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Supramolecular hydrogels emerge as a promising paradigm for sutureless wound management. However, their translation is still challenged by the insufficient mechanical robustness in the context of complex wounds in dynamic tissues. Herein, we report a tissue-adhesive supramolecular hydrogel membrane based on biocompatible precursors for dressing wounds in highly dynamic tissues, featuring robust mechanical resilience through programmable strain-adaptive entanglement among microdomains. Specifically, the hydrogels are synthesized by incorporating a long-chain polyurethane segment into a Schiff base-ligated short-chain oxidized cellulose/quaternized chitosan network via acylhydrazone bonding, which readily establishes interpenetrating entangled microdomains in dynamic cross-linked hydrogel matrices to enhance their tear and fatigue resistance against extreme mechanical stresses. After being placed onto dynamic tissues, the hydrogel dressing could efficiently absorb blood to achieve rapid hemostasis. Moreover, metal ions released from ruptured erythrocytes could be scavenged by the Schiff base linkers to form additional ionic bonds, which would trigger the cross-linking of the short-chain components and establish abundant crystalline microdomains, eventually leading to the in situ stiffening of the hydrogels to endure heavy mechanical loads. Benefiting from its hemostatic capacity and strain adaptable mechanical performance, this hydrogel wound dressing shows promise for the clinical management of various traumatic wounds.

Healing of large open wounds remains a major challenge in clinics due to the high risk of bacterial infection and slow healing rates, while excessive use of antibiotics would cause enhanced antibiotic resistance and reduced biocompatibility. Herein, we developed a multifunctional hydrogel dressing (GCNO) through embedding nitrosothiol-conjugated chitosan into a crosslinked gelatin methacrylate (GelMA) network via hydrogen bonding, which presented self-regulated nitric oxide (NO) release kinetics for temporally controlled bacterial elimination and wound repair. At early stages after implantation, the positively charged chitosan molecules of the GCNO hydrogel precursors and release of a high level of NO from the GCNO hydrogel demonstrated effective coordinated antimicrobial capability, thus preventing wound infection in the early stages of healing. While at later stages of wound healing, the hydrogel could sustainably release low levels of NO to stimulate the proliferation and migration of fibroblasts and endothelial cells, leading to accelerated angiogenesis and cell deposition at the wound area. GCNO hydrogels exhibited anti-bacterial and wound repair performance with excellent biocompatibility and biosafety. Overall, this antibiotic-free GCNO hydrogel could demonstrate self-adaptive NO release profiles to prevent bacterial infection in early stages of wound healing while accelerating skin regeneration at later stages, which may offer new insights for the clinical management of large open wounds in clinics.

