Biocide effect against SARS-CoV-2 and ESKAPE pathogens of a noncytotoxic silver-copper nanofilm.

Nanometric materials with biocidal properties effective against severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) and pathogenic bacteria could be used to modify surfaces, reducing the risk of touching transmission. In this work, we showed that a nanometric layer of bimetallic AgCu can be effectively deposited on polypropylene (PP) fibers. The virucidal properties of the AgCu nanofilm were evaluated by comparing the viral loads remaining on uncoated and coated PP after contact times between 2 and 24 h. Quantification of virion numbers for different initial concentrations indicated a reduction of more than 95% after 2 h of contact. The bactericidal action of the AgCu nanofilm was also confirmed by inoculating uncoated and coated PP with a pool of pathogenic bacteria associated with pneumonia (ESKAPE). Meanwhile, no cytotoxicity was observed for human fibroblasts and keratinocyte cells, indicating that the nanofilm could be in contact with human skin without threat. The deposition of the AgCu nanofilm on the nonwoven component of reusable cloth masks might help to prevent virus and bacterial infection while reducing the pollution burden related to the disposable masks. The possible mechanism of biocide contact action was studied by quantum chemistry calculations that show that the addition of Ag and/or Cu makes the polymeric fiber a better electron acceptor. This can promote the oxidation of the phospholipids present at both the virus and bacterial membranes. The rupture at the membrane exposes and damages the genetic material of the virus. More studies are needed to determine the mechanism of action, but the results reported here indicate that Cu and Ag ions are good allies, which can help protect us from the virus that has caused this disturbing pandemic.

Fabrication and in vitro behavior of dual-function chitosan/silver nanocomposites for potential wound dressing applications.

We report the synthesis and in vitro evaluation of dual-function chitosan-silver nanoparticles (CTS-AgNPs) films with potential applications as wound dressings. We attempted to formulate nanocomposite films with appropriate AgNPs concentrations to simultaneously display antibacterial activity and suitability for cell culture. Nanocomposites were obtained by CTS-mediated in situ chemical reduction of AgNO3. Circular-shape AgNPs (sizes ca. 7-50 nm) well distributed within the CTS matrices were obtained in concentrations from 0.018 to 0.573 wt%. Efficacy (bacteriostatic and bactericidal properties) of CTS-AgNPs films to decrease planktonic and biofilm bacterial growth was AgNPs concentration- and bacteria strain-dependent. Films showed significant antibacterial activity against Gram-negative E. coli and P. aeruginosa and Gram-positive S. aureus. Antibacterial activity against S. epidermidis was moderated. Films suitability for cell culture was characterized using primary human fibroblasts (HF). HF displayed cell viability higher than 90% and the characteristic fusiform morphology of adhered fibroblast upon culture on films with AgNPs concentration ≤ 0.036 wt%. HF cultured on these films also showed positive expression of tropoelastin, procollagen type I and Ki-67, characteristic proteins of extracellular matrix and proliferative cells, respectively. In vitro assays demonstrated that cytocompatibility/antibacterial properties decreased/increased as silver concentration increased, suggesting that CTS-AgNPS nanocomposite films with ≈0.04-0.20 wt% might be considered as potential temporary dual-function wound dressings.

Bismuth subsalicylate nanoparticles with anaerobic antibacterial activity for dental applications.

