Melanin-Based Nanoparticles for Lymph Node Tattooing: Experimental, Histopathological and Ultrastructural Study.

In breast cancer, Targeted Axillary Dissection (TAD) allows for the selective excision of the sentinel lymph node (SLN) during primary tumor surgery. TAD consists of the resection of labelled SLNs prior to neoadjuvant chemotherapy (NACT). Numerous clinical and preclinical studies have explored the use of carbon-based colloids for SLN tattooing prior to NACT. However, carbon vectors show varying degrees of inflammatory reactions and, in about one fifth of cases, carbon particles migrate via the lymphatic pathway to other nodes, causing the SLN to mismatch the tattooed node. To overcome these limitations, in this study, we explored the use of melanin as a staining endogenous pigment. We synthesized and characterized melanin-loaded polymeric nanoparticles (Mel-NPs) and used them to tattoo lymph nodes in pig animal models given the similarity in the size of the human and pig nodes. Mel-NPs tattooed lymph nodes showed high identification rates, reaching 83.3% positive identification 16 weeks after tattooing. We did not observe any reduction in the identification as time increased, implying that the colloid is stable in the lymph node tissue. In addition, we performed histological and ultrastructural studies to characterize the biological behavior of the tag. We observed foreign-body-like granulomatous inflammatory responses associated with Mel-NPs, characterized by the formation of multinucleated giant cells. In addition, electron microscopy studies showed that uptake is mainly performed by macrophages, and that macrophages undergo cellular damage associated with particle uptake.

PLGA nanoparticle-encapsulated lysostaphin for the treatment of Staphylococcus aureus infections.

Staphylococcus aureus possesses the ability to become pathogenic, leading to severe and life-threatening infections. Its methicillin-resistant variant MRSA has garnered high-priority status due to its increased morbidity and associated mortality. This emphasizes the urgency for novel anti-staphylococcal agents. The bacteriocin lysostaphin stands out for its remarkable bactericidal activity against S. aureus, including MRSA, outperforming conventional antibiotics. However, the clinical application of lysostaphin faces challenges, including enzymatic activity loss under physiological conditions and potential immunogenicity. This study introduces a novel approach by encapsulating lysostaphin within polylactic-co-glycolic acid (PLGA) nanoparticles, a biodegradable copolymer known for its biocompatibility and sustained drug release ability. The study assesses the antimicrobial activity of lysostaphin-loaded PLGA nanoparticles against different S. aureus strains, and we also used GFP-expressing S. aureus for facilitating its traceability in planktonic, biofilm, and intracellular infection models. The results showed the significant reduction in bacteria viability both in planktonic and biofilm states. The in vitro intracellular infection model demonstrated the significantly enhanced efficiency of the developed nanoparticles compared to the treatment with the free bacteriocin. This research presents lysostaphin encapsulation within PLGA nanoparticles and offers promising avenues for enhancing lysostaphin's therapeutic efficacy against S. aureus infections.

Antimicrobial Photodynamic Therapy Using Encapsulated Protoporphyrin IX for the Treatment of Bacterial Pathogens.

Herein, we report on the antimicrobial photodynamic effect of polymeric nanoparticles containing the endogenous photosensitizer protoporphyrin IX. Compared to equivalent doses of the free photosensitizer, we demonstrated that the photodynamic antimicrobial efficacy of PLGA (polylactic-co-glycolic acid) nanoparticles containing protoporphyrin IX (PpIX) against pathogenic Staphylococcus aureus (S. aureus) is preserved after encapsulation, while photobleaching is reduced. In addition, compared to equivalent doses of the free porphyrin, we show that a reduction in the cytotoxicity in mammalian cell cultures is observed when encapsulated. Therefore, the encapsulation of protoporphyrin IX reduces its photodegradation, while the released photosensitizer maintains its ability to generate reactive oxygen species upon light irradiation. The polymeric nanoencapsulation promotes aqueous solubility for the hydrophobic PpIX, improves its photostability and reduces the cytotoxicity, while providing an extended release of this endogenous photosensitizer.

Expert opinion on antimicrobial therapies: is there enough scientific evidence to state that targeted therapies outperform non-targeted ones?