Oxidative stress induced by reactive oxygen species (ROS) overproduction would hinder bone healing process at the interface of bone/implant, yet underlying mechanism remains to be explored. To endow titanium (Ti) substrates with antioxidant activity for enhanced bone formation, multilayered structure composing of chitosan-catechol (Chi-C), gelatin (Gel) and hydroxyapatite (HA) nanofibers was constructed on Ti substrates. Surface wettability and topography of multilayer coated Ti substrates were characterized by water contact angle measurement, scanning electron microscopy and atomic force microscopy, respectively. Chi-C containing multilayer on Ti surface effectively protected osteoblasts from ROS damage, which was revealed by high level of intracellular ROS scavenging activity and reduced oxidative damage on cellular level by regulating the expression of cell adhesion related genes (integrin αv, β3, CDH11 and CDH2). Moreover, it regulated the production of cell adhesive and anti-apoptotic related proteins (p-MYPT1, p-FAK, p-Akt and Bcl-2) and pro-apoptotic critical executioners (Bax and cleaved caspase 3). Beside, the composite multilayer of Chi-C/Gel/HA nanofibers on Ti substrates promoted osteoblasts differentiation, which was evidenced by high expression levels of alkaline phosphatase activity, collagen secretion, ECM mineralization and osteogenesis-related genes expression in vitro. The in vivo experiments of μ-CT analysis, push out test and histochemistry staining further confirmed that Chi-C multilayered implant had great potential for improved early bone healing. Overall, the study offers an effective strategy for the exploration of high quality Ti implants for orthopedic applications., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Orthopedic implants failures are generally related to poor osseointegration and/or bacterial infection in clinical application. Surface functionalization of an implant is one promising alternative for enhancing osseointegration and/or reducing bacterial infection, thus ensuring the long term survival of the implant. In this study, titanium (Ti) substrates were surface functionalized with a polydopamine (PDOP) film as an intermediate layer for post-immobilization of chitosan-lauric acid (Chi-LA) conjugate. Chi-LA conjugate was synthesized and characterized by Fourier transform infrared spectroscopy (FTIR) and hydrogen proton nuclear magnetic resonance (NMR) spectrometer, respectively. Lauric acid (LA), a natural saturated fatty acid, was used mainly due to its good antibacterial property. Scanning electron microscopy (SEM) and water contact angle measurements were employed to detect the morphology changes and surface wettability of Ti substrates. The results suggested that Chi-LA conjugate was successfully immobilized onto the surfaces of Ti substrates. In vitro tests confirmed that the cell adhesion, cell viability, intracellular alkaline phosphatase activity and mineralization capacity of osteoblasts were remarkably improved when cultured onto Chi-LA surface functionalized Ti substrates. Antibacterial assay against Staphylococcus aureus (S. aureus) and Pseudomonas aeruginosa (P. aeruginosa) showed that the Chi-LA modified Ti substrates efficiently inhibited the adhesion and growth of bacteria. Overall, this study developed a promising approach to fabricate functional Ti-based orthopedic implants, which could enhance the biological functions of osteoblasts and concurrently reduce bacteria adhesion., (Copyright © 2014 Elsevier B.V. All rights reserved.)

To develop Ti implants with potent antibacterial activity, a novel "sandwich-type" structure of sulfhydrylated chitosan (Chi-SH)/gelatin (Gel) polyelectrolyte multilayer films embedding silver (Ag) nanoparticles was coated onto titanium substrate using a spin-assisted layer-by-layer assembly technique. Ag ions would be enriched in the polyelectrolyte multilayer films via the specific interactions between Ag ions and -HS groups in Chi-HS, thus leading to the formation of Ag nanoparticles in situ by photo-catalytic reaction (ultraviolet irradiation). Contact angle measurement and field emission scanning electron microscopy equipped with energy dispersive X-ray spectroscopy were employed to monitor the construction of Ag-containing multilayer on titanium surface, respectively. The functional multilayered films on titanium substrate [Ti/PEI/(Gel/Chi-SH/Ag) n /Gel] could efficiently inhibit the growth and activity of Bacillus subtitles and Escherichia coli onto titanium surface. Moreover, studies in vitro confirmed that Ti substrates coating with functional multilayer films remained the biological functions of osteoblasts, which was reflected by cell morphology, cell viability and ALP activity measurements. This study provides a simple, versatile and generalized methodology to design functional titanium implants with good cyto-compatibility and antibacterial activity for potential clinical applications.

A new fabrication protocol is described to obtain heparin and chitosan conjugated magnetic nanocomposite as a blood purification material for removal of low-density lipoprotein (LDL) from blood plasma. The adsorbent could be easily separated with an external magnet for recyclable use since it had a magnetic core. The LDL level of plasma decreased by 67.3 % after hemoperfusion for 2 h. Moreover, the adsorbent could be recycled simply washing with NaCl solution. After eight cycles, the removal efficiency of the adsorbent was still above 50 %. The recyclable magnetic adsorbent had good blood compatibility due to the conjugation of heparin to the chitosan-coated magnetic nanocomposites. The fabricated magnetic adsorbent could be applied for LDL apheresis without side effects.

