ABSTRACT
INTRODUCTION: Tetracyclines (TCs) embrace a class of broad-spectrum antibiotics with unrelated effects at sub-antimicrobial levels, including an effective anti-inflammatory activity and stimulation of osteogenesis, allowing their repurposing for different clinical applications. Recently, sarecycline (SA)-a new-generation molecule with a narrower antimicrobial spectrum-was clinically approved due to its anti-inflammatory profile and reduced adverse effects verified with prolonged use. Notwithstanding, little is known about its osteogenic potential, previously verified for early generation TCs. MATERIALS AND METHODS: Accordingly, the present study is focused on the assessment of the response of human bone marrow-derived mesenchymal stromal cells (hBMSCs) to a concentration range of SA, addressing the metabolic activity, morphology and osteoblastic differentiation capability, further detailing the modulation of Wnt, Hedgehog, and Notch signaling pathways. In addition, an ex vivo organotypic bone development system was established in the presence of SA and characterized by microtomographic and histochemical analysis. RESULTS: hBMSCs cultured with SA presented a significantly increased metabolic activity compared to control, with an indistinguishable cell morphology. Moreover, RUNX2 expression was upregulated 2.5-fold, and ALP expression was increased around sevenfold in the presence of SA. Further, GLI2 expression was significantly upregulated, while HEY1 and HNF1A were downregulated, substantiating Hedgehog and Notch signaling pathways' modulation. The ex vivo model developed in the presence of SA presented a significantly enhanced collagen deposition, extended migration areas of osteogenesis, and an increased bone mineral content, substantiating an increased osteogenic development. CONCLUSION: Summarizing, SA is a promising candidate for drug repurposing within therapies envisaging the enhancement of bone healing/regeneration.
Subject(s)
Drug Repositioning , Hedgehogs , Humans , Animals , Osteogenesis , Cell Differentiation , Tetracyclines/pharmacology , Cells, Cultured , Bone Marrow CellsABSTRACT
Titanium (Ti) and its alloys are the most widely used metallic biomaterials in total joint replacement; however, increasing evidence supports the degradation of its surface due to corrosion and wear processes releasing debris (ions, and micro and nanoparticles) and contribute to particle-induced osteolysis and implant loosening. Cell-to-cell communication involving several cell types is one of the major biological processes occurring during bone healing and regeneration at the implant-bone interface. In addition to the internal response of cells to the uptake and intracellular localization of wear debris, a red flag is the ability of titanium dioxide nanoparticles (mimicking wear debris) to alter cellular communication with the tissue background, disturbing the balance between osseous tissue integrity and bone regenerative processes. This study aims to understand whether titanium dioxide nanoparticles (TiO2 NPs) alter osteoblast-derived exosome (Exo) biogenesis and whether exosomal protein cargos affect the communication of osteoblasts with human mesenchymal stem/stromal cells (HMSCs). Osteoblasts are derived from mesenchymal stem cells coexisting in the bone microenvironment during development and remodelling. We observed that TiO2 NPs stimulate immature osteoblast- and mature osteoblast-derived Exo secretion that present a distinct proteomic cargo. Functional tests confirmed that Exos derived from both osteoblasts decrease the osteogenic differentiation of HMSCs. These findings are clinically relevant since wear debris alter extracellular communication in the bone periprosthetic niche, contributing to particle-induced osteolysis and consequent prosthetic joint failure.
Subject(s)
Exosomes , Mesenchymal Stem Cells , Nanoparticles , Osteolysis , Humans , Osteogenesis , Titanium/adverse effects , Osteolysis/chemically induced , Exosomes/metabolism , Proteomics , Osteoblasts , Cell Differentiation , Immunologic Factors , Cell CommunicationABSTRACT
Development of multifunctional 3D patches with appropriate antibacterial and biocompatible properties is needed to deal with wound care regeneration. Combining gelatin-based hydrogel with a well-known natural antibacterial honey (Manuka honey, MH) in a 3D patch can provide improved printability and at the same time provide favourable biological effects that may be useful in regenerative wound treatment. In this study, an antibacterial Manuka-Gelatin 3D patches was developed by an extrusion-based printing process, with controlled porosity, high shape fidelity, and structural stability. It was demonstrated the antibacterial activity of Manuka-Gelatin 3D patches against both gram-positive bacteria (S. epidermidis and S. aureus) and gram-negative (E. coli), common in wound infection. The 3D Manuka-Gelatin base patches demonstrated antibacterial activity, and moreover enhanced the proliferation of human dermal fibroblasts and human epidermal keratinocytes, and promotion of angiogenesis. Moreover, the ease of printing achieved by the addition of honey, coupled with the interesting biological response obtained, makes this 3D patch a good candidate for wound healing applications.
