Search

The presence of a worn surface in the implanted material, as in the case of a replacement of a damaged osteoarticular joint, is the normal condition after implantation. This manuscript focuses precisely on the comparative study of the cellular behavior on worn CoCr surfaces, analyzing the effect of different surface modifications on macrophages’ responses. CoCr surfaces were modified by the deposition of electrochemically reduced graphene oxide (CoCrErGO), followed by additional surface functionalization with hyaluronic acid (CoCrErGOHA). After the wear corrosion processes, the macrophage response was studied. In addition, macrophage supernatants exposed to the surfaces, before and after wear, were also evaluated for osteoblast response through the analysis of the metabolic activity, plasma membrane damage, and phosphatase alkaline activity (ALP). The proteomic analysis and the quantitative TNF- /IL-10 ratios of the J774A.1 macrophages exposed to the surfaces under study showed a polarization shift from M0 (basal state) to M1, associated with the pro-inflammatory response of all surfaces. A lower M1 polarization was observed upon exposure to the surface modification with ErGO, whereas posterior HA functionalization attenuated, even more, the M1 polarization. The wear corrosion process contributed to inflammation and exacerbated the M1 polarization response on macrophages to CoCr, which was diminished for the ErGO and attenuated the most for the ErGOHA surfaces. Comparative proteomics showed that the pathways related to M1 polarization were downregulated on the surfaces of CoCrErGOHA, which suggests mechanisms for the observed attenuation of M1 polarization. The suitable immuno-modulatory potential induced by the ErGOHA surface, with and without wear, together with the stimulation of ALP activity in osteoblasts induced by macrophage supernatants, promotes the mineralization processes necessary for bone repair. This makes it feasible to consider the adsorption of ErGO

[EN] This is the experimental dataset used in the paper “Barrier Graphene Oxide on a CoCr Alloy via Silane/GO Covalent Bonding and Its Electrochemical Behavior in a Simulated Synovial Fluid Electrolyte". The article includes the experimental procedure carried out for the deposition of graphene oxide on CoCr surfaces: CoCr was hydroxylated through alkalization in NaOH 5 M for 2 h, (sample named CoCr-OH). Subsequently, silanization was carried out using APTES. APTES was pre-mixed at 2% vol in isopropanol-water (200:1 v/v) and stirred for 1 h. After this, CoCr-OH samples were immersed in the APTES solution at room temperature for 1 min. The silane-coated samples were kept under curing conditions at 45 °C (CoCr-OH-Si45 sample) and 75 °C (CoCr-OH-Si75 sample) for 24 h and 30 min, respectively. Finally, CoCr-OH-Si45 and CoCr-OH-Si75 surfaces were immersed in 4 g/L GO aqueous suspension at 60 °C for 12 h and 24 h to obtain CoCr-OH-Si45-GO12h, CoCr-OH-Si75-GO12h and CoCr-OH-Si45-GO24h, and CoCr-OH-Si75-GO24h. This data set includes the results of the electrochemical impedance data obtained for each CoCr surface: CoCr (CoCr, CoCr-OH, CoCr-OHSi45, CoCr-OH-Si75 CoCr-OH-Si45-GO12h, CoCr-OH-Si75-GO12h y CoCr-OH-Si45-GO24h, y CoCr-OH-Si75-GO24h, immersed in 3 g/L Hyaluronic acid solution for 7 days., [ES] Este es el conjunto de datos experimentales utilizado en el artículo "Barrera de Óxido de Grafeno en una Aleación CoCr mediante enlace covalente de Silano/GO y su comportamiento electroquímico en un electrolito de fluido sinovial simulado". El artículo incluye el procedimiento experimental llevado a cabo para la deposición de óxido de grafeno sobre superficies de CoCr: CoCr fue hidroxilado mediante alcalinización en NaOH 5 M durante 2 horas (muestra denominada CoCr-OH). Posteriormente, se llevó a cabo la silanización utilizando APTES. APTES fue premezclado al 2% en volumen en isopropanol-agua (200:1 v/v) y agitado durante 1 hora. Después, las muestras de CoCr-OH fueron sumergidas en la solución de APTES a temperatura ambiente durante 1 minuto. Las muestras recubiertas con silano se mantuvieron bajo condiciones de curado a 45 °C (muestra CoCr-OH-Si45) y 75 °C (muestra CoCr-OH-Si75) durante 24 horas y 30 minutos, respectivamente. Finalmente, las superficies CoCr-OH-Si45 y CoCr-OH-Si75 fueron sumergidas en suspensión acuosa de 4 g/L GO a 60 °C durante 12 y 24 horas obteniendo las superficies CoCr-OH-Si45-GO12h, CoCr-OH-Si75-GO12h y CoCr-OH-Si45-GO24h, y CoCr-OH-Si75-GO24h. Este conjunto de datos incluye los resultados de los datos de impedancia electroquímica obtenidos para cada superficie de CoCr (CoCr, CoCr-OH, CoCr-OHSi45, CoCr-OH-Si75 CoCr-OH-Si45-GO12h, CoCr-OH-Si75-GO12h y CoCr-OH-Si45-GO24h, y CoCr-OH-Si75-GO24h sumergidas en una solución de 3 g/L ácido hialurónico durante 7 días.

