Osteosarcoma cell death induced by innovative scaffolds doped with chemotherapeutics.

Osteosarcoma (OS) cancer treatments include systemic chemotherapy and surgical resection. In the last years, novel treatment approaches have been proposed, which employ a drug-delivery system to prevent offside effects and improves treatment efficacy. Locally delivering anticancer compounds improves on high local concentrations with more efficient tumour-killing effect, reduced drugs resistance and confined systemic effects. Here, the synthesis of injectable strontium-doped calcium phosphate (SrCPC) scaffold was proposed as drug delivery system to combine bone tissue regeneration and anticancer treatment by controlled release of methotrexate (MTX) and doxorubicin (DOX), coded as SrCPC-MTX and SrCPC-DOX, respectively. The drug-loaded cements were tested in an in vitro model of human OS cell line SAOS-2, engineered OS cell line (SAOS-2-eGFP) and U2-OS. The ability of doped scaffolds to induce OS cell death and apoptosis was assessed analysing cell proliferation and Caspase-3/7 activities, respectively. To determine if OS cells grown on doped-scaffolds change their migratory ability and invasiveness, a wound-healing assay was performed. In addition, the osteogenic potential of SrCPC material was evaluated using human adipose derived-mesenchymal stem cells. Osteogenic markers such as (i) the mineral matrix deposition was analysed by alizarin red staining; (ii) the osteocalcin (OCN) protein expression was investigated by enzyme-linked immunosorbent assay test, and (iii) the osteogenic process was studied by real-time polymerase chain reaction array. The delivery system induced cell-killing cytotoxic effects and apoptosis in OS cell lines up to Day 7. SrCPC demonstrates a good cytocompatibility and it induced upregulation of osteogenic genes involved in the skeletal development pathway, together with OCN protein expression and mineral matrix deposition. The proposed approach, based on the local, sustained release of anticancer drugs from nanostructured biomimetic drug-loaded cements is promising for future therapies aiming to combine bone regeneration and anticancer local therapy.

A model approach to show that monocytes can enter microporous ß-TCP ceramics.

ß-TCP ceramics are versatile bone substitute materials and show many interactions with cells of the monocyte-macrophage-lineage. The possibility of monocytes entering microporous ß-TCP ceramics has however not yet been researched. In this study, we used a model approach to investigate whether monocytes might enter ß-TCP, providing a possible explanation for the origin of CD68-positive osteoclast-like giant cells found in earlier works.We used flow chambers to unidirectionally load BC, PRP, or PPP into slice models of either 2 mm or 6 mm ß-TCP. Immunofluorescence for CD68 and live/dead staining was performed after the loading process.Our results show that monocytes were present in a relevant number of PRP and BC slices representing the inside of our 2 mm slice model and also present on the actual inside of our 6 mm model. For PPP, monocytes were not found beyond the surface in either model.Our results indicate the possibility of a new and so far neglected constituent in ß-TCP degradation, perhaps causing the process of ceramic degradation also starting from inside the ceramics as opposed to the current understanding. We also demonstrated flow chambers as a possible new in vitro model for interactions between blood and ß-TCP.

Dual release of daptomycin and BMP-2 from a composite of ß-TCP ceramic and ADA gelatin.

BACKGROUND: Antibiotic-containing carrier systems are one option that offers the advantage of releasing active ingredients over a longer period of time. In vitro sustained drug release from a carrier system consisting of microporous ß-TCP ceramic and alginate has been reported in previous works. Alginate dialdehyde (ADA) gelatin gel showed both better mechanical properties when loaded into a ß-TCP ceramic and higher biodegradability than pure alginate. METHODS: Dual release of daptomycin and BMP-2 was measured on days 1, 2, 3, 6, 9, 14, 21, and 28 by HPLC and ELISA. After release, the microbial efficacy of the daptomycin was verified and the biocompatibility of the composite was tested in cell culture. RESULTS: Daptomycin and the model compound FITC protein A (n = 30) were released from the composite over 28 days. A Daptomycin release above the minimum inhibitory concentration (MIC) by day 9 and a burst release of 71.7 ± 5.9% were observed in the loaded ceramics. Low concentrations of BMP-2 were released from the loaded ceramics over 28 days.