Subject(s)
Honey , Staphylococcus aureus , Humans , Gelatin , Microbial Sensitivity Tests , Escherichia coli , Wound Healing , Honey/analysis , Anti-Bacterial Agents/chemistry , Printing, Three-Dimensional , HydrogelsABSTRACT
STATEMENT OF PROBLEM: Microgap and bacterial microleakage at the implant-prosthetic abutment interface are recognized concerns for implant-supported restorations, leading to inflammation of the peri-implant tissues, with deleterious consequences for crestal bone levels. However, little is known regarding the interface established between the implant and the healing abutment or cover screw placed for the osseointegration phase. PURPOSE: The purpose of this in vitro study was to characterize the implant-cover screw and implant-healing abutment interfaces of a platform-switched implant system to determine the microgap and bacterial microleakage of the system and evaluate the biological response and functionality of an interface sealing agent. MATERIAL AND METHODS: The interfacial microgaps of the implant-healing abutment and implant-cover screw interfaces were characterized by scanning electron microscopy (n=10), and bacterial microleakage was evaluated after colonization with Enterococcus faecalis in a 30-day follow-up (n=10). The sealing efficacy and irritation potential of a silicone-based sealer were determined by using the hen's egg test on chorioallantoic membrane assay. The 2-sample t test was performed to compare means between groups, and data presented with the Kaplan-Meier method were compared statistically by using the log-rank test (α=.05). RESULTS: The interfacial microgap was less than 2.5 µm for both systems. Bacterial microleakage was noted in approximately 50% of the specimens, particularly at early time points, at both the healing abutment and cover screw interfaces. The silicone-based sealer prevented bacterial leakage in the experimental setting. CONCLUSIONS: The implant-healing abutment and implant-cover screw interfaces of the tested system, despite the low microgap, allowed for bacterial microleakage after internal colonization. The use of a nonirritating silicone-based sealing agent effectively sealed the system.
Subject(s)
Dental Implants , Osseointegration , Animals , Female , Chickens , Dental Abutments , Dental Implants/microbiology , Bacteria , Dental Implant-Abutment DesignABSTRACT
Silicone-based medical devices composed of polydimethylsiloxane (PDMS) are widely used all over the human body (e.g., urinary stents and catheters, central venous catheters stents) with extreme clinical success. Nevertheless, their abiotic surfaces, being prone to microorganism colonization, are often involved in infection occurrence. Improving PDMS antimicrobial properties by surface functionalization with biosurfactants to prevent related infections has been the goal of different works, but studies that mimic the clinical use of these novel surfaces are missing. This work aims at the biofunctional assessment of PDMS functionalized with rhamnolipids (RLs), using translational tests that more closely mimic the clinical microenvironment. Rhamnolipids were covalently bonded to PDMS, and the obtained surfaces were characterized by contact angle modification assessment, ATR-FTIR analysis and atomic force microscopy imaging. Moreover, a parallel flow chamber was used to assess the Staphylococcus aureus antibiofilm activity of the obtained surfaces under dynamic conditions, and an in vitro characterization with human dermal fibroblast cells in both direct and indirect characterization assays, along with an in vivo subcutaneous implantation assay in the translational rabbit model, was performed. A 1.2 log reduction in S. aureus biofilm was observed after 24 h under flow dynamic conditions. Additionally, functionalized PDMS lessened cell adhesion upon direct contact, while supporting a cytocompatible profile, within an indirect assay. The adequacy of the biological response was further validated upon in vivo subcutaneous tissue implantation. An important step was taken towards biofunctional assessment of RLs-functionalized PDMS, reinforcing their suitability for medical device usage and infection prevention.
