Calcium Phosphate Cements Combined with Blood as a Promising Tool for the Treatment of Bone Marrow Lesions.

The solid phase of a commercial calcium phosphate (Graftys® HBS) was combined with ovine or human blood stabilized either with sodium citrate or sodium heparin. The presence of blood delayed the setting reaction of the cement by ca. 7-15 h, depending on the nature of the blood and blood stabilizer. This phenomenon was found to be directly related to the particle size of the HBS solid phase, since prolonged grinding of the latter resulted in a shortened setting time (10-30 min). Even though ca. 10 h were necessary for the HBS blood composite to harden, its cohesion right after injection was improved when compared to the HBS reference as well as its injectability. A fibrin-based material was gradually formed in the HBS blood composite to end-up, after ca. 100 h, with a dense 3D organic network present in the intergranular space, thus affecting the microstructure of the composite. Indeed, SEM analyses of polished cross-sections showed areas of low mineral density (over 10-20 µm) spread in the whole volume of the HBS blood composite. Most importantly, when the two cement formulations were injected in the tibial subchondral cancellous bone in a bone marrow lesion ovine model, quantitative SEM analyses showed a highly significant difference between the HBS reference versus its analogue combined with blood. After a 4-month implantation, histological analyses clearly showed that the HBS blood composite underwent high resorption (remaining cement: ca. 13.1 ± 7.3%) and new bone formation (newly formed bone: 41.8 ± 14.7%). This was in sharp contrast with the case of the HBS reference for which a low resorption rate was observed (remaining cement: 79.0 ± 6.9%; newly formed bone: 8.6 ± 4.8%). This study suggested that the particular microstructure, induced by the use of blood as the HBS liquid phase, favored quicker colonization of the implant and acceleration of its replacement by newly formed bone. For this reason, the HBS blood composite might be worth considering as a potentially suitable material for subchondroplasty.

Electrochemical analysis of a microbial electrochemical snorkel in laboratory and constructed wetlands.

Microbial electrochemical snorkel (MES) is a short-circuited microbial fuel cell applicable to water treatment that does not produce energy but requires lower cost for its implementation. Few reports have already described its water treatment capabilities but no deeper electrochemical analysis were yet performed. We tested various materials (iron, stainless steel and porous graphite) and configurations of snorkel in order to better understand the rules that will control in a wetland the mixed potential of this self-powered system. We designed a model snorkel that was studied in laboratory and on the field. We confirmed the development of MES by identifying anodic and cathodic parts, by measuring the current between them and by analyzing microbial ecology in laboratory and field experiments. An important application is denitrification of surface water. Here we discuss the influence of nitrate on its electrochemical response and denitrification performances. Introducing nitrate caused the increase of the mixed potential of MES and of current at a potential value relatively more positive than for nitrate-reducing biocathodes described in the literature. The major criteria for promoting application of MES in artificial wetland dedicated to mitigation of non-point source nitrate pollution from agricultural water are considered.

Combination of biocompatible hydrogel precursors to apatitic calcium phosphate cements (CPCs): Influence of the in situ hydrogel reticulation on the CPC properties.

In the field of bone regenerative medicine, injectable calcium phosphate cements (CPCs) are used for decades in clinics, as bone void fillers. Most often preformed polymers (e.g., hyaluronic acid, collagen, chitosan, cellulose ethers…) are introduced in the CPC formulation to make it injectable and improve its cohesion. Once the cement has hardened, the polymer is simply trapped in the CPC structure and no organic subnetwork is present. By contrast, in this work a CPC was combined with organic monomers that reticulated in situ so that a continuous biocompatible 3D polymeric subnetwork was formed in the CPC microstructure, resulting in a higher permeability of the CPC, which might allow to accelerate its in vivo degradation. Two options were investigated depending on whether the polymer was formed before the apatitic inorganic network or concomitantly. In the former case, conditions were found to reach a suitable rheology for easy injection of the composite. In addition, the in situ formed polymer was shown to strongly affect the size, density, and arrangement of the apatite crystals formed during the setting reaction, thereby offering an original route to modulate the microstructure and porosity of apatitic cements.