In this work, a sequential covalent immobilization of graphene oxide (GO) and hyaluronic acid (HA) is performed to obtain a biocompatible wear-resistant nanocoating on the surface of the biomedical grade cobalt-chrome (CoCr) alloy. Nanocoated CoCr surfaces were characterized by Raman spectroscopy and electrochemical impedance spectroscopy (EIS) in 3 g/L HA electrolyte. Tribocorrosion tests of the nanocoated CoCr surfaces were carried out in a pin on flat tribometer. The biological response of covalently HA/GO biofunctionalized CoCr surfaces with and without wear-corrosion processes was studied through the analysis of the proteome of macrophages. Raman spectra revealed characteristic bands of GO and HA on the functionalized CoCr surfaces. The electrochemical response by EIS showed a stable and protective behavior over 23 days in the simulated biological environment. HA/GO covalently immobilized on CoCr alloy is able to protect the surface and reduce the wear volume released under tribocorrosion tests. Unsupervised classification analysis of the macrophage proteome via hierarchical clustering and principal component analysis (PCA) revealed that the covalent functionalization on CoCr enhances the macrophage biocompatibility in vitro. On the other hand, disruption of the HA/GO nanocoating by tribocorrosion processes induced a macrophage proteome which was differently clustered and was distantly located in the PCA space. In addition, tribocorrosion induced an increase in the percentage of upregulated and downregulated proteins in the macrophage proteome, revealing that disruption of the covalent nanocoating impacts the macrophage proteome. Although macrophage inflammation induced by tribocorrosion of HA/GO/CoCr surfaces is observed, it is ameliorated by the covalently grafting of HA, which provides immunomodulation by eliciting downregulations in characteristic pro-inflammatory signaling involved in inflammation and aseptic loosening of CoCr joint arthroplasties. Covalen

In this work, a sequential covalent co-immobilization of graphene oxide (GO) and hyaluronic acid (HA) is performed to obtain a biocompatible wear-resistant nanocoating on the surface of the biomedical grade Cobalt-Chrome (CoCr) alloy. Nanocoated CoCr surfaces were characterized by Raman spectroscopy and electrochemical impedance spectroscopy (EIS) in 3 g/L HA electrolyte. Tribocorrosion tests of the nanocoated CoCr surfaces were carried out in a pin on flat tribometer. The biological response of covalently HA/GO biofunctionalized CoCr surfaces with and without wear-corrosion processes was studied through the analysis of the proteome of macrophages. Raman spectra revealed characteristic bands of GO and HA on the functionalized CoCr surfaces. The electrochemical response by EIS showed a stable and protective behavior over 23 days in the simulated biological environment. HA/GO covalently immobilized on CoCr alloy is able to protect the surface and reduce the wear volume released under tribocorrosion tests. Unsupervised classification analysis of the macrophage proteome via Hierarchical clustering and Principal Component Analysis (PCA) revealed that the covalent functionalization on CoCr enhances the macrophage biocompatibility in vitro. On the other hand, disruption of the HA/GO nanocoating by tribocorrosion processes induced a macrophage proteome which was differently clustered and was distantly located in the PCA space. In addition, tribocorrosion induced an increase in the percentage of upregulated and downregulated proteins in the macrophage proteome, revealing that disruption of the covalent nanocoating impacts the macrophage proteome. Although macrophage inflammation induced by tribocorrosion of HA/GO/CoCr surfaces is observed, it is ameliorated by the covalently grafting of HA, which provides immunomodulation by eliciting downregulations in characteristic pro-inflammatory signaling involved in inflammation and aseptic loosening of CoCr joint arthroplasties. Cova

