Macrophage Activation in the Synovium of Healthy and Osteoarthritic Equine Joints.

Synovitis is a major component of osteoarthritis and is driven primarily by macrophages. Synovial macrophages are crucial for joint homeostasis (M2-like phenotype), but induce inflammation (M1-like) when regulatory functions become overwhelmed. Macrophage phenotypes in synovium from osteoarthritic and healthy joints are poorly characterized; however, comparative knowledge of their phenotypes during health and disease is paramount for developing targeted treatments. This study compared patterns of macrophage activation in healthy and osteoarthritic equine synovium and correlated histology with cytokine/chemokine profiles in synovial fluid. Synovial histology and immunohistochemistry for M1-like (CD86), M2-like (CD206, IL-10), and pan macrophage (CD14) markers were performed on biopsies from 29 healthy and 26 osteoarthritic equine joints. Synovial fluid cytokines (MCP-1, IL-10, PGE2, IL-1ß, IL-6, TNF-α, IL-1ra) and growth factors (GM-CSF, SDF-1α+ß, IGF-1, and FGF-2) were quantified. Macrophage phenotypes were not as clearly defined in vivo as they are in vitro. All macrophage markers were expressed with minimal differences between OA and normal joints. Expression for all markers increased proportionate to synovial inflammation, especially CD86. Synovial fluid MCP-1 was higher in osteoarthritic joints while SDF-1 and IL-10 were lower, and PGE2 concentrations did not differ between groups. Increased CD14/CD86/CD206/IL-10 expression was associated with synovial hyperplasia, consistent with macrophage recruitment and activation in response to injury. Lower synovial fluid IL-10 could suggest that homeostatic mechanisms from synovial macrophages became overwhelmed preventing inflammation resolution, resulting in chronic inflammation and OA. Further investigations into mechanisms of arthritis resolution are warranted. Developing pro-resolving therapies may provide superior results in the treatment of OA.

Autologous bone marrow mononuclear cells modulate joint homeostasis in an equine in vivo model of synovitis.

Osteoarthritis (OA) is characterized by macrophage-driven synovitis. Macrophages promote synovial health but become inflammatory when their regulatory functions are overwhelmed. Bone marrow mononuclear cells (BMNCs) are a rich source of macrophage progenitors used for treating chronic inflammation and produce essential molecules for cartilage metabolism. This study investigated the response to autologous BMNC injection in normal and inflamed joints. Synovitis was induced in both radiocarpal joints of 6 horses. After 8 h, 1 inflamed radiocarpal and 1 normal tarsocrural joint received BMNC injection. Contralateral joints were injected with saline. Synovial fluid was collected at 24, 96, and 144 h for cytology, cytokine quantification, and flow cytometry. At 144 h, horses were euthanatized, joints were evaluated, and synovium was harvested for histology and immunohistochemistry. Four days after BMNC treatment, inflamed joints had 24% higher macrophage counts with 10% more IL-10+ cells than saline-treated controls. BMNC-treated joints showed gross and analytical improvements in synovial fluid and synovial membrane, with increasing regulatory macrophages and synovial fluid IL-10 concentrations compared with saline-treated controls. BMNC-treated joints were comparable to healthy joints histologically, which remained abnormal in saline-treated controls. Autologous BMNCs are readily available, regulate synovitis through macrophage-associated effects, and can benefit thousands of patients with OA.-Menarim, B. C., Gillis, K. H., Oliver, A., Mason, C., Ngo, Y., Werre, S. R., Barrett, S. H., Luo, X., Byron, C. R., Dahlgren, L. A. Autologous bone marrow mononuclear cells modulate joint homeostasis in an equine in vivo model of synovitis.

Gold nanoparticles paper as a SERS bio-diagnostic platform.

Bioactive papers are usually challenged by four major limitations: sensitivity, selectivity, simplicity and strength (4S). Gold nanoparticles (AuNPs) treated paper has previously been demonstrated as a Surface Enhanced Raman Scattering (SERS) active substrate, capable of addressing the 4S issues. In this study, AuNPs on paper substrate were functionalized by a series of biomolecules to develop a generic SERS platform for antibody-antigen detection. The functionalization steps were performed by taking advantage of the high affinity association between Streptomyces avidinii-derived protein, streptavidin, and biotin. Streptavidin was firstly bound onto the AuNPs treated paper using biotinylated-thiol. Subsequently, desired biotinylated-antibody was bound onto the streptavidin. SERS spectra of each functionalization step were obtained to ensure specific adsorption of the bio-molecules. The binding interaction of the antibody with its specific antigen was detected using SERS. Shifts of Raman band associated with α-helix and ß-sheet structures indicated structural modification of the antibody upon interaction with its antigen. Predominant tryptophan and tyrosine residue bands were also detected, confirming the presence of antigen. Reproducible spectral features were quantified as AuNP papers were subjected to different concentrations of antigen; the spectra intensity increased as a function of the antigen concentration. The retention of AuNPs on paper remained constant after all the consecutive washing and functionalization steps. The feasibility of AuNPs paper as a low-cost and generic SERS platform for bio-diagnostic applications was demonstrated.

Formation of polyelectrolyte-gold nanoparticle necklaces on paper.