ABSTRACT
In clinic there is a demand to solve the drawback of medical devices multispecies related infections. Consequently, different biomaterial surfaces, such as vascular catheters, urgently need improvement regarding their antifouling/antimicrobial properties. In this work, we covalently functionalized medical grade polydimethylsiloxane (PDMS) with antimicrobial rhamnolipids to investigate the biomaterial surface activity towards mono and dual species biofilms. Preparation of surfaces with "piranha" oxidation, followed by APTES bonding and carbodiimide reaction with rhamnolipids effectively bonded these compounds to PDMS surface as confirmed by FTIR-ATR and XPS analysis. Generated surfaces were active towards S. aureus biofilm formation showing a 4.2 log reduction while with S. epidermidis and C. albicans biofilms a reduction of 1.2 and 1.0 log reduction, respectively, was observed. Regarding dual-species testing the higher biofilm log reduction observed was 1.9. Additionally, biocompatibility was assessed by cytocompatibility towards human fibroblastic cells, low platelet activation and absence of vascular irritation. Our work not only sheds light on using covalently bonded rhamnolipids towards dual species biofilms but also highlights the biocompatibility of the obtained PDMS surfaces.
Subject(s)
Anti-Infective Agents , Catheter-Related Infections , Anti-Bacterial Agents , Biocompatible Materials/pharmacology , Biofilms , Candida albicans , Catheter-Related Infections/drug therapy , Catheter-Related Infections/prevention & control , Dimethylpolysiloxanes/pharmacology , Glycolipids , Humans , Staphylococcus aureus/physiology , Staphylococcus epidermidisABSTRACT
The development of nanoparticles as antimicrobial agents against pathogenic bacteria has emerged as one of the leading global healthcare challenges. In this study, Mg(OH)2 NPs with controlled morphology and nanometric size, using two distinct counterions, chloride or nitrate, have been synthesized using Rosehip (RH) extract that has privileges beyond conventional chemical and physical methods. Various physicochemical techniques were used to characterize the RH-functionalized Mg-based NPs. They exhibited a spherical shape with a diameter of ~10 nm, low crystallinity compared to non-functionalized NPs, high polyphenol content, and negative zeta potential in three different media (H2O, TSB, and cell medium). The resulting RH-functionalized Mg-based NPs also exhibited an increased antibacterial activity against Gram-positive (S. Epidermis and S. aureus) and Gram-negative (E. Coli) bacteria compared to those prepared in pure water (0 % RH), an effect that was well evident with low NPs contents (250 µg/mL). A preliminary attempt to elucidate their mechanism of action revealed that RH-functionalized Mg-based NPs could disrupt cellular structures (bacterial cell wall and cytoplasmic membrane) and damage the bacterial cell, as confirmed by TEM imaging. Noteworthy is that Mg-based NPs exhibited higher toxicity to bacteria than to eukaryotic cells. More significantly, was their enhanced in vivo efficacy in a Galleria mellonella invertebrate animal model, when infected with S. aureus bacteria. Overall, our findings indicate that well-engineered Rosehip magnesium-based nanoparticles can be used as a green non-cytotoxic polyphenolic source in different antibacterial applications for the biomedical industry.
