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Most bone metastases are caused by primary breast or prostate cancer cells settling in the bone microenvironment, affecting normal bone physiology and function and reducing 5‐year survival rates to 10% and 6%, respectively. To expedite clinical availability of novel and effective bone metastases treatments, reliable and predictive in vitro models are urgently required to screen for novel therapies as current in vitro 2D planar mono‐culture models do not accurately predict the clinical efficacy. We herein engineered a novel human in vitro 3D co‐culture model based on spheroids to study dynamic cellular quantities of (breast or prostate) cancer cells and human bone marrow stromal cells and screen chemotherapeutic efficacy and specificity of the common anticancer drug cisplatin. Bone metastatic spheroids (BMSs) were formed rapidly within 24 h, while the morphology of breast versus prostate cancer BMS differed in terms of size and circularity upon prolonged culture periods. Prestaining cell types prior to BMS formation enabled confocal imaging and quantitative image analysis of in‐spheroid cellular dynamics for up to 7 days of BMS culture. We found that cancer cells in BMS proliferated faster and were less susceptible to cisplatin treatment compared to 2D control cultures. Based on these findings and the versatility of our methodology, BMS represent a feasible 3D in vitro model for screening of new bone cancer metastases therapies. [ABSTRACT FROM AUTHOR]

Abstract: Apatitic calcium phosphate cements (CPC) are frequently used to fill bone defects due to their favourable clinical handling and excellent bone response, but their lack of degradability inhibits complete bone regeneration. In order to render these injectable CaP cements biodegradable, hollow microspheres made of poly (d,l-lactic-co-glycolic) acid (PLGA) have been previously used as porogen since these microspheres were shown to be able to induce macroporosity upon degradation as well as to accelerate CPC degradation by release of acid degradation products. Recently, the capacity of PLGA microspheres to form porosity in situ in injectable CPCs was optimized by investigating the influence of PLGA characteristics such as microsphere morphology (dense vs. hollow) and end-group functionalization (acid terminated vs. end-capped) on acid production and corresponding porosity formation in vitro. The current study has investigated the in vivo bone response to CPCs containing two types of microspheres (hollow and dense) made of PLGA with two different end-group functionalizations (end capped and acid terminated). Microspheres were embedded in CPC and injected in the distal femoral condyle of New Zealand White Rabbits for 6 and 12 weeks. Histological results confirmed the excellent biocompatibility and osteoconductivity of all tested materials. Composites containing acid terminated PLGA microspheres displayed considerable porosity and concomitant bone ingrowth after 6 weeks, whereas end capped microspheres only revealed open porosity after 12 weeks of implantation. In addition, it was found that dense PLGA microspheres induced significantly more CPC degradation and bone tissue formation compared to hollow PLGA microspheres. In conclusion, it was shown that PLGA microspheres have a strong capacity to induce fast degradation of injectable CPC and concomitant replacement by bone tissue by controlled release of acid polymeric degradation products without compromising the excellent biocompatibility and osteoconductivity of the CPC matrix. [Copyright &y& Elsevier]

Abstract: For orthopedic and dental implants, the ultimate goal is to obtain a life-long secure anchoring of the implant in the native surrounding bone. To this end, nanoscale calcium phosphate (CaP) and collagen-CaP (col-CaP) composite coatings have been successfully deposited using the electrospray deposition (ESD) technique. In order to study to what extent the thickness of these coatings can be reduced without losing coating osteogenic properties, we have characterized the mechanical and biological coating properties using tape tests (ASTM D-3359) and in vitro cell culture experiments, respectively. Co-deposition of collagen significantly improved coating adhesive and cohesive strength, resulting in a remarkably high coating retention of up to 97% for coating thicknesses below 100nm. In vitro cell culture experiments showed that electrosprayed CaP and col-CaP composite coatings enhanced osteoblast differentiation, leading to improved mineral deposition. This effect was most pronounced upon co-deposition of collagen with CaP, and these coatings displayed osteogenic effects even for a coating thickness of below 100nm. [Copyright &y& Elsevier]

Biodegradable polymers that can be processed into injectable hydrogel matrices are promising candidates for bone-substituting purposes. Furthermore, by incorporating degradable calcium phosphate (CaP) particles and growth factors into these hydrogel matrices, a bone construct can be designed which stimulates the formation of new bone by the surrounding tissue, thereby compensating for the loss of structural integrity of the degrading synthetic bone-substitute. Generally, a major challenge in synthesis of nanoceramic-reinforced polymers is the achievement of a fine dispersion of nanoparticles throughout the polymer, since the unique properties of nanocomposites are lost when nanoparticles aggregate. In the current study, composite hydrogels consisting of oligo(poly(ethylene glycol)fumarate) (OPF) matrices and CaP dispersions of varying crystallinity were successfully developed using physical or chemical preparation strategies. Physical mixing of dried, micrometer-sized CaP powders resulted into formation of irreproducible composites with a highly heterogeneous dispersion of large and agglomerated CaP microparticles throughout the OPF matrix. On the contrary, reproducible and homogeneous hydrogels were fabricated using a chemical mixing strategy, whereby CaP crystals were formed in the presence of dissolved OPF macromers. This co-precipitation technique resulted into a much higher degree of dispersion of the CaP crystals, which can enable higher CaP contents in organic matrices such as OPF. By using these CaP suspensions instead of dried powders, the nanosized structure of separated CaP crystals was preserved, resulting into a higher reactivity of the CaP phase, as indicated by a reduced swelling behavior of these hydrogels. This effect was most likely caused by a physicochemical interaction between Ca2+ and unreacted COOH end-groups, thereby leading to increased physical cross-linking of the composite hydrogels. [ABSTRACT FROM AUTHOR]

Abstract: The dissolution and/or precipitation behaviour of porous calcium phosphate (CaP) coatings, deposited using electrostatic spray deposition (ESD), was investigated (a) in vitro after soaking in simulated body fluid (SBF) for several time periods (2, 4, 8, and 12 weeks), and (b) in vivo after subcutaneous implantation of CaP-coated implants in the back of goats for identical time periods. Physical and chemical properties of coatings were characterized before and after in vitro/vivo testing by means of scanning electron microscopy, X-ray diffraction, Fourier-transform infrared spectroscopy, and energy dispersive spectroscopy. Moreover, part of the explants was prepared for light microscopical evaluation of the tissue response. In vitro, all apatitic ESD-coatings induced the formation of homogeneous and adherent CaP precipitation layers. Amorphous CaP, however, displayed a delayed precipitation of poorly adherent CaP layers, whereas heterogeneous calcification was observed on top of β-TCP-coated substrates, indicating that β-TCP and amorphous CaP coatings exhibit a poor ability of inducing calcification in SBF as compared to crystalline apatitic coatings. In vivo, no adverse tissue reactions (toxic effects/inflammatory cells) were observed using light microscopy, and all coatings became surrounded by a dense, fibrous tissue capsule after implantation. All ESD-coatings degraded gradually at a dissolution rate depending on the chemical phase (order of relative solubility: amorphous CaP≈carbonate apatite>β-TCP>carbonated hydroxyapatite), thereby enabling synthesis of CaP coatings with a tailored degradation rate. [Copyright &y& Elsevier]