This work reports a simple method to form and visualize individual polyelectrolyte-nanoparticle necklace-like structures on paper, which is applicable to other porous surfaces. In this work, one-dimensional necklaces of negatively charged gold nanoparticles (AuNPs) have been electrostatically assembled along the backbone of a cationic polyacrylamide (CPAM) chain adsorbed on paper. The process involves rapidly passing a dilute CPAM solution through filter paper, adsorbing the polyelectrolyte on the surface, followed by the immediate filtration of an AuNP suspension through the same filter paper. The nanoparticles used were negatively charged, citrate ion capped AuNPs with an average diameter of 20 nm. Scanning electron microscopy images of the dried paper sample showed that the AuNP necklaces were adsorbed in a perpendicular direction to that of the cellulose fibers and along the length of the CPAM molecules which were draped over the fibers. The effects of CPAM polymer concentration, charge density, and molecular weight on such assembly of AuNPs were studied. This technique enables the visualization of polyelectrolyte molecules and the formation of very well organized and reproducible polyelectrolyte-nanoparticle necklaces on a porous, three-dimensional substrate.

Effect of cationic polyacrylamides on the aggregation and SERS performance of gold nanoparticles-treated paper.

This study examines and quantifies the effect of cationic polyelectrolyte adsorption on paper on the aggregation and retention of gold nanoparticles (AuNPs) to optimize their Surface Enhanced Raman Scattering (SERS) enhancement factor and sensitivity. Aggregation of metallic nanoparticles is known as a key factor for intense SERS enhancement. Paper substrates were treated with aqueous solutions of cationic polyacrylamide (CPAM) varying in concentration, charge density, and molecular weight to control the AuNPs' aggregate size distribution and surface coverage on paper. The Raman Enhancement Factor of AuNPs-CPAM paper was almost an order of magnitude greater than for the untreated AuNPs paper. The high loading and uniform distribution of AuNPs aggregates on CPAM pre-treated paper contributed toward the excellent SERS reproducibility, sensitivity, and high enhancement factor. This configuration of AuNP on paper was favoured by treating the substrate with CPAM solutions of higher concentrations, higher charge density, and greater molecular weight.

Gold nanoparticle-paper as a three-dimensional surface enhanced Raman scattering substrate.

This work investigates the effect of gold nanoparticle (AuNP) addition to paper substrate and examines the ability of these composite materials to amplify the surface enhanced Raman scattering (SERS) signal of a dye adsorbed. Paper has a three-dimensional (3D), porous, and heterogeneous morphology. The manner in which paper adsorbs the nanoparticles is crucial to its SERS properties, particularly with regards to aggregation. In this work, we sought to maintain the same degree of aggregation, while changing the concentration of nanoparticles deposited on paper. We achieved this by dipping paper into AuNP solutions of different, known concentration and found that the initial packing density of AuNPs in solutions was retained on paper with the same degree of aggregation. The surface coverage of AuNPs on paper was found to scale linearly to their concentration profile in solutions. The SERS performances of the AuNP-treated papers were evaluated with 4-aminothiophenol (4-ATP) as the Raman molecule, and their SERS intensities increased linearly with the AuNPs' concentration. Compared to AuNP-treated silicon, the Raman enhancement factor (EF) from paper was relatively higher due to a more uniform and greater degree of adsorption of AuNPs. The effect of the spatial distribution of AuNPs in their substrates on SERS activity was also investigated. In this experiment, the number of AuNPs was kept constant (a 1 µL droplet of AuNPs was deposited on all substrates), and the distribution profile of AuNPs was controlled by the nature of the substrate: paper, silicon, and hydrophobized paper. The AuNP droplet on paper showed the most reproducible and sensitive SERS signal. This highlighted the role of the z-distribution (through film) of AuNPs within the bulk of the paper, producing a 3D multilayer structure to allow inter- and intralayer plasmon coupling, and hence amplifying the SERS signal. The SERS performance of nanoparticle-functionalized paper can thus be optimized by controlling the 3D distribution of the metallic nanoparticles, and such control is critical if these systems are to be implemented as a low-cost and highly sensitive bioassay platform.

Paper surfaces functionalized by nanoparticles.

Nanomaterials with unique electronic, optical and catalytic properties have recently been at the forefront of research due to their tremendous range of applications. Taking gold, silver and titania nanoparticles as examples, we have reviewed the current research works on paper functionalized by these nanoparticles. The functionalization of paper with only a very small concentration of nanoparticles is able to produce devices with excellent photocatalytic, antibacterial, anti-counterfeiting, Surface Enhanced Raman Scattering (SERS) and Surface Plasmon Resonance (SPR) performances. This review presents a brief overview of the properties of gold, silver and titania nanoparticles which contribute to the major applications of nanoparticles-functionalized paper. Different preparation methods of the nanoparticles-functionalized paper are reviewed, focusing on their ability to control the morphology and structure of paper as well as the spatial location and adsorption state of nanoparticles which are critical in achieving their optimum applications. In addition, main applications of the nanoparticles-functionalized papers are highlighted and their critical challenges are discussed, followed by perspectives on the future direction in this research field. Whilst a few studies to date have characterized the distribution of nanoparticles on paper substrates, none have yet optimized paper as a nanoparticles' substrate. There remains a strong need to improve understanding on the optimum adsorption state of nanoparticles on paper and the heterogeneity effects of paper on the properties of these nanoparticles.