Enhanced model protein adsorption of nanoparticulate hydroxyapatite thin films on silk sericin and fibroin surfaces.

Hydroxyapatite coated metallic implants favorably combine the required biocompatibility with the mechanical properties. As an alternative to the industrial coating method of plasma spraying with inherently potential deleterious effects, sol-gel methods have attracted much attention. In this study, the effects of intermediate silk fibroin and silk sericin layers on the protein adsorption capacity of hydroxyapatite films formed by a particulate sol-gel method were determined experimentally. The preparation of the layered silk protein/hydroxyapatite structures on glass substrates, and the effects of the underlying silk proteins on the topography of the hydroxyapatite coatings were described. The topography of the hydroxyapatite layer fabricated on the silk sericin was such that the hydroxyapatite particles were oriented forming an oriented crystalline surface. The model protein (bovine serum albumin) adsorption increased to 2.62 µg/cm2 on the latter surface as compared to 1.37 µg/cm2 of hydroxyapatite on glass without an intermediate silk sericin layer. The BSA adsorption on glass (blank), glass/c-HAp, glass/m-HAp, glass/sericin/c-HAp, and glass/sericin/m-HAp substrates, reported as decrease in BSA concentration versus contact time.

Investigation of the influence of TiO2distribution on HA/TiO2composite wetting ability using the dispersant SDBS, high-temperature annealing, and ultrasonication.

The use of composites such as hydroxyapatite (HA)/TiO2in bioapplications has attracted increasing attention in recent years. Herein, for the enhancement wetting ability and biocompatibility, the HA/TiO2composite was subjected to different treatments to improve nanoparticle (NP) distribution and surface energy with an aim of mitigating nanotoxicity concerns. The treatments included ultrasonication, high-temperature annealing, and addition of a dispersant and surfactant, sodium dodecylbenzenesulfonate (SDBS). Contact angle measurement tests revealed the effect of SDBS addition on the distribution of TiO2NPs on the HA surface: a decrease in the contact angle and, thus, an increase in the wetting ability of the HA/TiO2composite were observed. The combination of annealing and SDBS addition treatments allowed for guest TiO2particles to be uniformly distributed on the surface of the host HA particles, showing a rapid conversion from a hydrophobic to superhydrophilic property.In vitroinvestigation suggested that the cell viabilities of annealed HA/TiO2, SDBS-added HA/TiO2, and SDBS-added and annealed HA/TiO2reached 89.7%, 94.7%, and 95.8%, respectively, while those of HA and untreated HA/TiO2were 80.3% and 86.9%, respectively. The modified composites exhibited lower cytotoxicities than the unmodified systems (HA and HA/TiO2). Furthermore, the cell adhesion behavior of the composites was confirmed through actin-4',6-Diamidino-2-phenylindole (DAPI) staining, which showed negligible changes in the cytoskeleton architecture of the cells. This study confirmed that a modified HA/TiO2composite has potential for bioapplications.

Nerve conduit based on HAP/PDLLA/PRGD for peripheral nerve regeneration with sustained release of valproic acid.

The nerve conduits have been developed for nerve defect repair. However, no artificial conduits have obtained comparable results to autografts to bridge the large gaps. A possible reason for this poor performance may be a lack of sustainable neurotrophic support for axonal regrowth. Previous studies suggested nanocomposite conduits can be used as a carrier for valproic acid (VPA), a common drug that can produce effects similar to the neurotrophic factors. Here, we developed the novel bioabsorbable conduits based on hydroxyapatite/poly d-l-lactic acid (PDLLA)/poly{(lactic acid)-co-[(glycolic acid)-alt-(l-lysine)]} with sustained release of VPA. Firstly, the sustained release of VPA in this conduit was examined by high-performance liquid chromatography. Then Schwann cells were treated with the conduit extracts. The cell metabolic activity and proliferation were assayed by 3-[4,5-dimethyl-2-thiazolyl]-2,5-diphenyl-2-tetrazolium bromide and bromodeoxyuridine staining. A 10-mm segment of rat sciatic nerve was resected and then repaired, respectively, using the VPA conduit (Group A), the PDLLA conduit (Group B), or the autografts (Group C). Nerve conduction velocities (NCVs), compound muscle action potentials (CMAPs), and histological staining were assayed following the surgery. The cell metabolic activity and proliferation were significantly increased (p < .05) by the extracts from VPA-conduit extract compared to others. NCVs and CMAPs were significantly higher in Groups A and C than Group B (p < .05). The nerve density of Groups A and C was higher than Group B. There was no significant difference between Groups A and C. Taken together, this study suggested the sustained-release VPA conduit promoted peripheral nerve regeneration that was comparable to the autografts. It holds potential for future use in nerve regeneration.