[EN] This is the experimental dataset used in the paper “Macrophage Proteomic Analysis of Covalent Immobilization of Hyaluronic Acid and Graphene Oxide on CoCr Alloy in a Tribocorrosive Environment". The article includes the experimental procedure carried out for the deposition of graphene oxide and posterior immobilization of hyaluronic acid on CoCr surfaces. CoCr was hydroxylated through alkalization in NaOH 5 M for 2 h. Subsequently, silanization was carried out using APTES. APTES was pre-mixed at 2% vol in isopropanol-water (200:1 v/v) and stirred for 1 h. After this, CoCr-OH samples were immersed in the APTES solution at room temperature for 1 min. The silane-coated samples were kept under curing conditions at 45 °C for 24 h. Then, silanized CoCr surfaces were immersed in 4 g/L GO aqueous suspension at 60 °C for 24 h. Carboxylic groups of graphene oxide in GO/CoCr samples were activated by immersing in distilled water (50 mL) with N-(3-dimethylaminopropyl)-N-ethylcarbodiimide hydrochloride (EDC, 250 mg) and N-hydroxysuccinimide (NHS, 750 mg) during 24 h under mild orbital saking. After that, the GO/CoCr surfaces with activated carboxyl groups were immersed in distilled water (100 mL) with dissolved adipic dihydrazide (ADH, 1.04 g) and let to react for another 24 h under mild orbital shaking. Pre-activated HA-(EDC/NHS) solution was obtained by dissolving HA (100 mg), EDC (253 mg) and NHS (754 mg) in 100 mL phosphate buffer saline (PBS) and stirred for 24h to activate the carboxylic acids of HA. After that, ADH/GO/CoCr surfaces were immersed in pre-activated HA-(EDC/NHS) solution and let to react for 24h under mild orbital motion (samples named HA/GO/CoCr). This data set includes the results of the final surface by Raman Spectroscopy and Electrochemical Impedance Spectroscopy. Raman was obtained from CoCr/GO/HA surfaces in comparison with GO/CoCr surfaces and HA powder. Electrochemical impedance data of CoCr/GO/HA were obtained under immersion in 3 g/L Hyaluronic, [ES] Este es el conjunto de datos experimentales utilizado en el artículo " Macrophage Proteomic Analysis of Covalent Immobilization of Hyaluronic Acid and Graphene Oxide on CoCr Alloy in a Tribocorrosive Environment". El artículo incluye el procedimiento experimental realizado para la deposición de óxido de grafeno y la posterior inmovilización de ácido hialurónico sobre superficies de CoCr. Las superficies de CoCr fueron hidroxiladas mediante alcalinización en NaOH 5 M durante 2 h. Posteriormente, se realizó la silanización utilizando APTES premezclado al 2% en volumen en isopropanol-agua (200:1 v/v) agitada durante 1 h. Las muestras de CoCr hidroxiladas se sumergieron en la solución de APTES a temperatura ambiente durante 1 minuto. Posteriormente, se mantuvieron en condiciones de curado a 45 °C durante 24 h. Transcurrido este tiempo, las superficies de CoCr silanizadas se sumergieron en una suspensión acuosa de 4g/L GO a 60 °C durante 24 h. Los grupos carboxílicos del óxido de grafeno en las muestras GO/CoCr se activaron mediante inmersión en 50 mL agua destilada que contenía hidrocloruro de N-(3-dimetilaminopropil)-N-etilcarbodiimida (EDC, 250 mg) y N-hidroxisuccinimida (NHS, 750 mg), durante 24 h en agitación orbital suave. Las superficies de GO/CoCr con grupos carboxílicos activados se sumergieron en 100 mL de agua destilada con dihidracida adipídica disuelta (ADH, 1.04 g) y se dejaron reaccionar durante otras 24 h en agitación orbital suave. La solución de HA preactivada (EDC/NHS) se obtuvo disolviendo HA (100 mg), EDC (253 mg) y NHS (754 mg) en 100 mL de solución salina de fosfato (PBS) y agitando durante 24 h para activar los ácidos carboxílicos de la HA. Después, las superficies de ADH/GO/CoCr se sumergieron en la solución de HA preactivada (EDC/NHS) y se dejaron reaccionar durante 24 h en movimiento orbital suave (muestras denominadas HA/GO/CoCr). Este conjunto de datos incluye los resultados de la caracterización superficial final mediante espectroscopía

In this work graphene-based solid lubricants biofunctionalized with hyaluronic acid were deposited on CoCr surfaces to decrease wear-corrosion phenomena. Graphene oxide was electrochemically reduced on CoCr in an aqueous suspension of graphene oxide 4 g/L by cyclic voltammetry. Biofunctionalization in phosphate buffer solution with HA 3 g/L (PBS-HA) for 24 h was performed. A pin-on-disk tribometer with an integrated cell was used for tribocorrosion tests. Wear-corrosion tests were performed against AISI316 steel balls applying normal load 1 N, 120 rpm for 500 m, in PBS and PBS-HA. Coefficients of friction in PBS were higher than in PBS-HA. ErGO and HA-ErGO increased wear rate in AISI316 balls, i.e., ErGO hardens the CoCr surface. Bio- functionalization reduces abrasive properties of ErGO deposits.