INTRODUCTION: Different active and passive strategies have been developed to fight against pathogenic bacteria. Those actions are undertaken to reduce the bacterial burden while minimizing the possibilities to develop not only antimicrobial resistance but also antimicrobial side-effects such as allergic or hypersensitivity reactions. AREAS COVERED: We have reviewed preclinical results that evidence that targeted antimicrobial therapies outperform non-targeted ones. Active selective targeting against pathogenic bacteria has been achieved through the functionalization of antimicrobials, either alone or encapsulated within micro- or nanocarriers, with various recognition moieties. These moieties include peptides, aptamers, antibodies, carbohydrates, extracellular vesicles, cell membranes, infective agents, and other affinity ligands with specific bacterial tropism. Those selective ligands increase retention and enhance effectiveness reducing the side-effects and the required dose to exert the antimicrobial action at the site of infection. EXPERT OPINION: When using targeted antimicrobial therapies not only reduced side-effects are observed, but also, compared to the administration of equivalent doses of the non-targeted drugs, a superior efficacy has been demonstrated against planktonic, sessile, and intracellular pathogenic bacterial persisters. The translation of those targeted therapies to subsequent phases of clinical development still requires the demonstration of a reduction in the probabilities for the pathogen to develop resistance when using targeted approaches.

Thioredoxin domain containing 5 is involved in the hepatic storage of squalene into lipid droplets in a sex-specific way.

Hepatic thioredoxin domain-containing 5 (TXNDC5) is a member of the protein disulfide isomerase family found associated with anti-steatotic properties of squalene and located in the endoplasmic reticulum and in lipid droplets. Considering that the latter are involved in hepatic squalene accumulation, the present research was aimed to investigate the role of TXNDC5 on hepatic squalene management in mice and in the AML12 hepatic cell line. Wild-type and TXNDC5-deficient (KO) mice were fed Western diets with or without 1% squalene supplementation for 6 weeks. In males, but not in females, absence of TXNDC5 blocked hepatic, but not duodenal, squalene accumulation. Hepatic lipid droplets were isolated and characterized using label-free LC-MS/MS analysis. TXNDC5 accumulated in this subcellular compartment of mice receiving squalene and was absent in TXNDC5-KO male mice. The latter mice were unable to store squalene in lipid droplets. CALR and APMAP were some of the proteins that responded to the squalene administration in all studied conditions. CALR and APMAP were positively associated with lipid droplets in the presence of squalene and they were decreased by the absence of TXNDC5. The increased squalene content was reproduced in vitro using AML12 cells incubated with squalene-loaded nanoparticles and this effect was not observed in an engineered cell line lacking TXNDC5. The phenomenon was also present when incubated in the presence of a squalene epoxidase inhibitor, suggesting a mechanism of squalene exocytosis involving CALR and APMAP. In conclusion, squalene accumulation in hepatic lipid droplets is sex-dependent on TXNDC5 that blocks its secretion.

Gold Nanoparticles Capped with a Novel Titanium(IV)-Containing Polyoxomolybdate Cluster: Selective and Enhanced Bactericidal Effect Against Escherichia coli.

Bacterial infections are a public health threat of increasing concern in medical care systems; hence, the search for novel strategies to lower the use of antibiotics and their harmful effects becomes imperative. Herein, the antimicrobial performance of four polyoxometalate (POM)-stabilized gold nanoparticles (Au@POM) against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) as Gram-negative and Gram-positive bacteria models, respectively, is studied. The bactericidal studies performed, both in planktonic and sessile forms, evidence the antimicrobial potential of these hybrid nanostructures with selectivity toward Gram-negative species. In particular, the Au@GeMoTi composite with the novel [Ti2 (HGeMo7 O28 )2 ]10- POM capping ligand exhibits outstanding bactericidal efficiency with a minimum inhibitory concentration of just 3.12 µm for the E. coli strain, thus outperforming the other three Au@POM counterparts. GeMoTi represents the fourth example of a water-soluble TiIV -containing polyoxomolybdate, and among them, the first sandwich-type structure having heteroatoms in high-oxidation state. The evaluation of the bactericidal mechanisms of action points to the cell membrane hyperpolarization, disruption, and subsequent nucleotide leakage and the low cytotoxicity exerted on five different cell lines at antimicrobial doses demonstrates the antibiotic-like character. These studies highlight the successful design and development of a new POM-based nanomaterial able to eradicate Gram-negative bacteria without damaging mammalian cells.

Real-time in vivo monitoring of the antimicrobial action of combination therapies in the management of infected topical wounds.