Preparation, characterization and preliminary pharmacokinetic study of pH-sensitive Hydroxyapatite/Zein nano-drug delivery system for doxorubicin hydrochloride.

OBJECTIVES: Zein nanoparticles (Zein NPs) were used as a hydroxyapatite (HA) biomineralization template to generate HA/Zein NPs. Doxorubicin hydrochloride (DOX) was loaded on HA/Zein NPs (HA/Zein-DOX NPs) to improve its pH-sensitive release, bioavailability and decrease cardiotoxicity. METHODS: HA/Zein-DOX NPs were prepared by phase separation and biomimetic mineralization method. Particle size, polydispersity index (PDI), Zeta potential, transmission electron microscope, X-ray diffraction and Fourier-transform infrared spectroscopy of HA/Zein-DOX NPs were characterized. The nanoparticles were then evaluated in vitro and in vivo. KEY FINDINGS: The small PDI and high Zeta potential demonstrated that HA/Zein-DOX NPs were a stable and homogeneous dispersed system and that HA was mineralized on Zein-DOX NPs. HA/Zein-DOX NPs showed pH-sensitive release. Compared with free DOX, HA/Zein-DOX NPs increased cellular uptake which caused 7 times higher in-vitro cytotoxicity in 4T1 cells. Pharmacokinetic experiments indicated the t1/2ß and AUC0- t of HA/Zein-DOX NPs were 2.73- and 3.12-fold higher than those of DOX solution, respectively. Tissue distribution exhibited HA/Zein-DOX NPs reduced heart toxicity with lower heart targeting efficiency (18.58%) than that of DOX solution (37.62%). CONCLUSION: In this study, HA/Zein-DOX NPs represented an antitumour drug delivery system for DOX in clinical tumour therapy with improved bioavailability and decreased cardiotoxicity.

Doxycycline containing hydroxyapatite ceramic microspheres as a bone-targeting drug delivery system.

Drug delivery technology is a promising way to enhance the therapeutic efficacy of drugs. The purpose of this study is to evaluate the physical and chemical properties of hydroxyapatite ceramic microspheres loaded with doxycycline (HADOX), their effects on in vitro osteoblast viability, and their antimicrobial activity, and to determine the effects of DOX on the healing of rat sockets after tooth extraction. The internal microsphere porosity was sensitive to the treatment used to adsorb DOX onto microsphere surface; HA microspheres without DOX presented 26% of pores, whereas HADOX0.15 microspheres presented 52.0%. An initial drug release of 49.15 µg/ml was observed in the first 24 hr. The minimal inhibitory concentration (MIC) tested against Enterococcus faecalis demonstrated that bacterial growth was inhibited for up to 7 days. Results of cell viability and cell proliferation did not indicate statistical differences in the metabolic activity of HADOX samples relative to HA without DOX microspheres (p > .05). After 1 week, a discreet inflammation reaction was observed in the control group, and after 6 weeks, newly-formed bone was observed in the HADOX0.15 (p < .05). The HADOX did not interfere in the bone repair and controlled the early inflammatory response. HADOX could be a promising biomaterial to promote bone repair in infected sites.