This study presents a novel multi-technique approach that integrates micro-CT and optical photothermal infrared spectroscopy (O-PTIR) to evaluate polymerisation differences, so-called spatio-temporal polymerisation properties, between flowable and sculptable dental resin-based composites. Ten commercially available dental composites were investigated, including flowable and sculptable counterparts from the same manufacturer. Eight parameters were evaluated: short-term polymerisation characteristics (degree of conversion after 5 min, maximum polymerisation rate, time to reach maximum polymerisation rate) was measured using ATR-FTIR with real-time monitoring; changes in the degree of conversion with depth were evaluated with O-PTIR, 3D visualisation of shrinkage patterns, overall volumetric shrinkage, depth-specific shrinkage, and porosity were measured using micro-CT; surface morphology with detailed measurements of elemental composition was characterised using SEM/EDX; light transmittance was analysed with a NIST-referenced spectrometer. The study found that the increase in filler weight and volume ratio reduced the degree of conversion and polymerisation shrinkage, while moderately influencing the maximum polymerisation rates. The time to reach maximum polymerisation rates and light transmittance were not dependent on the filler amount. O-PTIR assessed a depth-dependent decrease in the degree of conversion for both composite types, with flowable composites generally showing a greater decrease in the degree of conversion than sculptable composites, except for bulk-fill composites. Micro-CT scans showed significantly higher flowable shrinkage values than their sculptable counterparts, highlighting the performance differences between the two types of composites. The findings of this study have practical implications for the selection and use of dental composites. Flowable composites, despite their higher degrees of conversion and polymerisation rates, also exhibit higher volumetric shrinkage, which can be detrimental for clinical applications. The new measurement methods used in this study provide a comprehensive overview of the polymerisation behaviour of commercially available dental composites, offering valuable insights for material optimisation. [ABSTRACT FROM AUTHOR]

Dental resin composites' performance is intricately linked to their polymerisation shrinkage characteristics. This study compares polymerisation shrinkage using advanced 3D micro-computed tomography (micro-CT) and traditional 2D linear assessments. It delves into the crucial role of filler content on shrinkage and the degree of conversion in dental resin composites, providing valuable insights for the field. Five experimental dental composite materials were prepared with increasing filler contents (55–75 wt%) and analysed using either 3D micro-CT for volumetric shrinkage or a custom-designed linometer for 2D linear shrinkage. The degree of conversion was assessed using Optical Photothermal Infrared (O-PTIR) and Fourier-Transform Infrared (FTIR) spectroscopy. Light transmittance through a 2-mm layer was evaluated using a NIST-calibrated spectrometer. Scanning Electron Microscopy (SEM) and Energy-Dispersive X-ray Spectroscopy (EDX) examined surface morphology and elemental distribution. Correlation between the investigated parameters was determined using Spearman correlation analyses. The study found significant differences in polymerisation-related properties among different filler content categories, with volumetric shrinkage consistently demonstrating higher mean values than linear shrinkage across most groups. Volumetric shrinkage decreased with increasing curing depth, showing no direct correlation between filler content and shrinkage levels at different curing depths. The results highlighted a strong negative correlation between filler content and degree of conversion, volumetric and linear shrinkage, as well as maximum shrinkage rate. Light transmittance showed a moderate correlation with the filler content and a weak correlation with other tested parameters. This study underscores the importance of considering both volumetric and linear shrinkage in the design and analysis of dental composite materials. The findings advocate optimising filler content to minimise shrinkage and enhance material performance. Integrating micro-CT and O-PTIR techniques offers novel insights into dental composites' polymerisation behaviour, providing a foundation for future research to develop materials with improved clinical outcomes. • 3D micro-CT and O-PTIR spectroscopy provide novel insights into dental composite polymerisation. • Filler content significantly influences polymerisation shrinkage and degree of conversion. • Volumetric shrinkage was consistently higher than linear shrinkage in most experimental groups. • Increasing filler content reduced shrinkage and improved material performance. • Integration of micro-CT and O-PTIR techniques advances understanding of dental composites. [ABSTRACT FROM AUTHOR]

Novel self-healing resin-based composites containing microcapsules have been developed to improve the mechanical performance of dental restorations. However, the long-term fatigue behaviour of these self-healing composites has still been hardly investigated. Therefore, this manuscript studied the fatigue behaviour of self-healing composites containing microcapsules by subjecting the specimens to traditional staircase tests and ageing in a custom-designed chewing simulator (Rub&Roll) to simulate oral ageing physiologically relevant conditions. To prepare self-healing composite, poly(urea-formaldehyde) microcapsules containing acrylic self-healing liquids were synthesized. Subsequently, these microcapsules (10 wt%) and initiator (benzoyl peroxide, BPO, 2 wt%) were incorporated into a commercial flowable resin-based composite. Microcapsule-free resin-based composites with and without BPO were also prepared as control specimens. A three-point flexural test was used to measure the initial flexural strength (S initial). Subsequently, half of the specimens were used for fatigue testing using a common staircase approach to measure the fatigue strengths (FS). In addition, the other specimens were aged in the Rub&Roll machine for four weeks where after the final flexural strength (S final) was measured. Compared to S initial , FS of all tested specimens significantly decreased as measured through staircase testing. After 4 weeks of ageing in the Rub&Roll machine, S final was significantly reduced compared to S initial for microcapsule-free resin-based composites, but not for the self-healing composites (p = 0.3658). However, the self-healing composites are still in the experimental phase characterized by a low mechanical strength, which still impedes further clinical translation. Self-healing composites containing microcapsules exhibit improved fatigue resistance compared to microcapsule-free non-self-healing composites. • Fatigue tests on resin-based composites containing microcapsules. • Different fatigue tests to characterize fatigue resistance of resin-based composites. • Novel fatigue tests with a custom-designed wear machine to simulate physiological oral ageing. [ABSTRACT FROM AUTHOR]