Signal amplification by electro-oligomerisation for improved isoproturon detection.

A novel concept is introduced for signal amplification in electrochemical sensing: the electro-oligomerisation stripping voltammetry, which has been applied here to the improved detection of the isoproturon herbicide in spring waters as a proof-of-principle. It involves a potentiostatic accumulation step onto a glassy carbon electrode (at +1.5 V vs Ag/AgCl reference electrode for 300 s) leading to the formation of an oligomeric film, which is then detected by cathodic stripping square wave voltammetry (SWV). The presence and composition of the film are confirmed by confocal Raman spectroscopy. Its characterisation by cyclic voltammetry demonstrates the reversible nature of the electrodeposited material, confirming its interest for sensitive detection by SWV. Adding a mesoporous silica membrane with vertically oriented nanochannels further enhances the sensitivity of the sensor, exhibiting a linear response in the 10-100 µM concentration range. This effect was even more interesting for real media analysis thanks to the permselective properties of such nanoporous coating in rejecting interferences and/or surface fouling agents. The method should be applicable to other analytes that are usually not detectable by conventional accumulation/stripping voltammetry.

Mesoporous Silica Thin Films for Improved Electrochemical Detection of Paraquat.

An electrochemical method was developed for rapid and sensitive detection of the herbicide paraquat in aqueous samples using mesoporous silica thin film modified glassy carbon electrodes (GCE). Vertically aligned mesoporous silica thin films were deposited onto GCE by electrochemically assisted self-assembly (EASA). Cyclic voltammetry revealed effective response to the cationic analyte (while rejecting anions) thanks to the charge selectivity exhibited by the negatively charged mesoporous channels. Square wave voltametry (SWV) was then used to detect paraquat via its one electron reduction process. Influence of various experimental parameters (i.e., pH, electrolyte concentration, and nature of electrolyte anions) on sensitivity was investigated and discussed with respect to the mesopore characteristics and accumulation efficiency, pointing out the key role of charge distribution in such confined spaces on these processes. Calibration plots for paraquat concentration ranging from 10 nM to 10 µM were constructed at mesoporous silica modified GCE which were linear with increasing paraquat concentration, showing dramatically enhanced sensitivity (almost 30 times) as compared to nonmodified electrodes. Finally, real samples from Meuse River (France) spiked with paraquat, without any pretreatment (except filtration), were analyzed by SWV, revealing the possible detection of paraquat at very low concentration (10-50 nM). Limit of detection (LOD) calculated from real sample analysis was found to be 12 nM, which is well below the permissible limits of paraquat in drinking water (40-400 nM) in various countries.

Design and properties of a novel radiopaque injectable apatitic calcium phosphate cement, suitable for image-guided implantation.

An injectable purely apatitic calcium phosphate cement (CPC) was successfully combined to a water-soluble radiopaque agent (i.e., Xenetix® ), to result in an optimized composition that was found to be as satisfactory as poly(methyl methacrylate) (PMMA) formulations used for vertebroplasty, in terms of radiopacity, texture and injectability. For that purpose, the Xenetix dosage in the cement paste was optimized by injection of the radiopaque CPC in human cadaveric vertebrae under classical PMMA vertebroplasty conditions, performed by interventional radiologists familiar with this surgical procedure. When present in the cement paste up to 70 mg I mL-1 , Xenetix did not influence the injectability, cohesion, and setting time of the resulting composite. After hardening of the material, the same observation was made regarding the microstructure, mechanical strength and alpha-tricalcium phosphate to calcium deficient apatite transformation rate. Upon implantation in bone in a small animal model (rat), the biocompatibility of the Xenetix-containing CPC was evidenced. Moreover, an almost quantitative release of the contrast agent was found to occur rapidly, on the basis of in vitro static and dynamic quantitative studies simulating in vivo implantation. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 2786-2795, 2018.