The increasing prevalence of non-healing infected wounds has become a serious concern in the clinical practice, being associated to population aging and to the rising prevalence of several chronic conditions such as diabetes. Herein, the evaluation of the bactericidal and antibiofilm effects of the natural antiseptic terpenes thymol and farnesol standing alone or in combination with the standard care antiseptic chlorhexidine was carried out both in vitro and in vivo. The in vitro combinatorial treatment of chlorhexidine associated with those terpenes against Staphylococcus aureus in its planktonic and sessile forms demonstrated a superior antibacterial activity than that of chlorhexidine alone. Real-time in vivo monitoring of infection progression and antimicrobial treatment outcomes were evaluated using the bioluminescent S. aureus strain Xen36. In vivo studies on infected wound splinting murine models corroborated the superior bactericidal effects of the combinatorial treatments here proposed. Moreover, the encapsulation of thymol in electrospun Eudragit® S100 (i.e., a synthetic anionic copolymer of methacrylic acid and ethyl acrylate)-based wound dressings was also carried out in order to design efficient antimicrobial wound dressings.

Encapsulation of Iron-Saturated Lactoferrin for Proteolysis Protection with Preserving Iron Coordination and Sustained Release.

Lactoferrin (Lf) is a globular glycoprotein found mainly in milk. It has a very high affinity for iron(III) ions, and its fully saturated form is called holoLf. The antimicrobial, antiviral, anticancer, and immunomodulatory properties of Lf have been studied extensively for the past two decades. However, to demonstrate therapeutic benefits, Lf has to be efficiently delivered to the intestinal tract in its structurally intact form. This work aimed to optimize the encapsulation of holoLf in a system based on the versatile Eudragit® RS polymer to protect Lf against the proteolytic environment of the stomach. Microparticles (MPs) with entrapped holoLf were obtained with satisfactory entrapment efficiency (90-95%), high loading capacity (9.7%), and suitable morphology (spherical without cracks or pores). Detailed studies of the Lf release from the MPs under conditions that included simulated gastric or intestinal fluids, prepared according to the 10th edition of the European Pharmacopeia, showed that MPs partially protected holoLf against enzymatic digestion and ionic iron release. The preincubation of MPs loaded with holoLf under conditions simulating the stomach environment resulted in the release of 40% of Lf from the MPs. The protein released was saturated with iron ions at 33%, was structurally intact, and its iron scavenging properties were preserved.

Antibody-Functionalized Polymer Nanoparticles for Targeted Antibiotic Delivery in Models of Pathogenic Bacteria Infecting Human Macrophages.

The efficacy of antibody-functionalized poly(d,l-lactide-co-glycolide) (PLGA) nanoparticles (NPs), prepared by nanoprecipitation, carrying rifampicin (RIF) against planktonic, sessile, and intracellular Staphylococcus aureus and Escherichia coli is reported here. A biotinylated anti-S. aureus polyclonal antibody, which binds to structural antigens of the whole bacterium, was functionalized on the surface of RIF-loaded PLGA-based NPs by using the high-affinity avidin-biotin complex. This general strategy allows the binding of commercially available biotinylated antibodies. Coculture models of S. aureus ATCC 25923 and Escherichia coli S17 were used to demonstrate the preferential selectivity of the antibody-functionalized NPs against the Gram-positive bacterium only. At 0.2 µg/mL, complete S. aureus eradication was observed for the antibody-functionalized RIF-loaded NPs, whereas only a 5-log reduction was observed for the nontargeted RIF-loaded NPs. S. aureus is a commensal facultative pathogen having part of its live cycle intracellularly in both phagocytic and nonphagocytic cells. Those intracellular bacterial persisters, named small colony variants, have been postulated as reservoirs of relapsed episodes of infection and consequent treatment failure. At 0.5 µg/mL, the RIF-loaded NPs reduced in 2-log intracellular S. aureus-infecting human macrophages. The ability of those antibody-functionalized nanoparticles to prevent biofilm formation or to reduce the bacterial burden in already-formed mature biofilms is also reported here using S. aureus and E. coli single and cocultured biofilms. In the prevention of S. aureus biofilm formation, the antibody-functionalized NPs exerted a superior inhibition of bacterial growth (up to 2 logs) compared to the nonfunctionalized ones. This study demonstrates the selectivity of the synthesized immunonanoparticles and their antimicrobial efficacy in different scenarios, including planktonic cultures, sessile conditions, and even against intracellular infective pathogens.