This study aimed to comparatively evaluate the in vitro effect of nanosized hydroxyapatite and collagen (nHA/COL) based composite hydrogels (with different ratios of nHA and COL) on the behavior of human mesenchymal stromal cells (MSCs), isolated from either adipose tissue (AT-MSCs) or bone marrow (BM-MSCs). We hypothesized that (i) nHA/COL composite hydrogels would promote the osteogenic differentiation of MSCs in an nHA concentration dependent manner, and that (ii) AT-MSCs would show higher osteogenic potential compared to BM-MSCs, due to their earlier observed higher proliferation and osteogenic differentiation potential in 2D in vitro cultures [1]. The obtained results indicated that AT-MSCs show indeed high proliferation, differentiation and mineralization capacities in nHA/COL constructs compared to BM-MSCs, but this effect was irrespective of nHA concentration. Based on the results of alkaline phosphatase (ALP) activity and osteocalcin (OCN) protein level, the osteogenic differentiation of BM-MSCs started in the beginning of the culture period and for AT-MSCs at the end of the culture period. At a molecular level, both cell types showed high expression of osteogenic markers (bone morphogenic protein 2 [BMP2], runt-related transcription factor 2 [RUNX2], OCN or COL1) in both an nHA concentration and time dependent manner. In conclusion, AT-MSCs demonstrated higher osteogenic potential in nHA/COL based 3D micro-environments compared to BM-MSCs, in which proliferation and osteogenic differentiation were highly promoted in a time dependent manner, irrespective of nHA amount in the constructs. The fact that AT-MSCs showed high proliferation and mineralization potential is appealing for their application in future pre-clinical research as an alternative cell source for BM-MSCs. [ABSTRACT FROM AUTHOR]

Biomaterials with the ability to self-heal and recover their structural integrity offer many advantages for applications in biomedicine. The past decade has witnessed the rapid emergence of a new class of self-healing biomaterials commonly termed injectable, or printable in the context of 3D printing. These self-healing injectable biomaterials, mostly hydrogels and other soft condensed matter based on reversible chemistry, are able to temporarily fluidize under shear stress and subsequently recover their original mechanical properties. Self-healing injectable hydrogels offer distinct advantages compared to traditional biomaterials. Most notably, they can be administered in a locally targeted and minimally invasive manner through a narrow syringe without the need for invasive surgery. Their moldability allows for a patient-specific intervention and shows great prospects for personalized medicine. Injected hydrogels can facilitate tissue regeneration in multiple ways owing to their viscoelastic and diffusive nature, ranging from simple mechanical support, spatiotemporally controlled delivery of cells or therapeutics, to local recruitment and modulation of host cells to promote tissue regeneration. Consequently, self-healing injectable hydrogels have been at the forefront of many cutting-edge tissue regeneration strategies. This study provides a critical review of the current state of self-healing injectable hydrogels for tissue regeneration. As key challenges toward further maturation of this exciting research field, we identify (i) the trade-off between the self-healing and injectability of hydrogels vs their physical stability, (ii) the lack of consensus on rheological characterization and quantitative benchmarks for self-healing injectable hydrogels, particularly regarding the capillary flow in syringes, and (iii) practical limitations regarding translation toward therapeutically effective formulations for regeneration of specific tissues. Hence, here we (i) review chemical and physical design strategies for self-healing injectable hydrogels, (ii) provide a practical guide for their rheological analysis, and (iii) showcase their applicability for regeneration of various tissues and 3D printing of complex tissues and organoids. [ABSTRACT FROM AUTHOR]

Messenger RNA (mRNA) is increasingly gaining interest as a modality in vaccination and protein replacement therapy. In regenerative medicine, the mRNA-mediated expression of growth factors has shown promising results. In contrast to protein delivery, successful mRNA delivery requires a vector to induce cellular uptake and subsequent endosomal escape to reach its end destination, the ribosome. Current non-viral vectors such as lipid- or polymer-based nanoparticles have been successfully used to express mRNA-encoded proteins. However, to advance the use of mRNA in regenerative medicine, it is required to assess the compatibility of mRNA with biomaterials that are typically applied in this field. Herein, we investigated the complexation, cellular uptake and maintenance of the integrity of mRNA complexed with gelatin nanoparticles (GNPs). To this end, GNPs with positive, neutral or negative surface charge were synthesized to assess their ability to bind and transport mRNA into cells. Positively charged GNPs exhibited the highest binding affinity and transported substantial amounts of mRNA into pre-osteoblastic cells, as assessed by confocal microscopy using fluorescently labeled mRNA. Furthermore, the GNP-bound mRNA remained stable. However, no expression of mRNA-encoded protein was detected, which is likely related to insufficient endosomal escape and/or mRNA release from the GNPs. Our results indicate that gelatin-based nanomaterials interact with mRNA in a charge-dependent manner and also mediate cellular uptake. These results create the basis for the incorporation of further functionality to yield endosomal release. [ABSTRACT FROM AUTHOR]

The development of bone substitute materials (BSMs) intended for load-bearing bone defects is highly complicated, as biological and mechanical requirements are often contradictory. In recent years, biological BSMs have been developed which allow for a more efficient integration of the material with the surrounding osseous environment and, hence, a higher mechanical stability of the treated defect. However, while these materials are promising, they are still far from ideal. Consequently, extensive preclinical experimentation is still required. The current review provides a comprehensive overview of biomechanical considerations relevant for the design of biological BSMs. Further, the preclinical evaluation of biological BSMs intended for application in highly loaded skeletal sites is discussed. The selected animal models and implantation site should mimic the pathophysiology and biomechanical loading patterns of human bone as closely as possible. In general, sheep are among the most frequently selected animal models for the evaluation of biomaterials intended for highly loaded skeletal sites. Regarding the anatomical sites, segmental bone defects created in the limbs and spinal column are suggested as the most suitable. Furthermore, the outcome measurements used to assess biological BSMs for regeneration of defects in heavily loaded bone should be relevant and straightforward. The quantitative evaluation of bone defect healing through ex vivo biomechanical tests is a valuable addition to conventional in vivo tests, as it determines the functional efficacy of BSM-induced bone healing. Finally, we conclude that further standardization of preclinical studies is essential for reliable evaluation of biological BSMs in highly loaded skeletal sites. [ABSTRACT FROM AUTHOR]

Among various classes of biomaterials, the majority of non‐centrosymmetric crystalline materials exhibit piezoelectric properties, i.e., the accumulation of charge in response to applied mechanical stress or deformation. Due to the growing interest in nanomaterials, piezoelectric nano‐biomaterials have been widely investigated, leading to remarkable advancements throughout the last two decades. Piezoelectric properties, high surface energy, targeting properties, and intricate cell–material interactions render piezoelectric nanomaterials highly attractive for application in therapeutics as well as regenerative medicine. Herein, the major focus is to highlight the wide range of applications of piezoelectric nano‐biomaterials in drug delivery, theranostics, and tissue regeneration. After a brief introduction to piezoelectricity, an overview is provided on the major classes of piezoelectric biomaterials as well as a description of the origin of biopiezoelectricity in different tissues and macromolecules. Subsequently, relevant properties and postfabrication strategies of nanostructured piezoelectric biomaterials are discussed aiming to maximize piezoresponse. Finally, recent studies on nano‐ piezoceramics and piezopolymers are presented, with specific focus on barium titanate, zinc oxide, and polyvinylidene fluoride. [ABSTRACT FROM AUTHOR]