The Effect of Boron-Containing Nano-Hydroxyapatite on Bone Cells.

Metabolic diseases or injuries damage bone structure and self-renewal capacity. Trace elements and hydroxyapatite crystals are important in the development of biomaterials to support the renewal of bone extracellular matrix. In this study, it was assumed that the boron-loaded nanometer-sized hydroxyapatite composite supports the construction of extracellular matrix by controlled boron release in order to prevent its toxic effect. In this context, boron release from nanometer-sized hydroxyapatite was calculated by ICP-MS as in large proportion within 1 h and continuing release was provided at a constant low dose. The effect of the boron-containing nanometer-sized hydroxyapatite composite on the proliferation of SaOS-2 osteoblasts and human bone marrow-derived mesenchymal stem cells was evaluated by WST-1 and compared with the effects of nano-hydroxyapatite and boric acid. Boron increased proliferation of mesenchymal stem cells at high doses and exhibited different effects on osteoblastic cell proliferation. Boron-containing nano-hydroxyapatite composites increased osteogenic differentiation of mesenchymal stem cells by increasing alkaline phosphatase activity, when compared to nano-hydroxyapatite composite and boric acid. The molecular mechanism of effective dose of boron-containing hydroxyapatite has been assessed by transcriptomic analysis and shown to affect genes involved in Wnt, TGF-ß, and response to stress signaling pathways when compared to nano-hydroxyapatite composite and boric acid. Finally, a safe osteoconductive dose range of boron-containing nano-hydroxyapatite composites for local repair of bone injuries and the molecular effect profile in the effective dose should be determined by further studies to validation of the regenerative therapeutic effect window.

An oral drug delivery system with programmed drug release and imaging properties for orthotopic colon cancer therapy.

Oral drug delivery systems (ODDSs) have attracted considerable attention in relation to orthotopic colon cancer therapy due to certain popular advantages. Unfortunately, their clinical applications are generally limited by the side-effects caused by systemic drug exposure and poor real-time monitoring capabilities. Inspired by the characteristics of pH changes of the gastrointestinal tract (GIT) and specific enzymes secreted by the colonic microflora, we anchored polyacrylic acid (PAA) and chitosan (CS) on Gd3+-doped mesoporous hydroxyapatite nanoparticles (Gd-MHAp NPs) to realize programmed drug release and magnetic resonance imaging (MRI) at the tumor sites. In particular, the grafted PAA, as a pH-responsive switch, could effect controlled drug release in the colon. Further, CS is functionalized as the enzyme-sensitive moiety, which could be degraded by ß-glycosidase in the colon. Gadolinium is a paramagnetic lanthanide element used in chelates, working as a contrast medium agent for an MRI system. Interestingly, after oral administration, CS and PAA could protect the drug-loaded nanoparticles (NPs) against variable physiological conditions in the GIT, allowing the drug to reach the colon tumor sites, preventing premature drug release. Enhanced drug concentrations at the colon tumor sites were achieved via this programmed drug release, which subsequently ameliorated the therapeutic effect. In addition, encapsulating both chemotherapeutic (5-fluorouracil, 5-FU) and targeted therapy drug (gefitinib, Gef) within Gd-MHAp NPs produced a synergistic therapeutic effect. In summary, this study demonstrated that such a novel drug system (Gd-MHAp/5-FU/Gef/CS/PAA NPs) could protect, transport, and program drug release locally within the colonic environment; further, this system exhibited a worthwhile therapeutic effect, providing a promising novel treatment strategy for orthotopic colon cancer.

Can nano-hydroxyapatite permeate the oral mucosa? A histological study using three-dimensional tissue models.