Colistin-loaded aerosolizable particles for the treatment of bacterial respiratory infections.

Compared to parenteral administration of colistin, its direct pulmonary administration can maximize lung drug deposition while reducing systemic adverse side effects and derived nephrotoxicity. Current pulmonary administration of colistin is carried out by the aerosolization of a prodrug, colistin methanesulfonate (CMS), which must be hydrolized to colistin in the lung to produce its bactericidal effect. However, this conversion is slow relative to the rate of absorption of CMS, and thus only 1.4 % (w/w) of the CMS dose is converted to colistin in the lungs of patients receiving inhaled CMS. We synthesized several aerosolizable nanoparticle carriers loaded with colistin using different techniques and selected particles with sufficient drug loading and adequate aerodynamic behavior to efficiently deliver colistin to the entire lung. Specifically, we carried out (i) the encapsulation of colistin by single emulsion-solvent evaporation with immiscible solvents using polylactic-co-glycolic (PLGA) nanoparticles; (ii) its encapsulation using nanoprecipitation with miscible solvents using poly(lactide-co-glycolide)-block-poly(ethylene glycol) as encapsulating matrix; (iii) colistin nanoprecipitation using the antisolvent precipitation method and its subsequent encapsulation within PLGA nanoparticles; and (iv) colistin encapsulation within PLGA-based microparticles using electrospraying. Nanoprecipitation of pure colistin using antisolvent precipitation showed the highest drug loading (55.0 ± 4.8 wt%) and spontaneously formed aggregates with adequate aerodynamic diameter (between 3 and 5 µm) to potentially reach the entire lung. These nanoparticles were able to completely eradicate Pseudomonas aeruginosa in an in vitro lung biofilm model at 10 µg/mL (MBC). This formulation could be a promising alternative for the treatment of pulmonary infections improving lung deposition and, therefore, the efficacy of aerosolized antibiotics.

In Cellulo Bioorthogonal Catalysis by Encapsulated AuPd Nanoalloys: Overcoming Intracellular Deactivation.

Bioorthogonal metallocatalysis has opened up a xenobiotic route to perform nonenzymatic catalytic transformations in living settings. Despite their promising features, most metals are deactivated inside cells by a myriad of reactive biomolecules, including biogenic thiols, thereby limiting the catalytic functioning of these abiotic reagents. Here we report the development of cytocompatible alloyed AuPd nanoparticles with the capacity to elicit bioorthogonal depropargylations with high efficiency in biological media. We also show that the intracellular catalytic performance of these nanoalloys is significantly enhanced by protecting them following two different encapsulation methods. Encapsulation in mesoporous silica nanorods resulted in augmented catalyst reactivity, whereas the use of a biodegradable PLGA matrix increased nanoalloy delivery across the cell membrane. The functional potential of encapsulated AuPd was demonstrated by releasing the potent chemotherapy drug paclitaxel inside cancer cells. Nanoalloy encapsulation provides a novel methodology to develop nanoreactors capable of mediating new-to-life reactions in cells.

Elucidating the mechanisms of action of antibiotic-like ionic gold and biogenic gold nanoparticles against bacteria.

The antimicrobial action of gold depends on different factors including its oxidation state in the intra- and extracellular medium, the redox potential, its ability to produce reactive oxygen species (ROS), the medium components, the properties of the targeted bacteria wall, its penetration in the bacterial cytosol, the cell membrane potential, and its interaction with intracellular components. We demonstrate that different gold species are able to induce bacterial wall damage as a result of their electrostatic interaction with the cell membrane, the promotion of ROS generation, and the consequent DNA damage. In-depth genomic and proteomic studies on Escherichia coli confirmed the superior toxicity of Au (III) vs Au (I) based on the different molecular mechanisms analyzed including oxidative stress, bacterial energetic metabolism, biosynthetic processes, and cell transport. At equivalent bactericidal doses of Au (III) and Au (I) eukaryotic cells were not as affected as bacteria did, maintaining unaffected cell viability, morphology, and focal adhesions; however, increased ROS generation and disruption in the mitochondrial membrane potential were also observed. Herein, we shed light on the antimicrobial mechanisms of ionic and biogenic gold nanoparticles against bacteria. Under selected conditions antibiotic-like ionic gold can exert a strong antimicrobial activity while being harmless to human cells.