Immobilized Dipeptidase in Manganese Ion-Loaded Polyethylenimine-Induced Calcium Phosphate Nanocrystals for Carnosine Synthesis.

Carnosine is a natural bioactive dipeptide with important physiological functions widely used in food and medicine. Dipeptidase (PepD) from Serratia marcescens can catalyze the reverse hydrolytic reaction of ß-alanine with l-histidine to synthesize carnosine in the presence of Mn2+. However, it remains challenging to practice carnosine biosynthesis due to the low activity and high cost of the enzyme. Therefore, the development of biocatalysts with high activity and stability is of significance for carnosine synthesis. Here, we proposed to chelate Mn2+ to polyethylenimine (PEI) that induced rapid formation of calcium phosphate nanocrystals (CaP), and Mn-PEI@CaP was used for PepD immobilization via electrostatic interaction. Mn-PEI@CaP as the carrier enhanced the stability of the immobilized enzyme. Moreover, Mn2+ loaded in the carrier acted as an in situ activator of the immobilized PepD for facilitating the biocatalytic process of carnosine synthesis. The as-prepared immobilized enzyme (PepD-Mn-PEI@CaP) kept similar activity with free PepD plus Mn2+ (activity recovery, 102.5%), while exhibiting elevated thermal stability and pH tolerance. Moreover, it exhibited about two times faster carnosine synthesis than the free PepD system. PepD-Mn-PEI@CaP retained 86.8% of the original activity after eight cycles of batch catalysis without the addition of free Mn2+ ions during multiple cycles. This work provides a new strategy for the co-immobilization of PepD and Mn2+, which greatly improves the operability of the biocatalysis and demonstrates the potential of the immobilized PepD system for efficient carnosine synthesis.

Nucleation Domains in Biomineralization: Biomolecular Sequence and Conformational Features.

Biomolecules play a vital role in the regulation of biomineralization. However, the characteristics of practical nucleation domains are still sketchy. Herein, the effects of the representative biomolecular sequence and conformations on calcium phosphate (Ca-P) nucleation and mineralization are investigated. The results of computer simulations and experiments prove that the line in the arrangement of dual acidic/essential amino acids with a single interval (Bc (Basic) -N (Neutral) -Bc-N-Ac (Acidic)- NN-Ac-N) is most conducive to the nucleation. 2α-helix conformation can best induce Ca-P ion cluster formation and nucleation. "Ac- × × × -Bc" sequences with α-helix are found to be the features of efficient nucleation domains, in which process, molecular recognition plays a non-negligible role. It further indicates that the sequence determines the potential of nucleation/mineralization of biomolecules, and conformation determines the ability of that during functional execution. The findings will guide the synthesis of biomimetic mineralized materials with improved performance for bone repair.

Stratified control of chemical crystallization in a pellet fluidized bed for pH-Adjusted fluoride and phosphate reduction: An experimental study.

Chemical crystallization granulation in a fluidized bed offers an environmentally friendly technology with significant promise for fluoride removal. This study investigates the impact of stratified pH control in a crystallization granulation fluidized bed for the removal of fluoride and phosphate on a pilot scale. The results indicate that using dolomite as a seed crystal, employing sodium dihydrogen phosphate (SDP) and calcium chloride as crystallizing agents, and controlling the molar ratio n(F):n(P):n(Ca) = 1:5:10 with an upflow velocity of 7.52 m/h, effectively removes fluoride and phosphate. Stratified pH control-maintaining weakly acidic conditions (pH = 6-7) at the bottom and weakly alkaline conditions (pH = 7-8) at the top-facilitates the induction of fluoroapatite (FAP) and calcium phosphate crystallization. This approach reduces groundwater fluoride levels from 9.5 mg/L to 0.2-0.6 mg/L and phosphate levels to 0.1-0.2 mg/L. Particle size analysis, scanning electron microscopy-energy-dispersive X-ray spectroscopy, and X-ray diffraction physical characterizations reveal significant differences in crystal morphology between the top and bottom layers, with the lower layer primarily generating high-purity FAP crystals. Further analysis shows that dolomite-induced FAP crystallization offers distinct advantages. SDP not only dissolves on the dolomite surface to provide active sites for crystallization but also, under weakly acidic conditions, renders both dolomite and FAP surfaces negatively charged. This allows for the effective adsorption of PO43-, HPO42-, and F- anions onto the crystal surfaces. This study provides supporting data for the removal of fluoride from groundwater through induced FAP crystallization in a chemical crystallization pellet fluidized bed.