Nano-hydroxyapatite is used in oral care products worldwide. But there is little evidence yet whether nano-hydroxyapatite can enter systemic tissues via the oral epithelium. We investigated histologically the ability of two types of nano-hydroxyapatite, SKM-1 and Mi-HAP, to permeate oral epithelium both with and without a stratum corneum, using two types of three-dimensional reconstituted human oral epithelium, SkinEthic HGE and SkinEthic HOE respectively with and without a stratum corneum. Both types of nano-hydroxyapatite formed aggregates in solution, but both aggregates and primary particles were much larger for SKM-1 than for Mi-HAP. Samples of each tissue model were exposed to SKM-1 and Mi-HAP for 24 h at concentrations ranging from 1,000 to 50,000 ppm. After treatment, paraffin sections from the samples were stained with Dahl or Von Kossa stains. We also used OsteoSense 680EX, a fluorescent imaging agent, to test for the presence of HAP in paraffin tissue sections for the first time. Our results for both types of nano-hydroxyapatite showed that the nanoparticles did not penetrate the stratum corneum in SkinEthic HGE samples and penetrated only the outermost layer of cells in SkinEthic HOE samples without stratum corneum, and no permeation into the deeper layers of the epithelium in either tissue model was observed. In the non-cornified model, OsteoSense 680EX staining confirmed the presence of nano-hydroxyapatite particles in both the cytoplasm and extracellular matrix of outermost cells, but not in the deeper layers. Our results suggest that the stratum corneum may act as a barrier to penetration of nano-hydroxyapatite into the oral epithelium. Moreover, since oral epithelial cell turnover is around 5-7 days, superficial cells of the non-keratinized mucosa in which nanoparticles are taken up are likely to be deciduated within that time frame. Our findings suggest that nano-hydroxyapatite is unlikely to enter systemic tissues via intact oral epithelium.

Localized low-dose rhBMP-2 is effective at promoting bone regeneration in mandibular segmental defects.

At least 26% of recent battlefield injuries are to the craniomaxillofacial (CMF) region. Recombinant human bone morphogenetic protein 2 (rhBMP-2) is used to treat CMF open fractures, but several complications have been associated with its use. This study tested the efficacy and safety of a lower (30% recommended) dose of rhBMP-2 to treat mandibular fractures. rhBMP-2 delivered via a polyurethane (PUR) and hydroxyapatite/ß-tricalcium phosphate (Mastergraft®) scaffold was evaluated in a 2 cm segmental mandibular defect in minipigs. Bone regeneration was analyzed at 4, 8, and 12 weeks postsurgery using clinical computed tomography (CT) and rhBMP-2, and inflammatory marker concentrations were analyzed in serum and surgery-site drain effluent. CT scans revealed that pigs treated with PUR-Mastergraft® + rhBMP-2 had complete bone bridging, while the negative control group showed incomplete bone-bridging (n = 6). Volumetric analysis of regenerated bone showed that the PUR-Mastergraft® + rhBMP-2 treatment generated significantly more bone than control by 4 weeks, a trend that continued through 12 weeks. Variations in inflammatory analytes were detected in drain effluent samples and saliva but not in serum, suggesting a localized healing response. Importantly, the rhBMP-2 group did not exhibit an excessive increase in inflammatory analytes compared to control. Treatment with low-dose rhBMP-2 increases bone regeneration capacity in pigs with mandibular continuity defects and restores bone quality. Negative complications from rhBMP-2, such as excessive inflammatory analyte levels, were not observed. Together, these results suggest that treatment with low-dose rhBMP-2 is efficacious and may improve safety when treating CMF open fractures. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 1491-1503, 2019.

In vitro evaluation of a biomaterial-based anticancer drug delivery system as an alternative to conventional post-surgery bone cancer treatment.