Subject(s)
Metal Nanoparticles , Rosa , Animals , Anti-Bacterial Agents/chemistry , Anti-Bacterial Agents/pharmacology , Bacteria , Escherichia coli , Metal Nanoparticles/chemistry , Microbial Sensitivity Tests , Plant Extracts/chemistry , Plant Extracts/pharmacology , Staphylococcus aureusABSTRACT
Controlling bacterial biofilm formation on silicone-based bloodstream catheters is of great concern to prevent related-infections. In this study, rhamnolipids (RLs), glycolipid biosurfactants, specifically a RLs mixture and the purified di-RL (RhaRhaC10:0C10:0) were covalently bonded to silicone with the intention of reaching long-lasting antibiofilm surfaces. RLs mixture and di-RL were identified by an UHPLC-MS method that also allowed the confirmation of compound isolation by automated flash chromatography. Silicone surfaces underwent air-plasma treatment, inducing reactive oxygen radicals able to promote the RLs grafting that was confirmed by contact angle, FTIR-ATR and AFM measurements. The antibiofilm activity towards different Gram positive strains was evaluated by colony forming units (CFU) count and confocal laser microscopy. In addition, protein adsorption and biocompatibility were also investigated. RLs were successfully grafted onto silicone and RLs mixture and RhaRhaC10C10:0 functionalized specimens reduced the biofilm formation over 2.3 log units against methicillin sensitive Staphylococcus aureus. Additionally, a decrease of 1 log unit was observed against methicillin resistant S. aureus and S. epidermidis. Functionalized samples showed cytocompatibility towards human dermal fibroblasts, hemocompatibility and no vascular irritation potential. The results mentioned above revealed a synergy between the antimicrobial and the anti-adhesive properties of RLs, making these compounds good candidates for the improvement of the medical devices antibiofilm properties.
Subject(s)
Anti-Infective Agents , Methicillin-Resistant Staphylococcus aureus , Anti-Bacterial Agents/pharmacology , Biofilms , Catheters/microbiology , Dimethylpolysiloxanes , Glycolipids/pharmacology , Humans , Staphylococcus epidermidisABSTRACT
INTRODUCTION: Mineral trioxide aggregate (MTA)-based sealers are endodontic materials with widespread success in distinct clinical applications, potentially embracing direct contact with the bone tissue. Bone response to these materials has been traditionally addressed in vitro. Nonetheless, translational data are limited by the absence of native cell-to-cell and cell-to-matrix interactions that hinder the representativeness of the analysis. Ex vivo organotypic systems, relying on the culture of explanted biological tissues, preserve the cell/tissue composition, reproducing the spatial and organizational in situ complexity. This study was grounded on an innovative research approach, relying on the assessment of an ex vivo organotypic bone tissue culture system to address the osteogenic response to 3 distinct MTA-based sealers. METHODS: Embryonic chick femurs were isolated and grown ex vivo for 11 days in the presence of MTA Plus (Avalon Biomed Inc, Bradenton, FL), ProRoot MTA (Dentsply Tulsa Dental, Hohnson City, Germany), Biodentine (Septodont, Saint Maurdes Fosses, France), or AH Plus (Dentsply Sirona, Konstanz, Germany); the latter was used as a control material. Femurs were characterized by histologic, histochemical, and histomorphometric analysis. Gene expression assessment of relevant osteogenic markers was conducted by quantitative polymerase chain reaction. RESULTS: All MTA-based sealers presented an enhanced osteogenic performance compared with AH Plus. Histochemical and histomorphometric analyses support the increased activation of the osteogenic program by MTA-based sealers, with enhanced collagenous matrix deposition and tissue mineralization. Gene expression analysis supported the enhanced activation of the osteogenic program. Comparatively, ProRoot MTA induced the highest osteogenic functionality on the characterized femurs. CONCLUSIONS: MTA-based sealers enhanced the osteogenic activity within the assayed organotypic bone model, which was found to be a sensitive system for the assessment of osteogenic modulation mediated by endodontic sealers.
Subject(s)
Root Canal Filling Materials , Aluminum Compounds/pharmacology , Bone and Bones , Calcium Compounds/pharmacology , Drug Combinations , Materials Testing , Osteogenesis , Oxides/pharmacology , Root Canal Filling Materials/pharmacology , Silicates/pharmacologyABSTRACT
This cohort study aimed to characterize the oral microbiome of children with CLP, from two different age groups, and evaluate the effect of supervised or unsupervised toothbrushing on the microbiome of the cleft over time. Swab samples were collected from the cleft area at three different time points (A; no brushing, B; after 15 days and C; after 30 days) and were analyzed using next-generation sequencing to determine the microbial composition and diversity in these time points. Overall, brushing significantly decreased the abundance of the genera Alloprevotella and Leptotrichia in the two age groups examined, and for Alloprevotella this decrease was more evident for children (2-6 years old). In the preteen group (7-12 years old), a significant relative increase of the genus Rothia was observed after brushing. In this study, the systematic brushing over a period of thirty days also resulted in differences at the intra-individual bacterial richness.
