Osteoblasts MC3T3-E1 response in 2D and 3D cell cultures models to high carbon content CoCr alloy particles. Effect of metallic particles on vimentin expression

The study of the biocompability of the metallic materials is a priority treating to avoid the osteolysis and aseptic loosening of prosthesis. Wear debris is considered one of the main factors responsible for aseptic loosening of orthopedic endoprostheses. We examined the response of mouse osteoblasts MC3T3-E1 to high carbon cobalt-chrome (HCCoCr) particles obtained a) from wear-corrosion assays on a pin-on-disk tribometer using as pair an alumina ball and a disc of a HCCoCr alloy, and b) HCCoCr bulk particles obtained by nitrogen gas atomization from an alloy used in clinic for prostheses application. Mitochondrial activity and lactate dehydrogenase activity assayed in 2D and 3D osteoblasts cell culture models were used to evaluate the cellular response to size, shape, and chemical composition of the metallic particles. 2D cell model was used to study the direct interaction of cells with particles and 3D cell cultures was used to more closely mimic in vivo conditions.The results showed that vimentin was overexpressed in the 2D osteoblasts cultures in presence of metal particles.This might be related to the appearance of pseudotumor in the peri-prosthetic vicinity described in some implanted patients.Financial support was received through the projects MAT2011-29152-C02-02 and MAT2011-29152-C02-01.RMLP and BTPM wish to thank the Ministerio de Economía y Competitividad from Spain for the financial support given by the project: MAT2011-29152-C02-02 and MAT2015-67750-C3-2-R.Peer reviewe

Corrosion Behaviour and J774A.1 Macrophage Response to Hyaluronic Acid Functionalization of Electrochemically Reduced Graphene Oxide on Biomedical Grade CoCr

Improvements in the lubrication of metal¿metal joint prostheses are of great clinical interest in order to minimize the particles released during wear¿corrosion processes. In this work, electrochemically reduced graphene oxide (ErGO) on CoCr was functionalized with hyaluronic acid (ErGOHA). Functionalization was carried out by soaking for 24 h in phosphate buffer saline (PBS) solution containing 3 g/L hyaluronic acid (HA). The corrosion performance of CoCrErGO and CoCrErGOHA surfaces was studied by electrochemical impedance spectroscopy (EIS) for 7 days in PBS. Biocompatibility and cytotoxicity were studied in mouse macrophages J774A.1 cell line by the measurement of mitochondrial activity (WST-1 assay) and plasma membrane damage (LDH assay). The inflammatory response was examined through TNF-¿ and IL-10 cytokines in macrophages culture supernatants, used as indicators of pro-inflammatory and anti-inflammatory responses, respectively. EIS diagrams of CoCrErGOHA revealed two time constants: the first one, attributed to the hydration and diffusion processes of the HA layer adsorbed on ErGO, and the second one, the corrosion resistance of ErGOHA/CoCr interface. Macrophage assays showed better behavior on CoCrErGOHA than CoCr and CoCrErGO surfaces based on their biocompatible, cytotoxic, and inflammatory responses. Comparative analysis of IL-10 showed that functionalization with HA induces higher values of anti-inflammatory cytokine, suggesting an improvement in inflammatory behavior.Financial support received through the RTI2018-101506-B-C31 and RTI2018-101506-B-C33 from the Ministerio de Ciencia, Innovación y Universidades (MICIU/FEDER), and through the MAT2015-67750-C3-2-R, MAT2015-67750-C3-1-R from the Ministerio de Economía y Competitividad (MINECO/FEDER) from Spain

Wettability, corrosion resistance, and osteoblast response to reduced graphene oxide on CoCr functionalized with hyaluronic acid

14 p.-8 fig.-2 tab. Blanca Teresa Pérez-Maceda who participated actively in this research with great enthusiasm has passed away.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 repairFinancial support is received through the RTI2018-101506-B-C31 and RTI2018-101506-B-C33 from the Ministerio de Ciencia, Innovación (MICINN/FEDER), and through the MAT2015-67750-C3-2-R, MAT2015-67750-C3-1-R from the Ministerio de Economía y Competitividad (MINECO/FEDER) from Spain.Peer reviewe