Patients diagnosed with osteosarcoma are currently treated with intravenous injections of anticancer agents after tumor resection. However, due to remaining neoplastic cells at the site of tumor removal, cancer recurrence often occurs. Successful bone regeneration combined with the control of residual cancer cells presents a challenge for tissue engineering. Cyclodextrins loaded with chemotherapeutic drugs reversibly release the drugs over time. Hydroxyapatite bone biomaterials coated with doxorubicin-loaded cyclodextrin should release the drug with time after implantation directly at the original tumor site and may be a way to eliminate residual neoplastic cells. In the present study, we have carried out in vitro studies to evaluate such a drug-delivery system and have shown that doxorubicin released from cyclodextrin-coated hydroxyapatite retained biological activity and exhibited longer and higher cytotoxic effects on both cancer (osteosarcoma cells) and healthy cells (primary osteoblasts and endothelial cells) compared to biomaterials without cyclodextrin loaded with doxorubicin. Furthermore, doxorubicin released from biomaterials with cyclodextrin moderately induced the expression of tumor suppressor protein p53 whereas p21 expression was similar to control cells. In addition, hypoxic conditions, which occur after implantation until blood-flow to the area is regenerated, protected endothelial cells and primary osteoblasts from doxorubicin-induced cytotoxicity. This chemo-protective effect was far less prominent for the osteosarcoma cells. These findings indicate that a hydroxyapatite-cyclodextrin-doxorubicin chemotherapeutic strategy may enhance the drug-targeting effect on tumor cells while protecting the more sensitive healthy cells for a period of time after implantation. A successful integration of such a drug delivery system might allow healthy cells to initially survive during the doxorubicin exposure period, while eliminating residual neoplastic cells.

Mesoporous and hollow hydroxyapatite nanostructured particles as a drug delivery vehicle for the local release of ibuprofen.

The high risk of infection caused by implantation of orthopedic bio-metals is still a daunting challenge for surgeons as it can lead to implant failure. One approach to overcome this issue is the local release of antibacterial drug through coating on the surface of a metallic implant. One ideal carrier for this purpose is hydroxyapatite (HA) particles which are bioactive, biodegradable, biocompatible and have the potential to bond to bone. In the current study, highly crystalline mesoporous HA nanostructure particles were successfully synthesized in a low-temperature solvent process with the aid of an inorganic CaCO3 template and then fully characterized. The specific surface area and the average size of the cavities of the nanostructured mesoporous HA particles were 85 m2/g and 20 nm, respectively. The feasibility of the prepared HA mesoporous nanostructures for drug delivery, using ibuprofen as a model drug, was also investigated. The as-prepared HA mesoporous nanostructures showed a high drug-loading capacity, as well as sustained drug release in a phosphate buffered saline (PBS) at a pH of 7.4. Overall, results show that HA mesoporous nanostructures gave great potential in bone regeneration and local delivery of either drugs or biomolecules.

A combined low-frequency electromagnetic and fluidic stimulation for a controlled drug release from superparamagnetic calcium phosphate nanoparticles: potential application for cardiovascular diseases.

Alternative drug delivery approaches to treat cardiovascular diseases are currently under intense investigation. In this domain, the possibility to target the heart and tailor the amount of drug dose by using a combination of magnetic nanoparticles (NPs) and electromagnetic devices is a fascinating approach. Here, an electromagnetic device based on Helmholtz coils was generated for the application of low-frequency magnetic stimulations to manage drug release from biocompatible superparamagnetic Fe-hydroxyapatite NPs (FeHAs). Integrated with a fluidic circuit mimicking the flow of the cardiovascular environment, the device was efficient to trigger the release of a model drug (ibuprofen) from FeHAs as a function of the applied frequencies. Furthermore, the biological effects on the cardiac system of the identified electromagnetic exposure were assessed in vitro and in vivo by acute stimulation of isolated adult cardiomyocytes and in an animal model. The cardio-compatibility of FeHAs was also assessed in vitro and in an animal model. No alterations of cardiac electrophysiological properties were observed in both cases, providing the evidence that the combination of low-frequency magnetic stimulations and FeHAs might represent a promising strategy for controlled drug delivery to the failing heart.

Box-Behnken Design of Experiment Assisted Development and Optimization of Bendamustine HCl loaded Hydroxyapatite Nanoparticles.

