Anti-inflammatory Surface Coatings Based on Polyelectrolyte Multilayers of Heparin and Polycationic Nanoparticles of Naproxen-Bearing Polymeric Drugs.

Immune response to biomaterials can produce chronic inflammation and fibrosis leading to implant failure, which is related to the surface properties of the biomaterials. This work describes the preparation and characterization of polyelectrolyte multilayer (PEM) coatings that combine the anti-inflammatory activity of heparin as polyanion with the potential release of Naproxen, a nonsteroidal anti-inflammatory drug from polymeric nanoparticles (NP) with cationic surface charge. The polyelectrolyte multilayers were characterized by physical methods to estimate multilayer growth, thickness, zeta potential, and topography. It was found that multilayers with NP had negative zeta potentials and expressed a viscoelastic behavior, while studies of topography showed that nanoparticles formed continuous surface coatings. THP-1-derived macrophages were used to study short-term anti-inflammatory activity (time scale 48 h), showing that PEM that contained heparin reduced cell adhesion and IL1-ß secretion, when compared to those with polystyrenesulfonate, used as alternative polyanion in multilayer formation. On the other hand, the presence of NP in PEM was related to a reduced foreign body giant cell formation after 15 days, when compared to PEM that contained chitosan as alternative polycation, which suggests a long-term anti-inflammatory effect of Naproxen-containing nanoparticles. It was also shown that macrophages were able to take up NP from multilayers, which indicates a release of Naproxen by digestion of NP in the lysosomal compartment. These findings indicate that surface coatings composed of heparin and Naproxen-based NP on implants such as biosensors have the potential to attenuate foreign body reaction after implantation, which may improve the long-term functionality of implants.

Theoretical analysis of single-cycle self-compression of near infrared pulses using high-spatial modes in capillary fibers.

Soliton self-compression is demonstrated during the propagation of high spatial modes in hollow core fibers in the near-infrared spectral region, taking advantage of their negative dispersion response. We have found that there is always an optimum spatial mode to observe this phenomenon, compressing the pulses down to the single-cycle regime without needing any external compression device and with a consequent increase in the output peak power. Our result is relevant for any ultrashort laser application in which few- or single-cycle pulses are crucial.

Theoretical analysis of single-cycle self-compression of near infrared pulses using high-spatial modes in capillary fibers: erratum.

The first equation in [Opt. Express26, 6345 (2018)] contains an error and is corrected in this erratum.

Strategies for achieving intense single-cycle pulses with in-line post-compression setups.

Intense few- and single-cycle pulses are powerful tools in different fields of science Today, third- and higher-order terms in the remnant spectral phase of the pulses remain a major obstacle for obtaining high-quality few- and single-cycle pulses from in-line post-compression setups. In this Letter, we show how input pulse shaping can successfully be applied to standard post-compression setups to minimize the occurrence of high-order phase components during nonlinear propagation and to directly obtain pulses with durations down to 3 fs. Furthermore, by combining this pulse shaping of the input pulse with new-generation broadband chirped mirrors and material addition for remnant third-order phase correction, pulses down to 2.2 fs duration have been measured.

Nonperturbative Twist in the Generation of Extreme-Ultraviolet Vortex Beams.

High-order harmonic generation (HHG) has been recently proven to produce extreme-ultraviolet (XUV) vortices from the nonlinear conversion of infrared twisted beams. Previous works have demonstrated a linear scaling law of the vortex charge with the harmonic order. We demonstrate that this simple law hides an unexpectedly rich scenario for the buildup of orbital angular momentum (OAM) due to the nonperturbative behavior of HHG. The complexity of these twisted XUV beams appears only when HHG is driven by nonpure vortex modes, where the XUV OAM content is dramatically increased. We explore the underlying mechanisms for this diversity and derive a general conservation rule for the nonperturbative OAM buildup. The simple scaling found in previous works corresponds to the collapse of this scenario for the particular case of pure (single-mode) OAM driving fields.

Anticancer and antiangiogenic activity of surfactant-free nanoparticles based on self-assembled polymeric derivatives of vitamin E: structure-activity relationship.

