Autologous patient-derived exhausted nano T-cells exploit tumor immune evasion to engage an effective cancer therapy.

BACKGROUND: Active targeting by surface-modified nanoplatforms enables a more precise and elevated accumulation of nanoparticles within the tumor, thereby enhancing drug delivery and efficacy for a successful cancer treatment. However, surface functionalization involves complex procedures that increase costs and timelines, presenting challenges for clinical implementation. Biomimetic nanoparticles (BNPs) have emerged as unique drug delivery platforms that overcome the limitations of actively targeted nanoparticles. Nevertheless, BNPs coated with unmodified cells show reduced functionalities such as specific tumor targeting, decreasing the therapeutic efficacy. Those challenges can be overcome by engineering non-patient-derived cells for BNP coating, but these are complex and cost-effective approaches that hinder their wider clinical application. Here we present an immune-driven strategy to improve nanotherapeutic delivery to tumors. Our unique perspective harnesses T-cell exhaustion and tumor immune evasion to develop a groundbreaking new class of BNPs crafted from exhausted T-cells (NExT) of triple-negative breast cancer (TNBC) patients by specific culture methods without sophisticated engineering. METHODS: NExT were generated by coating PLGA (poly(lactic-co-glycolic acid)) nanoparticles with TNBC-derived T-cells exhausted in vitro by acute activation. Physicochemical characterization of NExT was made by dynamic light scattering, electrophoretic light scattering and transmission electron microscopy, and preservation and orientation of immune checkpoint receptors by flow cytometry. The efficacy of chemotherapy-loaded NExT was assessed in TNBC cell lines in vitro. In vivo toxicity was made in CD1 mice. Biodistribution and therapeutic activity of NExT were determined in cell-line- and autologous patient-derived xenografts in immunodeficient mice. RESULTS: We report a cost-effective approach with a good performance that provides NExT naturally endowed with immune checkpoint receptors (PD1, LAG3, TIM3), augmenting specific tumor targeting by engaging cognate ligands, enhancing the therapeutic efficacy of chemotherapy, and disrupting the PD1/PDL1 axis in an immunotherapy-like way. Autologous patient-derived NExT revealed exceptional intratumor accumulation, heightened chemotherapeutic index and efficiency, and targeted the tumor stroma in a PDL1+ patient-derived xenograft model of triple-negative breast cancer. CONCLUSIONS: These advantages underline the potential of autologous patient-derived NExT to revolutionize tailored adoptive cancer nanotherapy and chemoimmunotherapy, which endorses their widespread clinical application of autologous patient-derived NExT.

pH-Dependent Adsorption Release of Doxorubicin on MamC-Biomimetic Magnetite Nanoparticles.

New biomimetic magnetite nanoparticles (hereafter BMNPs) with sizes larger than most common superparamagnetic nanoparticles were produced in the presence of the recombinant MamC protein from Magnetococcus marinus MC-1 and functionalized with doxorubicin (DOXO) intended as potential drug nanocarriers. Unlike inorganic magnetite nanoparticles, in BMNPs the MamC protein controls their size and morphology, providing them with magnetic properties consistent with a large magnetic moment per particle; moreover, it provides the nanoparticles with novel surface properties. BMNPs display the isoelectric point at pH 4.4, being strongly negatively charged at physiological pH (pH 7.4). This allows both (i) their functionalization with DOXO, which is positively charged at pH 7.4, and (ii) the stability of the DOXO-surface bond and DOXO release to be pH dependent and governed by electrostatic interactions. DOXO adsorption follows a Langmuir-Freundlich model, and the coupling of DOXO to BMNPs (binary biomimetic nanoparticles) is very stable at physiological pH (maximum release of 5% of the drug adsorbed). Conversely, when pH decreases, these electrostatic interactions weaken, and at pH 5, DOXO is released up to ∼35% of the amount initially adsorbed. The DOXO-BMNPs display cytotoxicity on the GTL-16 human gastric carcinoma cell line in a dose-dependent manner, reaching about ∼70% of mortality at the maximum amount tested, while the nonloaded BMNPs are fully cytocompatible. The present data suggest that BMNPs could be useful as potential drug nanocarriers with a drug adsorption-release governed by changes in local pH values.