BACKGROUND: Bendamustine HCl, an antineoplastic drug, has a very short life of about 40 minutes which necessitates administration of large doses which leads to increased side effects as well as costs. OBJECTIVE: The present work describes the fabrication, optimization, and evaluation of bioactive hydroxyapatite nanoparticles to achieve sustained delivery of bendamustine HCl. METHODS: Hydroxyapatite nanoparticles (NPs) were prepared by the wet chemical precipitation method by reacting a calcium and phosphate precursor and the reaction was optimized via Box-Behnken DOE. The drug was loaded on particles by physical adsorption. Various analytical studies were performed on the fabricated nanoparticles in addition to biodistribution studies to establish the physicochemical and biological characteristics of the designed formulation. RESULTS: pH of the reactant solution was found to have a more profound effect on the particle size and size distribution in comparison to reactant concentration. The particles were found to have a spherical morphology by SEM. Size of the blank and drug-loaded nanoparticles was found to be 130±20 nm by TEM. Energy Dispersive X-ray Spectroscopy (EDS) studies confirmed the presence of hydroxyapatite as the dominant phase while DSC studies indicated the presence of the drug in its amorphous form after its adsorption on NPs. Tissue distribution studies further suggested that the majority of drug concentration was released in blood rather than the other organs implying low organ toxicity. CONCLUSION: Bendamustine loaded hydroxyapatite nanoparticles were successfully optimized and fabricated. Favorable results were obtained in in vitro, in vivo, and analytical studies.

Behavior of new hydroxyapatite/glucan composite in human serum.

Biomaterials for bone tissue regeneration, including polymer-based composites, are typically evaluated in vitro prior to the clinical trials. However, such composites tested in vivo may behave different due to the specific body conditions. For example, some composites implanted into the tissue acidified due to transient postoperative inflammation may unexpectedly swell which delays the wound healing. Such massive swelling in acidic medium was previously observed for new elastic hydroxyapatite (HAp)/ß-glucan biomaterial. However, in further clinical cases concerning the composite implantation in patients without significant inflammation indicators, no side effects were observed. Therefore, it was reasonable to test the effect of human serum of neutral pH (typical for noninflamed tissues) on the composite parameters, in particular volume changes. Thus, this article shows the characterization of physicochemical parameters of the composite after incubation (5 days) in human serum of neutral pH by means of weight and volume measurement, scanning electron microscopy, X-ray diffraction, Fourier-transform infrared spectroscopy, microcomputed tomography, mercury intrusion, and biochemical techniques. Results showed that human serum collected from healthy people caused no uncontrolled changes in weight and volume, porosity and mechanical properties of the composite. Therefore, this suggests the lack of volume change-related side effects of HAp/glucan composite in bone defects treatment if postoperative inflammation is prevented. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 2653-2664, 2018.

Hydroxyapatite decreases cytotoxicity of a glass ionomer cement by calcium fluoride uptake in vitro.

BACKGROUND: Glass ionomer cements (GICs) are widely used in dentistry because of their remineralizing and cariostatic potential induced by fluoride. In vitro studies have reported cell toxicity triggered by GICs; however, the influence of hydroxyapatite (HAp) must be considered. The aim of this study was to evaluate the effect of HAp in decreasing the cytotoxicity of the GIC 3M Vitrebond in vitro. METHODS: Samples of 3M Vitrebond (powder, liquid and light-cured) were incubated in Dulbecco's modified Eagle's medium-Ham's F12 (DMEM-F12) for 24 hours at 37°C. Subsequently, the light-cured medium was treated with 100 mg/mL of HAp overnight. Toxicity of conditioned media diluted 1:2, 1:4, 1:8 and 1:20 was analyzed on human gingival fibroblasts (HGFs) using light microscopy and the fluorometric microculture cytotoxicity assay. The amounts of calcium fluoride (CaF2) were determined by the alizarin red S method. RESULTS: The exposure of HGFs to light-cured induced cell death and morphological changes such as chromatin condensation, pyknotic nuclei and cytoplasmic modifications. Exposure to light-cured treated with HAp, significantly increased cell viability leading to mostly spindle-shaped cells (p<0.001). The concentration of CaF2 released by the light-cured was 200 ppm, although, in the light-cured/HAp conditioned medium, this quantity decreased to 88 ppm (p<0.01). CONCLUSIONS: These data suggest that HAp plays a protective role, decreasing the cytotoxic effect of 3M Vitrebond induced by CaF2.