The presence of a worn surface in the implanted material, as in the case of a replacement of a damaged osteoarticular joint, is the normal condition after implantation. This manuscript focuses precisely on the comparative study of the cellular behavior on worn CoCr surfaces, analyzing the effect of different surface modifications on macrophages¿ responses. CoCr surfaces were modified by the deposition of electrochemically reduced graphene oxide (CoCrErGO), followed by additional surface functionalization with hyaluronic acid (CoCrErGOHA). After the wear corrosion processes, the macrophage response was studied. In addition, macrophage supernatants exposed to the surfaces, before and after wear, were also evaluated for osteoblast response through the analysis of the metabolic activity, plasma membrane damage, and phosphatase alkaline activity (ALP). The proteomic analysis and the quantitative TNF-¿/IL-10 ratios of the J774A.1 macrophages exposed to the surfaces under study showed a polarization shift from M0 (basal state) to M1, associated with the pro-inflammatory response of all surfaces. A lower M1 polarization was observed upon exposure to the surface modification with ErGO, whereas posterior HA functionalization attenuated, even more, the M1 polarization. The wear corrosion process contributed to inflammation and exacerbated the M1 polarization response on macrophages to CoCr, which was diminished for the ErGO and attenuated the most for the ErGOHA surfaces. Comparative proteomics showed that the pathways related to M1 polarization were downregulated on the surfaces of CoCrErGOHA, which suggests mechanisms for the observed attenuation of M1 polarization. The suitable immuno-modulatory potential induced by the ErGOHA surface, with and without wear, together with the stimulation of ALP activity in osteoblasts induced by macrophage supernatants, promotes the mineralization processes necessary for bone repair. This makes it feasible to consider the adsorption of ErGO

In this work, impermeable and ultrathin surface nanomodifications for joint applications based on graphene oxide (GO) are assembled on CoCr surfaces via covalent immobilization between GO nanosheets and silane monolayers. Two silane curing temperatures, 45 °C for 24 h and 75 °C for 30 min, on CoCr surfaces and two incubation times for GO suspension, 12 h and 24 h, on silanized CoCr surfaces are prepared. Electrochemical characterization is performed using electrochemical impedance spectroscopy (EIS) in a 3 g/L hyaluronic acid solution. Results show that GO nanosheets immobilized with silane covalent bonding confer impermeability of sp2 networks on GO and strong interfacial adhesion of GO sheets anchored to silanized CoCr via organosilane chemistry, which prevents the permeation of oxidant species at the metal interface. At short GO incubation times (12 h), the Rs values decrease with the immersion time, indicating that small species, such as metal ions, are able to diffuse through the interlayer gaps of nanolayers. Longer GO incubation times (24 h) favor the formation of bonds between the GO and the silane, thus slowing downdiffusion and metal ion release into the medium. EIS data confirm the impermeability of GO nanocoatings with lengthening GO incubation time for medical application of metallic implants.

Actualmente, una de las patologías más comunes y que afecta a un elevado porcentaje de la población de edad avanzada es la artritis. Es por ello, que en el campo de la medicina se han realizado grandes avances, principalmente estos últimos 20 años, respecto al desarrollo y mejora de prótesis osteoarticulares. Sin embargo, existen grandes desafíos en cuanto al aumento de la durabilidad de las prótesis, afectadas por procesos de desgastecorrosión que provocan la liberación de partículas e iones al organismo y que pueden llegar a resultar nocivos. Por ello, la investigación centrada en la mejora del comportamiento frente a tribocorrosión de los implantes osteoarticulares es de gran interés. Además del proceso de degradación continuo de la superficie de las prótesis, se ha detectado la presencia de unas capas triboquímicas carbonosas que pueden tener un efecto lubricante sobre la superficie metálica disminuyendo la tasa de desgaste y por tanto la disminución del número de iones y partículas metálicas liberadas al entorno del implante. El principal objetivo del proyecto fin de grado es la síntesis, caracterización y el estudio del comportamiento a desgaste-corrosión de un recubrimiento base grafeno aplicado capa por capa sobre una aleación de CoCr utilizadas en articulaciones por sus buenas propiedades frente al desgaste, a la corrosión y su buena biocompatibilidad

Degradation is a mechano-chemical phenomenon which impacts all materials, e.g. solar panels, wind and marine turbines, cultural heritage, biomaterials, etc. These materials are affected by degradation under the environmental conditions where they are employed, depending on the purpose of the application. The loss of material properties and its replacement is expensive; therefore, both the durability and the green recovery of materials are paramount for a circular economy in the modern industry. Concerning metallic materials working in the environment, the deterioration is produced as a consequence of a corrosive process carried out at room temperature, which consists in an electrochemical process in which two reactions occur: an anodic oxidation reaction and a cathodic reduction reaction in an electrical conducting electrolyte. If the purpose is to increase the durability and therefore increase the operational lifespan of these materials, first of all it is required to determine the material degradation kinetics through precise techniques of high accuracy and sensibility levels like electrochemical techniques. Electrochemical methods provide robust data regarding the current material properties which are used to determine the functionality status of the material and the possible need of material replacement or the viability of suitable protection methods to enhance its durability. Specifically, in this lecture, metallic biomaterials will be used as an example to understand the material deterioration via electrochemical techniques such as electrochemical impedance spectroscopy as well as the proposal of application of protective techniques to improve the operational durability of the materials via surface modification and functionalization strategies.