Monoclonal antibody-targeted fluorescein-5-isothiocyanate-labeled biomimetic nanoapatites: a promising fluorescent probe for imaging applications.

Multifunctional biomimetic nanoparticles (NPs) are acquiring increasing interest as carriers in medicine and basic research since they can efficiently combine labels for subsequent tracking, moieties for specific cell targeting, and bioactive molecules, e.g., drugs. In particular, because of their easy synthesis, low cost, good biocompatibility, high resorbability, easy surface functionalization, and pH-dependent solubility, nanocrystalline apatites are promising candidates as nanocarriers. This work describes the synthesis and characterization of bioinspired apatite nanoparticles to be used as fluorescent nanocarriers targeted against the Met/hepatocyte growth factor receptor, which is considered a tumor associated cell surface marker of many cancers. To this aim the nanoparticles have been labeled with Fluorescein-5-isothiocyanate (FITC) by simple isothermal adsorption, in the absence of organic, possibly toxic, molecules, and then functionalized with a monoclonal antibody (mAb) directed against such a receptor. Direct labeling of the nanoparticles allowed tracking the moieties with spatiotemporal resolution and thus following their interaction with cells, expressing or not the targeted receptor, as well as their fate in vitro. Cytofluorometry and confocal microscopy experiments showed that the functionalized nanocarriers, which emitted a strong fluorescent signal, were rapidly and specifically internalized in cells expressing the receptor. Indeed, we found that, once inside the cells expressing the receptor, mAb-functionalized FITC nanoparticles partially dissociated in their two components, with some mAbs being recycled to the cell surface and the FITC-labeled nanoparticles remaining in the cytosol. This work thus shows that FITC-labeled nanoapatites are very promising probes for targeted cell imaging applications.

pH-responsive collagen fibrillogenesis in confined droplets induced by vapour diffusion.

A novel methodology for the assembly of collagen fibrils in microliter drops is proposed. It consists in the gradual increase of pH by means of vapour diffusion coming from the decomposition of NH4HCO3 solutions. The pH increase rate as well as the final steady pH of solutions containing collagen can be adjusted by varying the concentration of NH4HCO3. Both parameters are of predominant importance in collagen fibrillogenesis. The effect of these parameters on the kinetic of the fibrillogenesis process and on the fibrils morphology was studied. We found that both the kinetic and the morphology are mainly driven by electrostatic interactions. A gradual increase of pH slows down the formation of collagen fibres and favours the lateral interaction between fibrils producing broader fibres. On the other hand, a rapid increase of pH reduces the lateral electrostatic interactions favouring the formation of thinner fibres. The formation of the D-band periodicity is also a pH-dependent process that occurs after fibrillogenesis when the most stable state of fibres formation has been reached.

Nanocrystalline and Amorphous Calcium Carbonate from Waste Seashells by Ball Milling Mechanochemistry Processes.

Nanocrystalline calcium carbonate (CaCO3) and amorphous CaCO3 (ACC) are materials of increasing technological interest. Nowadays, they are mainly synthetically produced by wet reactions using CaCO3 reagents in the presence of stabilizers. However, it has recently been discovered that ACC can be produced by ball milling calcite. Calcite and/or aragonite are the mineral phases of mollusk shells, which are formed from ACC precursors. Here, we investigated the possibility to convert, on a potentially industrial scale, the biogenic CaCO3 (bCC) from waste mollusk seashells into nanocrystalline CaCO3 and ACC. Waste seashells from the aquaculture species, namely oysters (Crassostrea gigas, low-Mg calcite), scallops (Pecten jacobaeus, medium-Mg calcite), and clams (Chamelea gallina, aragonite) were used. The ball milling process was carried out by using different dispersing solvents and potential ACC stabilizers. Structural, morphological, and spectroscopic characterization techniques were used. The results showed that the mechanochemical process produced a reduction of the crystalline domain sizes and formation of ACC domains, which coexisted in microsized aggregates. Interestingly, bCC behaved differently from the geogenic CaCO3 (gCC), and upon long milling times (24 h), the ACC reconverted into crystalline phases. The aging in diverse environments of mechanochemically treated bCC produced a mixture of calcite and aragonite in a species-specific mass ratio, while the ACC from gCC converted only into calcite. In conclusion, this research showed that bCC can produce nanocrystalline CaCO3 and ACC composites or mixtures having species-specific features. These materials can enlarge the already wide fields of applications of CaCO3, which span from medical to material science.