ABSTRACT
Implant surfaces with cytocompatible and antibacterial properties are extremely desirable for the prevention of implant's infection and the promotion of osseointegration. In this work, both micro-arc oxidation (MAO) and DC magnetron sputtering techniques were combined in order to endow tantalum-based surfaces with osteoblastic cytocompatibility and antibacterial activity. Porous Ta2O5 layers containing calcium (Ca) and phosphorous (P) were produced by MAO (TaCaP) to mimic the bone tissue morphology and chemical composition (Ca/P ratio close to 1.67). Furthermore, zinc (Zn) nanoparticles were deposited onto the previous surfaces by DC magnetron sputtering without or with an additional thin carbon layer deposited over the nanoparticles (respectively, TaCaP-Zn and TaCaP-ZnC) to control the Zn ions (Zn2+) release. Before osteoblastic cell seeding, the surfaces were leached for three time-points in PBS. All modified samples were cytocompatible. TaCaP-Zn slightly impaired cell adhesion but this was improved in the samples leached for longer immersion times. The initial cell adhesion was clearly improved by the deposition of the carbon layer on the Zn nanoparticles, which also translated to a higher proliferation rate. Both Zn-containing surfaces presented antibacterial activity against S. aureus. The two surfaces were active against planktonic bacteria, and TaCaP-Zn also inhibited sessile bacteria. Attributing to the excellent in vitro performance of the nanostructured Ta surface, with osteoconductive elements by MAO followed by antimicrobial nanoparticles incorporation by magnetron sputtering, this work is clearly a progress on the strategy to develop a new generation of dental implants.
Subject(s)
Dental Implants , Nanoparticles , Anti-Bacterial Agents/pharmacology , Coated Materials, Biocompatible/pharmacology , Oxides , Porosity , Staphylococcus aureus , Surface Properties , Tantalum , TitaniumABSTRACT
Polymeric platforms obtained by three-dimensional (3D) printing are becoming increasingly important as multifunctional therapeutic systems for bone treatment applications. In particularly, researchers aim to control bacterial biofilm on these 3D-platforms and enhance re-growing bone tissue, at the same time. This study aimed to fabricate a 3D-printed polylactic acid platform loaded with hydroxyapatite (HA), iron oxide nanoparticles (IONPs) and an antibiotic (minocycline) with tuneable properties and multistimuli response. IONPs were produced by a facile chemical co-precipitation method showing an average diameter between 11 and 15 nm and a superparamagnetic behaviour which was preserved when loaded into the 3D-platforms. The presence of two types of nanoparticles (IONPs and HA) modify the nanomorphological/nanotopographical feature of the 3D-platforms justifying their adequate bioactivity profile and in vitro cellular effects on immortalized and primary osteoblasts, including cytocompatibility and increased osteogenesis-related gene expression (RUNX2, BGLAP and SPP1). Disk diffusion assays and SEM analysis confirmed the effect of the 3D-platforms loaded with minocycline against Staphylococcus aureus. Altogether results showed that fabricated 3D-platforms combined the exact therapeutic antibiofilm dose of the antibiotic against S. aureus, with the enhanced osteogenic stimulation of the HA and IONPs nanoparticles which is a disruptive approach for bone targeting applications.