A bone-on-a-chip collagen hydrogel-based model using pre-differentiated adipose-derived stem cells for personalized bone tissue engineering.

Mesenchymal stem cells have contributed to the continuous progress of tissue engineering and regenerative medicine. Adipose-derived stem cells (ADSC) possess many advantages compared to other origins including easy tissue harvesting, self-renewal potential, and fast population doubling time. As multipotent cells, they can differentiate into osteoblastic cell linages. In vitro bone models are needed to carry out an initial safety assessment in the study of novel bone regeneration therapies. We hypothesized that 3D bone-on-a-chip models containing ADSC could closely recreate the physiological bone microenvironment and promote differentiation. They represent an intermedium step between traditional 2D-in vitro and in vivo experiments facilitating the screening of therapeutic molecules while saving resources. Herein, we have differentiated ADSC for 7 and 14 days and used them to fabricate in vitro bone models by embedding the pre-differentiated cells in a 3D collagen matrix placed in a microfluidic chip. Osteogenic markers such as alkaline phosphatase activity, calcium mineralization, changes on cell morphology, and expression of specific proteins (bone sialoprotein 2, dentin matrix acidic phosphoprotein-1, and osteocalcin) were evaluated to determine cell differentiation potential and evolution. This is the first miniaturized 3D-in vitro bone model created from pre-differentiated ADSC embedded in a hydrogel collagen matrix which could be used for personalized bone tissue engineering.

New insights in osteoarthritis diagnosis and treatment: Nano-strategies for an improved disease management.

Osteoarthritis (OA) is a common chronic joint pathology that has become a predominant cause of disability worldwide. Even though the origin and evolution of OA rely on different factors that are not yet elucidated nor understood, the development of novel strategies to treat OA has emerged in the last years. Cartilage degradation is the main hallmark of the pathology though alterations in bone and synovial inflammation, among other comorbidities, are also involved during OA progression. From a molecular point of view, a vast amount of signaling pathways are implicated in the progression of the disease, opening up a wide plethora of targets to attenuate or even halt OA. The main purpose of this review is to shed light on the recent strategies published based on nanotechnology for the early diagnosis of the disease as well as the most promising nano-enabling therapeutic approaches validated in preclinical models. To address the clinical issue, the key pathways involved in OA initiation and progression are described as the main potential targets for OA prevention and early treatment. Furthermore, an overview of current therapeutic strategies is depicted. Finally, to solve the drawbacks of current treatments, nanobiomedicine has shown demonstrated benefits when using drug delivery systems compared with the administration of the equivalent doses of the free drugs and the potential of disease-modifying OA drugs when using nanosystems. We anticipate that the development of smart and specific bioresponsive and biocompatible nanosystems will provide a solid and promising basis for effective OA early diagnosis and treatment. This article is categorized under: Diagnostic Tools > In Vivo Nanodiagnostics and Imaging Implantable Materials and Surgical Technologies > Nanotechnology in Tissue Repair and Replacement.

Optimization of superfluorinated PLGA nanoparticles for enhanced cell labelling and detection by 19F-MRI.

Fluorine-19 (19F) Magnetic Resonance Imaging (MRI) is an emergent imaging technique for molecular imaging and cell tracking. Lack of intrinsic 19F signals in tissues allows unambiguous in vivo detection of exogenous fluorinated probes, complementary to the anatomical and multiparametric information obtained by standard 1H-MRI. However, the intrinsic low sensitivity of MRI technique requires the need of designing increasingly effective fluorinated tracers. PERFECTA, with its 36 magnetically equivalent 19F atoms and a designed branched molecular structure, represents an excellent superfluorinated tracer. In this paper, we report the development of PERFECTA loaded PLGA NPs stabilized by different coatings as promising 19F-MRI probes. The results clearly show the optimal cellular uptake of the produced colloidally stable PERFECTA loaded PLGA NPs without impact on cells viability. Importantly, NPs stabilization with the anionic surfactant sodium cholate (NaC) clearly enhances NPs internalization within cells with respect to PVA-coated NPs. Moreover, the optimized NPs are characterized by shorter T1 relaxation times with respect to other PERFECTA formulations that would allow the increase of 19F-MRI sensitivity with fast imaging acquisitions.