Stearate-Coated Biogenic Calcium Carbonate from Waste Seashells: A Sustainable Plastic Filler.

Waste seashells from aquaculture are a massive source of biogenic calcium carbonate (bCC) that can be a potential substitute for ground calcium carbonate and precipitated calcium carbonate. These last materials find several applications in industry after a surface coating with hydrophobic molecules, with stearate as the most used. Here, we investigate for the first time the capability of aqueous stearate dispersions to coat bCC powders from seashells of market-relevant mollusc aquaculture species, namely the oyster Crassostrea gigas, the scallop Pecten jacobaeus, and the clam Chamelea gallina. The chemical-physical features of bCC were extensively characterized by different analytical techniques. The results of stearate adsorption experiments showed that the oyster shell powder, which is the bCC with a higher content of the organic matrix, showed the highest adsorption capability (about 23 wt % compared to 10 wt % of geogenic calcite). These results agree with the mechanism proposed in the literature in which stearate adsorption mainly involves the formation of calcium stearate micelles in the dispersion before the physical adsorption. The coated bCC from oyster shells was also tested as fillers in an ethylene vinyl acetate compound used for the preparation of shoe soles. The obtained compound showed better mechanical performance than the one prepared using ground calcium. In conclusion, we can state that bCC can replace ground and precipitated calcium carbonate and has a higher stearate adsorbing capability. Moreover, they represent an environmentally friendly and sustainable source of calcium carbonate that organisms produce by high biological control over composition, polymorphism, and crystal texture. These features can be exploited for applications in fields where calcium carbonate with selected features is required.

Analysis of the structural integrity of SU-8-based optofluidic systems for small-molecule crystallization studies.

The use of SU-8-based optofluidic systems (OFS) is validated as an affordable and easy alternative to expensive glass device manufacturing for small-molecule crystallization studies and, in comparison with other polymers, able to withstand most organic solvents. A comparison between two identical OFS (using SU-8 and poly(dimethylsiloxane), PDMS) against the 36 most commonly used organic solvents for small-molecule crystallization studies have confirmed both the structural and optical stability of the SU-8, whereas PDMS suffered from unsealing or tearing in most cases. In order to test its compatibility, measurements before and after 24 h of continued exposure against solvents have been pursued. Here, three aspects have been considered: in the macroscale, swelling has been determined by analyzing the variations in the optical path in the OFS. For determining compatibility at microscale, fabricated SU-8 micropatterns were solvent-etched and subsequently characterized by scanning electron microscopy (SEM). Roughness of the polymer has also been studied through atomic force microscopy (AFM) measurements at the nanoscale. Experimental measurements of PDMS swelling were in accordance with previously reported observations, while SU-8 displayed a great stability against all the tested solvents. Through this experimental procedure we also show that the OFS are suitable for real-time, on-chip, UV-vis spectroscopy. Micro- and nanoscale observations did not show apparent corrosion on SU-8 surface. Also, two commonly used carrier fluids for microdroplet generation (FC-70 Fluorinert oil and silicone oil) were also tested against the different solvents with the aim of providing useful information for later microbatch experiments.

Cell surface receptor targeted biomimetic apatite nanocrystals for cancer therapy.