The effect of normal load and rotational rate on the coefficient of friction (COF) of CoCr sliding against alumina was evaluated using a pin on disk tribometer. Tribological tests were carried out by using CoCr disks of 38 mm diameter and alumina balls of 6 mm diameter as counterpart (pin). An aqueous solution containing 3g/L of HA, concentration equivalent of that found in the synovial fluid, was used as lubricant and it was in continuous recirculation throughout the test. The COF values for two normal loads, 1N and 5N, and two rotational rates, 60 rpm and 120 rpm, have been obtained for a constant sliding distance of 500 m. The tribological tests were performed in duplicate in order to verify the reproducibility of the results. The results show that the COF varies with the load and with the rotational rate, so that, for the same load, the COF increases as the rotational rate decreases. This effect is more significant at lower loads. Meanwhile, the COF values are higher for loads of 1N than for 5N regardless of the rotational rate. The wear tracks were observed by optical microscopy and the images revealed the widest tracks at 5N and 60 rpm.

Upon implantation of any biomaterial, a biomaterial-based foreign response (B-FBR) is initiated. Immune cells are recruited to the implant site, being macrophages one of the key innate immune cell regulators of the biomaterial-based FBR. Implanted metallic materials may trigger cellular stress and they may induce adaptive responses in macrophages such as alteration of biological processes, depending on the cytotoxicity potential of the metallic compounds [1]. Moreover, upon implantation, metallic joint prostheses are subjected to degradation due to wear and corrosion, releasing metallic debris (metallic micro/nanoparticles and ions) which causes the risk of morbidity of biological and immunological responses. CoCr particles can cause local toxicity such as adverse local tissue reactions (ALTR, which includes aseptic lymphocyte-dominated vasculitis-associated lesion, necrosis, osteolysis and implant loosening, pseudo-tumors) and systemic toxicity (including cardiomyopathy, polycythemia, hypothyroidism, and neurological disorders) [2]. In this study, CoCr surfaces were subjected to wear-corrosion to generate metallic debris, and the worn surfaces impacted the macrophage response. Since an exhaustive characterization of macrophages in response to biomaterials is required, advances in new omics technologies, which allow massive expression analyses, may reveal new insights in the altered biological processes upon macrophage-biomaterial interactions [3]. Macrophage responses were characterized by a high-throughput proteomic approach. With this proteomic tool the biological responses of macrophages to worn CoCr surfaces were analyzed, elucidating the effect of wear-corrosion process on J774A.1 macrophages. The differential proteomics study allowed us to determine that: wear-corrosion process on CoCr surface impacted the macrophage response by inducing an increase in protein expression of Heme-oxygenase 1 (Hmox1) a well-known antioxidant protein involved in redox and metal

Electrochemically reduced graphene oxide (ErGO) films on a biomedical grade CoCr alloy have been generated and the macrophage cellular response was here analyzed through a proteomic approach for its possible biomedical application. CoCr discs of 38 mm diameter and 2 mm thickness were polished on successive abrasive papers in silicon carbide grade from 600 to 2000 grain size. Immediately after, polished CoCr disks were immersed in an aqueous suspension of graphene oxide (GO) at 4g/L in order to perform direct electrochemical reduction of graphene oxide on the CoCr surface. An Autolab potentiostat/galvanostat was used for the electrochemical reduction of GO, which was performed in a three-electrode electrochemical cell consisting of a graphite bar used as counter electrode, Ag/AgCl as reference electrode and CoCr discs as working electrode. The electrochemical reduction method used was cyclic voltammetry (CV), carried out from -2.1V to -0.5V (vs. Ag/AgCl) at a scan rate of 10 mV/min. Study of cell-material interactions was focused on the innate immune system response by analyzing the macrophage proteome, since it may reveal new insights in molecular mechanisms and signaling pathways being trigger upon biomaterial interactions. Mouse macrophages (J774A.1) were seeded on tissue culture dishes on CoCr (Control group), and on CoCrErGO surfaces (CoCrErGO group). After 72h and 96h of cell culture with the biomaterial, cells were lysed in RIPA buffer containing protease inhibitors. Samples of same protein concentration were taken for SDS/PAGE, excised and trypsin digested, and peptides were analyzed in Q-Exactive. Mass spectra were searched against the Mus musculus database using Mascot search engine through Proteome Discoverer. Peptide identification was performed at a FDR of 1% Annotated MS data were filtered with Matlab 2020b based on maintained patterns of expression observed both at 72h and 96h. Protein abundance was normalized following data flow in (1). Differentially

Improvements regarding durable lubrication together with minimized wear are mandatory for obtaining long-term, functioning metallic joint prostheses. For this goal, CoCr surface was functionalized with Graphene Oxide (GO), characterized by FTIR and XPS, and tested at tribocorrosion. Deposition of graphene oxide was carried out by consecutive steps: alkalinization of CoCr; intermediate coupling via 3-aminopropyltriethoxysilane (APTES) which was cured at 45 °C for 24 h; and final assemble of GO layers at 60 °C for 24 h upon exposure with silane-coated CoCr. FTIR displayed C=C band confirming the assembly of GO on silane-coated CoCr surfaces. XPS elucidated reaction between primary amines of APTES and epoxy groups of GO, free -OH groups in APTES and carboxyl groups in GO, and reaction between APTES primary amines and -OH from carboxyl groups of GO. COF values for GO incubated on silane-coated CoCr disks show stationary values near 0.2. CoCr surfaces in as-reception state show higher COF values increasing at longer testing times.