α-Tocopheryl succinate (α-TOS) is a well-known mitochondrially targeted anticancer compound, however, it is highly hydrophobic and toxic. In order to improve its activity and reduce its toxicity, new surfactant-free biologically active nanoparticles (NP) were synthesized. A methacrylic derivative of α-TOS (MTOS) was prepared and incorporated in amphiphilic pseudoblock copolymers when copolymerized with N-vinylpyrrolidone (VP) by free radical polymerization (poly(VP-co-MTOS)). The selected poly(VP-co-MTOS) copolymers formed surfactant-free NP by nanoprecipitation with sizes between 96 and 220 nm and narrow size distribution, and the in vitro biological activity was tested. In order to understand the structure-activity relationship three other methacrylic monomers were synthesized and characterized: MVE did not have the succinate group, SPHY did not have the chromanol ring, and MPHY did not have both the succinate group and the chromanol ring. The corresponding families of copolymers (poly(VP-co-MVE), poly(VP-co-SPHY), and poly(VP-co-MPHY)) were synthesized and characterized, and their biological activity was compared to poly(VP-co-MTOS). Both poly(VP-co-MTOS) and poly(VP-co-MVE) presented triple action: reduced cell viability of cancer cells with little or no harm to normal cells (anticancer), reduced viability of proliferating endothelial cells with little or no harm to quiescent endothelial cells (antiangiogenic), and efficiently encapsulated hydrophobic molecules (nanocarrier). The anticancer and antiangiogenic activity of the synthesized copolymers is demonstrated as the active compound (vitamin E or α-tocopheryl succinate) do not need to be cleaved to trigger the biological action targeting ubiquinone binding sites of complex II. Poly(VP-co-SPHY) and poly(VP-co-MPHY) also formed surfactant-free NP that were also endocyted by the assayed cells; however, these NP did not selectively reduce cell viability of cancer cells. Therefore, the chromanol ring of the vitamin E analogues has an important role in the biological activity of the copolymers. Moreover, when succinate moiety is substituted and vitamin E is directly linked to the macromolecular chain through an ester bond, the biological activity is maintained.

Chitosan-gelatin biopolymers as carrier substrata for limbal epithelial stem cells.

The aim of this work was to evaluate semi-synthetic biopolymers based on chitosan (CH) and gelatin (G) as potential in vitro carrier substrata for human limbal epithelial cells (hLECs). To that end, human corneal epithelial cells (HCE) were cultured onto different CH-G membranes. None of the polymers were cytotoxic and cell proliferation was higher when CH was functionalized with G. Expression levels of corneal epithelial markers (K3, K12, E-caherin, desmoplakin, and zonula occludens (ZO)-1) were better maintained in HCE cells grown on CH-G 20:80 membranes than other proportions. Consequently, CH-G 20:80 was chosen for the subsequent expansion of hLECs. Cells derived from limbal explants were successfully expanded on CH-G 20:80 membranes using a culture medium lacking components of non-human animal origin. The expression levels found for corneal (K3 and K12) and limbal epithelial stem cells (K15) specific markers were similar to or higher than those found in limbal cells grown onto the control substratum. Our results demonstrate that CH-G 20:80 membranes are suitable for the expansion and maintenance of stem cells derived from the limbal niche. These results strongly support the use of polymers as alternative substrata for the transplantation of cultivated limbal cells onto the ocular surface.

Necklace-structured high-harmonic generation for low-divergence, soft x-ray harmonic combs with tunable line spacing.

The extreme nonlinear optical process of high-harmonic generation (HHG) makes it possible to map the properties of a laser beam onto a radiating electron wave function and, in turn, onto the emitted x-ray light. Bright HHG beams typically emerge from a longitudinal phased distribution of atomic-scale quantum antennae. Here, we form a transverse necklace-shaped phased array of linearly polarized HHG emitters, where orbital angular momentum conservation allows us to tune the line spacing and divergence properties of extreme ultraviolet and soft x-ray high-harmonic combs. The on-axis HHG emission has extremely low divergence, well below that obtained when using Gaussian driving beams, which further decreases with harmonic order. This work provides a new degree of freedom for the design of harmonic combs-particularly in the soft x-ray regime, where very limited options are available. Such harmonic beams can enable more sensitive probes of the fastest correlated charge and spin dynamics in molecules, nanoparticles, and materials.

Enhancement of filamentation postcompression by astigmatic focusing.

The energy scaling up of pulse postcompression is still an open issue. In this work we analyze the use of astigmatic focusing to improve the output pulses in a filamentation based postcompression setup. Unlike spherical conditions, astigmatic focusing enhances the output energy and the spectral broadening of the filament. This is due to the increase of critical power, allowing a considerable improvement of the postcompression energy and stability in a simple way. We demonstrated compression from FWHM 100 fs, 10 nm, 3 mJ input pulses to 13 fs, 142 nm, near 1 mJ pulses.

Antimitogenic polymer drugs based on AMPS: monomer distribution-bioactivity relationship of water-soluble macromolecules.