Platinum-based chemotherapeutics exhibit excellent antitumor properties. However, these drugs cause severe side effects including toxicity, drug resistance, and lack of tumor selectivity. Tumor-targeted drug delivery has demonstrated great potential to overcome these drawbacks. Herein, we aimed to design radioactive bisphosphonate-functionalized platinum (195mPt-BP) complexes to confirm preferential accumulation of these Pt-based drugs in metabolically active bone. In vitro NMR studies revealed that release of Pt from Pt BP complexes increased with decreasing pH. Upon systemic administration to mice, Pt-BP exhibited a 4.5-fold higher affinity to bone compared to platinum complexes lacking the bone-seeking bisphosphonate moiety. These Pt-BP complexes formed less Pt-DNA adducts compared to bisphosphonate-free platinum complexes, indicating that in vivo release of Pt from Pt-BP complexes proceeded relatively slow. Subsequently, radioactive 195mPt-BP complexes were synthesized using 195mPt(NO3)2(en) as precursor and injected intravenously into mice. Specific accumulation of 195mPt-BP was observed at skeletal sites with high metabolic activity using micro-SPECT/CT imaging. Furthermore, laser ablation-ICP-MS imaging of proximal tibia sections confirmed that 195mPt BP co-localized with calcium in the trabeculae of mice tibia. [ABSTRACT FROM AUTHOR]

Bone adhesive biomaterials are currently being developed to replace conventional biomaterials that do not adhere to bone. Recently, poly (2-oxazoline) polymers have been functionalized with calcium-binding alendronate groups (POx-Ale) to render these polymers bone adhesive. These polymers hold promise for application as bone fixation biomaterials, but their effects on cells present in bone tissue has not been studied yet. This study evaluated the in vitro cytocompatibility of POx-Ale for cell types that are relevant for applications in bone reconstructive surgery. Cell viability of murine pre-osteoblasts (MC3T3-E1), fibroblasts (NIH3T3) and pre-osteoclasts derived from macrophages (RAW264.7) is quantified as a function of POx-Ale polymer concentrations ranging between 5.0 and 0.3 mg mL−1. In addition, the effect of POx-Ale on cellular function is assessed by performing mineralization, gelatin zymography as well as TRAP activity anad TRAP staining assays. The results show that POx-Ale has in vitro cytotoxic effects on murine pre-osteoblasts (MC3T3-E1), fibroblasts (NIH3T3) and pre-osteoclasts (RAW264.7) when cultured in media with polymer concentrations ≥2.5 mg mL−1, and inhibits mineralization of osteoblasts and MMP activity of fibroblasts at lower polymer concentrations. However, POx-Ale did not negatively affect Tartrate-resistant acid phosphate (TRAP) activity in osteoclasts. [Display omitted] • Cellular viability of bone cells in response to bone adhesive alendronate-functionalized poly (2-oxazoline). Van Erk et al. • POx-Ale reduces in vitro viability of cells at concentrations ≥2.5 mg mL−1 • POx-Ale affects MMP activity in fibroblasts. • POx-Ale affects the calcium deposition in differentiated osteoblasts. • POx-Ale did not negatively affect Tartrate- TRAP activity in osteoclasts. [ABSTRACT FROM AUTHOR]

Abstract Hydrogels are the preferred material choice for various strategies in regenerative medicine. Nevertheless, due to their high water content and soft nature, these materials are often mechanically weak, which limits their applicability. This study demonstrates mechanical reinforcement of colloidal gels at microscale using discrete polyester fibers, as confirmed by rheological, compression and nanoindentation tests. This reinforcement strategy results in injectable and moldable colloidal gels with improved mechanical performance. The fully organic gels presented here are cytocompatible and can maintain their mechanical integrity under physiological conditions. Consequently, these gels exhibit a strong potential for applications in tissue engineering and regenerative medicine. Graphical abstract Unlabelled Image Highlights • Colloidal gels are mechanically reinforced using discrete polyester fibers. • The resulting gels are injectable, moldable, and cytocompatible. • These gels exhibit a strong potential for applications in regenerative medicine. [ABSTRACT FROM AUTHOR]

Nanoparticles (NPs) are widely studied for biomedical applications. Generally, NPs and NP-based materials should be effectively sterilized without altering their structure and function before clinical translation can be considered. However, research on industrially scalable sterilization methods of nanoparticles is still scarce. Therefore, we explored if treatment of gelatin nanoparticles (GNPs) using supercritical carbon dioxide (scCO 2) resulted in effective inactivation of Staphylococcus aureus as frequent pathogen causing infections. After scCO 2 -treatment the morphology, size and surface charge of GNPs remained unchanged, although a slight decrease in crosslinking density was observed. ScCO 2 successfully inactivated Staphylococcus aureus in GNPs, as evidenced by a 12-log reduction in bacterial burden. However, cytocompatibility of GNPs was lower after scCO 2 treatment, indicating that the process still requires further optimization. Nevertheless, the work presented herein highlights the strong potential of scCO 2 for inactivation of pathogens in nanoparticles. [Display omitted] • Supercritical carbon dioxide does not affect the morphology of gelatin nanoparticles. • Gelatin nanoparticles still form gels after supercritical carbon dioxide treatment. • Supercritical carbon dioxide treatment reduces bacteria load by 12-log reduction. [ABSTRACT FROM AUTHOR]

Injectable composite colloidal gels are developed for regeneration of osteoporotic bone defects through a bottom-up assembly from bisphosphonate-functionalized gelatin and bioactive glass particles. Upon bisphosphonate functionalization, gelatin nanoparticles show superior adhesion toward bioactive glass particles, resulting in elastic composite gels. By tuning their composition, these composite colloidal gels combine mechanical robustness with self-healing ability. The composite colloidal gels support cell proliferation and differentiation in vitro without requiring any osteogenic supplement. In vivo evaluation of the composite colloidal gels reveals their capacity to support the regeneration of osteoporotic bone defects. Furthermore, the bisphosphonate modification of gelatin induces a therapeutic effect on the peri-implantation region by enhancing the bone density of the osteoporotic bone tissue. Consequently, these composite colloidal gels offer new therapeutic opportunities for treatment of osteoporotic bone defects. [ABSTRACT FROM AUTHOR]