A straightforward approach to enhance the textural, mechanical and biological properties of injectable calcium phosphate apatitic cements (CPCs): CPC/blood composites, a comprehensive study.

Two commercial formulations of apatitic calcium phosphate cements (CPCs), Graftys® Quickset (QS) and Graftys® HBS (HBS), similar in composition but with different initial setting time (7 and 15min, respectively), were combined to ovine whole blood. Surprisingly, although a very cohesive paste was obtained after a few minutes, the setting time of the HBS/blood composite dramatically delayed when compared to its QS analogue and the two blood-free references. Using solid state NMR, scanning electron microscopy and high frequency impedance measurements, it was shown that, in the particular case of the HBS/blood composite, formation of a reticulated and porous organic network occurred in the intergranular space, prior to the precipitation of apatite crystals driven by the cement setting process. The resulting microstructure conferred unique biological properties to this material upon implantation in bone defects, since its degradation rate after 4 and 12weeks was more than twice that for the three other CPCs, with a significant replacement by newly formed bone. STATEMENT OF SIGNIFICANCE: A major challenge in the design of bone graft substitutes is the development of injectable, cohesive, resorbable and self-setting calcium phosphate cement (CPC) that enables rapid cell invasion with initial mechanical properties as close as bone ones. Thus, we describe specific conditions in CPC-blood composites where the formation of a 3D clot-like network can interact with the precipitated apatite crystals formed during the cement setting process. The resulting microstructure appears more ductile at short-term and more sensitive to biological degradation which finally promotes new bone formation. This important and original paper reports the design and in-depth chemical and physical characterization of this groundbreaking technology.

Sorption of polluting metal ions on a palm tree frond sawdust studied by the means of modified carbon paste electrodes.

Water remediation by adsorption of the metal ions on a low cost sorbent is the frame of the present study. The metal ions adsorption properties of sawdust of palm tree fronds (PTF sawdust) are investigated by both equilibrium measurements and modified carbon paste electrode. The ability to adsorb Cu(II), Cr(VI) and As(III) in significant quantities is demonstrated. Carbon paste electrodes modified by incorporation of PTF sawdust (PTF-CPE) or, for comparison, an organically modified silica for the detection of copper(II) are investigated in term of sensitivity, estimation of number of possible reuses, repeatability and interference effect. A detection limit for Cu(II) analysis of 1.0×10(-8) M has been achieved after 5 min preconcentration and a single PTF-CPE can be used for up to 10 preconcentration-analysis-regeneration cycles. The relative standard deviation (n=9) for the determination of a 10(-6) M Cu(II) solution (pH=5) was about 26%. The effects of Ca(II), As(III) and Cr(VI) on the copper detection are investigated: calcium ions were shown to compete with copper on the same adsorption sites, arsenic(III) has no effect on the copper detection whereas chromium(VI) was shown to enhance the copper detection.

Immobilization of membrane-bounded (S)-mandelate dehydrogenase in sol-gel matrix for electroenzymatic synthesis.

Membrane-bounded (S)-mandelate dehydrogenase has been immobilized on the surface of glassy carbon and carbon felt electrodes by encapsulation in a silica film obtained by sol-gel chemistry. Such bioelectrochemical system has been used for the first time for electroenzymatic conversion of (S)-mandelic acid to phenylglyoxylic acid. Apparent Km in this sol-gel matrix was 0.7 mM in the presence of ferrocenedimethanol, a value in the same order of magnitude as reported previously for vesicles in solution with other electron acceptors, i.e., Fe(CN)6(3-) or 2,6-dichloroindophenol. The bioelectrode shows very good operational stability for more than 6 days. This stability was definitively improved by comparison to a bioelectrode prepared by simple adsorption of the proteins on the electrode surface (fast activity decrease during the first 15 h of experiment). Optimal electroenzymatic reaction was achieved at pH9 and 40 °C. Apparent Km of the protein activity was 3 times higher in carbon felt electrode than on glassy carbon surface, possibly because of transport limitations in the porous architecture of the carbon felt. A good correlation was found between electrochemical data and chromatographic characterization of the reaction products in the bioelectrochemical reactor.