Calcium phosphate coating enhances osteointegration of melt electrowritten scaffold by regulating macrophage polarization.

The osteoimmune microenvironment induced by implants plays a significant role in bone regeneration. It is essential to efficiently and timely switch the macrophage phenotype from M1 to M2 for optimal bone healing. This study examined the impact of a calcium phosphate (CaP) coating on the physiochemical properties of highly ordered polycaprolactone (PCL) scaffolds fabricated using melt electrowritten (MEW). Additionally, it investigated the influence of these scaffolds on macrophage polarization and their immunomodulation on osteogenesis. The results revealed that the CaP coated PCL scaffold exhibited a rougher surface topography and higher hydrophilicity in comparison to the PCL scaffold without coating. Besides, the surface morphology of the coating and the release of Ca2+ from the CaP coating were crucial in regulating the transition of macrophages from M1 to M2 phenotypes. They might activate the PI3K/AKT and cAMP-PKA pathways, respectively, to facilitate M2 polarization. In addition, the osteoimmune microenvironment induced by CaP coated PCL could not only enhance the osteogenic differentiation of bone marrow-derived mesenchymal stem cells (BMSCs) in vitro but also promote the bone regeneration in vivo. Taken together, the CaP coating can be employed to control the phenotypic switching of macrophages, thereby creating a beneficial immunomodulatory microenvironment that promotes bone regeneration.

Magnesium malate-modified calcium phosphate bone cement promotes the repair of vertebral bone defects in minipigs via regulating CGRP.

Active artificial bone substitutes are crucial in bone repair and reconstruction. Calcium phosphate bone cement (CPC) is known for its biocompatibility, degradability, and ability to fill various shaped bone defects. However, its low osteoinductive capacity limits bone regeneration applications. Effectively integrating osteoinductive magnesium ions with CPC remains a challenge. Herein, we developed magnesium malate-modified CPC (MCPC). Incorporating 5% magnesium malate significantly enhances the compressive strength of CPC to (6.18 ± 0.49) MPa, reduces setting time and improves disintegration resistance. In vitro, MCPC steadily releases magnesium ions, promoting the proliferation of MC3T3-E1 cells without causing significant apoptosis, proving its biocompatibility. Molecularly, magnesium malate prompts macrophages to release prostaglandin E2 (PGE2) and synergistically stimulates dorsal root ganglion (DRG) neurons to synthesize and release calcitonin gene-related peptide (CGRP). The CGRP released by DRG neurons enhances the expression of the key osteogenic transcription factor Runt-related transcription factor-2 (RUNX2) in MC3T3-E1 cells, promoting osteogenesis. In vivo experiments using minipig vertebral bone defect model showed MCPC significantly increases the bone volume fraction, bone density, new bone formation, and proportion of mature bone in the defect area compared to CPC. Additionally, MCPC group exhibited significantly higher levels of osteogenesis and angiogenesis markers compared to CPC group, with no inflammation or necrosis observed in the hearts, livers, or kidneys, indicating its good biocompatibility. In conclusion, MCPC participates in the repair of bone defects in the complex post-fracture microenvironment through interactions among macrophages, DRG neurons, and osteoblasts. This demonstrates its significant potential for clinical application in bone defect repair.

Biomimetic bone-periosteum scaffold for spatiotemporal regulated innervated bone regeneration and therapy of osteosarcoma.