Star-shaped poly(ethylene glycol) (PEG) chain termini were functionalized with alendronate to create transient networks with reversible cross-links upon addition of calcium ions. The gelation ability of alendronate-functionalized PEG was greatly dependent on the number of arms and arm molecular weight. After mixing polymer and calcium solutions, the formed hydrogels could be cut and then brought back together without any visible interface. After 2 min of contact, their connection was strong enough to allow for stretching without tearing through the previous fracture surface. Oscillatory rheology showed that the hydrogels recovered between 70 and 100% of the original storage and loss modulus after rupture. Frequency sweep measurements revealed a liquidlike behavior at lower frequencies and solidlike at high frequencies. Shifting frequency curves obtained at different calcium and polymer concentrations, all data collapsed in a single common master curve. This time-concentration superposition reveals a common relaxation mechanism intrinsically connected to the calcium-bisphosphonate complexation equilibrium. [ABSTRACT FROM AUTHOR]

In order to prevent hemorrhage during surgical procedures, a wide range of hemostatic agents have been developed. However, their efficacy is variable; hemostatic devices that use bioactive components to accelerate coagulation are dependent on natural sources, which limits reproducibility. Hybrid devices in which chain-end reactive poly(ethylene glycol) is employed as active component sometimes suffer from irregular cross-linking and dissolution of the polar PEG when blood flow is substantial. Herein, we describe a synthetic, nonbioactive hemostatic product by coating N-hydroxysuccinimide ester (NHS)-functional poly(2-oxazoline)s (POx-NHS) onto gelatin patches, which acts by formation of covalent cross-links between polymer, host blood proteins, gelatin and tissue to seal the wound site and prevent hemorrhage during surgery. We studied different process parameters (including polymer, carrier, and coating technique) in direct comparison with clinical products (Hemopatch and Tachosil) to obtain deeper understanding of this class of hemostatic products. In this work, we successfully prove the hemostatic efficacy of POx-NHS as polymer powders and coated patches both in vitro and in vivo against Hemopatch and Tachosil, demonstrating that POx-NHS are excellent candidate polymers for the development of next generation hemostatic patches. [ABSTRACT FROM AUTHOR]

Calcium phosphates and bioactive glass ceramics have been considered promising biomaterials for use in surgeries. However, their moldability should be further enhanced. We here thereby report the handling, physicochemical features, and morphological characteristics of formulations consisting of carboxymethylcellulose–glycerol and hydroxyapatite-tricalcium phosphate or Biosilicate® particles. We hypothesized that combining either material with carboxymethylcellulose–glycerol would improve handling properties, retaining their bioactivity. In addition to scanning electron microscopy, cohesion, mineralization, pH, and viscoelastic properties of the novel formulations, cell culture experiments were performed to evaluate the cytotoxicity and cell proliferation. Putty-like formulations were obtained with improved cohesion and moldability. Remarkably, mineralization in simulated body fluid of hydroxyapatite-tricalcium phosphate/carboxymethylcellulose–glycerol formulations was enhanced compared to pure hydroxyapatite-tricalcium phosphate. Cell experiments showed that all formulations were noncytotoxic and that HA-TCP60 and BGC50 extracts led to an increased cell proliferation. We conclude that combining carboxymethylcellulose–glycerol with either hydroxyapatite-tricalcium phosphate or Biosilicate® allows for the generation of moldable putties, improves handling properties, and retains the ceramic bioactivity. [ABSTRACT FROM AUTHOR]

Herein the preparation of Sr 2 + -doped calcium phosphate (CaP) coatings on poly(etheretherketone) (PEEK) substrates by pulsed electron deposition (PED) was investigated. Hydroxyapatites (HA) substituted with tailored amounts of Sr 2 + up to 9 wt% were synthesized to generate the deposition targets. Dense and uniform amorphous CaP films having Sr/Ca molar ratios close to that of targets were achieved. The roughness and the wettability of the bare PEEK were significantly enhanced after coating deposition. The presence of Sr 2 + led to a slight increase of the particles size while it did not mainly affect the RMS of the coatings that consisted of nanostructured aggregates of globular shape. After a mild annealing treatment at 130 °C for 6 h, the amorphous coatings transformed into nanocrystalline HA films incorporating Sr 2 + having Sr/Ca molar ratios close to those of the as-deposited films. The annealing did not affect the topography and the roughness of the coatings, while improved the hardness of the films un-doped and doped at low-extent. This study shows that PED is feasible technique to coat PEEK implants with CaP films with control over the Sr/Ca ratio, which could improve bone fixation and in vivo stability especially under osteoporotic conditions thanks to the anabolic effect of Sr 2 + . [ABSTRACT FROM AUTHOR]

Electrophoretic deposition (EPD) is an effective technique to construct coatings onto the surface of medical devices. These coatings can act as reservoirs for local delivery of therapeutic agents, including metal ions and metal-free drugs. This manuscript presents a comprehensive overview of the fundamentals and application of EPD coatings for local delivery of therapeutic agents, with a specific focus on the mechanism and control over loading and release of therapeutic agents. After a brief introduction of the EPD coating process, the matrices that are most commonly deposited using EPD for drug delivery purposes are summarized including the mechanism of their deposition without additives and co-deposition with additives. Subsequently, three strategies towards loading therapeutic agents are discussed, including i) co-deposition of therapeutic agents with coating matrices, ii) pre-loading therapeutic agents into microcarriers, and iii) post-loading therapeutic agents into prepared coatings. Furthermore, factors influencing loading (e.g., EPD processing parameters and additives) are identified. Subsequently, the release mechanisms as well as the influence of both intrinsic (e.g., additives, layered structures, building blocks, and microcarrier properties) and extrinsic processing parameters (e.g., pH, electric, magnetic and photothermal stimuli) used to tune the release kinetics are presented. Additionally, the applications of therapeutic agent loaded EPD coatings are summarized. Finally, the remaining challenges and future perspectives related to the use of EPD coatings for drug delivery purposes are reviewed. [ABSTRACT FROM AUTHOR]

Hydrogels are widely recognized as promising candidates for various biomedical applications, such as tissue engineering. Recently, extensive research efforts have been devoted to the improvement of the biological and mechanical performance of hydrogel systems by incorporation of functional groups and/or inorganic particles in their composition. Bisphosphonates are a class of drugs, commonly used for treatment of osteoporosis, which exhibit a strong binding affinity for hydroxyapatite. In this study, the binding affinity of a bisphosphonate-functionalized polymer, hyaluronan, toward a bioactive glass (i.e., 45S5 Bioglass) is evaluated using force-distance measurements with atomic force microscopy. The strong interaction between bisphosphonate and bioactive glass is then exploited to develop organic-inorganic composite hydrogels and the viscoelastic and self-healing ability of these materials are investigated. Finally, the stability and mineralization behavior of these hydrogels are evaluated in simulated body fluid. Following this approach, injectable, bioactive and self-healing organic-inorganic composite hydrogels are produced, which mineralize abundantly and rapidly in simulated body fluid. These properties render these composite gels suitable for applications in bone-tissue engineering. [ABSTRACT FROM AUTHOR]