In this study, chitosan-atorvastatin (Chi-AT) conjugate was immobilized onto the surfaces of titanium substrates to reduce inflammation responses and improve cytocompatibility. Polydopamine film was initially formed onto the titanium surfaces as the intermediate layer for the successful immobilization of Chi-AT, which was confirmed by scanning electron microscopy, X-ray photoelectron spectroscopy, and contact angle measurements, respectively. Endothelial cells grown onto Chi-AT immobilized titanium substrates displayed significantly higher (p < 0.01) cell viability and statistically lower (p < 0.01) lactate dehydrogenase production than those of native titanium substrates (control) after culture for 4 days and 7 days, respectively. Furthermore, macrophages cells cultured onto Chi-AT immobilized titanium substrates demonstrated significantly lower (p < 0.01) production levels of nitric oxide, acid phosphatase, reactive oxygen species, pro-inflammatory cytokines of tumor necrosis factor α, and interleukin-1 β than those of controls. All results indicated that the immobilization of Chi-AT conjugate onto titanium substrates was beneficial for improving their cytocompatibility and inhibiting pro-inflammatory responses. The study thus presents an alternative to fabricate bio-functionalized titanium-based implants for further clinical application., (Copyright © 2012 Wiley Periodicals, Inc.)

To exploit titanium materials with anticancer and antibacterial properties, TiO(2) nanotubes arrays as nanoreservoirs for deposition of selenium were generated onto titanium substrates and then covered with chitosan layer. The deposition of selenium in TiO(2) nanotubes was performed with electrodeposition. The physical properties (surface morphologies, chemical compositions and wettability) of the substrates were characterized by field-emission scanning electron microscopy (FE-SEM) equipped with energy dispersive X-ray spectroscopy (EDS) and contact angle measurement, respectively. The growth behaviors of both healthy osteoblasts and cancerous osteoblasts on the substrates were investigated in vitro. The selenium-deposited and chitosan-coated TiO(2) nanotubes substrates (TiO(2) nanotubes-Se-Chi) demonstrated great potential for promoting the proliferation of healthy osteoblasts and inhibiting the growth of cancerous osteoblasts. Meanwhile, the TiO(2) nanotubes-Se-Chi substrates displayed a sustained release of selenium for 21 days. The antibacterial properties of the prepared substrates were evaluated with Escherichia coli (E. coli). The result showed that TiO(2) nanotubes-Se-Chi substrates had long term antibacterial capacity. The approach in this study provides an alternative to fabricate anticancer and antibacterial titanium-based implants for potential clinical application., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Chitosan (Chi) and poly (styrene sulfonate) (PSS) were employed to surface modify titanium thin film via electrostatic self-assembly (ESA) technique in order to improve its biocompatibility. The surface chemistry, wettability and surface topography of the coated films with different number of deposited layers were investigated by using X-ray photoelectron spectroscopy (XPS), water contact angle measurement and atomic force microscopy (AFM), respectively. The results indicated that a full surface coverage for the outmost layer was achieved at least after deposition of five layers, i.e., PEI/(PSS/Chi)2 on the titanium films. The formed multi-layered structure of PEI(PSS/Chi)x (x > or = 2) on the titanium film was stable in air at room temperature and in phosphate buffered solution (PBS) for at least 3 weeks. Cell proliferation, cell viability, DNA synthesis as well as differentiation function (alkaline phosphatase) of osteoblasts on chitosan-modified titanium film (PEI/(PSS/Chi)6) and control sample were investigated, respectively. Osteoblasts cultured on chitosan-modified titanium film displayed a higher proliferation tendency than that of control (p < 0.01). Cell viability, alkaline phosphatase as well as DNA synthesis measurements indicated that osteoblasts on chitosan-modified titanium films were greater (p < 0.01) than those for the control, respectively. These results suggest that surface modification of titanium film was successfully achieved via deposition of PEI/(PSS/Chi)x layers, which is useful to enhance the biocompatibility of the titanium film.

In an effort to surface engineering of poly(D,L-lactic acid) (PDLLA), layer-by-layer (LbL) self-assembly of chitosan (Chi) and deoxyribonucleic acid (DNA) were employed to build up multilayered films. The formation of multilayers was monitored by using Fourier transform infrared spectroscopy (FTIR), X-ray photoelectron spectroscopy (XPS), water contact-angle measurement, and atomic force microscopy (AFM), respectively. A full coverage of Chi/DNA pair film was formed only after the fifth sequential deposition (PEI/(DNA/Chi)2), which was revealed by contact-angle measurement. Surface chemistry and topography of multilayered films were directly related to the corresponding outmost layer component. Discernable island-like structures on PEI/(DNA/Chi)5/DNA layered PDLLA film was observed. Lysozyme-mediated multilayer degradation and DNA-releasing measurement suggested that DNA was gradually released into the incubation medium over a period of up to 32 h. The approach presented here may be exploited to develop controlled administration of functional DNA constructs from the surfaces of biomedical materials and devices in situ., ((c) 2007 Wiley Periodicals, Inc. J Biomed Mater Res, 2008.)