The complexity of repairing large segment defects and eradicating residual tumor cell puts the osteosarcoma clinical management challenging. Current biomaterial design often overlooks the crucial role of precisely regulating innervation in bone regeneration. Here, we develop a Germanium Selenium (GeSe) co-doped polylactic acid (PLA) nanofiber membrane-coated tricalcium phosphate bioceramic scaffold (TCP-PLA/GeSe) that mimics the bone-periosteum structure. This biomimetic scaffold offers a dual functionality, combining piezoelectric and photothermal conversion capabilities while remaining biodegradable. When subjected to ultrasound irradiation, the US-electric stimulation of TCP-PLA/GeSe enables spatiotemporal control of neurogenic differentiation. This feature supports early innervation during bone formation, promoting early neurogenic differentiation of Schwann cells (SCs) by increasing intracellular Ca2+ and subsequently activating the PI3K-Akt and Ras signaling pathways. The biomimetic scaffold also demonstrates exceptional osteogenic differentiation potential under ultrasound irradiation. In rabbit model of large segment bone defects, the TCP-PLA/GeSe demonstrates promoted osteogenesis and nerve fibre ingrowth. The combined attributes of high photothermal conversion capacity and the sustained release of anti-tumor selenium from the TCP-PLA/GeSe enable the synergistic eradication of osteosarcoma both in vitro and in vivo. This strategy provides new insights on designing advanced biomaterials of repairing large segment bone defect and osteosarcoma.

Porous Nano-Fiber Structure of Modified Electrospun Chitosan GBR Membranes Improve Osteoblast Calcium Phosphate Deposition in Osteoblast-Fibroblast Co-Cultures.

Desirable characteristics of electrospun chitosan membranes (ESCM) for guided bone regeneration are their nanofiber structure that mimics the extracellular fiber matrix and porosity for the exchange of signals between bone and soft tissue compartments. However, ESCM are susceptible to swelling and loss of nanofiber and porous structure in physiological environments. A novel post-electrospinning method using di-tert-butyl dicarbonate (tBOC) prevents swelling and loss of nanofibrous structure better than sodium carbonate treatments. This study aimed to evaluate the hypothesis that retention of nanofiber morphology and high porosity of tBOC-modified ESCM (tBOC-ESCM) would support more bone mineralization in osteoblast-fibroblast co-cultures compared to Na2CO3 treated membranes (Na2CO3-ESCM) and solution-cast chitosan solid films (CM-film). The results showed that only the tBOC-ESCM retained the nanofibrous structure and had approximately 14 times more pore volume than Na2CO3-ESCM and thousands of times more pore volume than CM-films, respectively. In co-cultures, the tBOC-ESCM resulted in a significantly greater calcium-phosphate deposition by osteoblasts than either the Na2CO3-ESCM or CM-film (p < 0.05). This work supports the study hypothesis that tBOC-ESCM with nanofiber structure and high porosity promotes the exchange of signals between osteoblasts and fibroblasts, leading to improved mineralization in vitro and thus potentially improved bone healing and regeneration in guided bone regeneration applications.

Pore graded borosilicate bioactive glass scaffolds: in vitro dissolution and cytocompatibility.

3D borosilicate bioactive glass (1393B20 and B12.5MgSr) scaffolds were prepared by robocasting, with and without a dense layer at the top. Pore graded scaffolds are promising as they allow for membrane deposition and could limit the risk of soft tissue infiltration. In vitro dissolution was studied in tris(hydroxymethyl)aminomethane (TRIS) and Simulated Body Fluid (SBF). 1393B20 scaffolds dissolved faster than B12.5MgSr in TRIS whereas they dissolved slower in SBF. The difference in dissolution profiles, as a function of the medium used, is assigned to the different rates of precipitation of hydroxyapatite (HA). While the precipitation of calcium phosphate (CaP) in the form of HA, first sign of bioactivity, was confirmed by ICP, FTIR-ATR and SEM-EDX analysis for both compositions, 1393B20 was found to precipitate HA at a faster rate. The presence of a dense top layer did not significantly impact the dissolution rate and CaP precipitation. In vitro cell culture was performed using human adipose-derived stem cells (hADSCs). Prior to cell plating, a preincubation of 3 days was found optimum to prevent burst ion release. In direct contact, cells proliferate and spread on the scaffolds while maintaining characteristic spindle morphology. Cell plated on 1393B20 scaffolds showed increased viability when compared to cell plated on B12.5MgSr. The lower cell viability, when testing B12.5MgSr, was assigned to the depletion of Ca2+ ions from culture medium and higher pH. Static cell culture leads to believe that the scaffold produced from the 1393B20 glass composition are promising in bone regeneration applications.

Biological and mechanical performance of calcium phosphate cements modified with phytic acid.