Cellular Uptake and Delivery-Dependent Effects of Tb3+-Doped Hydroxyapatite Nanorods.

With the increasing interest in hydroxyapatite (HA) nanostructures for use in biomedicine, the systematic evaluation of their potential effects on biological systems is becoming critically important. In this work, we report the in vitro cellular uptake, in vivo tissue distributions and toxicity of Tb3+-doped HA (HA-Tb) after short-, intermediate-, and long-term exposure. Transmission electron microscopy analysis indicated that HA-Tb was taken up by cells via vesicle endocytosis. Cell proliferation and cytotoxicity assay, combined with confocal laser scanning microscopy, indicated excellent cell viability with no changes in cell morphology at the examined doses. Three HA-Tb delivery methods (intraperitoneal, intragastric, and intravenous) resulted in similar time-dependent tissue distributions, while intraperitoneal injection produced the highest bioavailability. HA-Tb initially accumulated in livers and intestines of rats (4 h to one day after administration), then became increasingly distributed in the kidney and bladder (seven days), and finally decreased in all tissues after 30 to 90 days. No histopathological abnormalities or lesions related to treatment with HA-Tb were observed. These results suggest that HA-Tb has minimal in vitro and in vivo toxicity, regardless of the delivery mode, time, and dose. The findings provide a foundation for the design and development of HA for biological applications.

Sustained release of simvastatin from hollow carbonated hydroxyapatite microspheres prepared by aspartic acid and sodium dodecyl sulfate.

Hollow carbonated hydroxyapatite (HCHAp) microspheres as simvastatin (SV) sustained-release vehicles were fabricated through a novel and simple one-step biomimetic strategy. Firstly, hollow CaCO3 microspheres were precipitated through the reaction of CaCl2 with Na2CO3 in the presence of aspartic acid and sodium dodecyl sulfate. Then, the as-prepared hollow CaCO3 microspheres were transformed into HCHAp microspheres with a controlled anion-exchange method. The HCHAp microspheres were 3-5µm with a shell thickness of 0.5-1µm and were constructed of short needle nanoparticles. The HCHAp microspheres were then loaded with SV, exhibiting excellent drug-loading capacity and sustained release properties. These results present a new material synthesis strategy for HCHAp microspheres and suggest that the as-prepared HCHAp microspheres are promising for applications in drug delivery.

Toxic effect of cadmium adsorbed by different sizes of nano-hydroxyapatite on the growth of rice seedlings.

Information regarding the toxic effects of cadmium (Cd) adsorbed by nano-hydroxyapatite (NHAP-Cd) on the growth of crop plants remain limited. We investigated the mechanism of NHAP-Cd (diameters, 20 and 40nm; NHAP20-Cd and NHAP40-Cd, respectively) phytotoxicity. Rice seedlings treated with Cd and NHAP20-Cd showed more severe growth retardation compared to those treated with NHAP40-Cd, for the same Cd concentration. Transmission electron microscopy revealed NHAP in the seedlings. The nanoparticles entered the rice seedlings with no Cd2+ signals in the NHAP treatments compared to -0.47pmolcm-2s-1 of Cd2+ fluxes in the Cd treatment. The higher Cd2+ content in the leaves and mesocotyl of NHAP20-Cd-treated rice seedlings suggested that smaller NHAP-Cd can translocate easily to the aboveground parts. Further, NHAP-Cd increased oxidative stress, which was determined as catalase activity changes in this study. Thus, NHAP-Cd particles in the growth medium can be transported to rice seedlings and cause toxicity.