Nanosized drug carriers functionalized with moieties specifically targeting tumor cells are promising tools in cancer therapy, due to their ability to circulate in the bloodstream for longer periods and their selectivity for tumor cells, enabling the sparing of healthy tissues. Because of its biocompatibility, high bioresorbability, and responsiveness to pH changes, synthetic biomimetic nanocrystalline apatites are used as nanocarriers to produce multifunctional nanoparticles, by coupling them with the chemotherapeutic drug doxorubicin (DOXO) and the DO-24 monoclonal antibody (mAb) directed against the Met/Hepatocyte Growth Factor receptor (Met/HGFR), which is over-expressed on different types of carcinomas and thus represents a useful tumor target. The chemical-physical features of the nanoparticles are fully investigated and their interaction with cells expressing (GTL-16 gastric carcinoma line) or not expressing (NIH-3T3 fibroblasts) the Met/HGFR is analyzed. Functionalized nanoparticles specifically bind to and are internalized in cells expressing the receptor (GTL-16) but not in the ones that do not express it (NIH-3T3). Moreover they discharge DOXO in the targeted GTL-16 cells that reach the nucleus and display cytotoxicity as assessed in an MTT assay. Two different types of ternary nanoparticles are prepared, differing for the sequence of the functionalization steps (adsorption of DOXO first and then mAb or vice versa), and it is found that the ones in which mAb is adsorbed first are more efficient under all the examined aspects (binding, internalization, cytotoxicity), possibly because of a better mAb orientation on the nanoparticle surface. These multifunctional nanoparticles could thus be useful instruments for targeted local or systemic drug delivery, allowing a reduction in the therapeutic dose of the drug and thus adverse side effects. Moreover, this work opens new perspectives in the use of nanocrystalline apatites as a new platform for theranostic applications in nanomedicine.

Monitoring picoliter sessile microdroplet dynamics shows that size does not matter.

We monitor the dissolution of arrayed picoliter-size sessile microdroplets of the aqueous phase in oil, generated using a recently developed fluidic device. Initial pinning of the microdroplet perimeter leads to a nearly constant contact diameter, thus contraction proceeds via microdroplet (micrometer-diameter) height and contact angle reductions. This confirms that picoliter microdroplets contraction or dissolution due to the selective diffusion of water in oil has comparable dynamics with microliter droplet evaporation in air. We observe a constant microdroplet dissolution rate in different aqueous solutions. The application of this simple model to solvent-diffusion-driven crystallization experiments in confined volumes, for instance, would allow us to determine precisely the concentration in the microdroplet during an experiment and particularly at nucleation.

pH-responsive delivery of doxorubicin from citrate-apatite nanocrystals with tailored carbonate content.

In this work, the efficiency of bioinspired citrate-functionalized nanocrystalline apatites as nanocarriers for delivery of doxorubicin (DOXO) has been assessed. The nanoparticles were synthesized by thermal decomplexing of metastable calcium/citrate/phosphate solutions both in the absence (Ap) and in the presence (cAp) of carbonate ions. The presence of citrate and carbonate ions in the solution allowed us to tailor the size, shape, carbonate content, and surface chemistry of the nanoparticles. The drug-loading efficiency of the two types of apatite was evaluated by means of the adsorption isotherms, which were found to fit a Langmuir-Freundlich behavior. A model describing the interaction between apatite surface and DOXO is proposed from adsorption isotherms and ζ-potential measurements. DOXO is adsorbed as a dimer by means of a positively charged amino group that electrostatically interacts with negatively charged surface groups of nanoparticles. The drug-release profiles were explored at pHs 7.4 and 5.0, mimicking the physiological pH in the blood circulation and the more acidic pH in the endosome-lysosome intracellular compartment, respectively. After 7 days at pH 7.4, cAp-DOXO released around 42% less drug than Ap-DOXO. However, at acidic pH, both nanoassemblies released similar amounts of DOXO. In vitro assays analyzed by confocal microscopy showed that both drug-loaded apatites were internalized within GTL-16 human carcinoma cells and could release DOXO, which accumulated in the nucleus in short times and exerted cytotoxic activity with the same efficiency. cAp are thus expected to be a more promising nanocarrier for experiments in vivo, in situations where intravenous injection of nanoparticles are required to reach the targeted tumor, after circulating in the bloodstream.

Metformin-NSAIDs Molecular Salts: A Path towards Enhanced Oral Bioavailability and Stability.

According to the World Health Organization, more than 422 million people worldwide have diabetes. The most common oral treatment for type 2 diabetes is the drug metformin (MTF), which is usually formulated as a hydrochloride to achieve higher water solubility. However, this drug is also highly hygroscopic, thus showing stability problems. Another kind of worldwide prescribed drug is the non-steroidal anti-inflammatory drug (NSAID). These latter, on the contrary, show a low solubility profile; therefore, they must be administered at high doses, which increases the probability of secondary effects. In this work, novel drug-drug pharmaceutical solids combining MTF-NSAIDs have been synthesized in solution or by mechanochemical methods. The aim of this concomitant treatment is to improve the physicochemical properties of the parent active pharmaceutical ingredients. After a careful solid-state characterization along with solubility and stability studies, it can be concluded that the new molecular salt formulations enhance not only the stability of MTF but also the solubility of NSAIDs, thus giving promising results regarding the development of these novel pharmaceutical multicomponent solids.

