasta obtener especímenes de 3mm de grosor. Las muestras fueron pretratadas con NaOCl y EDTA, incubadas en octadeciltriclosorilano (OTS) al 1 y 4% y octadeciltrietoxisilano (TEOS) 3 y 6% por 5min y 8h. Después, las muestras fueron sumergidas en ácido cítrico por 2 meses. Las muestras fueron analizadas con ángulo de contacto, espectroscopía infrarroja, microscopía electrónica de barrido, atómica y confocal, antes y después de tratamiento con ácido cítrico. Resultados. Las muestras cubiertas con OTS 1 y 4% por 5min y 8h mantuvieron el agente silanizante sobre la superficie después de 2 meses en ácido cítrico. El tratamiento con TEOS fue efectivo al 6% y con un tiempo de reacción de 5min. Conclusiones. La modificación con 1 y 4% de OTS proteje la superficie del esmalte dental contra la desmineralización en un medio ácido. Estos resultados indican que el tratamiento con OTS 4% es efectivo desde 5min de aplicación, lo cual es apropiado en la práctica clínica.

Synthesis and Characterization of Innovative Microgels Based on Polyacrylic Acid and Microalgae Cell Wall and Their Potential as Antigen Delivery Vehicles.

In this study, hybrid polyacrylic acid and Schizochytrium sp. microalgae (PAA/Schizo) microgels were synthesized by inverse emulsion assisted by ultrasound using the cell wall fraction as crosslinker. Physicochemical characterization of PAA/Schizo microgels revealed polymeric spherical particles (288 ± 39 nm) and were deemed stable and negatively charged. The produced microgels are not inherently toxic as cell viability was sustained above 80% when mice splenocytes were exposed to concentrations ranging 10-900 µg/mL. PAA/Schizo microgels were evaluated as antigen delivery nanovehicle by adsorbing bovine serum albumin (BSA); with a loading efficiency of 72% and loading capacity of 362 µg/mg. Overall, intranasally-immunized BALB/c mice showed null IgG or IgA responses against PAA/Schizo microgel-BSA, whereas soluble BSA induced significant humoral responses in systemic and mucosal compartments. Splenocytes proliferation assay upon BSA stimulus revealed positive CD4+ T cells-proliferation response in PAA/Schizo microgels-BSA group. Thus, PAA/Schizo microgels constitute functional antigen delivery vehicles of simple and ecofriendly synthesis. Moreover, the use of cell wall fraction as cross-linker agent provides an alternative use for the generation of high-value products using residual algae biomass from the oil industry. Our data suggests that the PAA/Schizo microgels are potential antigen delivery vehicles for immunotherapy development.

Synthesis and immunogenicity assessment of a gold nanoparticle conjugate for the delivery of a peptide from SARS-CoV-2.

The development of vaccines is a crucial response against the COVID-19 pandemic and innovative nanovaccines could increase the potential to address this remarkable challenge. In the present study a B cell epitope (S461-493) from the spike protein of SARS-CoV-2 was selected and its immunogenicity validated in sheep. This synthetic peptide was coupled to gold nanoparticles (AuNP) functionalized with SH-PEG-NH2 via glutaraldehyde-mediated coupling to obtain the AuNP-S461-493 candidate, which showed in s.c.-immunized mice a superior immunogenicity (IgG responses) when compared to soluble S461-493; and led to increased expression of relevant cytokines in splenocyte cultures. Interestingly, the response triggered by AuNP-S461-493 was similar in magnitude to that induced using a conventional strong adjuvant (Freund's adjuvant). This study provides a platform for the development of AuNP-based nanovaccines targeting specific SARS-CoV-2 epitopes.

Optimized microwave-assisted functionalization and quantification of superficial amino groups on porous silicon nanostructured microparticles.

This work presents an optimized microwave (MW)-assisted method for the chemical functionalization of porous silicon particles (PSip). 3-(Aminopropyl)triethoxysilane (APTES) was grafted on previously stabilized PSip. The functionalization efficiency was studied and optimized in terms of reaction time (Rt) and reaction temperature (RT) using a central composite design (CCD). The effect of MW irradiation on the surface coverage was found to strongly depend on the PSip surface chemistry, Rt, RT, and percentage of APTES. Quantification of grafted amino groups was performed by the ninhydrin method (NHIM); confirming the results by thermogravimetric analysis (TGA). Reacting with 5% APTES solution at 95 °C for 26 min was the best functionalization conditions. The efficiency of PSip-APTES prepared under the optimized conditions was compared to those functionalized by the traditional method; MW irradiation increases by 39% the number of functional groups grafted onto the PSip surfaces with the additional benefit of having a drastic reduction in Rt.