In recent years, nanomaterials have been used in the medical-dental field as new alternative antimicrobial agents. Bismuth subsalicylate (BSS) has been used as an antimicrobial agent, but the effect of BSS in the form of nanoparticles (BSS-nano) as a potential antimicrobial agent has not been tested, in specific against bacteria responsible for periodontal disease. The aim of this study was to evaluate the antibacterial effect of BSS-nano against oral anaerobic bacteria and to assess the safety of BSS-nano by evaluating their cytotoxicity in human gingival fibroblast (HGF-1) cells. BSS-nano were synthesized by laser ablation and were previously physico-chemically characterized using in vitro assays. The antibacterial activity was measured using the tetrazolium-based XTT assay, and cytotoxicity was determined using lactate dehydrogenase (LDH) and MTS assays in HGF-1 cells. Transmission electron microscopy of HGF-1 exposed to BSS-nano was also performed. BSS-nano was shown to have a primary size of 4-22 nm and a polygonal shape. Among the tested bacterial strains, those with a greater sensitivity to BSS-nano (highest concentration of 21.7 µg ml-1) were A. actinomycetemcomitans, C. gingivalis, and P. gingivalis. BSS-nano at a concentration of 60 µg ml-1 showed low cytotoxicity (6%) in HFG-1 cells and was mainly localized intracellularly in acidic vesicles. Our results indicate that the concentration of BSS-nano used as an effective antibacterial agent does not induce cytotoxicity in mammalian cells; thus, BSS-nano can be applied as an antibacterial agent in dental materials or antiseptic solutions.

The effect of simulated inflammatory conditions on the surface properties of titanium and stainless steel and their importance as biomaterials.

This work compares the surface modifications induced by the immersion in solutions that simulate inflammatory conditions of pure titanium (cpTi) and medical grade stainless steel (SS). The inflammatory conditions were simulated using a mixture of Hartman solution and 50mM of hydrogen peroxide (H2O2) at pH=5.2. The samples were immersed by 7days refreshing the solution every day to keep the reactivity of the H2O2. The surface characteristics that were investigated were: elemental composition by X-ray photoelectron spectroscopy (XPS); topography by atomic force microscopy (AFM) and profilometry; wettability and surface energy by sessile drop contact angle and point of zero charge by titration. Moreover, the variations in the electrochemical response were evaluated by open circuit potential (OCP), electrochemical impedance spectroscopy (EIS) and potentiodynamic polarization (PP) performed before and after the treatment using the Hartman solution as the electrolyte. The XPS results indicated that for both metallic samples, oxidation of the surface was promoted and/or the oxide layer was thicker after the immersion. The roughness and the solid-liquid surface energy were increased; the samples showed a more hydrophilic character after the treatment. However, the surface energy of the solid estimated using the Van Oss-Chaudhury-Good approach showed different trends between the cpTi and the SS surfaces; the polar component decreased for cpTi, while it increased for SS. Finally, the electrochemical results indicated that the corrosion resistance (Rcor) and the pore resistance (Rpo) significantly decreased for cpTi, while both resistances were not significantly different for the SS. This is indicative of a higher dissolution of the cpTi compared to SS and the lower Rpo means that the species are easily transported through the surface layer, which can be explained in terms of the formation of a porous TiOx layer, not observed on the SS. The cpTi surface suffered from a dissolution/oxidation process that allows its integration with the surrounding media, while the SS remained completely passive and this different response might be related to their distinguished clinical outcome.

Role of the N-terminal peptide of amelogenin on osteoblastic differentiation of human mesenchymal stem cells.

Porcine enamel matrix derivative (pEMD), a complex mixture of proteins and peptides including full-length amelogenin protein, splice variants, and proteolytic peptides, is used clinically with a carrier to regenerate supportive tissue around teeth. During application, pEMD self-assembles as nanospheres and precipitates as a three-dimensional matrix to facilitate cell migration and differentiation. Amelogenin, the primary constituent of pEMD, stimulates osteoblast differentiation, but it is unclear what specific roles other components of pEMD play in determining biological response. This study examined the potential of one constituent of pEMD, the N-terminal amelogenin peptide (NTAP), to promote osteoblastic differentiation of human mesenchymal stem cells (MSCs) and to elucidate possible signaling pathways involved. Effects of porcine NTAP on MSC cultures were compared to those of recombinant human amelogenin. While amelogenin induced MSC osteoblastic differentiation, a more robust osteoblastic response was seen after NTAP treatment. A phospho-kinase proteasome array measuring phosphorylation of 35 proteins indicated that protein kinase C (PKC), extracellular signal-regulated kinase 1/2 (ERK1/2), and ß-catenin were highly phosphorylated by NTAP. This was confirmed by measuring PKC activity and levels of phospho-ERK1/2 and ß-catenin. Both amelogenin and NTAP increased PKC, but NTAP induced higher phosho-ERK1/2 and phospho-ß-catenin than amelogenin. ERK1/2 inhibition blocked both amelogenin- and NTAP-induced increases in RUNX2, ALP, OCN, COL1, and BMP2. The results demonstrate that NTAP induces osteogenic differentiation of MSCs via PKC and ERK1/2 activation and ß-catenin degradation. NTAP may be an active bone regeneration component of amelogenin, and may play this role in pEMD-stimulated periodontal regeneration.