Hydrothermal Transformation of Eggshell Calcium Carbonate into Apatite Micro-Nanoparticles: Cytocompatibility and Osteoinductive Properties.

The eggshell is a biomineral consisting of CaCO3 in the form of calcite phase and a pervading organic matrix (1-3.5 wt.%). Transforming eggshell calcite particles into calcium phosphate (apatite) micro-nanoparticles opens the door to repurposing the eggshell waste as materials with potential biomedical applications, fulfilling the principles of the circular economy. Previous methods to obtain these particles consisted mainly of two steps, the first one involving the calcination of the eggshell. In this research, direct transformation by a one-pot hydrothermal method ranging from 100-200 °C was studied, using suspensions with a stoichiometric P/CaCO3 ratio, K2HPO4 as P reagent, and eggshells particles (Ø < 50 µm) both untreated and treated with NaClO to remove surface organic matter. In the untreated group, the complete conversion was achieved at 160 °C, and most particles displayed a hexagonal plate morphology, eventually with a central hole. In the treated group, this replacement occurred at 180 °C, yielding granular (spherulitic) apatite nanoparticles. The eggshell particles and apatite micro-nanoparticles were cytocompatible when incubated with MG-63 human osteosarcoma cells and m17.ASC murine mesenchymal stem cells and promoted the osteogenic differentiation of m17.ASC cells. The study results are useful for designing and fabricating biocompatible microstructured materials with osteoinductive properties for applications in bone tissue engineering and dentistry.

Apatite-coated outer layer eggshell membrane: A novel osteoinductive biohybrid composite for guided bone/tissue regeneration.

Hybrid biomimetic materials aim to replicate the organic-inorganic constructs of mineralized tissues. During eggshell formation, the outer surface of the eggshell membrane (ESM) promotes calcium carbonate nucleation, while the inner one prevents mineralization toward the egg white and yolk. In the current study, the outer surface of the ESM acted as a heteronucleant in calcium phosphate precipitation by the vapor diffusion sitting drop method, while the inner one remained unmineralized. The aim was to fabricate a 2D biomaterial with dual functions, osteoinductive on one side and protective against cell invasion on the other side. The microstructural, physicochemical, morphological, and mechanical properties of the mineralized ESM were characterized by XRD, TGA, XPS, FTIR/Raman, HR-SEM, and mechanical testing techniques. The cytocompatibility and osteoinductive ability were assessed by biological assays of cell viability, proliferation, and osteogenic differentiation on human mesenchymal stromal cells (hMSCs). Results indicate that the outer surface of the ESM induces the heterogeneous precipitation of carbonate-apatite phase depicting biomimetic features. In addition, the apatite/ESM shows a much higher cytocompatibility than the pristine ESM and promotes the osteogenic differentiation of hMSCs more efficiently. Overall, the apatite/ESM composite exhibits compositional, crystalline, mechanical, and biological properties that resemble those of mineralized tissues, rendering it an approachable and novel material especially useful in guided tissue/bone regeneration.

Rational Coformer Selection in the Development of 6-Propyl-2-thiouracil Pharmaceutical Cocrystals.

Pharmaceutical multicomponent solids have proved to efficiently modulate the physicochemical properties of active pharmaceutical ingredients. In this context, polyphenols are interesting coformers for designing pharmaceutical cocrystals due to their wide safety profile and interesting antioxidant properties. The novel 6-propyl-2-thiouracil multicomponent solids have been obtained by mechanochemical synthesis and fully characterized by powder and single-crystal X-ray diffraction methods. The analysis of supramolecular synthons has been further performed with computational methods, with both results revealing a robust supramolecular organization influenced by the different positions of the hydroxyl groups within the polyphenolic coformers. All novel 6-propyl-2-thiouracil cocrystals show an enhanced solubility profile, but unfortunately, their thermodynamic stability in aqueous media is limited to 24 h.