A number of polysulfonated molecules have demonstrated their interaction with fibroblast growth factor (FGF), hampering their binding to its receptors (low affinity heparan sulfate proteoglycans (HSPG) and high affinity tyrosine kinase FGF receptors) and inhibiting the intracellular signaling and mitogenic response in cultured endothelial cells. The aim of this work was the synthesis and characterization of new copolymers based on 2-acrylamido-2-methylpropane sulfonic acid (AMPS) with antiproliferative activity for antitumoral applications. N-Vinylpyrrolidone (VP) or butyl acrylate (BA) was copolymerized with the sulfonated monomer to obtain macromolecules with different hydrophilic/hydrophobic balance and distribution of the sulfonated groups within the macromolecules. In vitro cell culture proliferative assays showed that monomer distribution affected the inhibition of the proliferative action of FGF. Reactivity ratios of the systems were determined following the free radical copolymerization by in situ (1)H NMR, and the correlation of the monomer sequence distribution with the bioactivity is discussed.

Chitosan hydrogels functionalized with either unfractionated heparin or bemiparin improve diabetic wound healing.

Diabetes mellitus causes severe impairment in the cutaneous wound healing process, which has led to extensive research striving to establish new treatments. In this work, we describe the effects of chitosan hydrogels functionalized with either unfractionated heparin or bemiparin (a low molecular weight heparin, LMWH) as topical treatments in an experimental diabetic wound healing model. Although wound morphometry showed similar values at the end of the study, microscopic analyses revealed impaired healing in diabetic animals in terms of inflammation and tissue formation. However, both types of loaded hydrogels accelerated inflammation resolution and improved the epithelialization process, while showing a neodermal thickness similar to that of nondiabetic animals. Immunohistochemistry analyses revealed an intermediate response in macrophage evolution between diabetic and nondiabetic controls in the treated groups, as well as enhanced collagenization and myofibroblast progression patterns. However, these changes were not accompanied by differences among groups in collagen I, III and TGF-ß1 gene expression. Functionalized hydrogels improved diabetes-associated impaired wound healing, thus promoting the progression of the process and inducing the formation of high-quality cicatricial tissue. Although the beneficial healing effect observed after topical treatment with chitosan hydrogels loaded with bemiparin or unfractionated heparin was similar, the chitosan hydrogel loaded with bemiparin is the preferred choice as it exhibited high-quality tissue in the neoformed dermal tissue.

Evaluation of Glycerylphytate Crosslinked Semi- and Interpenetrated Polymer Membranes of Hyaluronic Acid and Chitosan for Tissue Engineering.

In the present study, semi- and interpenetrated polymer network (IPN) systems based on hyaluronic acid (HA) and chitosan using ionic crosslinking of chitosan with a bioactive crosslinker, glycerylphytate (G1Phy), and UV irradiation of methacrylate were developed, characterized and evaluated as potential supports for tissue engineering. Semi- and IPN systems showed significant differences between them regarding composition, morphology, and mechanical properties after physicochemical characterization. Dual crosslinking process of IPN systems enhanced HA retention and mechanical properties, providing also flatter and denser surfaces in comparison to semi-IPN membranes. The biological performance was evaluated on primary human mesenchymal stem cells (hMSCs) and the systems revealed no cytotoxic effect. The excellent biocompatibility of the systems was demonstrated by large spreading areas of hMSCs on hydrogel membrane surfaces. Cell proliferation increased over time for all the systems, being significantly enhanced in the semi-IPN, which suggested that these polymeric membranes could be proposed as an effective promoter system of tissue repair. In this sense, the developed crosslinked biomimetic and biodegradable membranes can provide a stable and amenable environment for hMSCs support and growth with potential applications in the biomedical field.

Innovative approach for producing injectable, biodegradable materials using chitooligosaccharides and green chemistry.

Although there are a number of injectable biomaterials currently under development, they present some drawbacks such as being based on synthetic polymers, needing toxic or aggressive synthesis procedures or using raw materials with low availability and/or high production costs. Having this in mind, a novel injectable biomaterial using chitooligosaccharides as starting materials was developed. This system uses a widely available and cheap polymer from marine biomass (chitosan), which can be turned into an injectable material by water-based and ecologically friendly reactions. Chitooligosaccharides were functionalized with methacrylic groups, to allow in situ cross-linking. The degree of substitution, as determined by (1)H NMR, varied between 5 and 50%. The system was characterized in terms of kinetics of gel formation, rheology, degradation behavior and in vitro cytotoxicity. The gelation time could be easily tailored between 1.5 and 60 min by changing the conditions of the methacrylation reaction, and the final gel presented rheological properties typical of strong gels, that is, shear stresses in the kPa range. The cross-linked gel was degradable and nontoxic, presenting indeed an interesting cytokinetic effect. Injectable materials based on chitooligosaccharides are, therefore, an innovative system combining adequate biological performance, ease of preparation, and an ecologically friendly concept of production.