Biocompatibility and tissue regenerating capacity are essential for biomaterials that used in tissue engineering. The aim of this study was to histologically assess the tissue reactions and bone conductivities of surface modified three dimensional (3-D) poly (D, L-lactic acid) (PDLLA) scaffolds, which were coated with chitosan via a physical entrapment method. The native PDLLA scaffold was prepared via thermally induced phrase separation technique and was characterized by scanning electron microscopy (SEM) and differential scanning calorimetry (DSC). Osteocalcin assay, a method to evaluate the bone formation potential, has shown that the osteocalcin production in chitosan-modified 3-D PDLLA scaffold group was significantly higher (p < 0.05) than that of in control. The tissue reactions and bone conductivities between surface modified PDLLA and native PDLLA scaffolds were evaluated using a rabbit radialis defect model in vivo and compared at different implantation intervals (2, 4, 8 and 12 weeks). The histological results have shown a higher bone formation potential and better biocompatibility of chitosan-modified 3-D PDLLA scaffolds as compared with the control group scaffolds.

To improve the surface biocompatibility of titanium films, a layer-by-layer (LBL) self-assembly technique, based on the polyelectrolyte-mediated electrostatic adsorption of chitosan (Chi) and gelatin (Gel), was used leading to the formation of multilayers on the titanium thin film surfaces. The film growth was initialized by deposition of one layer of positively charged poly(ethylene imine) (PEI). Then the thin film was formed by the alternate deposition of negatively charged Gel and positively charged Chi utilizing electrostatic interactions. The LBL film growth was monitored by several techniques. The chemical composition, surface topography as well as wettability were investigated by using X-ray photoelectron spectroscopy (XPS), atomic force microscopy (AFM), confocal laser scanning microscopy (CLSM) and water contact angle measurement, respectively. Quantitative XPS analysis showed the alternative change of C/N ratio after four sequential cycles coating of Ti/PEI/Gel/Chi/Gel, which indicated the discrete layer structure of coatings. Uncoated titanium (control sample) displayed a smooth surface morphology (root mean square (RMS) roughness was around 2.5 nm). A full coverage of coating with Gel/Chi layers was achieved on the titanium surface only after the deposition layers of PEI/(Gel/Chi)2. The PEI/Gel/(Chi/Gel)3 layer displayed a rough surface morphology with a tree-like structure (RMS roughness is around 82 nm). These results showed that titanium films could be modified with Chi/Gel which may affect the biocompatibility of the modified titanium films. To confirm this hypothesis, cell proliferation and cell viability of osteoblasts on LBL-modified titanium films as well as control samples were investigated in vitro. The proliferation of osteoblasts on modified titanium films was found to be greater than that on control (p<0.05) after 1 and 7 days culture, respectively. Cell viability measurement showed that the Chi/Gel-modified films have higher cell viability (p<0.05) than the control. These data suggest that Chi/Gel were successfully employed to surface engineer titanium via LBL technique, and enhanced its cell biocompatibility. The approach presented here may be exploited for fabrication of titanium-based implant surfaces.

Abstract: The extracellular microenvironment plays a key role in the regulation of cellular behavior. To mimic the natural extracellular microenvironment, TiO2 nanotube (TNT) arrays as drug nanoreservoirs for loading of bone morphogenetic protein 2 (BMP2) were constructed on titanium substrates and then covered with multilayered coatings of gelatin/chitosan (Gel/Chi) for controlled drug release. The multilayered coatings were constructed via a spin-assisted layer-by-layer assembly technique. The successful fabrication of this system was monitored by field emission scanning electron microscopy, atomic force microscopy, X-ray photoelectron spectroscopy and contact angle measurements. Multilayered coating with Gel/Chi retained the drug bioactivity and release properties, which were revealed by superoxide dismutase activity measurement. In addition, cytoskeleton observation and wound healing assay confirmed that BMP2-loaded and multilayer-coated TNT arrays were able to stimulate motogenic responses of mesenchymal stem cells (MSCs). More importantly, the system demonstrated that it was capable of promoting the osteoblastic differentiation of MSCs. This study may have potential impact on the development of bone implants for enhanced bone osseointegration. [Copyright &y& Elsevier]

