Time-Lapse Evaluation of Interactions Between Biodegradable Mg Particles and Cells.

Mg-based implants have promising applications as biodegradable materials in medicine for orthopedic, dental, and cardiovascular therapies. During wear and degradation microdebris are released. Time-lapse multidimensional microscopy (MM) is proposed here as a suitable tool to follow, in fixed intervals over 24-h periods, the interaction between cells and particles. Results of MM show interactions of macrophages (J774) with the magnesium particles (MgPa) that led to modifications of cell size and morphology, a decrease in duplication rate, and cell damage. Corrosion products were progressively formed on the surface of the particles and turbulence was generated due to hydrogen development. Changes were more significant after treating MgPa with potassium fluoride. In order to complement MM observations, membrane damage as detected by a lactase dehydrogenase (LDH) assay and mitochondrial activity as detected by a WST-1 assay with macrophages and osteoblasts (MC3T3-E1) were compared. A more significant concentration-dependent effect was detected for macrophages exposed to MgPa than for osteoblasts. Accordingly, complementary data showed that viability and cell cycle seem to be more altered in macrophages. In addition, protein profiles and expression of proteins associated with the adhesion process changed in the presence of MgPa. These studies revealed that time-lapse MM is a helpful tool for monitoring changes of biodegradable materials and the biological surrounding in real time and in situ. This information is useful in studies related to biodegradable biomaterials.

Dendritic cell-specific ICAM-3-grabbing nonintegrin expression on M2-polarized and tumor-associated macrophages is macrophage-CSF dependent and enhanced by tumor-derived IL-6 and IL-10.

Dendritic cell-specific ICAM-3-grabbing nonintegrin (DC-SIGN; CD209) is a human pathogen-attachment C-type lectin with no obvious murine ortholog and for which ligation leads to enhanced anti-inflammatory cytokine release and altered proinflammatory cytokine production. Although induced by IL-4 in monocytes and considered as a DC marker, DC-SIGN expression on human APCs under homeostatic conditions is so far unexplained. We report in this study that M-CSF enhances DC-SIGN expression on in vitro derived anti-inflammatory macrophages and that M-CSF mediates the induction of DC-SIGN by fibroblast- and tumor cell-conditioned media. The M-CSF-inducible DC-SIGN expression along monocyte-to-macrophage differentiation is dependent on JNK and STAT3 activation, potentiated by STAT3-activating cytokines (IL-6, IL-10), and abrogated by the M1-polarizing cytokine GM-CSF. In pathological settings, DC-SIGN expression is detected in tumor tissues and on ex vivo-isolated CD14(+) CD163(+) IL-10-producing tumor-associated macrophages. Importantly, DC-SIGN Abs reduced the release of IL-10 from macrophages exposed to Lewis(x)-expressing SKBR3 tumor cells. These results indicate that DC-SIGN is expressed on both wound-healing (IL-4-dependent) and regulatory (M-CSF-dependent) alternative (M2) macrophages and that DC-SIGN expression on tumor-associated macrophages might help tumor progression by contributing to the maintenance of an immunosuppressive environment.

Wettability, Corrosion Resistance, and Osteoblast Response to Reduced Graphene Oxide on CoCr Functionalized with Hyaluronic Acid.

The durability of metal-metal prostheses depends on achieving a higher degree of lubrication. The beneficial effect of hyaluronic acid (HA) on the friction and wear of both natural and artificial joints has been reported. For this purpose, graphene oxide layers have been electrochemically reduced on CoCr surfaces (CoCrErGO) and subsequently functionalized with HA (CoCrErGOHA). These layers have been evaluated from the point of view of wettability and corrosion resistance in a physiological medium containing HA. The wettability was analyzed by contact angle measurements in phosphate buffer saline-hyaluronic acid (PBS-HA) solution. The corrosion behavior of functionalized CoCr surfaces was studied with electrochemical measurements. Biocompatibility, cytotoxicity, and expression of proteins related to wound healing and repair were studied in osteoblast-like MC3T3-E1 cell cultures. All of the reported results suggest that HA-functionalized CoCr surfaces, through ErGO layers in HA-containing media, exhibit higher hydrophilicity and better corrosion resistance. Related to this increase in wettability was the increase in the expressions of vimentin and ICAM-1, which favored the growth and adhesion of osteoblasts. Therefore, it is a promising material for consideration in trauma applications, with improved properties in terms of wettability for promoting the adhesion and growth of osteoblasts, which is desirable in implanted materials used for bone repair.