Glycerylphytate as an ionic crosslinker for 3D printing of multi-layered scaffolds with improved shape fidelity and biological features.

The fabrication of intricate and long-term stable 3D polymeric scaffolds by a 3D printing technique is still a challenge. In the biomedical field, hydrogel materials are very frequently used because of their excellent biocompatibility and biodegradability, however the improvement of their processability and mechanical properties is still required. This paper reports the fabrication of dual crosslinked 3D scaffolds using a low concentrated (<10 wt%) ink of gelatin methacryloyl (GelMA)/chitosan and a novel crosslinking agent, glycerylphytate (G1Phy) to overcome the current limitations in the 3D printing field using hydrogels. The applied methodology consisted of a first ultraviolet light (UV) photopolymerization followed by a post-printing ionic crosslinking treatment with G1Phy. This crosslinker provides a robust framework and avoids the necessity of neutralization with strong bases. The blend ink showed shear-thinning behavior and excellent printability in the form of a straight and homogeneous filament. UV curing was undertaken simultaneously to 3D deposition, which enhanced precision and shape fidelity (resolution ≈150 µm), and prevented the collapse of the subsequent printed layers (up to 28 layers). In the second step, the novel G1Phy ionic crosslinker agent provided swelling and long term stability properties to the 3D scaffolds. The multi-layered printed scaffolds were mechanically stable under physiological conditions for at least one month. Preliminary in vitro assays using L929 fibroblasts showed very promising results in terms of adhesion, spreading, and proliferation in comparison to other phosphate-based traditional crosslinkers (i.e. TPP). We envision that the proposed combination of the blend ink and 3D printing approach can have widespread applications in the regeneration of soft tissues.

Poly(N-isopropylacrylamide) surface-grafted chitosan membranes as a new substrate for cell sheet engineering and manipulation.

The immobilization of poly(N-isopropylacrylamide) (PNIPAAm) on chitosan membranes was performed in order to render membranes with thermo-responsive surface properties. The aim was to create membranes suitable for cell culture and in which confluent cell sheets can be recovered by simply lowering the temperature. The chitosan membranes were immersed in a solution of the monomer that was polymerized via radical initiation. The composition of the polymerization reaction solvent, which was a mixture of a chitosan non-solvent (isopropanol) and a solvent (water), provided a tight control over the chitosan membranes swelling capability. The different swelling ratio, obtained at different solvent composition of the reaction mixture, drives simultaneously the monomer solubility and diffusion into the polymeric matrix, the polymerization reaction rate, as well as the eventual chain transfer to the side substituents of the pyranosyl groups of chitosan. A combined analysis of the modified membranes chemistry by proton nuclear magnetic resonance ((1)H-NMR), Fourier transform spectroscopy with attenuated total reflection (FTIR-ATR) and X-ray photoelectron spectroscopy (XPS) showed that it was possible to control the chitosan modification yield and depth in the solvent composition range between 75% and 100% of isopropanol. Plasma treatment was also applied to the original chitosan membranes in order to improve cell adhesion and proliferation. Chitosan membranes, which had been previously subjected to oxygen plasma treatment, were then modified by means of the previously described methodology. A human fetal lung fibroblast cell line was cultured until confluence on the plasma-treated thermo-responsive chitosan membranes and cell sheets were harvested lowering the temperature.

Controlling the polarization and vortex charge of attosecond high-harmonic beams via simultaneous spin-orbit momentum conservation.

Optical interactions are governed by both spin and angular momentum conservation laws, which serve as a tool for controlling light-matter interactions or elucidating electron dynamics and structure of complex systems. Here, we uncover a form of simultaneous spin and orbital angular momentum conservation and show, theoretically and experimentally, that this phenomenon allows for unprecedented control over the divergence and polarization of extreme-ultraviolet vortex beams. High harmonics with spin and orbital angular momenta are produced, opening a novel regime of angular momentum conservation that allows for manipulation of the polarization of attosecond pulses-from linear to circular-and for the generation of circularly polarized vortices with tailored orbital angular momentum, including harmonic vortices with the same topological charge as the driving laser beam. Our work paves the way to ultrafast studies of chiral systems using high-harmonic beams with designer spin and orbital angular momentum.