Improvements in durable lubrication together with minimized wear are essential for obtaining long-term, functioning metallic joint prostheses. To achieve this objective, CoCr surface was functionalized with Graphene Oxide (GO) and characterized by FTIR and XPS. CoCr alloy was subjected to alkalinization in order to generate high hydroxyl content on the surface. FTIR and XPS revealed reactive OH groups, enabling intermediate coupling via (3-aminopropyl) triethoxysilane (APTES), which was cured at 45 °C and 75 °C for 24 h and 30 min, respectively. FTIR revealed cross-linked films (Si–O–Si), inferring condensation and self-assembly of silane layers, while XPS revealed the presence of NH, enabling chemical binding of GO. Silane-coated CoCr disks were immersed in GO solution at 60 °C for 12h and 24h, respectively. FTIR displayed C[dbnd]C band confirming the assembly of GO on silane-coated CoCr surfaces. XPS revealed three possible surface mechanisms: (1) reaction between primary amines of APTES and epoxy groups of GO; (2) free –OH groups in APTES and carboxyl groups in GO; and (3) reaction between APTES primary amines and –OH from carboxyl groups of GO. Overall, the multilayer system CoCr–OH–Si45-GO24h showed covalent functionalization of metal substrate with GO to a large extension of surface area among all the multilayer systems studied.

Background: The discrepancy among the in vivo results found in the literature regarding graphene’s side effects led us to conduct an in vivo study with graphene. Methods: In vivo tests involving intraperitoneal inoculation of graphene and graphene oxide nanosheets in rats were carried out to assess potential changes in the blood and organs after 15 and 30 days. Graphene and graphene oxide nanosheets at a concentration of 4 mg per kilogram were suspended in an aqueous solution of 0.9% NaCl at a 1:1 proportion (graphene or graphene oxide), i.e., 1 mg/mL. Results: Optical microscopy of liver, kidney, spleen, and lung tissues revealed no visible histological changes. However, particle traces were found in the peritoneal cavity. Thirty days after inoculation, blood samples were collected for hematological analysis. The blood analysis showed changes indicating a hepatic inflammatory process. Hematological changes after 30 days consisted of alterations to the red series, including microcytosis or higher mean hemoglobin concentrations. In addition, changes in prothrombin and thromboplastin caused longer coagulation times. Conclusion: This study contributes to further clarifying the possible toxicity of graphene and its potential biomedical applications.

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

CoCr alloys are extensively used in biomedical implants for joint replacements due to their excellent wear-corrosion resistance properties and good biocompatibility. Nevertheless, long term prostheses irretrievably undergo wear-corrosion phenomena causing debris and metallic dissolution that can induce adverse cellular reactions such as inflammatory complications, macrophage activation or bone resorption. Several studies can be found in the literature related to biological responses to debris and metallic particles, however, scarce works are addressed towards the biological response of cells contacting CoCr surfaces after tribocorrosion processes. In this study, the in vitro immune response on worn CoCr surfaces after tribocorrosion tests of modified graphenebased compounds on CoCr alloy in simulated physiological solutions (PBS plus 0.3% hyaluronic acid) has been studied. Graphene oxide was electrochemically reduced on CoCr surfaces (CoCrErGO) and further functionalized with hyaluronic acid (CoCrErGOHA). Wear-corrosion tests were carried out by using disks of 38 mm diameter in contact with a cylinder of 7 mm diameter as counterpart (pin). A normal load of 5N was applied on the counterpart at a rotation rate of 120 rpm for 30 Km. Wear-corrosion tests were performed in a phosphate Buffer Saline (PBS) solution with 3 g/L hyaluronic acid. Immune response and inflammatory balance induced by the exposition to the different surfaces modifications of CoCr disks, before and after wear-corrosion assays, was here analyzed in supernatants of mouse macrophage cell cultures (J774A.1).

Surface modification of CoCr biomedical implants arises for increasing durability of mechanical and corrosion properties in order to minimize adverse biological reactions in the human body. In this study, we focus on the reduction of graphene oxide as promising method of tailoring CoCr surfaces for further functionalization in order to regulate the responses of macrophages and osteoblasts contacting modified CoCr surfaces. Reduction of graphene oxide (GO) was performed on biomedical grade CoCr surfaces by using both electrochemical and chemical methods. Electrochemical reduction of GO was carried out by cyclic voltammetry after immersion of silanized CoCr surfaces in the aqueous solution of graphene oxide (4 mg/ml) at room temperature. Cyclic voltammetry was performed from -2,1 V to -0,5 V, vs. Ag/Ag/Cl, for 5 scans at 10 mV/min. Chemical reduction of GO was performed through ascorbic acid by maintaining a controlled temperature below 100ºC. The final suspension was dropped onto the silanized CoCr surfaces and allowed drying at room temperature. An extensive characterization of the modified CoCr surfaces has been performed by comparing both reduction methods. The corrosion performance of modified CoCr surfaces was assessed by the measurement of the corrosion potential and Electrochemical Impedance Spectroscopy (EIS) in 3 G/L hyaluronic acid solution.