Calcium phosphate cements, primarily brushite cements, require the addition of setting retarders to ensure adequate processing time and processability. So far, citric acid has been the primary setting retarder used in this context. Due to the poor biocompatibility, it is crucial to explore alternative options for better processing. In recent years, the setting retarder phytic acid (IP6) has been increasingly investigated. This study investigates the biological behaviour of calcium phosphate cements with varying concentrations of IP6, in addition to their physical properties. Therefore cytocompatibility in vitro testing was performed using osteoblastic (MG-63) and osteoclastic (RAW 264.7 differentiated with RANKL) cells. We could demonstrate that the physical properties like the compressive strength of specimens formed with IP6 (brushite_IP6_5 = 11.2 MPa) were improved compared to the reference (brushite = 9.8 MPa). In osteoblast and osteoclast assays, IP6 exhibited significantly better cytocompatibility in terms of cell activity and cell number for brushite cements up to 11 times compared to the brushite reference. In contrast, the calcium-deficient hydroxyapatite (CDHA) cements produced similar results for IP6 (CDHA_IP6_0.25 = 27.0 MPa) when compared to their reference (CDHA = 21.2 MPa). Interestingly, lower doses of IP6 were found to be more effective than higher doses with up to 3 times higher. Additionally, IP6 significantly increased degradation in both passive and active resorption. For these reasons, IP6 is emerging as a strong new competitor to established setting retarders such as citric acid. These cements have potential applications in bone augmentation, the stabilisation of non-load bearing fractures (craniofacial), or the cementation of metal implants.

3D-printed polycaprolactone scaffolds coated with beta tricalcium phosphate for bone regeneration.

BACKGROUND/PURPOSE: 3D-printing technology is an important tool for the bone tissue engineering (BTE). The aim of this study was to investigate the interaction of polycaprolactone (PCL) scaffolds and modified mesh PCL coated with beta TCP (PCL/ß-TCP) scaffolds with MG-63. METHODS: This study used the fused deposition modeling (FDM) technique with the 3D printing technique to fabricate the thermoplastic polymer and composite scaffolds. Scaffold structure and coating quality were observed under a scanning electron microscope (SEM). MG-63 cells were injected and attached to the mesh-manufactured PCL scaffolds. The biocompatibility of mesh structured PCL and PCL/ß-TCP scaffolds could be examined by measuring the viability of MG-63 cells of MTT assay. Bone cell differentiation was evaluated ALP activity by mineralization assay. RESULTS: The results showed that both mesh PCL scaffolds and PCL/ß-TCP scaffolds were non-toxic to the cells. The ALP activities of cells in PCL/ß-TCP scaffolds groups were significant differences and better than PCL groups in all groups at all experimental dates. The mineralization process was time-dependent, and significantly higher mineralization of osteosarcoma cells was observed on PCL/ß-TCP scaffolds at experimental dates. CONCLUSION: We concluded that both meshes structured PCL and PCL/ß-TCP scaffolds could promote the MG-63 cell growth, and PCL/ß-TCP was better than the PCL scaffolds for the outcome of MG63 cell differentiation and mineralization.

Biomimetic Mineral Synthesis by Nanopatterned Supramolecular-Block Copolymer Templates.

Supramolecular structures of matrix proteins in mineralizing tissues are known to direct the crystallization of inorganic materials. Here we demonstrate how such structures can be synthetically directed into predetermined patterns for which functionality is maintained. The study employs block copolymer lamellar patterns with alternating hydrophilic and hydrophobic regions to direct the assembly of amelogenin-derived peptide nanoribbons that template calcium phosphate nucleation by creating a low-energy interface. Results show that the patterned nanoribbons retain their ß-sheet structure and function and direct the formation of filamentous and plate-shaped calcium phosphate with high fidelity, where the phase, amorphous or crystalline, depends on the choice of mineral precursor and the fidelity depends on peptide sequence. The common ability of supramolecular systems to assemble on surfaces with appropriate chemistry combined with the tendency of many templates to mineralize multiple inorganic materials implies this approach defines a general platform for bottom-up-patterning of hybrid organic-inorganic materials.

Obtaining Polyvinylpyrrolidone Fibers Using the Electroforming Method with the Inclusion of Microcrystalline High-Temperature Phosphates.