Abstract: In this work, multilayered and gene-functionalized titanium films composed of chitosan (Chi) and plasmid DNA (pEGFP–hBMP2, pGB) were employed to investigate the surface mediated in situ differentiation of mesenchymal stem cells (MSCs). The Chi/pGB multilayered structures were fabricated by layer-by-layer (LbL) assembly technique and degraded to release plasmid DNA complexes depending on bilayer numbers over 7 days. Therefore, the differentiation behaviors of MSCs cultured onto Chi/pGB multilayered titanium films surface were investigated. Chi/pGB LbL-modified titanium films show significant higher (p<0.01) transfection efficiency than those of other groups transfected by lipofectamine 2000 regarding the expression of green fluorescent protein (GFP). Reverse transcription-polymerase chain reaction (RT-PCR) assay revealed that MSCs adhered onto Chi/pGB LbL-modified titanium films could still express hBMP2 mRNA over 7 days culture. Compared with control groups, MSCs cultured onto Chi/pGB LbL-modified titanium films display significantly higher (p<0.01 or p<0.05) production levels of alkaline phosphatase (ALP) and osteocalcin over 7 days and 14 days culture, respectively. These results demonstrate that Chi/pGB LbL-modified titanium films are beneficial for sustained in situ inducing osteoprogenitor cells to differentiate into mature osteoblasts over long time. The approach presented here has potential applications in the development of gene-stimulating biomaterials and implant technology. [Copyright &y& Elsevier]

Poly(D,L-lactic acid) (PDLLA) was modified with alkylated chitosan (N-butyl chitosan and N-cetyl chitosan), and the effects of modified films on the functions of rat osteoblasts were investigated. The characteristics of surfaces(both modified and control) were examined by water contact angle measurement and electron spectroscopy for chemical analysis (ESCA). Cell morphologies on these surfaces were taken using scanning electron microscopy (SEM). Cell attachment and proliferation were used to assess cell behavior on modified surface and control. MTT assay was used to determined cell viability, and alkaline phosphatase (ALP) activity was taken to evaluate differentiated cell function. Compared with the untreated films, no significant difference in cell attachment of osteoblasts was found on the modified films at a period of 8 h (p > 0.05). However, cell proliferation of N-butyl chitosan rather than N-cetyl chitosan modified PDLLA films was significantly higher than that found on control one (p < 0.05) at the end of the 4th and 7th days. The cell viability of osteoblasts on N-butyl chitosan modified PDLLA films were found higher than that on control (p < 0.05). These results suggested that N-butyl chitosan contributed greater than N-cetyl chitosan when used to modify PDLLA films for improving its biocompatibility. [ABSTRACT FROM AUTHOR]

In this study, the functions of rat osteoblasts on o-carboxymethyl chitosan-modified poly(D,L-lactic acid) (PDLLA) films were investigated in vitro. The surface characterization was measured by contact angle and electron spectroscopy for chemical analysis (ESCA). Cell adhesion and proliferation were used to assess cell behavior on the modified surface and control. The MTT assay was used to determined cell viability and alkaline phosphatase (ALP) activity was performed to evaluate differentiated cell function. Compared to the control films, cell adhesion of osteoblasts on o-carboxymethyl chitosan-modified PDLLA films was significantly higher (p < 0.05) after 6 and 8 h culture, and osteoblast proliferation was also significantly hlgher (p < 0.01) between 4 and 7 days. The MTT assay suggested cell viability of osteoblasts cultured on o-carboxymethyl chitosan modified PDLLA films was significantly greater (p < 0.05) than that seeded on control one, and the ALP activity of cells cultured on modified PDLLA films was significantly higher (p < 0.01) than that found on control. These results give the first evidence that o-carboxymethyl chitosan could be used to modify PDLLA surface for improving biocompatibility. [ABSTRACT FROM AUTHOR]

