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Objective: Fracture-related infection (FRI) remains a major concern in orthopaedic trauma. Functionalizing implants with antibacterial coatings are a promising strategy in mitigating FRI. Numerous implant coatings have been reported but the preventive and therapeutic effects vary. This systematic review aimed to provide a comprehensive overview of current implant coating strategies to prevent and treat FRI in animal fracture and bone defect models., Methods: A literature search was performed in three databases: PubMed, Web of Science and Embase, with predetermined keywords and criteria up to 28 February 2023. Preclinical studies on implant coatings in animal fracture or defect models that assessed antibacterial and bone healing effects were included., Results: A total of 14 studies were included in this systematic review, seven of which used fracture models and seven used defect models. Passive coatings with bacteria adhesion resistance were investigated in two studies. Active coatings with bactericidal effects were investigated in 12 studies, four of which used metal ions including Ag + and Cu 2+ ; five studies used antibiotics including chlorhexidine, tigecycline, vancomycin, and gentamicin sulfate; and the other three studies used natural antibacterial materials including chitosan, antimicrobial peptides, and lysostaphin. Overall, these implant coatings exhibited promising efficacy in antibacterial effects and bone formation., Conclusion: Antibacterial coating strategies reduced bacterial infections in animal models and favored bone healing in vivo . Future studies of implant coatings should focus on optimal biocompatibility, antibacterial effects against multi-drug resistant bacteria and polymicrobial infections, and osseointegration and osteogenesis promotion especially in osteoporotic bone by constructing multi-functional coatings for FRI therapy., The Translational Potential of This Paper: The clinical treatment of FRI is complex and challenging. This review summarizes novel orthopaedic implant coating strategies applied to FRI in preclinical studies, and offers a perspective on the future development of orthopaedic implant coatings, which can potentially contribute to alternative strategies in clinical practice., Competing Interests: A conflict of interest occurs when an individual’s objectivity is potentially compromised by a desire for financial gain, prominence, professional advancement or a successful outcome. The Editors of the Journal of Orthopaedic Translation strive to ensure that what is published in the Journal is as balanced, objective and evidence-based as possible. Since it can be difficult to distinguish between an actual conflict of interest and a perceived conflict of interest, the Journal requires authors to disclose all and any potential conflicts of interest., (© 2024 The Authors.)

Biomimetic cell culture systems are required to provide more physiologically relevant microenvironments for bone cells. Here, a simple 2.5D culture platform is proposed, combining adjustable stiffness and surface features that mimic bone topography by using sandpaper grits as master molds with two stiffness formulations of polydimethylsiloxane (PDMS). The subsequent replicas perfectly conform the grits and reproduce the corresponding negative relief with cavities separated by convex edges. Biomimicry is also provided by an extracellular matrix (ECM)-like thin film coating, using the layer-by-layer (LbL) method. The topographical features, alternating concave, and convex structures drive preosteoblasts organization and morphology. Strikingly, curvature orchestrates the commitment of preosteoblasts, with i) maturation to active osteoblasts able to produce a dense collagenous matrix that ultimately mineralizes in the cavities, and ii) edges hosting quiescent cells that synthetize a very thin immature collagen layer with no mineralization. In summary, the present in vitro culture system model offers a cell-instructive 2.5D microenvironment that controls preosteoblasts fate, leading to two coexisting subpopulations: mature osteoblasts and bone lining cells (BLC). This promising culture system opens new avenues to advanced tissue-engineered modeling and can be applied to precellularized bone biomaterials., (© 2023 Wiley-VCH GmbH.)

Anthropogenic atmospheric pollution and immune response regularly expose bacteria to toxic nitrogen oxides such as NO • and NO 2 . These reactive molecules can damage a wide variety of biomolecules such as DNA, proteins and lipids. Several components of the bacterial envelope are susceptible to be damaged by reactive nitrogen species. Furthermore, the hydrophobic core of the membranes favors the reactivity of nitrogen oxides with other molecules, making membranes an important factor in the chemistry of nitrosative stress. Since bacteria are often exposed to endogenous or exogenous nitrogen oxides, they have acquired protection mechanisms against the deleterious effects of these molecules. By exposing bacteria to gaseous NO 2 , this work aims to analyze the physiological effects of NO 2 on the cell envelope of the airborne bacterium Pseudomonas fluorescens MFAF76a and its potential adaptive responses. Electron microscopy showed that exposure to NO 2 leads to morphological alterations of the cell envelope. Furthermore, the proteomic profiling data revealed that these cell envelope alterations might be partly explained by modifications of the synthesis pathways of multiple cell envelope components, such as peptidoglycan, lipid A, and phospholipids. Together these results provide important insights into the potential adaptive responses to NO 2 exposure in P. fluorescens MFAF76a needing further investigations., (© 2022. The Author(s).)