The results of the synthesis of microcrystalline calcium phosphates such as hydroxoapatite, pyrophosphate, and tricalcium phosphate are presented herein. The influence of the addition of polyvinylpyrrolidone (PVP) on the phase characteristics of the resulting high-temperature ceramic sample is considered. The X-ray results show that hydroxyapatite (HAp) consists of a Ca5(PO4)3(OH) phase, while the sample with the addition of polyvinylpyrrolidone contains ß-Ca3(PO4)2 (65.5%) and ß-Ca2P2O7 (34.5%) phases calcium phosphates (CPs). IR spectroscopy was used to characterize the compositions of the samples. An important characteristic of the obtained samples is the elemental Ca/P ratio, which was determined via energy-dispersive analysis. The data obtained are consistent with the composition of dental enamel apatites, namely, in the CPs (1.27) and HAp (1.40). SEM was used to study the morphology of the surfaces of hydroxyapatite particles. Polyvinylpyrrolidone polymer fibers were obtained using the electroforming method with the inclusion of CPs in the composition. The fibers were oriented randomly, and nanoscale hydroxyapatite particles were incorporated into the fiber structure. Solubility data of the HAp, CPs, and Fibers in a physiological solution at room temperature and human body temperature were obtained. The solubility of the resulting HAp turned out to be higher than the solubility of the CPs. In turn, the concentration of Ca2+ in a physiological solution of PVP composite fibers with the inclusion of CPs was lower than that in powdered CPs.

Characterization of Porous ß-Type Tricalcium Phosphate Ceramics Formed via Physical Foaming with Freeze-Drying.

Porous ß-tricalcium phosphate (Ca3(PO4)2; ß-TCP) was prepared via freeze-drying and the effects of this process on pore shapes and sizes were investigated. Various samples were prepared by freezing ß-TCP slurries above a liquid nitrogen surface at -180 °C with subsequent immersion in liquid nitrogen at -196 °C. These materials were then dried under reduced pressure in a freeze-dryer, after which they were sintered with heating. Compared with conventional heat-based drying, the resulting pores were more spherical, which increased both the mechanical strength and porosity of the ß-TCP. These materials had a wide range of pore sizes from 50 to 200 µm, with the mean and median values both approximately 100 µm regardless of the freeze-drying conditions. Mercury porosimetry data showed that the samples contained small, interconnected pores with sizes of 1.24 ± 0.25 µm and macroscopic, interconnected pores of 25.8 ± 4.7 µm in size. The effects of nonionic surfactants having different hydrophilic/lipophilic balance (HLB) values on foaming and pore size were also investigated. Materials made with surfactants having lower HLB values exhibited smaller pores and lower porosity, whereas higher HLB surfactants gave higher porosity and slightly larger macropores. Even so, the pore diameter could not be readily controlled solely by adjusting the HLB value. The findings of this work indicated that high porosity (>75%) and good compressive strength (>2 MPa) can both be obtained in the same porous material and that foaming agents with HLB values between 12.0 and 13.5 were optimal.

Development of Rapid Bioactivity-Expressed Zr-50Ti Alloys by Surface Treatment with Modified Simulated Body Fluid.

Zr-50Ti alloys are promising biomaterials due to their excellent mechanical properties and low magnetic susceptibility. However, Zr-50Ti alloys do not inherently bond well with bone. This study aims to enhance the bioactivity and bonding strength of Zr-50Ti alloys for orthopedic implant materials. Initially, the surface of Zr-50Ti alloys was treated with a sulfuric acid solution to create a microporous structure, increasing surface roughness and area. Subsequently, low crystalline calcium phosphate (L-CaP) precipitation was controlled by adding Mg2+ and/or CO32- ions in modified simulated body fluid (m-SBF). The treated Zr-50Ti alloys were then subjected to cold isostatic pressing to force m-SBF into the micropores, followed by incubation to allow L-CaP formation. The apatite-forming process was tested in simulated body fluid (SBF). The results demonstrated that the incorporation of Mg2+ and/or CO32- ions enabled the L-CaP to cover the entire surface of Zr-50Ti alloys within only one day. After short-term soaking in SBF, the L-CaP layer, modulated by Mg2+ and/or CO32- ions, formed a uniform hydroxyapatite (HA) coating on the surface of the Zr-50Ti alloys, showing potential for optimized bone integration. After soaking in SBF for 14 days, the bonding strength between the apatite layer and alloy has the potential to meet the orthopedic application requirement of 22 MPa. This study demonstrates an effective method to enhance the bioactivity and bonding strength of Zr-50Ti alloys for orthopedic applications.