Abiotic process for Fe(II) oxidation and green rust mineralization driven by a heterotrophic nitrate reducing bacteria (Klebsiella mobilis).

Green rusts (GRs) are mixed Fe(II)-Fe(III) hydroxides with a high reactivity toward organic and inorganic pollutants. GRs can be produced from ferric reducing or ferrous oxidizing bacterial activities. In this study, we investigated the capability of Klebsiella mobilis to produce iron minerals in the presence of nitrate and ferrous iron. This bacterium is well-known to reduce nitrate using an organic carbon source as electron donor but is unable to enzymatically oxidize Fe(II) species. During incubation, GR formation occurred as a secondary iron mineral precipitating on cell surfaces, resulting from Fe(II) oxidation by nitrite produced via bacterial respiration of nitrate. For the first time, we demonstrate GR formation by indirect microbial oxidation of Fe(II) (i.e., a combination of biotic/abiotic processes). These results therefore suggest that nitrate-reducing bacteria can potentially contribute to the formation of GR in natural environments. In addition, the chemical reduction of nitrite to ammonium by GR is observed, which gradually turns the GR into the end-product goethite. The nitrogen mass-balance clearly demonstrates that the total amount of ammonium produced corresponds to the quantity of bioreduced nitrate. These findings demonstrate how the activity of nitrate-reducing bacteria in ferrous environments may provide a direct link between the biogeochemical cycles of nitrogen and iron.

High Frequency Impedance Measurement as a Relevant Tool for Monitoring the Apatitic Cement Setting Reaction.

This work reports the development of a relevant and general method based on high frequency impedance measurements, for the in situ monitoring of the alpha-tricalcium phosphate (α-TCP) to calcium-deficient hydroxyapatite (CDA) transformation which is the driving force of the hardening processes of some calcium phosphate cements (CPC) used as bone substitutes. The three main steps of the setting reaction are identified in a non invasive way through the variation of dielectric permittivity and dielectric losses. The method is also likely to characterize the effect of the incorporation of additives (i.e, antiosteoporotic bisphosphonate drugs such as Alendronate) in the CPC formulation on the hydration process. It allows not only to confirm the retarding effect of bisphosphonate by an accurate determination of setting times, but also to assess the phenomena taking place whether alendronate is added in the liquid phase or combined to the solid phase of the cement composition. Compared to the conventional Gillmore needle test, the present method offers the advantage of accurate, user-independent, in situ and real-time determination of the initial and final times of the chemical hardening process, which are important parameters when considering surgical applications.

Investigation of alendronate-doped apatitic cements as a potential technology for the prevention of osteoporotic hip fractures: critical influence of the drug introduction mode on the in vitro cement properties.

Combination of a bisphosphonate (BP) anti-osteoporotic drug, alendronate, with an apatitic calcium phosphate cement does not significantly affect the main properties of the biomaterial, in terms of injectability and setting time, provided that the BP is introduced chemisorbed onto calcium-deficient apatite, one of the components of the cement. In contrast to other modes of introducing the BP into the cement formulation, this mode allows to minimize alendronate release in the cement paste, thus limiting the setting retardant effect of the BP. An original approach based on high frequency impedance measurements is found to be a convenient method for in situ monitoring of the cement setting reaction. The release profile of the drug from a cement block under continuous flow conditions can be well described using a coupled chemistry/transport model, under simulated in vivo conditions. The results show that the released alendronate concentration is expected to be much lower than the cytotoxic concentration.