Some removable medical devices such as catheters and cardiovascular biomaterials require antiadhesive properties towards both prokaryotic and eukaryotic cells in order to prevent the tissues from infections upon implantation and, from alteration upon removal. In order to inhibit cell adhesion, we developed ultrathin hydrated Layer-by-Layer (LbL) coatings composed of biocompatible polyelectrolytes, namely chondroitin sulfate A (CSA) and poly-l-lysine (PLL). The coatings were crosslinked with genipin (GnP), a natural and biocompatible crosslinking agent, to increase their resistance against environmental changes. In order to confer antibacterial activity to the coatings, we proceeded to the electrostatically-driven immobilization of nisin Z, an antimicrobial peptide (AMP) active against gram-positive bacteria. The nisin-enriched coatings had a significantly increased anti-proliferative impact on fibroblasts, as well as a strong contact-killing activity against Staphylococcus aureus in the short and long term., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Implanted biomaterials can be regarded in a cornerstone in the domain of bone surgery. Their surfaces are expected to fulfil two particular requirements: preventing the settlement and the development of bacteria, and stimulating bone cells in view to foster osseointegration. Therefore, a modern approach consists in the design of dual functional coatings with both antibacterial and osteogenic features. To this end, we developed ultrathin Layer-by-Layer (LbL) coatings composed of biocompatible polyelectrolytes, namely chondroitin sulfate A (CSA) and poly-l-lysine (PLL). The coatings were crosslinked with genipin (GnP), a natural and biocompatible crosslinking agent, to increase their resistance against environmental changes, and to confer them adequate mechanical properties with regards to bone cell behaviors. Antibacterial activity was obtained with nisin Z, an antimicrobial peptide (AMP), which is active against gram-positive bacteria. The coatings had a significant bactericidal impact upon Staphylococcus aureus, with fully maintained bone cell adhesion, proliferation and osteogenic differentiation., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Neuroblastoma is the third most common pediatric cancer composed of malignant immature cells that are usually treated pharmacologically by all trans-retinoic acid (ATRA) but sometimes, they can spontaneously differentiate into benign forms. In that context, biomimetic cell culture models are warranted tools as they can recapitulate many of the biochemical and biophysical cues of normal or pathological microenvironments. Inspired by that challenge, we developed a neuroblastoma culture system based on biomimetic LbL films of physiological biochemical composition and mechanical properties. For that, we used chondroitin sulfate A (CSA) and poly-L-lysine (PLL) that were assembled and mechanically tuned by crosslinking with genipin (GnP), a natural biocompatible crosslinker, in a relevant range of stiffness (30-160 kPa). We then assessed the adhesion, survival, motility, and differentiation of LAN-1 neuroblastoma cells. Remarkably, increasing the stiffness of the LbL films induced neuritogenesis that was strengthened by the combination with ATRA. These results highlight the crucial role of the mechanical cues of the neuroblastoma microenvironment since it can dramatically modulate the effect of pharmacologic drugs. In conclusion, our biomimetic platform offers a promising tool to help fundamental understanding and pharmacological screening of neuroblastoma differentiation and may assist the design of translational biomaterials to support neuronal regeneration. STATEMENT OF SIGNIFICANCE: Neuroblastoma is one of the most common pediatric tumor commonly treated by the administration of all-trans-retinoic acid (ATRA). Unfortunately, advanced neuroblastoma often develop ATRA resistance. Accordingly, in the field of pharmacological investigations on neuroblastoma, there is a tremendous need of physiologically relevant cell culture systems that can mimic normal or pathological extracellular matrices. In that context, we developed a promising matrix-like cell culture model that provides new insights on the crucial role of mechanical properties of the microenvironment upon the success of ATRA treatment on the neuroblastoma maturation. We were able to control adhesion, survival, motility, and differentiation of neuroblastoma cells. More broadly, we believe that our system will help the design of in vitro pharmacological screening strategy., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

In the present study, new polymer microspheres of amphotericin B (AmB) were prepared by a spray drying technique using cyclodextrin polymers (Poly-CD) to improve the solubility and dissolution of AmB, to prevent in vivo toxic AmB aggregations. Formulations were characterized through scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FT-IR), differential scanning calorimetry (DSC), thermal analysis, Raman spectroscopy, particle size, drug purity test and in vitro release studies. The analysis indicated that the chemical structure of AmB remained unchanged in the amorphous solid dispersion, but the structure was changed from crystalline to amorphous. AmB was completely release from such optimized formulations in dissolution media in 40 min. This work may contribute to a new generation of spherical amorphous solid dispersion using a cyclodextrin polymer, which has implications for the possibility of drug development for oral utilization or as powder aerosols for pulmonary administration.