ABSTRACT Polymers that possess lower critical solution temperature behavior such as poly(2-alkyl-2-oxazoline)s (PAOx) are interesting for their application as stimulus-responsive materials, for example in the biomedical field. In this work, we discuss the scalable and controlled synthesis of a library of pH- and temperature-sensitive 2- n-propyl-2-oxazoline P( nPropOx) based copolymers containing amine and carboxylic acid functionalized side chains by cationic ring opening polymerization and postpolymerization functionalization strategies. Using turbidimetry, we found that the cloud point temperature (CP) is strongly dependent on both the polymer concentration and the polymer charge (as a function of pH). Furthermore, we observed that the CP decreased with increasing salt concentration, whereas the CP increased linearly with increasing amount of carboxylic acid groups. Finally, turbidimetry studies in PBS-buffer indicate that CPs of these polymers are close to body temperature at biologically relevant polymer concentrations, which demonstrates the potential of P( nPropOx) as stimulus-responsive polymeric systems in, for example, drug delivery applications. © 2016 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2016, 54, 1573-1582 [ABSTRACT FROM AUTHOR]

Artificial 3-dimensional (3D) cell culture systems, which mimic the extracellular matrix (ECM), hold great potential as models to study cellular processes under controlled conditions. The natural ECM is a 3D structure composed of a fibrous hydrogel that provides both mechanical and biochemical cues to instruct cell behavior. Here we present an ECM-mimicking genetically engineered protein-based hydrogel as a 3D cell culture system that combines several key features: (1) Mild and straightforward encapsulation meters (1) ease of ut I am not so sure.encapsulation of the cells, without the need of an external crosslinker. (2) Supramolecular assembly resulting in a fibrous architecture that recapitulates some of the unique mechanical characteristics of the ECM, i.e. strain-stiffening and self-healing behavior. (3) A modular approach allowing controlled incorporation of the biochemical cue density (integrin binding RGD domains). We tested the gels by encapsulating MG-63 osteoblastic cells and found that encapsulated cells not only respond to higher RGD density, but also to overall gel concentration. Cells in 1% and 2% (weight fraction) protein gels showed spreading and proliferation, provided a relative RGD density of at least 50%. In contrast, in 4% gels very little spreading and proliferation occurred, even for a relative RGD density of 100%. The independent control over both mechanical and biochemical cues obtained in this modular approach renders our hydrogels suitable to study cellular responses under highly defined conditions. [ABSTRACT FROM AUTHOR]

The integration of bone implants within native bone tissue depends on periprosthetic bone quality, which is severely decreased in osteoporotic patients. In this work, we have synthesized bone-like hydroxyapatite nanocrystals (nHA) using an acid–base neutralization reaction and analysed their physicochemical properties. Subsequently, we have functionalized the nHA with alendronate (nHA ALE ), a well-known bisphosphonate drug used for the treatment of osteoporosis. An in vitro osteoclastogenesis test was carried out to evaluate the effect of nHA ALE on the formation of osteoclast-like cells from monocytic precursor cells (i.e. RAW264.7 cell line) showing that nHA ALE significantly promoted apoptosis of osteoclast-like cells. Subsequently, nHA and nHA ALE were deposited on titanium disks using electrospray deposition (ESD), for which characterisation of the deposited coatings confirmed the presence of alendronate in nHA ALE coatings with nanoscale thickness of about 700 nm. These results indicate that alendronate linked to hydroxyapatite nanocrystals has therapeutic potential and nHA ALE can be considered as an appealing coating constituent material for orthopaedic and oral implants for application in osteoporotic patients. [ABSTRACT FROM AUTHOR]

Abstract: Non-covalent interactions are often regarded as insufficient to construct macroscopic materials of substantial integrity and cohesion. However, the low binding energy of such reversible interactions can be compensated by increasing their number to work in concert to create strong materials. Here we present the successful development of an injectable, cohesive nanocomposite hydrogel based on reversible bonds between calcium phosphate nanoparticles and bisphosphonate-functionalized hyaluronic acid. These nanocomposites display a capacity for self-healing as well as adhesiveness to mineral surfaces such as enamel and hydroxyapatite. Most importantly, these non-covalently cross-linked composites are surprisingly robust yet biodegradable upon extensive in vitro and in vivo testing and show bone interactive capacity evidenced by bone ingrowth into material remnants. The herein presented method provides a new methodology for constructing nanoscale composites for biomedical applications, which owe their integrity to reversible bonds. [Copyright &y& Elsevier]

Abstract: The prevalence of osteoporosis will increase within the next decades due to the aging world population, which can affect the bone healing response to dental and orthopedic implants. Consequently, local drug targeting of peri-implant bone has been proposed as a strategy for the enhancement of bone-implant integration in osteoporotic conditions. In the present study, an established in-vivo femoral condyle implantation model in osteoporotic and healthy bone is used to analyze the osteogenic capacity of titanium implants coated with bisphosphonate (BP)-loaded calcium phosphate nanoparticles (nCaP) under compromised medical conditions. After 4 weeks of implantation, peri-implant bone volume (%BV; by μCT) and bone area (%BA; by histomorphometry) were significantly increased within a distance of 500 μm from implant surfaces functionalized with BP compared to control implants in osteoporotic and healthy conditions. Interestingly, the deposition of nCaP/BP coatings onto implant surfaces increased both peri-implant bone contact (%BIC) and volume (%BV) compared to the deposition of nCaP or BP coatings individually, in osteoporotic and healthy conditions. The results of real-time PCR revealed similar osteogenic gene expression levels to all implant surfaces at 4-weeks post-implantation. In conclusion, simultaneous targeting of bone formation (by nCaP) and bone resorption (by BP) using nCaP/BP surface coatings represents an effective strategy for synergistically improvement of bone-implant integration, especially in osteoporotic conditions. [Copyright &y& Elsevier]