Effect of degradation products of iron-bioresorbable implants on the physiological behavior of macrophages in vitro.

The degradation of bioresorbable metals in vivo changes the physicochemical properties in the environment of an implant, such as a stent in the artery wall, and may induce the alteration of the functions of the surrounding cells. The Fe-degradation, from bioresorbable stents, is a particularly intricate process because it leads to the release of soluble (SDP) and insoluble degradation products (IDP) of varied composition. Macrophages are involved in the resorption of the exogenous agents coming from degradation of these materials. In the present work an Fe0 ring, made with a pure Fe wire, in contact with macrophage cell cultures was used to simulate the behaviour of a biodegradable Fe-based implant in a biological environment. Non-invasive time-lapse optical microscopy was applied to obtain images of macrophages exposed to Fe-degradation products, without using staining to avoid distortions and artefacts. It was noticed that as metal degraded, the IDP formed in situ accumulated close to the Fe0 ring. In this zone, the macrophages showed a dynamic process of uptake of dark Fe-containing products, confirmed by SEM-EDX. These macrophages showed alterations in the morphology and decrease in the motility and viability. The inability of the macrophages to move and to degrade the engulfed products caused a long persistence of IDP in the zone closest to the metal. The deleterious effects of IDP accumulated close to the ring, were significantly worse than those observed in the experiments made with (1) concentrated salt solutions (Fe3+ salt 3 mM), with the same amount of precipitates but uniformly distributed in the well, and (2) diluted salt solutions (Fe3+ salt 1 mM) with mainly soluble species. The results were confirmed by standard staining protocols that revealed dead cells close to the Fe0 ring and oxidative stress in cells exposed to both soluble and insoluble species.

Macrophage Biocompatibility of CoCr Wear Particles Produced under Polarization in Hyaluronic Acid Aqueous Solution.

Macrophages are the main cells involved in inflammatory processes and in the primary response to debris derived from wear of implanted CoCr alloys. The biocompatibility of wear particles from a high carbon CoCr alloy produced under polarization in hyaluronic acid (HA) aqueous solution was evaluated in J774A.1 mouse macrophages cultures. Polarization was applied to mimic the electrical interactions observed in living tissues. Wear tests were performed in a pin-on-disk tribometer integrating an electrochemical cell in phosphate buffer solution (PBS) and in PBS supplemented with 3 g/L HA, an average concentration that is generally found in synovial fluid, used as lubricant solution. Wear particles produced in 3 g/L HA solution showed a higher biocompatibility in J774A.1 macrophages in comparison to those elicited by particles obtained in PBS. A considerable enhancement in macrophages biocompatibility in the presence of 3 g/L of HA was further observed by the application of polarization at potentials having current densities typical of injured tissues suggesting that polarization produces an effect on the surface of the metallic material that leads to the production of wear particles that seem to be macrophage-biocompatible and less cytotoxic. The results showed the convenience of considering the influence of the electric interactions in the chemical composition of debris detached from metallic surfaces under wear corrosion to get a better understanding of the biological effects caused by the wear products.

Response of MC3T3-E1 osteoblasts, L929 fibroblasts, and J774 macrophages to fluoride surface-modified AZ31 magnesium alloy.

The present work evaluates the biocompatibility of a fluoride surface-modified AZ31 magnesium alloy (AZ31HF) with different cell lines that coexist in the implant environment to test its potential use as a biodegradable and absorbable biomaterial for bone repair. A clear stimulation of cell proliferation and an enhancement of the mitochondrial respiratory activity were observed when mouse osteoblasts (MC3T3-E1), fibroblasts (L929), and macrophages (J774) cell lines were cultured with the modified alloy. No significant change in apoptosis or viability rates was observed when osteoblasts and fibroblasts cultures were grown in the presence of this alloy. A proteomic analysis of the MC3T3-E1 cell extracts cultured in the presence of AZ31HF showed an overexpression of proteins related with the mineralization process, which is a necessary step for bone repair. An increase in the lactate dehydrogenase activity was observed in the MC3T3-E1 and J774 cell cultures that could be a response of the oxidative stress produced by the presence of the material. This stress could be related to the increase observed in the respiratory mitochondrial activity or respiratory burst measured in theses cultures that indicate damage in the cell membranes and subsequently some cell death. Results reported here, for and against AZ31HF, should be taken into account when considering the potential use of this modified alloy in bone repair applications.