Eggshell Membrane as a Biomaterial for Bone Regeneration.

The physicochemical features of the avian eggshell membrane play an essential role in the process of calcium carbonate deposition during shell mineralization, giving rise to a porous mineralized tissue with remarkable mechanical properties and biological functions. The membrane could be useful by itself or as a bi-dimensional scaffold to build future bone-regenerative materials. This review focuses on the biological, physical, and mechanical properties of the eggshell membrane that could be useful for that purpose. Due to its low cost and wide availability as a waste byproduct of the egg processing industry, repurposing the eggshell membrane for bone bio-material manufacturing fulfills the principles of a circular economy. In addition, eggshell membrane particles have has the potential to be used as bio-ink for 3D printing of tailored implantable scaffolds. Herein, a literature review was conducted to ascertain the degree to which the properties of the eggshell membrane satisfy the requirements for the development of bone scaffolds. In principle, it is biocompatible and non-cytotoxic, and induces proliferation and differentiation of different cell types. Moreover, when implanted in animal models, it elicits a mild inflammatory response and displays characteristics of stability and biodegradability. Furthermore, the eggshell membrane possesses a mechanical viscoelastic behavior comparable to other collagen-based systems. Overall, the biological, physical, and mechanical features of the eggshell membrane, which can be further tuned and improved, make this natural polymer suitable as a basic component for developing new bone graft materials.

A sustainable one-pot method to transform seashell waste calcium carbonate to osteoinductive hydroxyapatite micro-nanoparticles.

We have developed a straightforward, one-pot, low-temperature hydrothermal method to transform oyster shell waste particles (bCCP) from the species Crassostrea gigas (Mg-calcite, 5 wt% Mg) into hydroxyapatite (HA) micro/nanoparticles. The influence of the P reagents (H3PO4, KH2PO4, and K2HPO4), P/bCCP molar ratios (0.24, 0.6, and 0.96), digestion temperatures (25-200 °C), and digestion times (1 week-2 months) on the transformation process was thoroughly investigated. At 1 week, the minimum temperature to yield the full transformation significantly reduced from 160 °C to 120 °C when using K2HPO4 instead of KH2PO4 at a P/bCCP ratio of 0.6, and even to 80 °C at a P/bCCP ratio of 0.96. The transformation took place via a dissolution-reprecipitation mechanism driven by the favorable balance between HA precipitation and bCCP dissolution, due to the lower solubility product of HA than that of calcite at any of the tested temperatures. Both the bCCP and the derived HA particles were cytocompatible for MG-63 human osteosarcoma cells and m17.ASC murine mesenchymal stem cells, and additionally, they promoted the osteogenic differentiation of m17.ASC, especially the HA particles. Because of their physicochemical features and biological compatibility, both particles could be useful osteoinductive platforms for translational applications in bone tissue engineering.

Preparation of core-shell poly(L-lactic) acid-nanocrystalline apatite hollow microspheres for bone repairing applications.

In this paper, hybrid inorganic-organic core-shell hollow microspheres, made of poly(L-lactic acid) (PLLA) and biomimetic nano apatites (HA), were prepared from biodegradable and biocompatible substances, suitable for bone tissue applications. Preparation is started from Pickering emulsification, i.e., solid particle-stabilized emulsions in the absence of any molecular surfactant, where solid particles adsorbed to an oil-water interface. Stable oil-in-water emulsions were produced using biomimetic 20 nm sized HA nanocrystals as particulate emulsifier and a dichloromethane (CH(2)Cl(2)) solution of PLLA as oil phase. Hybrid hollow PLLA microspheres at three different HA nanocrystals surface coverage, ranging from 10 to 50 µm, were produced. The resulting materials were completely characterized with spectroscopic, calorimetric and microscopic techniques and the cytocompatibility was established by indirect contact tests with both fibroblasts and osteoblasts and direct contact with these latter. They displayed a high level of cytocompatibility and thus represent promising materials for drug delivery systems, cell carriers and scaffolds for regeneration of bone useful in the treatment of orthopaedic, maxillofacial and dental fields.