This article reports the synthesis and functionalization of a novel CuO@SiO 2 -APTES@Ag 0 core-shell-shell material using a simple and low-cost process. The growth, design strategies and synthesis approach are the key factors for the development of CuO@SiO 2 -APTES@Ag 0 as efficient material with enhanced antibacterial activity. We investigated the morphology, surface charge, structure and stability of our new core-shell-shell by atomic force microscopy, scanning electron microscopy, energy dispersive X-ray, Fourier transform infrared and UV-visible spectroscopies, zeta potential measurements, and differential scanning calorimetry. The covalent surface grafting of APTES (3-(aminopropyl)triethoxysilane) onto CuO@SiO 2 involving electrostatic interactions was confirmed. Size measurements and Scanning electron images showed that both APTES grafting and SiO 2 /Ag shells dropped on the surface of CuO produced structural compaction. UV-Vis spectroscopy proved to be a fast and convenient way to optically detect SiO 2 shell on the surface of colloids. Additionally, the Ag-decorated CuO@SiO 2 -APTES surfaces were found to possess antibacterial activity and thermally more stable than undecorated surfaces. CuO@SiO 2 -APTES@Ag 0 core-shell had antibacterial properties against Gram-positive bacteria making it a promising candidate for antibacterial applications., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Thin films mimicking the structure and composition of the extra-cellular matrix (ECM) are potentially attractive as biomaterials for cell contacting applications. Layer-by-layer (LbL) assembly of a biological polycation, poly(l-lysine) (PLL), and a common ECM protein, fibronectin (Fn), was employed here to construct nanoscale, ECM mimicking films. Incremental film thickness and interfacial charge magnitude are observed to diminish with layer number, resulting in sub-linear film growth scaling and saturation after about 10 layers. Infrared spectroscopy and electron microscopy together reveal the formation of Fn containing aggregates, whose presence correlates with diminished charge reversal and suppressed LbL assembly. PLL-Fn films induce a significantly greater murine MC3T3-E1 pre-osteoblastic cell proliferation, while maintaining a much higher proportion of Fn in the molecular (as opposed to fibrillar) state, compared to a Fn monolayer, suggesting the enhanced Fn content of these ECM-mimicking films to significantly, and positively, affect cell behavior., (Copyright © 2017 Elsevier B.V. All rights reserved.)

We combined topographical and chemical surface modifications of Ti-6Al-4V (TA6V) to improve its osteogenic potential. By acid-etching, we first generated topomimetic surface features resembling, in size and roughness, bone cavities left by osteoclasts. Next, we coated these surfaces with biomimetic Layer-by-Layer films (LbL), composed of chondroitin sulfate A and poly-l-lysine that were mechanically tuned after a post-treatment with genipin. The structural impact of each surface processing step was thoroughly inspected. The desired nano/microrough topographies of TA6V were maintained upon LbL deposition. Whereas no significant promotion of adhesion and proliferation of MC3T3-E1 preosteoblasts were detected after independent or combined modifications of the topography and the chemical composition of the substrates, osteogenic maturation was promoted when both surface treatments were combined, as was evidenced by significant long-term matrix mineralization. The results open promising route toward improved osseointegration of titanium-based implants. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1988-2000, 2016., (© 2016 Wiley Periodicals, Inc.)

The design of biomimetic coatings capable of improving the osseointegration of bone biomaterials is a current challenge in the field of bone repair. Toward this end, layer-by-layer (LbL) films composed of natural components are suitable candidates. Chondroitin sulfate A (CSA), a natural glycosaminoglycan (GAG), was used as the polyanionic component because it promotes osteoblast maturation in vivo. In their native state, GAG-containing LbL films are generally cytophobic because of their low stiffness. To stiffen our CSA-based LbL films, genipin (GnP) was used as a natural cross-linking agent, which is much less cytotoxic than conventional chemical cross-linkers. GnP-cross-linked films display an original combination of microscale topography and tunable mechanical properties. Structural characterization was partly based on a novel donor/acceptor Förster resonance energy transfer (FRET) couple, namely, FITC/GnP, which is a promising approach for further inspection of any GnP-cross-linked system. GnP-cross-linked films significantly promote adhesion, proliferation, and early and late differentiation of preosteoblasts.