Subject(s)
Magnetite Nanoparticles , Nanoparticles , Anti-Bacterial Agents/pharmacology , Bone Regeneration , Bone and Bones , Osteogenesis , Printing, Three-Dimensional , Staphylococcus aureus , Tissue ScaffoldsABSTRACT
The development of biomaterials that mimicking the hydroxyapatite nanoparticles existent in the immature bone tissue is crucial, especially to accelerate the bone remodeling and regeneration. In this work, it was developed for the first time, hydroxyapatite nanoparticles (NPs) incorporating citrate and zinc (cit-Zn-Hap) in their composition towards a one-step hydrothermal procedure. For comparison purposes, hydroxyapatite NPs incorporating only zinc (Zn-Hap) or citrate (cit-Hap), as well as hydroxyapatite without any of these elements (Hap) were synthesised. The physicochemical characterization was carried out reveling that, the presence of zinc on hydroxyapatite (cit-Zn-Hap), reduced the size of nanoparticles, changed the phosphate environment and decreased the surface charge when compared with cit-Hap nanoparticles. The osteogenic potential of cit-Zn-Hap NPs was analysed in human bone marrow-derived stromal cells (BMSCs), in the absence of osteoinductive factors. NPs were internalized by endocytosis appearing trapped in endosomes and lysosomes scattered through the cytoplasm. Exposure to these NPs resulted in a significant induction of ALP activity, extracellular matrix mineralization, and gene expression of early and later osteogenic transcription factors, as well as of osteoblastic markers. The osteoinductive effect might be regulated, at least in part, by the increased signalling through the canonical WNT pathway. Evaluation of the cell behaviour following exposure to Zn-Hap and cit-Hap strongly suggested a synergistic effect of citrate and Zn in cit-Zn-Hap NPs towards the induction of the osteogenic commitment and functionality of BMSCs. These findings will allow the design of new biomimetic hydroxyapatite nanoparticles with great potential for bone regeneration.
Subject(s)
Bone Regeneration/drug effects , Durapatite/pharmacology , Mesenchymal Stem Cells/drug effects , Osteogenesis/drug effects , Cells, Cultured , Citrates/chemistry , Durapatite/chemistry , Gene Expression Regulation/drug effects , Humans , Mesenchymal Stem Cells/cytology , Nanotubes , Particle Size , Zinc/chemistryABSTRACT
The clinical demand for bone scaffolds as an alternative strategy for bone grafting has increased exponentially and, up to date, numerous formulations have been proposed to regenerate the bone tissue. However, most of these structures lack at least one of the fundamental/ideal properties of these materials (e.g., mechanical resistance, interconnected porosity, bioactivity, biodegradability, etc.). In this work, we developed innovative composite scaffolds, based on crosslinked chitosan with glutaraldehyde (GA), combined with different atomized calcium phosphates (CaP) granules - hydroxyapatite (HA) or biphasic mixtures of HA and ß - tricalcium phosphate (ß-TCP), with improved biomechanical behavior and enhanced biological response. This innovative combination was designed to improve the scaffolds' functionality, in which GA improved chitosan mechanical strength and stability, whereas CaP granules enhanced the scaffolds' bioactivity and osteoblastic response, further reinforcing the scaffolds' structure. The biological assessment of the composite scaffolds showed that the specimens with 0.2% crosslinking were the ones with the best biological performance. In addition, the inclusion of biphasic granules induced a trend for increase osteogenic activation, as compared to the addition of HA granules. In conclusion, scaffolds produced in the present work, both with HA granules or the biphasic ones, and with low concentrations of GA, have shown adequate properties and enhanced biological performance, being potential candidates for application in bone tissue engineering.
Subject(s)
Bone and Bones/metabolism , Chitosan/chemistry , Durapatite/chemistry , Osteoblasts/metabolism , Osteogenesis , Tissue Scaffolds/chemistry , Bone and Bones/cytology , Cell Line, Tumor , Cross-Linking Reagents/chemistry , Glutaral/chemistry , Humans , Osteoblasts/cytologyABSTRACT
Herein the quantitative synthesis of eight new mono- and dianionic Organic Salts and Ionic Liquids (OSILs) from alendronic acid (ALN) is reported by following two distinct sustainable and straightforward methodologies, according to the type of cation. The prepared ALN-OSILs were characterized by spectroscopic techniques and their solubility in water and biological fluids was determined. An evaluation of the toxicity towards human healthy cells and also human breast, lung and bone (osteosarcoma) cell lines was performed. Globally, it was observed that the monoanionic OSILs showed lower toxicity than the corresponding dianionic structures to all cell types. The highest cytotoxic effect was observed in OSILs containing a [C2OHMIM] cation, in particular [C2OHMIM][ALN]. The latter showed an improvement in IC50 values of ca. three orders of magnitude for the lung and bone cancer cell lines as well as fibroblasts in comparison with ALN. The development of OSILs with high cytotoxicity effect towards the tested cancer cell types, and containing an anti-resorbing molecule such as ALN may represent a promising strategy for the development of new pharmacological tools to be used in those pathological conditions.