Structural and surface studies of luminescent Ca/Eu phosphate nanomaterials: From the bulk to surface features.

Three luminescent Eu-containing phosphate materials (Ca-doped europium phosphate monohydrate, Eu-doped carbonated-apatite, and europium phosphate monohydrate) were prepared and analyzed on the level of bulk structure and surface properties and compared to the biomimetic non-luminescent counterpart hydroxyapatite. Europium-containing phosphate materials exhibited nanosized dimensions but different luminescence emissions and luminescence lifetimes depending on their crystalline structures (i.e., lanthanide phosphate or apatites) and chemical composition. The introduction of Eu in the crystal lattice leads to a notable decrease in the overall Lewis acidity of the surface cationic sites detected by CO probing. Further, the mixed Eu/Ca-containing materials surfaces were found to be very similar to the reference hydroxyapatite in terms of water adsorption energy, while the pure europium phosphate resulted to have the notably higher energy values of direct interaction of water molecules with the surface cations with no detected propagation of this effect towards water overlayers.

Recovering and Exploiting Aragonite and Calcite Single Crystals with Biologically Controlled Shapes from Mussel Shells.

Control over the shape and morphology of single crystals is a theme of great interest in fundamental science and for technological application. Many synthetic strategies to achieve this goal are inspired by biomineralization processes. Indeed, organisms are able to produce crystals with high fidelity in shape and morphology utilizing macromolecules that act as modifiers. An alternative strategy can be the recovery of crystals from biomineralization products, in this case, seashells. In particular, waste mussel shells from aquaculture are considered. They are mainly built up of single crystals of calcite fibers and aragonite tablets forming an outer and an inner layer, respectively. A simple mechanochemical treatment has been developed to separate and recover these two typologies of single crystals. The characterization of these single crystals showed peculiar properties with respect to the calcium carbonate from quarry or synthesis. We exploited these biomaterials in the water remediation field using them as substrate adsorbing dyes. We found that these substrates show a high capability of adsorption for anionic dye, such as Eosin Y, but a low capability of adsorption for cationic dyes, such as Blue Methylene. The adsorption was reversible at pH 5.6. This application represents just an example of the potential use of these biogenic single crystals. We also envision potential applications as reinforcing fillers and optical devices.

Luminescent Citrate-Functionalized Terbium-Substituted Carbonated Apatite Nanomaterials: Structural Aspects, Sensitized Luminescence, Cytocompatibility, and Cell Uptake Imaging.

This work explores the preparation of luminescent and biomimetic Tb3+-doped citrate-functionalized carbonated apatite nanoparticles. These nanoparticles were synthesized employing a citrate-based thermal decomplexing precipitation method, testing a nominal Tb3+ doping concentration between 0.001 M to 0.020 M, and a maturation time from 4 h to 7 days. This approach allowed to prepare apatite nanoparticles as a single hydroxyapatite phase when the used Tb3+ concentrations were (i) ≤ 0.005 M at all maturation times or (ii) = 0.010 M with 4 h of maturation. At higher Tb3+ concentrations, amorphous TbPO4·nH2O formed at short maturation times, while materials consisting of a mixture of carbonated apatite prisms, TbPO4·H2O (rhabdophane) nanocrystals, and an amorphous phase formed at longer times. The Tb3+ content of the samples reached a maximum of 21.71 wt%. The relative luminescence intensity revealed an almost linear dependence with Tb3+ up to a maximum of 850 units. Neither pH, nor ionic strength, nor temperature significantly affected the luminescence properties. All precipitates were cytocompatible against A375, MCF7, and HeLa carcinogenic cells, and also against healthy fibroblast cells. Moreover, the luminescence properties of these nanoparticles allowed to visualize their intracellular cytoplasmic uptake at 12 h of treatment through flow cytometry and fluorescence confocal microscopy (green fluorescence) when incubated with A375 cells. This demonstrates for the first time the potential of these materials as nanophosphors for living cell imaging compatible with flow cytometry and fluorescence confocal microscopy without the need to introduce an additional fluorescence dye. Overall, our results demonstrated that Tb3+-doped citrate-functionalized apatite nanoparticles are excellent candidates for bioimaging applications.