Croscarmellose Sodium as Pelletization Aid in Extrusion-Spheronization.

Only few excipients are known to be suitable as pelletization aids. In this study, the potential use of croscarmellose sodium (CCS) as pelletization aid was investigated. Furthermore, the impact of cations on extrusion-spheronization (ES) of CCS was studied and different grades of CCS were tested. The influence of different cations on the swelling of CCS was investigated by laser diffraction. Mixtures of CCS with lactose monohydrate as filler with or without the inclusion of different cations were produced. The mixtures were investigated by mixer torque rheometry and consequently extruded and spheronized. Resulting pellets were analyzed by dynamic image analysis. In addition, mixtures of different CCS grades with dibasic calcium phosphate anhydrous (DP) and a mixture with praziquantel (PZQ) as filler were investigated. Calcium and magnesium cations caused a decrease of the swelling of CCS and influenced the use of CCS as pelletization aid since they needed to be included for successful ES. Aluminum, however, led to an aggregation of the CCS particles and to failure of extrusion. The inclusion of cations decreased the uptake of water by the mixtures which also reduced the liquid-to-solid-ratio (L/S) for successful ES. This was shown to be dependent on the amount of divalent cations in the mixture. With DP or PZQ as filler, no addition of cations was necessary for a successful production of pellets, however the optimal L/S for ES was dependent on the CCS grade used. In conclusion, CCS can be used as a pelletization aid.

Avoiding and allowing apatite precipitation in oxygenic photolithotrophs.

The essential elements Ca and P, taken up and used metabolically as Ca2+ and H2 PO4 - /HPO4 2- respectively, could precipitate as one or more of the insoluble forms calcium phosphate (mainly apatite) if the free ion concentrations and pH are high enough. In the cytosol, chloroplast stroma, and mitochondrial matrix, the very low free Ca2+ concentration avoids calcium phosphate precipitation, apart from occasionally in the mitochondrial matrix. The low free Ca2+ concentration in these compartments is commonly thought of in terms of the role of Ca2+ in signalling. However, it also helps avoids calcium phosphate precipitation, and this could be its earliest function in evolution. In vacuoles, cell walls, and xylem conduits, there can be relatively high concentrations of Ca2+ and inorganic orthophosphate, but pH and/or other ligands for Ca2+ , suggests that calcium phosphate precipitates are rare. However, apatite is precipitated under metabolic control in shoot trichomes, and by evaporative water loss in hydathodes, in some terrestrial flowering plants. In aquatic macrophytes that deposit CaCO3 on their cell walls or in their environment as a result of pH increase or removal of inhibitors of nucleation or crystal growth, phosphate is sometimes incorporated in the CaCO3 . Calcium phosphate precipitation also occurs in some stromatolites.

Calcium Phosphate Delivery Systems for Regeneration and Biomineralization of Mineralized Tissues of the Craniofacial Complex.

Calcium phosphate (CaP)-based materials have been extensively used for mineralized tissues in the craniofacial complex. Owing to their excellent biocompatibility, biodegradability, and inherent osteoconductive nature, their use as delivery systems for drugs and bioactive factors has several advantages. Of the three mineralized tissues in the craniofacial complex (bone, dentin, and enamel), only bone and dentin have some regenerative properties that can diminish due to disease and severe injuries. Therefore, targeting these regenerative tissues with CaP delivery systems carrying relevant drugs, morphogenic factors, and ions is imperative to improve tissue health in the mineralized tissue engineering field. In this review, the use of CaP-based microparticles, nanoparticles, and polymer-induced liquid precursor (PILPs) amorphous CaP nanodroplets for delivery to craniofacial bone and dentin are discussed. The use of these various form factors to obtain either a high local concentration of cargo at the macroscale and/or to deliver cargos precisely to nanoscale structures is also described. Finally, perspectives on the field using these CaP materials and next steps for the future delivery to the craniofacial complex are presented.