Bioactive implants intended for rapid, robust, and durable bone tissue regeneration are presented. The implants are based on nanofibrous 3D-scaffolds of bioresorbable poly-ϵ-caprolactone mimicking the fibrillar architecture of bone matrix. Layer-by-layer nanoimmobilization of the growth factor BMP-2 in association with chitosan (CHI) or poly-L-lysine over the nanofibers is described. The osteogenetic potential of the scaffolds coated with layers of CHI and BMP-2 is demonstrated in vitro, and in vivo in mouse calvaria, through enhanced osteopontin gene expression and calcium phosphate biomineralization. The therapeutic strategy described here contributes to the field of regenerative medicine, as it proposes a route toward efficient repair of bone defects at reduced risk and cost level., (© 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Implants triggering rapid, robust and durable tissue regeneration are needed to shorten recovery times and decrease risks of postoperative complications for patients. Here, we describe active living collagen implants with highly promising bone regenerative properties. Bioactivity of the implants is obtained through the protective and stabilizing layer-by-layer immobilization of a protein growth factor in association with a polysaccharide (chitosan), within the form of nanocontainers decorating the collagen nanofibers. All components of the implants are US FDA approved. From both in vitro and in vivo evaluations, the sophisticated strategy described here should enhance, at a reduced cost, the safety and efficacy of the therapeutic implants in terms of large bone defects repair compared with current simplistic approaches based on the soaking of the implants with protein growth factor.

Nanobiotechnology enables the emergence of entirely new classes of bioactive devices intended for targeted intracellular delivery for more efficacies and less toxicities. Among organic and inorganic approaches currently developed, controlled release from polymer matrices promises utmost clinical impact. Here, a unique nanotechnology strategy is used to entrap, protect, and stabilize therapeutic agents into polymer coatings acting as nanoreservoirs enrobing nanofibers of implantable membranes. Upon contact with cells, therapeutic agents become available through enzymatic degradation of the nanoreservoirs. As cells grow, divide, and infiltrate deeper into the porous membrane, they trigger slow and progressive release of therapeutic agents that, in turn, stimulate further cell proliferation. This constitutes the first instance of a smart living nanostructured hybrid membrane for regenerative medicine. The cell contact-dependent bioerodable nanoreservoirs described here will permit sustained release of drugs, genes, growth factors, etc., opening a general route to the design of sophisticated cell-therapy implants capable of robust and durable regeneration of a broad variety of tissues., (© 2011 American Chemical Society)

Layer-by-Layer (LbL) coatings are promising tools for the biofunctionalization of biomaterials, as they allow stress-free immobilization of proteins. Here, we explore the possibility to immobilize phosvitin, a highly phosphorylated protein viewed as a model of bone phosphoproteins and, as such, a potential promotive agent of surface-directed biomineralization, into biomimetic LbL architectures. Two immobilization protocols are attempted, first, using phosvitin as the polyanionic component of phosvitin/poly-(L-lysine) films and, second, adsorbing it onto preformed chondroitin sulfate/poly-(L-lysine) films. Surprisingly, it is neither possible to embed phosvitin as the constitutive polyanion of the LbL architectures nor to adsorb it atop preformed films. Instead, phosvitin triggers instant massive film disassembly. This unexpected, incidentally detected behavior constitutes the first example of destructive interactions between LbL films and a third polyelectrolyte, a fortiori a protein, which might open a route toward new stimuli-responsive films for biosensing or drug delivery applications. Interestingly, additional preliminary results still indicate a promotive effect of phosvitin-containing remnant films on calcium phosphate deposition., (© 2011 American Chemical Society)

Polyelectrolyte multilayers (PEM) are well established nanoarchitectures with numerous potential applications, in particular as biomaterial coatings. They may exhibit specific biological properties in terms of controlled cell activation or local drug delivery. Here, in a new approach for bone metastasis prevention, we employed poly-l-lysine covalently grafted with beta-cyclodextrin as a polycationic vector (PLL-CD) for the antitumor bisphosphonate drug risedronate (RIS). Molar ratio for maximum loading of the PLL-CD vector with RIS was determined by Raman microspectroscopy. The efficacy of RIS at inhibiting cancer cell invasion in vitro was strongly enhanced upon complexation, whatever PLL-CD:RIS complexes were in solution or embedded into PEM nanoarchitectures. Complexes in solution also clearly prevented cancer-induced bone metastasis in animals. Incorporation of the complexes into PEM nanoarchitectures covering bone implants appears of interest for in situ prevention of bone metastasis after ablation.