Abstract: The objective of the current study was to present a simple and standardized system as a preliminary attempt to assess the confinement of polymer-based injectable bone substitutes (IBSs) in vitro. Four different types of polymer-based IBSs were selected as model compounds, a thermosensitive collagen gel, a colloidal gelatin gel, a covalently crosslinked oligo(poly(ethylene glycol)fumarate (OPF) gel and a OPF-calcium phosphate composite. A ceramic-based IBS (i.e. a self-setting calcium phosphate cement) was used as reference. The confinement of all IBSs was tested under three different conditions: (1) no flow (as control), (2) dynamic flow after injection and (3) dynamic flow during injection. The results presented herein confirmed that the proposed test can be used to quantify the confinement of various IBSs within artificial defects under static or dynamic flow conditions, thereby offering a potential tool for predictive quantitative determination of the confinement of IBSs in vivo. [Copyright &y& Elsevier]

Abstract: Synthetic inorganic and organic materials have been extensively investigated in the field of bone regeneration in an attempt to mimic the composition and structure of the extracellular matrix (ECM) of bone tissue with the ultimate aim of generating suitable synthetic bone substitute materials and modifying the surface of bone implants. For load-bearing applications, bone implants are generally made of a bioinert metal with appropriate mechanical properties and a modified surface (i.e. roughened, coated or a combination thereof), to enhance the surface biocompatibility and osteoconductivity. Currently, biomaterials research is evolving from the use of bioinert and biologically passive implants toward interactive implants that stimulate tissue regeneration. Therefore, surface physico-chemical properties of bone implants need to be optimal and capable to biologically instruct and stimulate the regeneration of bone tissue. This review is focused on surface modification approaches in the field of bone regeneration for load-bearing bone implants with emphasis on the use of inorganic and organic coating compounds that actively participate in the biological processes that occur upon implantation. The review shows that appropriate surface modifications can be helpful to achieve desired tissue responses in healthy as well as compromised conditions. [Copyright &y& Elsevier]

The design of bioactive three-dimensional (3D) scaffolds is a major focus in bone tissue engineering. Incorporation of growth factors into bioprinted scaffolds offers many new possibilities regarding both biological and architectural properties of the scaffolds. This study investigates whether the sustained release of bone morphogenetic protein 2 (BMP-2) influences osteogenicity of tissue engineered bioprinted constructs. BMP-2 loaded on gelatin microparticles (GMPs) was used as a sustained release system, which was dispersed in hydrogel-based constructs and compared to direct inclusion of BMP-2 in alginate or control GMPs. The constructs were supplemented with goat multipotent stromal cells (gMSCs) and biphasic calcium phosphate to study osteogenic differentiation and bone formation respectively. BMP-2 release kinetics and bioactivity showed continuous release for three weeks coinciding with osteogenicity. Osteogenic differentiation and bone formation of bioprinted GMP containing constructs were investigated after subcutaneous implantation in mice or rats. BMP-2 significantly increased bone formation, which was not influenced by the release timing. We showed that 3D printing of controlled release particles is feasible and that the released BMP-2 directs osteogenic differentiation in vitro and in vivo. [ABSTRACT FROM AUTHOR]

Abstract: Thermosensitive chitosan hydrogels containing sodium beta-glycerophosphate (β-GP), whose gelation is induced by increasing temperature to body temperature, were functionalized by incorporation of alkaline phosphatase (ALP), an enzyme involved in mineralization of bone. ALP incorporation led to acceleration of gelation upon increase of temperature for four different chitosan preparations of differing molecular weight, as demonstrated by rheometric time sweeps at 37°C. Hydrogels containing ALP were subsequently incubated in calcium glycerophosphate (Ca-GP) solution to induce their mineralization with calcium phosphate (CaP) in order to improve their suitability as materials for bone replacement. Incorporated ALP retained its bioactivity and induced formation of CaP mineral, as confirmed by SEM, FTIR, Raman spectroscopy, XRD, ICP-OES, and increases in dry mass percentage, which rose with increasing ALP concentration and incubation time in Ca-GP solution. The results demonstrate that ALP accelerates formation of thermosensitive chitosan/β-GP hydrogels and induces their mineralization with CaP, which paves the way for applications as injectable bone replacement materials. [Copyright &y& Elsevier]

Abstract: During the process of bone regeneration, a multitude of morphogenetic signaling factors regulate cellular behavior and ultimately tissue response. These factors are presented to cells under strong spatial and temporal control, which stresses the relevance of controlled delivery of multiple growth factors for bone tissue regeneration. This demand for biomimetic delivery has prompted the development of a novel generation of biomaterials that is capable of delivering multiple growth factors in a controlled manner. Therefore, the current study has exploited the strong capacity of colloidal gels solely made of oppositely charged gelatin nanospheres to obtain controlled release of angiogenic and osteogenic growth factors. The release kinetics of dual delivery of osteogenic bone morphogenetic protein-2 (BMP-2) and angiogenic basic fibroblast growth factor (bFGF) were investigated in vitro by radiolabeling the respective growth factors and monitoring their release in vitro. Furthermore, the effect of single or dual delivery of BMP-2 and bFGF on bone regeneration was evaluated in vivo using a rat femoral condyle defect model. The in vitro results confirmed that the delivery kinetics of BMP-2 and/or bFGF are more dependent on the degree of crosslinking than on the type of gelatin. Sequential release characterized by rapid release of angiogenic bFGF and more sustained release of BMP-2 was obtained by loading bFGF onto cationic nanospheres of low crosslinking density and BMP-2 onto anionic nanospheres of high crosslinking density. The in vivo study demonstrated the biocompatibility and biodegradability of bare colloidal gelatin gels, and did not show any adverse effects on the process of bone healing after 4week of implantation since the volumes of new bone formation were comparable to empty control defects. An obvious stimulatory effect on bone regeneration was observed for the colloidal gels loaded with BMP-2, whereas bFGF-loaded colloidal gelatin gels did not influence the rate of bone regeneration. In contrast, the combined delivery of BMP-2 and bFGF resulted into an inhibitory effect on osteogenesis under the current experimental conditions. Summarizing, the current study proved that nanostructured colloidal gelatin gels are suitable carriers for programmed and sustained release of multiple therapeutic proteins for tissue regeneration. [Copyright &y& Elsevier]