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.

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.

Electrochemically reduced graphene oxide (ErGO) films on a biomedical grade CoCr alloy have been generated and characterized in order to study their possible application for use on joint prostheses. The electrodeposition process was performed by cyclic voltammetry. The characterization of the ErGO films on CoCr alloys by XPS revealed sp2 bonding and the presence of C[dbnd]O and C[sbnd]O residual groups in the graphene network. Biocompatibility studies were performed with mouse macrophages J774A.1 cell cultures measured by the ratio between lactate dehydrogenase and mitochondrial activities. An enhancement in the biocompatibility of the CoCr with the ErGO films was obtained, a result that became more evident as exposure time increased. Macrophages on the CoCr with the ErGO were well-distributed and conserved the characteristic cell shape. In addition, vimentin expression was unaltered in comparison with the control, results that indicated an improvement in the CoCr biocompatibility with the ErGO on the material surface. The in vivo response of graphene and graphene oxide was assessed by intraperitoneal injection in wistar rats. Red blood cells are one of the primary interaction sites so hemocompatibility tests were carried out. Rats inoculated with graphene and graphene oxide showed red blood cells of smaller size with a high content in hemoglobin.

A. PRESENTACIÓN DE ÁREAS DE INVESTIGACIÓN: CORROSIÓN EN BIOMATERIALES METÁLICOS B. CASO PRÁCTICO: ESTUDIO DEL COMPORTAMIENTO FRENTE A LA CORROSIÓN DE DIFERENTES MATERIALES METÁLICOS UTILIZADOS EN APLICACIONES BIOMÉDICAS. C. PROCEDIMIENTO EXPERIMENTAL: a). MATERIALES: Cu, Ti, CoCrMo, acero inoxidable, acero al carbono, Zn, Al b). PREPARACIÓN SUPERFICIE DE MUESTRAS: - Desbaste con papel de lija de diferentes granulometrías (600, 1200), - Cálculo de la superficie de ensayo c). Disoluciones D). METODOLOGÍA a) Ensayo gravimétrico: Zn, acero b) Ensayos electroquímicos: - Montaje de celdas electroquímicas de corrosión - Determinación de parámetros electroquímicos en las distintas soluciones (ver anexo) - Potencial de corrosión (Ecorr). - Resistencia a la polarización (Rp) - Morfología de ataque de corrosión después de los ensayos (corrosión generalizada, corrosión por picaduras,…)

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.

Actuar con criterios de seguridad e higiene en sus actividades en el puesto de trabajo. - Responder adecuadamente en condiciones de emergencia simuladas. - Planificar las actuaciones a realizar para llevar a cabo los ensayos u operaciones de proceso de acuerdo con los procedimientos establecidos o según el manual de calidad de la empresa. - Obtener productos químicos aplicando operaciones de formulación y transformación realizando de forma autónoma los ensayos correspondientes. - Comportarse de forma responsable en el centro de trabajo e integrarse en el sistema de relaciones técnico-sociales de la empresa.

Estudio del comportamiento frente a la corrosión de diferentes materiales metálicos utilizados en aplicaciones biomédicas. METODOLOGÍA a) Ensayo gravimétrico: Zn, acero b) Ensayos electroquímicos: - Montaje de celdas electroquímicas de corrosión - Determinación de parámetros electroquímicos en las distintas soluciones (ver anexo) - Potencial de corrosión (Ecorr). - Resistencia a la polarización (Rp) - Morfología de ataque de corrosión después de los ensayos (corrosión generalizada, corrosión por picaduras)

In memory of Dr. Blanca Teresa Perez-Maceda whose lovely life, plenty of fortitude, joy and spiritual values, we had the honour of sharing. Electrochemically reduced graphene oxide (ErGO) films on a biomedical grade CoCr alloy have been generated and characterized in order to study the biocompatibility and in vivo behavior for its possible biomedical application. The electrodeposition process was performed by cyclic voltammetry from -2.1 V to -0.5 V at a scanning rate of 10 mV s-1 for five cycles. ErGO/CoCr surface analysis was studied by XPS. Electrochemical techniques such as the measurement of the corrosion potential, Ecorr, and electrochemical impedance spectroscopy (EIS) were applied to assess the corrosion behavior. ErGO films on CoCr were further functionalized with hyaluronic acid (ErGOHA) after 24 hours soaking in a solution containing 3 g/L, a concentration equivalent to the content of the synovial fluid. Biocompatibility and cytotoxicity of CoCr, ErGO/CoCr, and ErGOHA/CoCr studies were performed in mouse macrophages J774A.1 cell cultures by measuring mitochondrial activity (WST-1 assay), plasma membrane damage (LDH assay) and the ratio LDH/WST-1, as relate plasma membrane damage index to the number of metabolically active cells in culture. Pattern distribution and morphology of macrophages on ErGO/CoCr and ErGOHA/CoCr alloys were performed by optical microscopy. The in vivo response of graphene oxide used in the electrochemical reduction was assessed by intraperitoneal injection in wistar rats. Graphene was also used as reference. Graphene (G) and graphene oxide (GO) nanosheets at a concentration of 4 mg per kilogram of weight were suspended into an aqueous solution of 0.9% NaCl in a proportion 1:1 (PBS:G or GO), i.e., 1 mg/ml. After 30 day¿s post-inoculation, blood extraction was performed in the rats in order to carry out an hematological analysis. XPS revealed sp2 bonding and the presence of C=O and C-O residual groups in the graphene network. ErGO fil