Influence of the activator in an acrylic bone cement on an array of cement properties.

In all but one of the acrylic bone cement brands used in cemented arthroplasties, N,N-dimethyl-4-toluidine (DMPT) serves as the activator of the polymerization reaction. However, many concerns have been raised about this activator, all related to its toxicity. Thus, various workers have assessed a number of alternative activators, with two examples being N,N-dimethylamino-4-benzyl laurate (DMAL) and N,N-dimethylamino-4-benzyl oleate (DMAO). The results of limited characterization of cements that contain DMAL or DMAO have been reported in the literature. The present work is a comprehensive comparison of cements that contain one of these three activators, in which the values of a large array of their properties were determined. These properties range from the setting time and maximum exotherm temperature of the curing cement to the variation of the loss elastic modulus of the cured cement with frequency of the applied indenting force in dynamic nanoindentation tests. The present results, taken in conjunction with those presented in previous reports by the present authors and co-workers on other properties of these cements, indicate that both DMAL and DMPT are suitable alternatives to DMPT.

Development of wollastonite-poly(ethylmethacrylate co-vinylpyrrolidone) based materials for multifunctional devices.

The manufacturing of a composite made of a synthetic bioactive ceramic, pseudowollastonite (psW), and a bioresorbable copolymer ethylmethacrylate-vinylpyrrolidone (EMA/VP) is presented in this article. psW porous blocks were produced by dipping an open porous polyurethane foam in a psW containing slurry. A 40/60 wt % EMA/VP monomers mixture was poured on the blocks, and free radical polymerization initiated by azobis(isobutyronitrile) at 50 degrees C. Disks of 1 mm height were obtained by cutting the composite with a diamond saw, and bioresorption and bioactivity of the specimens were tested by immersion of the disks into SBF. A ceramic/polymer weight ratio of 72/28, greater than the usually achievable ratio by polymeric solidification of slurries of monomers charged with a powdered solid component, has been obtained. The system is bioactive and does not change the pH of the medium during the degradation test.

Chitosan nanoparticles for combined drug delivery and magnetic hyperthermia: From preparation to in vitro studies.

Chitosan nanoparticles (CSNPs) ionically crosslinked with tripolyphosphate salts (TPP) were employed as nanocarriers in combined drug delivery and magnetic hyperthermia (MH) therapy. To that aim, three different ferrofluid concentrations and a constant 5-fluorouracil (5-FU) concentration were efficiently encapsulated to yield magnetic CSNPs with core-shell morphology. In vitro experiments using normal cells, fibroblasts (FHB) and cancer cells, human glioblastoma A-172, showed that CSNPs presented a dose-dependent cytotoxicity and that they were successfully uptaken into both cell lines. The application of a MH treatment in A-172 cells resulted in a cell viability of 67-75% whereas no significant reduction of cell viability was observed for FHB. However, the A-172 cells showed re-growth populations 4h after the application of the MH treatment when CSNPs were loaded only with ferrofluid. Finally, a combined effect of MH and 5-FU release was observed with the application of a second MH treatment for CSNPs exhibiting a lower amount of released 5-FU. This result demonstrates the potential of CSNPs for the improvement of MH therapies.

Injectable and self-curing composites of acrylic/bioactive glass and drug systems. A histomorphometric analysis of the behaviour in rabbits.

Injectable self-curing systems based on PMMA, phosphate-free bioactive glasses and the drug fosfosal, a phosphate derivative of salicylic acid with analgesic and moderate anti-inflammatory properties, have been tested in vivo to evaluate their biocompatibility. The model consisted of the injection of dough of cement into a defect created in the femur of rabbits, and the cure of the cement in situ after implantation. The biological response was studied in the short and long terms by macroscopic, radiological and histopathological examination, and quantitatively by histomorphometric and statistical analysis considering the most representative variables at the bone-cement interface: cement, bone marrow, newly formed bone and connective tissue. All bioactive formulations presented resorption of the cement at the end of the experiment in contrast to the control of PMMA, due to the presence of resorbable components. The presence or absence of the phosphate group added by the drug fosfosal influenced mainly on the new bone formation process. The cement formulated with bioactive glasses and in absence of fosfosal produced the maximum amount of neoformed bone at 2 weeks, and then it resorbed at 4 weeks to give a higher amount of neoformed bone at the end of the experiment, compared with the formulation containing only fosfosal. The presence of fosfosal and bioactive glass together affected the ossification process strongly. The osseous tissue was produced more gradually but it continuously increased giving rise to a more stable bone at the end of the experiment.