Synthesis of Bamboo-like Multiwall Carbon Nanotube-Poly(Acrylic Acid-co-Itaconic Acid)/NaOH Composite Hydrogel and its Potential Application for Electrochemical Detection of Cadmium(II).

A poly(acrylic acid-co-itaconic acid) (PAA-co-IA)/NaOH hydrogel containing bamboo-type multiwall carbon nanotubes (B-MWCNTs) doped with nitrogen (PAA-co-IA/NaOH/B-MWCNTs) was synthesized and characterized by SEM, absorption of water, point of zero charges, infrared spectroscopy, thermogravimetric analysis, and differential scanning calorimetry. The possible use of the PAA-co-IA/NaOH/B-MWCNT hydrogel as an electrode modifier and pre-concentrator agent for Cd(II) sensing purposes was then evaluated using carbon paste electrodes via differential pulse voltammetry. The presence of the B-MWCNTs in the hydrogel matrix decreased its degree of swelling, stabilized the structure of the swollen gel, and favored the detection of 3 ppb Cd(II), which is comparable to the World Health Organization's allowable maximum value in drinking water. A calibration curve was obtained in the concentration range of 2.67 × 10-8 to 6.23 × 10-7 M (i.e., 3 and 70 ppb) to determine a limit of detection (LOD) of 19.24 µgL-1 and a sensitivity of 0.15 µC ppb-1. Also, the Zn(II), Hg(II), Pb(II) and Cu(II) ions interfered moderately on the determination of Cd(II).

Can nanotechnology help in the fight against COVID-19?

INTRODUCTION: The current COVID-19 pandemic caused by the SARS-CoV-2 virus demands the development of strategies not only to detect or inactivate the virus, but to treat it (therapeutically and prophylactically). COVID-19 is not only a critical threat for the population with risk factors, but also generates a dramatic economic impact in terms of morbidity and the overall interruption of economic activities. AREAS COVERED: Advanced materials are the basis of several technologies that could diminish the impact of COVID-19: biosensors might allow early virus detection, nanosized vaccines are powerful agents that could prevent viral infections, and nanosystems with antiviral activity could bind the virus for inactivation or destruction upon application of an external stimulus. Herein all these methods are discussed under the light of cutting-edge technologies and the previously reported prototypes targeting enveloped viruses similar to SARS-CoV-2. This analysis was derived from an extensive scientific literature search (including pubmed) performed on April 2020. EXPERT OPINION: Perspectives on how biosensors, vaccines, and antiviral nanosystems can be implemented to fight COVID-19 are envisioned; identifying the approaches that can be implemented in the short term and those that deserve long term research to cope with respiratory viruses-related pandemics in the future.

An overview of nanogel-based vaccines.

Introduction: The development of more efficacious vaccines, especially subunit vaccines administered via non-invasive routes, is a priority in vaccinology. Nanogels are materials that can meet the requirements to serve as efficient vaccine delivery vehicles (in terms of thermo-sensitivity, biocompatibility, and pH-responsiveness; among others); thus there is a growing interest in exploring the potential of nanogels for vaccine development. Areas covered: Herein, a critical analysis of nanogel synthesis methodologies is presented and nanogel-based vaccines under development are summarized and placed in perspective. Promising vaccine candidates based on nanogels have been reported for cancer, obesity, and infectious diseases (mainly respiratory diseases). Some of the candidates were administered by mucosal routes which are highly attractive in terms of simple administration and induction of protective responses at both mucosal and systemic levels. Expert opinion: The most advanced models of nanogel-based vaccines comprise candidates against cancer, based on cholesteryl pullulan nanogels evaluated in clinical trials with promising findings; as well as some vaccines against respiratory pathogens tested in mice thus far. Nonetheless, the challenge for this field is advancing in clinical trials and proving the protective potential in test animals for many other candidates. Implementing green synthesis approaches for nanogels is also required.

Biosynthesis of ß-d-glucan­gold nanoparticles, cytotoxicity and oxidative stress in mouse splenocytes.

This study reports biosynthesis of gold-nanoparticles (AuNPs) by using ß-d-glucans isolated from the yeast Yarrowia lypolitica D1. ß-d-glucans serve as reducing and stabilizing mediators that induce the formation of AuNPs on the outer surface of the own ß-d-glucan. The systems were physicochemically characterized by ultraviolet visible (UV-Vis) spectroscopy, high-resolution transmission electron microscopy (HR-TEM), scanning electron microscope (SEM), energy-dispersive X-ray spectroscopy (EDS), and dynamic light scattering (DLS) analyses. The results revealed the generation of AuNPs with quasi-spherical shape or large one dimension (1D) gold-nanostructures (AuNSs) depending on the HAuCl4 concentration. A cytotoxic study was assessed in mouse splenocytes. Contrary to that expected, important cytotoxicity was found in all ß-d-gluc+AuNPs systems by an oxidative stress increase. This study discusses the cytotoxic mechanism, suggesting that the resulting ß-d-gluc+AuNPs systems may not be candidates for the formulation of immunostimulants or nanocarriers for biomedical applications.