Difference in release kinetics of unwashed and washed platelet-released supernatants from bone substitute materials: the impact of platelet preparation modalities.

BACKGROUND AND OBJECTIVE: In regenerative dentistry, platelet preparations are applied to stimulate bone healing and periodontal regeneration. Here, we pursue a strategy where bone substitutes are used as carriers for platelet-released supernatants. The mitogenic capacity and release kinetics of loaded bone substitutes were assessed. MATERIAL AND METHODS: Platelet-released supernatants of washed platelets (washed PRS) and platelet-released supernatants of unwashed platelets (unwashed PRS) were lyophilized onto the bone substitutes deproteinized bovine bone mineral, hydroxyapatite and ß-tricalcium phosphate. Scanning electron microscopy images were taken. Supernatants of bone substitutes were collected at hours 1, 3, 6, 24, and 48 and medium was replaced. We evaluated the protein content with the bicinchoninic acid assay and the effect on proliferation using bioassays with human periodontal fibroblasts. Release of growth factors from the loaded bone substitutes was measured based on the platelet-derived growth factor isoform (PDGF-BB) and thrombin immunoassays. Furthermore, we assessed DNA and RNA content of washed PRS and unwashed PRS. RESULTS: Unwashed PRS showed higher total protein concentrations than washed PRS, while the concentration of PDGF-BB, thrombin, DNA, RNA and their mitogenic effect was not significantly different. The bone substitute materials adsorbed protein over time but no significant changes in overall appearance was found. Supernatants collected from unwashed PRS-loaded bone substitute after 1 h induced a potent mitogenic response in periodontal fibroblasts. This pro-mitogenic capacity of the supernatants decreased over the observation period. Supernatants of washed PRS-loaded bone substitutes did not induce a substantial mitogenic effect. Levels of PDGF-BB, thrombin and protein were higher in supernatants of unwashed PRS-loaded bone substitutes than of washed PRS-loaded bone substitutes. CONCLUSION: Bone substitutes loaded with unwashed PRS, but not bone substitutes loaded with washed PRS show continuously declining release kinetics. These data suggest that plasma components in platelet preparations can modify the release kinetics profile.

Enhanced osteogenesis and angiogenesis by mesoporous hydroxyapatite microspheres-derived simvastatin sustained release system for superior bone regeneration.

Biomaterials with both excellent osteogenic and angiogenic activities are desirable to repair massive bone defects. In this study, simvastatin with both osteogenic and angiogenic activities was incorporated into the mesoporous hydroxyapatite microspheres (MHMs) synthesized through a microwave-assisted hydrothermal method using fructose 1,6-bisphosphate trisodium salt (FBP) as an organic phosphorous source. The effects of the simvastatin-loaded MHMs (S-MHMs) on the osteogenic differentiation of rat bone marrow mesenchymal stem cells (rBMSCs) and angiogenesis in EA.hy926 cells were investigated. The results showed that the S-MHMs not only enhanced the expression of osteogenic markers in rBMSCs but also promoted the migration and tube formation of EA.hy926 cells. Furthermore, the S-MHMs were incorporated into collagen matrix to construct a novel S-MHMs/collagen composite scaffold. With the aid of MHMs, the water-insoluble simvastatin was homogenously incorporated into the hydrophilic collagen matrix and presented a sustained release profile. In vivo experiments showed that the S-MHMs/collagen scaffolds enhanced the bone regeneration and neovascularization simultaneously. These results demonstrated that the water-insoluble simvastatin could be incorporated into the MHMs and maintained its biological activities, more importantly, the S-MHMs/collagen scaffolds fabricated in this study are of immense potential in bone defect repair by enhancing osteogenesis and angiogenesis simultaneously.