Electrostatic self-assembly approach in the deposition of bio-functional chitosan-based layers enriched with caffeic acid on Ti-6Al-7Nb alloys by alternate immersion.

Tailoring surface properties by layer-by-layer (LBL) deposition directed on the construction of complex multilayer coatings with nanoscale precision enables the development of novel structures and devices with desired functional properties (i.e., osseointegration, bactericidal activity, biocorrosion protection). Herein, electrostatic self-assembly was applied to fabricate biopolymer-based coatings involving chitosan (CSM) and alginate (AL) enriched with caffeic acid (CA) on Ti-6Al-7Nb alloyed surfaces. The method of CA grafting onto the chitosan backbone (CA-g-CSM) as well as all used reagents for implant functionalization were chosen as green and sustainable approach. The final procedure of surface modification of the Ti-6Al-7Nb alloy consists of three steps: (i) chemical treatment in Piranha solution, (ii) plasma chemical-activation of the Ti alloy surface in a RF CVD (Radio Frequency Chemical Vapour Deposition) reactor using Ar, O2 and NH3 gaseous precursors, and (iii) a multi-step deposition of bio-functional coatings via dip-coating method. Corrosion tests have revealed that the resulting chitosan-based coatings, also these involving CA, block the specimen surface and hinder corrosion of titanium alloy. Furthermore, the antioxidant layers are characterized by beneficial level of roughness (Ra up ca. 350 nm) and moderate hydrophilicity (59°) with the dispersion part of conducive surface energy ca. 30 mJ/m2. Noteworthy, all coatings are biocompatible as the intact morphology of cultured eukaryotic cells ensured proper growth and proliferation, while exhibit bacteriostatic character, particularly in contact with Gram-(-) bacteria (E. coli). The study indicates that the applied simple sustainable strategy has contributed significantly to obtaining homogeneous, stable, and biocompatible while antibacterial biopolymer-based coatings.

Silver Nanoparticles-Polyethyleneimine-Based Coatings with Antiviral Activity against SARS-CoV-2: A New Method to Functionalize Filtration Media.

The use of face masks and air purification systems has been key to curbing the transmission of SARS-CoV-2 aerosols in the context of the current COVID-19 pandemic. However, some masks or air conditioning filtration systems are designed to remove large airborne particles or bacteria from the air, being limited their effectiveness against SARS-CoV-2. Continuous research has been aimed at improving the performance of filter materials through nanotechnology. This article presents a new low-cost method based on electrostatic forces and coordination complex formation to generate antiviral coatings on filter materials using silver nanoparticles and polyethyleneimine. Initially, the AgNPs synthesis procedure was optimized until reaching a particle size of 6.2 ± 2.6 nm, promoting a fast ionic silver release due to its reduced size, obtaining a stable colloid over time and having reduced size polydispersity. The stability of the binding of the AgNPs to the fibers was corroborated using polypropylene, polyester-viscose, and polypropylene-glass spunbond mats as substrates, obtaining very low amounts of detached AgNPs in all cases. Under simulated operational conditions, a material loss less than 1% of nanostructured silver was measured. SEM micrographs demonstrated high silver distribution homogeneity on the polymer fibers. The antiviral coatings were tested against SARS-CoV-2, obtaining inactivation yields greater than 99.9%. We believe our results will be beneficial in the fight against the current COVID-19 pandemic and in controlling other infectious airborne pathogens.

Pharmacokinetic control on the release of antimicrobial drugs from pH-responsive electrospun wound dressings.

The acidic pH of healthy skin changes during wound healing due to the exposure of the inner dermal and subcutaneous tissue and due to the potential colonization of pathogenic bacteria. In chronic non-healing wounds, the pH values vary in a wide pH range but the appearance of an alkaline shift is common. After a wound is incurred, neutral pH in the wound bed is characteristic of the activation of the cascade of regenerative and remodeling processes. In order to adjust drug release to the specific pH of the wound, herein, drug-loaded wound dressings having pH-responsiveness containing antiseptics and antibiotics and exerting different release kinetics in order to have a perfect match between the drug release kinetics, and the pH conditions of each wound type, were developed. We have fabricated drug-loaded electrospun nanofibers loaded with the antiseptic chlorhexidine, with the broad-spectrum antibiotic rifampicin, and with the antimicrobial of natural origin thymol, using the pH-dependent methacrylic acid copolymer Eudragit® L100-55, which dissolves at pH > 5.5; those drugs were loaded within Eudragit® S100, which dissolves at pH > 7 and, finally, within the methacrylic ester copolymer Eudragit® RS100 which is pH independent and slowly erodes and releases its contained cargo. The antibacterial action of those advanced wound dressings has been evaluated against methicillin-sensitive S. aureus Newman strain expressing the coral green fluorescent protein (cGFP), as a model of a Gram-positive bacteria, and against E. coli S17 strain as a model of a Gram-negative bacteria. It was demonstrated that those combinational products integrate in one device the required characteristics for a wound dressing with the therapeutic action of a contained active principle and can be selected depending on the wound acidic or alkaline status for its appropriated management.