Mesoporous Silicon Particles Favor the Induction of Long-Lived Humoral Responses in Mice to a Peptide-Based Vaccine.

Vaccinology faces the challenge of developing improved immunization approaches that are able to induce long-term immunity with the desired Th profile according to the pathology. In this context, new vehicles for efficient antigen delivery that exert adjuvant effects play a critical role in addressing this goal. Herein, mesoporous silicon particles (PSiP) were assessed as carriers for a peptide-based vaccine targeting the receptor for advanced glycation end products (RAGE), which is a relevant receptor in Alzheimer´s disease and other diseases. A RAGE peptide was adsorbed onto PSiP (PSiP vaccine) and administered to BALB/c mice, leading to immune responses that were similar in magnitude to those induced by the soluble peptide. However, the response induced by PSiP lasted for a significantly longer period when compared with the behavior of the group immunized with the peptide alone. Therefore, PSiP are proposed as carriers to enhance immune memory, which is critical in vaccination. This study opens interesting perspectives related to the application of PSiP in vaccinology.

Toxicity evaluation of high-fluorescent rare-earth metal nanoparticles for bioimaging applications.

Research on nanometer-sized luminescent semiconductors and their biological applications in detectors and contrasting agents is an emergent field in nanotechnology. When new nanosize technologies are developed for human health applications, their interaction with biological systems should be studied in depth. Rare-earth elements are used in medical and industrial applications, but their toxic effects are not known. In this work, the biological interaction between terbium-doped gadolinium oxysulfide nanoparticles (GOSNPs) with human peripheral blood mononuclear cells (PBMC), human-derived macrophages (THP-1), and human cervical carcinoma cell (HeLa) were evaluated. The GOSNPs were synthetized using a hydrothermal method to obtain monodisperse nanoparticles with an average size of 91 ± 9 nm. Characterization techniques showed the hexagonal phase of the Gd2 O2 S:Tb3+ free of impurities, and a strong green emission at λemi = 544 nm produced by Tb3+ was observed. Toxic effects of GOSNPs were evaluated using cell viability, apoptosis, cell-cycle progression, and immunological response techniques. In addition, an Artemia model was used to assess the toxicity in vivo. Results indicated cell apoptosis in both types of cells with less sensitivity for PBMC cells compared to HeLa cells. In addition, no toxic effects were observed in the in vivo model of Artemia. Moreover, GOSNPs significantly reduced the activation and cell-cycle progression of PBMC and HeLa cells, respectively. Interestingly, an increase in proinflammatory cytokines was not observed. Our data suggest that fluorescence applications of GOSNPs for biolabeling are not toxic in primary immune cells and they may have an immunomodulatory effect. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 605-615, 2017.

An overview on the role of silica-based materials in vaccine development.

INTRODUCTION: Although vaccination has prevented millions of deaths, the development of highly immunogenic subunit vaccines is still required. Since the number of adjuvants approved for human use is limited, the new paths for the development of delivery vehicles offered by nanotechnology are of key relevance. Areas covered: Herein, the potential of silica nanoparticles (SP) as both adjuvants and vaccine delivery vehicles is discussed based on the analysis of the current biomedical literature. Expert commentary: SP are reported not only as biodegradable and biocompatible material but also as easy to modify and with a low production cost. Additionally, several reports suggest that SP enhance the immune response. Therefore, SP are a promising delivery vehicle and/or adjuvant in vaccines. However, knowledge on the industrial production and specific aspects of immunity are still required.

A turn-on fluorescent solid-sensor for Hg(II) detection.

A rhodamine organosilane derivative (Rh-UTES) has been obtained by one-pot synthesis. The chemical structure of Rh-UTES was confirmed by nuclear magnetic resonance (NMR) and infrared (FTIR) techniques. To obtain an inorganic-organic hybrid sensor, Rh-UTES was covalently immobilized on a porous silicon microcavity (PSiMc) via triethoxysilane groups. The attachment of the organic derivative into PSiMc was confirmed by FTIR, specular reflectance, and scanning electron microscopy (SEM). The optical performance of Rh-UTES receptor for Hg(2+) detection was investigated by fluorescent spectroscopy and microscopy. Upon the addition of increasing amounts of Hg(2+) ions, a remarkable enhancement in emission intensity was produced in both systems. In the solid phase, an increase of integrated fluorescent emission of 0.12- and 0.15-fold after Hg(2+) receptor coordination was observed. The light harvesting capability of PSiMc devices allowed obtaining an enhanced fluorescent emission after Rh-UTES immobilization (277-fold). The fluorescence microscopy of hybrid PSiMc sensor provided an optical qualitative test for Hg(2+) detection.