Abstract: Colloidal gels have recently emerged as a promising new class of materials for regenerative medicine by employing micro- and nanospheres as building blocks to assemble into integral scaffolds. To this end, physically crosslinked particulate networks are formed that are injectable yet cohesive. By varying the physicochemical properties of different particle populations, the suitability of colloidal gels for programmed delivery of multiple therapeutic proteins is superior over conventional monolithic gels that lack this strong capacity for controlled drug release. Colloidal gels made of biodegradable polymer micro- or nanospheres have been widely investigated over the past few years, but a direct comparison between micro- vs. nanostructured colloidal gels has not been made yet. Therefore, the current study has compared the viscoelastic properties and capacity for drug release of colloidal gels made of oppositely charged gelatin microspheres vs. nanospheres. Viscoelastic properties of the colloidal gelatin gels were characterized by rheology and simple injectability tests, and in vitro release of two selected osteogenic proteins (i.e. bone morphogenetic protein-2 (BMP-2) and alkaline phosphatase (ALP)) from the colloidal gelatin gels was evaluated using radiolabeled BMP-2 and ALP. Nanostructured colloidal gelatin gels displayed superior viscoelastic properties over microsphere-based gels in terms of elasticity, injectability, structural integrity, and self-healing behavior upon severe network destruction. In contrast, microstructured colloidal gelatin gels exhibited poor gel strength and integrity, unfavorable injectability, and did not recover after shearing, resulting from the poor gel cohesion due to insufficiently strong interparticle forces. Regarding the capacity for drug delivery, sustained growth factor (BMP-2) release was obtained for both micro- and nanosphere-based gels, the kinetics of which were mainly depending on the particle size of gelatin spheres with the same crosslinking density. Therefore, the optimal gelatin carrier for drug delivery in terms of particle size and crosslinking density still needs to be established for specific clinical indications that require either short-term or long-term release. It can be concluded that nanostructured colloidal gelatin gels show great potential for sustained delivery of therapeutic proteins, whereas microstructured colloidal gelatin gels are not sufficiently cohesive as injectables for biomedical applications. [Copyright &y& Elsevier]

Abstract: Current state of the art reconstruction of bony defects in the craniomaxillofacial (CMF) area involves transplantation of autogenous or allogenous bone grafts. However, the inherent drawbacks of this approach strongly urge clinicians and researchers to explore alternative treatment options. Currently, a wide interest exists in local delivery of biomolecules from synthetic biomaterials for CMF bone regeneration, in which small biomolecules are rapidly emerging in recent years as an interesting adjunct for upgrading the clinical treatment of CMF bone regeneration under compromised healing conditions. This review highlights recent advances in the local delivery small and large biomolecules for the clinical treatment of CMF bone defects. Further, it provides a perspective on the efficacy of biomolecule delivery in CMF bone regeneration by reviewing presently available reports of pre-clinical studies using various animal models. [Copyright &y& Elsevier]

Objectives: Electrostatic spray deposition (ESD) is a new technique to deposit calcium phosphate (CaP) coatings. The aim of the present study was to evaluate the bone behavior of ESD CaP-coated implants with various degrees of crystallinities in the trabecular bone of the femoral condyle of goats. Material and methods: Using the ESD technique, thin porous CaP coatings were deposited on tapered, conical, screw-shaped titanium implants. Three different heat-treatments were applied, resulting in amorphous CaP (400°C, ESD1), crystalline carbonate apatite (500°C, ESD2), and crystalline carbonated hydroxyapatite (700°C, ESD3). Implants were inserted into the trabecular bone of the femoral condyles of goats for 12 weeks, and titanium (Ti) implants served as controls. Results: The results showed that ESD-derived coatings are osteocompatible. Histomorphometrical analysis showed that the application of a CaP coating resulted in more bone contact along the press-fit area of the implant compared with the Ti implants. Moreover, the percentage bone contact of the ESD3-coated implants was increased, compared with the Ti control group. Regarding the other coatings, no differences were found compared with the control group. Conclusion: Crystalline carbonated hydroxyapatite ESD-coated implants positively influenced the biological performance compared with Ti control implants. [ABSTRACT FROM AUTHOR]

To investigate the delivery of next-generation macromolecular drugs, such as engineered proteins and mRNA-containing nanoparticles, there is an increasing push towards the use of physiologically relevant disease models that incorporate human cells and do not face ethical dilemmas associated with animal use. Here, we illustrate the versatility and ease of use of a microfluidic platform for studying drug delivery using high-resolution microscopy in 3D. Using this microfluidic platform, we successfully demonstrate the specific targeting of carbonic anhydrase IX (CAIX) on cells overexpressing the protein in a tumor-mimicking chip system using affibodies, with CAIX-negative cells and non-binding affibodies as controls. Furthermore, we demonstrate this system's feasibility for testing mRNA-containing biomaterials designed to regenerate bone defects. To this end, peptide- and lipid-based mRNA formulations were successfully mixed with colloidal gelatin in microfluidic devices, while translational activity was studied by the expression of a green fluorescent protein. This microfluidic platform enables the testing of mRNA delivery from colloidal biomaterials of relatively high densities, which represents a first important step towards a bone-on-a-chip platform. Collectively, by illustrating the ease of adaptation of our microfluidic platform towards use in distinct applications, we show that our microfluidic chip represents a powerful and flexible way to investigate drug delivery in 3D disease-mimicking culture systems that recapitulate key parameters associated with in vivo drug application. [ABSTRACT FROM AUTHOR]

In the present study we have studied the incorporation and release of selenite ions (SeO 3 2−) in hydroxyapatite nanoparticles for the treatment of bone tumors. Two types of selenium-doped hydroxyapatite (HASe) nanoparticles (NPs) with a nominal Se/(P + Se) molar ratio ranging from 0.01 up to 0.40 have been synthesized by a new and mild wet method. The two series of samples were thoroughly characterized and resulted to be slightly different in chemical composition, but they had similar properties in terms of morphology and degree of crystallinity. Selenium release from HASe was investigated under neutral and acidic conditions to simulate both healthy tissues and the low-pH environment surrounding a tumor mass, respectively. The comparison of the release profiles at two pH values clearly showed the possibility of modulating the Se release by simply changing the amount of Se in the HASe particles. The correlation between the physicochemical properties of HASe and their dissolution as a function of pH has been also investigated to facilitate future application of the NPs as chemotherapeutic adjuvant agents. Finally, the cytotoxic activity of HASe was evaluated using prostate (PC3) and breast (MDA-MB-231) cancer cells as well as healthy human bone marrow stem cells (hBMSc). HASe NPs exerted a good cytocompatibility at low concentration of Se but, with high Se doping concentration, they displayed strong cytotoxicity. Selenium-doped hydroxyapatite (HASe) nanoparticles for potential treatment of bone tumors were effectively doped with Se up to 13%wt by a mild and green synthesis. In vitro ions' release and cytotoxicity studies have shown that HASe with high Se-doping exert cytotoxicity on cancer but also on healthy cells. Unlabelled Image • Se-doped hydroxyapatite (HASe) nanoparticles for potential treatment of bone tumors. • Hydroxyapatite was effectively doped with Se up to 13%wt by a mild and green synthesis. • HASe with low crystallinity resembling the physico-chemical properties of bone mineral. • HASe with high Se doping exerts cytotoxicity on cancer cells but also on healthy cells. [ABSTRACT FROM AUTHOR]