Trojan pH-Sensitive Polymer Particles Produced in a Continuous-Flow Capillary Microfluidic Device Using Water-in-Oil-in-Water Double-Emulsion Droplets.

A facile and robust microfluidic method to produce nanoparticle-in-microparticle systems (Trojan systems) is reported as a delivery vector for the oral administration of active pharmaceutical ingredients. The microfluidic system is based on two coaxial capillaries that produce monodisperse water-in-oil-in-water (W/O/W) double emulsions in a highly controlled fashion with precise control over the resulting particle structure, including the core and shell dimensions. The influence of the three phase flow rates, pH and drying process on the formation and overall size is evaluated. These droplets are then used as templates for the production of pH-sensitive Trojan microparticles after solvent evaporation. The shell of Trojan microparticles is made of Eudragit®, a methacrylic acid-ethyl acrylate copolymer that would enable the Trojan microparticle payload to first pass through the stomach without being degraded and then dissolve in the intestinal fluid, releasing the inner payload. The synthesis of the pH-sensitive Trojan microparticles was also compared with a conventional batch production method. The payloads considered in this work were different in nature: (1) fluorescein, to validate the feasibility of the polymeric shell to protect the payload under gastric pH; (2) poly(D,L-lactic acid/glycolic acid)-PLGA nanoparticles loaded with the antibiotic rifampicin. These PLGA nanoparticles were produced also using a microfluidic continuous process and (3) PLGA nanoparticles loaded with Au nanoparticles to trace the PLGA formulation under different environments (gastric and intestinal), and to assess whether active pharmaceutical ingredient (API) encapsulation in PLGA is due efficiently. We further showed that Trojan microparticles released the embedded PLGA nanoparticles in contact with suitable media, as confirmed by electron microscopy. Finally, the results show the possibility of developing Trojan microparticles in a continuous manner with the ability to deliver therapeutic nanoparticles in the gastrointestinal tract.

In Vitro Hydrolytic Degradation of Polyester-Based Scaffolds under Static and Dynamic Conditions in a Customized Perfusion Bioreactor.

Creating biofunctional artificial scaffolds could potentially meet the demand of patients suffering from bone defects without having to rely on donors or autologous transplantation. Three-dimensional (3D) printing has emerged as a promising tool to fabricate, by computer design, biodegradable polymeric scaffolds with high precision and accuracy, using patient-specific anatomical data. Achieving controlled degradation profiles of 3D printed polymeric scaffolds is an essential feature to consider to match them with the tissue regeneration rate. Thus, achieving a thorough characterization of the biomaterial degradation kinetics in physiological conditions is needed. Here, 50:50 blends made of poly(ε-caprolactone)-Poly(D,L-lactic-co-glycolic acid (PCL-PLGA) were used to fabricate cylindrical scaffolds by 3D printing (⌀ 7 × 2 mm). Their hydrolytic degradation under static and dynamic conditions was characterized and quantified. For this purpose, we designed and in-house fabricated a customized bioreactor. Several techniques were used to characterize the degradation of the parent polymers: X-ray Photoelectron Spectroscopy (XPS), Gel Permeation Chromatography (GPC), Scanning Electron Microscopy (SEM), evaluation of the mechanical properties, weigh loss measurements as well as the monitoring of the degradation media pH. Our results showed that flow perfusion is critical in the degradation process of PCL-PLGA based scaffolds implying an accelerated hydrolysis compared to the ones studied under static conditions, and up to 4 weeks are needed to observe significant degradation in polyester scaffolds of this size and chemical composition. Our degradation study and characterization methodology are relevant for an accurate design and to tailor the physicochemical properties of polyester-based scaffolds for bone tissue engineering.