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

24 p.-7 fig. (In memory of Dr Blanca Teresa Perez-Maceda whose lovely life, plenty of fortitude, unwavering strength, joy and spiritual values, we had the honour of sharing.)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.CONICET (P-UE 22920170100100CO), ANPCyT (PICT 2015-0232, PICT 2016-1424), UNLP(11/I221), MICINN (MAT2011-29152-C02-02), MINECO (MAT2015-67750-C3-2-R), RTI2018-101506-B-C33.Peer reviewe

In vitro performance of magnesium processed by different routes for bone regeneration applications

4 páginas, 3 figuras, 1 tabla -- PAGS nros. 3020-3023Magnesium processed by powder metallurgy (PM) and casting was investigated for potential use in bone regeneration applications. To reduce its corrosion rate, magnesium was surface-modified by a chemical conversion treatment in hydrofluoric acid (HF). In vitro behaviour was evaluated in terms of biodegradation and osteoblastic cell response. The metallurgical route used to produce magnesium has more significant consequences on biodegradation and biocompatibility than the effects of the surface coating. The coated magnesium processed by casting exhibits the best in vitro performanceThe authors wish to thank MICINN, Spain, for the financial support given by projects MAT2008-06719-C03-01-02-03. JAdV wishes to thank MEC for financial support of the project MAT 2006-02672. MC thanks CSIC for a JAE-Doc contractPeer reviewe

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.Funding: Financial support received through the MAT2015-67750-C3-2-R, MAT2015-67750-C3-1-R, MAT2011-29152-C02-01 and the MAT2011-29152-C02-02 projects from the Ministerio de Economía y Competitividad (MINECO/FEDER) from Spain

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.Financial support received through the MAT2015-67750-C3-2-R, MAT2015-67750-C3-1-R, MAT2011 -29152-C02-01 and the MAT2011-29152-C02-02 projects from the Ministerio de Economía y Competitividad (MINECO/FEDER) from Spain

Reducción electroquímica de óxido de grafeno sobre aleación de CoCr de grado biomédico para aplicaciones endoprotésicas: ensayos de biocompatibilidad en macrófagos

Trabajo presentado en el VIII Congreso Internacional de Materiales, celebrado en Arequipa (Perú), del 21 al 23 de noviembre de 2018Peer reviewe

In vivo and macrophage response of graphene and its derivatives

The possible application in the biomedical field of reduced Graphene oxide (ErGO) on CoCr alloy has been studied. Biocompatibility tests were carried out on ErGO/CoCr alloy. The discrepancy of in vivo results found in the literature regarding the side effects of graphene led to perform an in vivo study with graphene. Biocompatibility tests of ErGO/CoCr were evaluated in J774A.1 mouse macrophages cultures. Mitochondrial activity (WST-1 assay) and plasma membrane damage (LDH assay) were measured to evaluate biocompatibility and cytotoxicity, respectively. The ratio of LDH/WST-1 activities was used as an index of biocompatibility as relates cell death and cell number, reaching a low value on ErGO/CoCr. Morphological analyses of macrophages cultures revealed different cell distribution and morphology on CoCr and ErGO/CoCr, after 48 h exposure. Optical microscopy and secondary electron microscopy images showed macrophages on the ErGO/CoCr well-distributed and conserved characteristic cell shape. These results show an improvement in the CoCr biocompatibility due to ErGO films. In vivo tests of graphene and graphene oxide nanosheets were carried out by intraperitoneal inoculation in rats to evaluate possible changes in the blood line and organs after 15 and 30 days. Optical microscopy of liver, kidney, spleen or lung, revealed no visible histological alterations. However, traces of particles were found in the peritoneal cavity. The blood analysis showed alterations indicative of the hepatic inflammatory process. Haematological changes after 30 days consisted of alterations of the red series as microcytosis with a higher concentration of mean haemoglobin. In addition, alteration in prothrombin and thromboplastin caused a longer coagulation time.The authors acknowledge the financial support from MAT2015-67750-C3