The prevention of nonspecific protein adsorption is a crucial prerequisite for many biomedical and biotechnological applications. Therefore, the design of robust and versatile methods conferring optimal protein-resistance properties to surfaces has become a challenging issue. Here we report the unexpected case of polycation-ending polyelectrolyte multilayers (PEM) that efficiently prevented the adsorption of a negatively charged model protein, glucose oxidase (GOX). PEM films were based on two typical weak poyelectrolytes: poly(allylamine hydrochloride) (PAH) and poly(acrylic acid) (PAA). No chemical modification of the polyelectrolytes was required and tunable GOX adsorption was possible by simply changing the buildup pH conditions. Protein-resistance properties are attributed to high film hydration becoming the predominant factor over electrostatic interactions. We explain this effect by oscillations of the internal PAA ionization state throughout the buildup, which results in an excess of carboxylic acid groups within the film. This excess acts as a reservoir of potential carboxylate groups compensating the outer PAH positive charges. Partial results indicated that the system was also resistant to the adsorption of a positively charged protein, lysozyme. Control of the internal ionization of weak polyelectrolyte multilayers might open a route toward simple tuning of protein adsorption. These results should help to rationalize the design of biomaterials, biosensors, or protein separation devices.

Recently [Jessel N, Schwinte P, Donohue R, Lavalle P, Boulmedais F, Darcy R, et al. Pyridylamino-beta-cyclodextrin as a molecular chaperone for lipopolysaccharide embedded in a multilayered polyelectrolyte architecture. Adv Funct Mater 2004;14:963-9], we demonstrated the biological activity of a lipopolysaccharide from Escherichia coli incorporated into layer-by-layer films made of poly (l-lysine) and poly (l-glutamic acid) and containing a polycationic beta-cyclodextrin (CD) with chaperone properties. Here we develop innovative architectures containing a complex made of a charged beta-cyclodextrin and a lipid A antagonist (LAA) as potential systems for local endotoxin antagonistic activity. We examine the biological activity of these architectures. The CD-LAA complex adsorbed on top, or embedded into the polyelectrolyte films keeps its LPS antagonistic activity on both murine and human macrophages for at least 24h.

We applied a colloidal force microscopy technique to measure the spreading and retraction forces generated by protrusions (pseudopodia) of vegetative amoeboid cells (Dictyostelium discoideum) adhering on soft tissue analogues composed of 2-mm thick hydrogels of hyaluronic acid exhibiting Young's moduli between 10 and 200 Pa. Local shear deformations of the polymer films evoked by magnetic tweezers and by cellular protrusions were determined by analyzing the deflections of colloidal beads randomly deposited on the surface of the polymer cushions, which enabled us to measure forces generated by advancing ("pushing" forces) and retracting ("pulling" forces) protrusions in a direct way. We found that the maximum amplitudes generated by the advancing protrusions (pushes) decrease with increasing stiffness of the HA substrate while the amplitudes of the retractions do not show such a dependence. The maximum forces transmitted by the advancing and retracting protrusions increase with increasing stiffness of the HA films (from 0.02 to 1 nN for the case of pushing). The protrusions spread or retract with constant velocities which are higher for retractions (100 nm s(-1)) than for spreadings (50 nm s(-1)) and are not significantly influenced by the substrate rigidity. We provide evidence that elastic equilibrium during protrusion formation and retraction is maintained by local elastic dipole fields generated at the rim of the protrusions. A model of protrusion force transmission by coupling of growing actin gel in the cytoplasm of the protrusions to cell surface receptors through talin clutches is proposed.

Surface modification by deposition of ordered protein systems constitutes one of the major objectives of bio-related chemistry and biotechnology. In this respect a concept has recently been reported aimed at fabricating multilayers by the consecutive adsorption of positively and negatively charged polyelectrolytes. We investigate the adsorption processes between polyelectrolyte multilayers and a series of positively and negatively charged proteins. The film buildup and adsorption experiments were followed by Scanning Angle Reflectometry (SAR). We find that proteins strongly interact with the polyelectrolyte film whatever the sign of the charge of both the multilayer and the protein. When charges of the multilayer and the protein are similar, one usually observes the formation of protein monolayers, which can become dense. We also show that when the protein and the multilayer become oppositely charged, the adsorbed amounts are usually larger and the formation of thick protein layers extending up to several times the largest dimension of the protein can be observed. Our results confirm that electrostatic interactions dominate protein/polyelectrolyte multilayer interactions.