ABSTRACT
The processing of traditional poultry- and pork-based semidried fermented smoked sausages needs to be modernized to improve product quality and further extend its shelf life. The aim of the present study was to apply different combinations of high pressure (300 to 600 MPa) and time (154 to 1,800 s) on the sausages using an experimental design based on response surface methodology. The chemical, microbial, and sensory characteristics of sausages treated with high-pressure processing (HPP) were investigated. HPP application to semidried fermented sausages resulted in color changes, which could be dependent on the ingredients, formulation, and smoking conditions used. Nevertheless, none of the HPP treatments applied resulted in detectable changes in sensory properties, as tested in a triangle test and confirmed by the analysis of focus groups assessment. Significant differences were detected for lactic acid bacteria (LAB) counts from 344 MPa and 1,530 s onward, with a marked decrease for the combination of 600 MPa and 960 s (P < 0.05). Coagulase-negative staphylococci showed higher tolerance to the increase in pressure than LAB. HPP induced a microbial reduction on Enterobacteriaceae, molds, and yeasts, minimizing the production of the main biogenic amines. However, the polyamines (spermine and spermidine) increased since their metabolic use by microorganisms did not occur. Given the reduction of the main spoilage microbial indicators with no detectable sensory changes observed with the binomial condition of 600 MPa and 960 s, this was chosen as the optimal combination to be further applied. PRACTICAL APPLICATION: The results from sensory analysis revealed that any of the HPP treatments applied resulted in detectable changes in sensory properties, as tested in a triangle test and confirmed by the analysis of the focus groups speeches.
Subject(s)
Biogenic Amines/chemistry , Hydrostatic Pressure , Meat Products/analysis , Animals , Bacteria/classification , Bacteria/isolation & purification , Colony Count, Microbial , Fermentation , Food Microbiology , Poultry , SwineABSTRACT
Scaffolds based on aligned and non-aligned poly (L-lactic acid) (PLLA)/polycaprolactone (PCL) fibers obtained by electrospinning, associated to electrosprayed hydroxyapatite (HA) for tissue engineering applications were developed and their performance was compared in terms of their morphology and biological and mechanical behaviors. The morphological results assessed by scanning electron microscopy showed a mesh of PLLA/PCL fibers (random and perfectly aligned) associated with aggregates of nanophased HA. Fourier transform infrared spectrometry confirmed the homogeneity in the blends and the presence of nanoHA in the scaffold. As a result of fiber alignment a 15-fold increase in Young's Modulus and an 8-fold increase in tensile strength were observed when compared to non-aligned fibers. In PLLA/PCL/HA scaffolds, the introduction of nanoHA caused a remarkable improvement of the mechanical strength of this material acting as a reinforcement, enhancing the response of these constructs to tensile stress. In vitro testing was evaluated using osteoblast (MC3T3-E1) cells. The results showed that both fibrous scaffolds were able to support osteoblast cell adhesion and proliferation and that fiber alignment induced increased cellular metabolic activity. In addition, the adhesion and proliferation of Staphylococcus aureus were evaluated and a lower number of colony forming units (CFUs) was obtained in the scaffolds with aligned fibers.