Influence of topography and hydrophilicity on initial oral biofilm formation on microstructured titanium surfaces in vitro.

OBJECTIVES: The aim of this study was to analyse the influence of the microtopography and hydrophilicity of titanium (Ti) substrates on initial oral biofilm formation. MATERIALS AND METHODS: Nine bacterial species belonging to the normal oral microbiota, including: Aggregatibacter actinomycetemcomitans, Actinomyces israelii, Campylobacter rectus, Eikenella corrodens, Fusobacterium nucleatum, Parvimonas micra, Porphyromonas gingivalis, Prevotella intermedia, and Streptococcus sanguinis were tested on Ti surfaces: pretreatment (PT [R(a) <0.2 µm]), acid-etched (A [R(a) <0.8 µm]), A modified to be hydrophilic (modA), sand-blasted/acid-etched (SLA [R(a) =4 µm]), and hydrophilic SLA (modSLA). Disks were incubated for 24 h in anaerobic conditions using a normal culture medium (CM) or human saliva (HS). The total counts of bacteria and the proportion of each bacterial species were analysed by checkerboard DNA-DNA hybridization. RESULTS: Higher counts of bacteria were observed on all surfaces incubated with CM compared with the samples incubated with HS. PT, SLA, and modSLA exhibited higher numbers of attached bacteria in CM, whereas SLA and modSLA had a significant increase in bacterial adhesion in HS. The proportion of the species in the initial biofilms was also influenced by the surface properties and the media used: SLA and modSLA increased the proportion of species like A. actinomycetemcomitans and S. sanguinis in both media, while the adhesion of A. israelii and P. gingivalis on the same surfaces was affected in the presence of saliva. CONCLUSIONS: The initial biofilm formation and composition were affected by the microtopography and hydrophilicity of the surface and by the media used.

Oral bacterial adhesion on amorphous carbon and titanium films: effect of surface roughness and culture media.

Implant infections can cause severe problems from malfunctioning to dangerous sepsis affecting the health of the patient. For many years, titanium has been the most common material used on dental implants due to their mechanical and biocompatibility properties. Recent studies suggest that amorphous carbon (a-C) films can be possible candidates for coating dental implants, improving some important features like biocompatibility and bone formation. In the oral cavity, the risk of an implant infection is high due to multiple species are capable to colonize this site and these biofilm infections can limit the use of these medical devices. The purpose of this study was to evaluate the influence of the surface chemistry, roughness, and culture media in the bacterial colonization process. To achieve this, a-C and Ti films were deposited on rough and smooth surfaces and cultured with different microorganisms belonging to the oral microbiota with mycoplasma medium (MM) or human saliva (HS). Samples were incubated for 24 h, after this, samples were sonicated and the number of attached bacteria was determined by counting the colony-forming units (CFU's) from each sample. The proportion of the species in the biofilms was determined using checkerboard DNA-DNA hybridization. Data were analyzed by Student's t test using Bonferroni's modification of Student's t test and differences on the proportion of the bacterial species attached to each surface were determined using the Mann-Whitney test. Results show an increased number of CFU's on rough surfaces, especially on the a-C surfaces. The incubation media were an important factor on the adhesion of certain taxa, whereas other species were more sensitive to surface chemistry and others to surface roughness.