In this work, synthesis of copper/reduced graphene oxide composites to increase the electrical conductivity of copper matrix has been carried out. Pure copper particles were mixed with 0.28 wt%, 1.1 wt% and 2.5 wt% graphene oxide (GO) contents. Different reduction methods were chosen to synthetize the Cu/rGO composite material: chemical reduction with hydrazine, thermal treatment at 815 °C in argon atmosphere. Characterization of the Cu/rGO composite disks was carried out by optical microscopy, scanning electron microscopy, X-ray photoelectron spectroscopy and Raman spectroscopy. Vickers Hardness and the electrical conductivity measured by Van der Pauw method of the Cu/rGO composites were analyzed. All the Cu/rGO disks showed lower hardness values than the Cu matrix. Microstructure of Cu/rGO composites is dominated by the presence of pores where GO agglomerates are accumulated and whose distribution and magnitude depends on the weight percentage of GO and the reduction process. Porosity percentage and pore size of Cu/rGO disks increase with GO content. Combination of chemical reduction with thermal treatment provides the most homogeneous microstructures with the smallest pore size and porosity percentage. Combination of both reduction treatments promotes more pathways for carrier transport within the reduced graphene oxide network structure, expanding areas of long-range conjugated structures and, consequently, increasing the electrical conductivity in the rGO/copper composites. Our research evidences that the Cu/rGO composites after chemical and thermal reduction increases the electrical conductivity, independently on the GO content, with respect to the pure copper matrix.

Flame Retardants (FR) are a group of anthropogenic environmental contaminants used at a relatively high concentration in many applications. Currently, the largest marked group of FRs is halogenated FR, and many of them are considered toxic, persistent and bio accumulative. Non-halogenated alternatives are a possible solution for the problem, but there is a lack of knowledge concerning environmental impact, health risks during the production process and final use. The main objective of the LIFE-FLAREX project that supports this work, is the mitigation of the environmental and human health impact of flame retardants used in textiles, looking for new efficient more ecological and healthy alternatives, able to replace the most common FR’s that include toxic compounds like halogens, formaldehyde and antimony. The aim of this work is the determination of the effect of conventional and ecological flame retardants on cotton and polyester fabrics by the application of differential scanning calorimetry DSC and thermogravimetric analysis TGA. Results have been compared with those given by the best FR applied to cotton/polyester blended fabric. The application of DSC up to 550ºC and TGA up to 600ºC in N2 and O2 atmospheres give results that are in accordance with those yielded by the micro-scale combustion calorimeter. Onset temperatures of decomposition, steps of loss of mass by temperature and final residues, enable to evaluate the thermal efficiency of the different flame retardants. The considered flame retardants are based on expandable graphite, dialkyl phosphono carboxylic acid amide and decabrome diphenyl ethane with antimony trioxide. The results have been compared with those given by the application of ammonium polyphosphate and guanidine phosphate on cotton/polyester 50/50 blend.

In the present work, the characterization of Electrochemically Reduced Graphene Oxide (ErGO) films on a biomedical grade CoCr alloy has been performed. The direct electrodeposition process was carried out by means of two electrochemical techniques: chronoamperometry and cyclic voltammetry experiments, both taking into account the influence of oxygen in the solution. Characterization of the resulting ErGO films on CoCr alloys was carried out by SEM, AFM, Raman spectroscopy and XPS. Both electrochemical reduction procedures led to the deposition of graphene-based films on the CoCr surfaces after the partial removal of the oxygenated functional groups present in the graphene network of the starting material. The deconvolution of the XPS high-resolution C1s and O1s spectra mainly showed Csp2 bonding and the presence of CO and CO residual groups covering the CoCr surface. Moreover, the ErGO films that were stacked on the CoCr surfaces exhibited non-uniform thickness reaching values corresponding to the presence of 80 layers of the graphene material. In addition, Raman spectroscopy revealed a certain structural disorder in the ErGO films deposited by means of the different electrochemical techniques investigated. This fact was more evident on the film from the chronoamperometric experiments, as the fastest electrochemical reduction rate provided by the application of a high reduction potential (−2.1 Vvs. Ag/AgCl) could induce the development of defects in the resulting film, so the peaks corresponding to the CoCr substrate could be observed.