ABSTRACT
Periodontal diseases remain a challenge due to a complex interplay of factors involving a chronic inflammatory activation and bacteria internalization in periodontal cells. In this work, chitosan-nanoparticles loaded with minocycline (MH-NPs), a tetracycline with antimicrobial and anti-inflammatory effects, were developed for in situ delivery in the periodontal milieu aiming to improve drug effectiveness. A general cytocompatibility evaluation and a detailed approach to address the cellular uptake process, trafficking pathways and the modulation of relevant inflammatory gene expression was conducted using human gingival fibroblasts. Results show that MH-NPs with an adequate cytocompatible profile can be internalized by distinct endocytic processes (macropinocytosis and clathrin-mediated endocytosis). The ability to modulate autophagy with the delivery within the same endosomal/lysosomal pathway as periodontal pathogens was observed, which increases the intracellular drug effectiveness. Porphyromonas gingivalis LPS-stimulated cultures, grown in the presence of MH-NPs, were found to express significantly reduced levels of inflammation-related markers (IL-1b, TNFα, CXCL-8, NFKB1). These nanoparticles can be potentially used in periodontal disease treatment conjoining the ability of intracellular drug targeting with significant anti-inflammatory effects.
Subject(s)
Anti-Bacterial Agents/administration & dosage , Anti-Inflammatory Agents/administration & dosage , Chitosan/chemistry , Minocycline/administration & dosage , Anti-Bacterial Agents/pharmacology , Anti-Inflammatory Agents/pharmacology , Cells, Cultured , Drug Delivery Systems , Fibroblasts/drug effects , Fibroblasts/pathology , Gingiva/cytology , Gingiva/drug effects , Gingiva/microbiology , Humans , Inflammation/drug therapy , Inflammation/pathology , Minocycline/pharmacology , Nanoparticles , Periodontal Diseases/drug therapy , Porphyromonas gingivalis/drug effectsABSTRACT
Novel ionic liquids and organic salts based on mono- or dianionic zoledronate and protonated superbases, choline and n-alkylmethylimidazolium cations, were prepared and characterized by spectroscopic and thermal analyses. Most of the prepared salts display amorphous structures and very high solubility in water and saline solutions, especially the dianionic salts. Among the zoledronate-based ionic compounds, those containing choline [Ch] and methoxyethylmethylimidazolium [C3 OMIM] cations appear to have significant cytotoxicity against human osteosarcoma cells (MG63) and low toxicity toward healthy skin fibroblast cells. Because osteosarcoma is a bone pathology characterized by an increase in bone turnover rate, the results presented herein may be a promising starting point for the development of new ionic pharmaceutical drugs against osteosarcoma.
Subject(s)
Antineoplastic Agents/pharmacology , Diphosphonates/pharmacology , Fibroblasts/drug effects , Ionic Liquids/pharmacology , Zoledronic Acid/pharmacology , Antineoplastic Agents/chemistry , Cell Line, Tumor , Cell Proliferation/drug effects , Diphosphonates/chemistry , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor , Humans , Ionic Liquids/chemistry , Molecular Structure , Salts/chemistry , Salts/pharmacology , Structure-Activity Relationship , Zoledronic Acid/chemistryABSTRACT
Nano-hydroxyapatite has been used as an oral care ingredient, being incorporated in several products for the treatment of dental hypersensitivity and enamel remineralisation. Despite its promising results, regulatory and safety concerns have been discussed and questioned by the European Scientific Committee on Consumer Safety (SCCS) regarding the usage of hydroxyapatite nanoparticles in oral care products. In this work, a commercially available nano-hydroxyapatite was characterized and its cytocompatibility towards human gingival fibroblasts was evaluated, as well as its irritation potential using the in vitro HET-CAM assay. All the conditions chosen in this study tried to simulate the tooth brushing procedure and the hydroxyapatite nanoparticles levels normally incorporated in oral care products. The commercial hydroxyapatite nanoparticles used in this study exhibited a rod-like morphology and the expected chemical and phase composition. The set of in vitro cytotoxicity parameters accessed showed that these nanoparticles are highly cytocompatible towards human gingival fibroblasts. Additionally, these nanoparticles did not possess any irritation potential on HET-CAM assay. This study clarifies the issues raised by SCCS and it concludes that this specific nano-hydroxyapatite is cytocompatible, as these nanoparticles did not alter the normal behaviour of the cells. Therefore, they are safe to be used in oral care products.