Tunable resonance transmission modes in hybrid heterostructures based on porous silicon.

: In this work, we report the experimental results and theoretical analysis of strong localization of resonance transmission modes generated by hybrid periodic/quasiperiodic heterostructures (HHs) based on porous silicon. The HHs are formed by stacking a quasiperiodic Fibonacci (FN) substructure between two distributed Bragg reflectors (DBRs). FN substructure defines the number of strong localized modes that can be tunable at any given wavelength and be unfolded when a partial periodicity condition is imposed. These structures show interesting properties for biomaterials research, biosensor applications and basic studies of adsorption of organic molecules. We also demonstrate the sensitivity of HHs to material infiltration.

Light-harvesting bio-nanomaterial using porous silicon and photosynthetic reaction center.

Porous silicon microcavity (PSiMc) structures were used to immobilize the photosynthetic reaction center (RC) purified from the purple bacterium Rhodobacter sphaeroides R-26. Two different binding methods were compared by specular reflectance measurements. Structural characterization of PSiMc was performed by scanning electron microscopy and atomic force microscopy. The activity of the immobilized RC was checked by measuring the visible absorption spectra of the externally added electron donor, mammalian cytochrome c. PSi/RC complex was found to oxidize the cytochrome c after every saturating Xe flash, indicating the accessibility of specific surface binding sites on the immobilized RC, for the external electron donor. This new type of bio-nanomaterial is considered as an excellent model for new generation applications of silicon-based electronics and biological redox systems.

Porous silicon/photosynthetic reaction center hybrid nanostructure.

The purified photosynthetic reaction center protein (RC) from Rhodobacter sphaeroides R-26 purple bacteria was bound to porous silicon microcavities (PSiMc) either through silane-glutaraldehyde (GTA) chemistry or via a noncovalent peptide cross-linker. The characteristic resonance mode in the microcavity reflectivity spectrum red shifted by several nanometers upon RC binding, indicating the protein infiltration into the porous silicon (PSi) photonic structure. Flash photolysis experiments confirmed the photochemical activity of RC after its binding to the solid substrate. The kinetic components of the intraprotein charge recombination were considerably faster (τ(fast) = 14 (±9) ms, τ(slow) = 230 (±28) ms with the RC bound through the GTA cross-linker and only τ(fast) = 27 (±3) ms through peptide coating) than in solution (τ(fast) = 120 (±3) ms, τ(slow) = 1387 (±2) ms), indicating the effect of the PSi surface on the light-induced electron transfer in the protein. The PSi/RC complex was found to oxidize the externally added electron donor, mammalian cytochrome c, and the cytochrome oxidation was blocked by the competitive RC inhibitor, terbutryne. This fact indicates that the specific surface binding sites on the PSi-bound RC are still accessible to external cofactors and an electronic interaction with redox components in the aqueous environment is possible. This new type of biophotonic material is considered to be an excellent model for new generation applications at the interface of silicon-based electronics and biological redox systems designed by nature.

Effect of 45 nm silver nanoparticles (AgNPs) upon the smooth muscle of rat trachea: role of nitric oxide.

AgNPs have been used to manufacture nanomaterials with new biophysical properties and functions. However, few experimental approaches have been used to assess their potential toxic or beneficial effects on human health, in association with the size, concentration, and biological target. The aim of this work was to evaluate the effects of the AgNPs on the smooth muscle of rat trachea. A single administration of AgNPs did not modify the smooth muscle tone, but, when the trachea rings were pre-treated with acetylcholine (ACh), AgNPs produced a contractile effect. Simultaneous administration of AgNPs and ACh resulted in a slight increase of smooth muscle contractility induced by ACh. AgNPs pretreatment followed by ACh administration showed that AgNPs exerted an important contraction effect induced by ACh after which muscle tone did not return to the basal level. This effect was associated with an increase in the production of nitric oxide (NO). The contractile response of the AgNPs induced by ACh was completely blocked when the rings were incubated, after the ACh but before the AgNPs administration, with 1400 W (NO blocker). The contractile effect was also abolished by atropine, which suggests that AgNPs alter ACh muscarinic receptor signaling. These data also show that AgNPs modify the contractile action of ACh through NO production and possibly induce hyper-reactivity of tracheal smooth muscle.