In this study, a type of bacteria enzyme-triggered antibacterial surface with a controlled release of Ag ions was developed. Firstly, chitosan-silver nanocomposites (Chi@Ag NPs) were in situ synthesized via using ascorbic acid as reducing agent. Chi@Ag NPs were characterized by transmission electron microscopy, ultraviolet-visible spectroscopy, X-ray diffraction and X-ray photoelectron spectroscopy. Subsequently, Chi@Ag NPs and hyaluronic acid (HA) were used to fabricate antibacterial composite coating via Layer-by-Layer (LBL) self-assembly method. The successful construction of Chi@Ag NPs/HA composite coating was confirmed by scanning electron microscopy, energy dispersive spectroscopy and contact angle measurements, respectively. Then, the amount of released Ag ion was analyzed by inductively coupled plasma atomic emission spectrometry, which demonstrated that the release of Ag ions from the surface could be triggered by enzyme (e.g. hyaluronidase). A series of antibacterial tests in vitro, including zone of inhibition test, bacterial viability assay, antibacterial rate measurement and bacteria adhesion observation, demonstrated that the enzyme-responsive surface could inhibit the growth of bacteria. On the whole, this study provides an alternative approach for the fabrication of antibacterial surfaces on synthetic materials in various fields with the minimal side effects on surrounding environment and human body. [ABSTRACT FROM AUTHOR]

The insufficient osseointegration and bacterial infection of titanium and its alloys remain the key challenges in their clinic applications, which may result in failure implantation. To improve osteogenetic and antibacterial properties, TiO 2 nanotube arrays were fabricated on titanium substrates for loading of antibacterial drug. Then, TiO 2 nanotube arrays were covered with chitosan/sodium alginate multilayer films. The successful construction of this system was verified via scanning electron microscopy and contact angle measurement. The cytocompatibility evaluation in vitro , including cytoskeleton observation, cell viability measurement, and alkaline phosphatase activity assay, confirmed that the present system was capable of accelerating the growth of osteoblasts. In addition, bacterial adhesion and viability assay verified that treated Ti substrates were capable of reducing the adhesion of bacteria. This study may provide an alternative to develop titanium-based implants for enhanced bone osseointegration and reduced bacterial infection. [ABSTRACT FROM AUTHOR]

To mimic extracellular microenvironment of endothelial cell, a bioactive multilayered structure of gelatin/chitosan pair, embedding with vascular endothelial growth factor (VEGF), was constructed onto NiTi alloy substrate surface via a layer-by-layer assembly technique. The successful fabrication of the multilayered structure was demonstrated by scanning electron microscopy, atomic force microscopy, contact angle measurement, attenuated total reflection-fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy, respectively. The growth behaviors of endothelial cells on various NiTi alloy substrates were investigated in vitro. Cytoskeleton observation, MTT assay, and wound healing assay proved that the VEGF-embedded multilayer structure positively stimulated adhesion, proliferation and motogenic responses of endothelial cells. More importantly, the present system promoted the nitric oxide production of endothelial cells. The approach affords an alternative to construct extracellular microenvironment for improving surface endothelialization of a cardiovascular implant. [ABSTRACT FROM AUTHOR]

Abstract: To enhance corrosion resistance of WE43 magnesium alloy, chitosan (CHI) and poly (styrene sulfonate) (PSS) polyelectrolyte multilayers were fabricated on micro-arc oxidation (MAO) treated magnesium alloy substrate via layer-by-layer (LBL) self-assembly technique. Surface morphology and chemical composition of the MAO/LBL composite coating were investigated by scanning electron microscopy (SEM), atomic force microscope (AFM) and X-ray photoelectron spectroscopy (XPS), respectively. Potentiodynamic polarization test and hydrogen evolution measurement showed that the formed MAO/LBL coating significantly enhanced corrosion resistance of WE43 magnesium alloy in simulated body fluid (SBF). The approach presented here affords an effective alternative for surface modification of magnesium-based materials to meet the requirements of biomaterials applications. [Copyright &y& Elsevier]