Cytotoxinic Mechanism of Hydroxyapatite Nanoparticles on Human Hepatoma Cell Lines

Stable and single dispersed HAP nanoparticles were synthesized with chemical method assisted by ultrasonic treatment.HAP nanoparticles were surveyed by AFM and Zataplus.The effect on the Bel-7402 human hepatoma cell lines treated with HAP nanoparticles was investigated by the MTT methods and observation of morphology,and the mechanism was studied in changes of cell cycle and ultrastructure.The result shows that inhibition of HAP nanoparticles on the Bel 7402 human hepatoma cell lines is obviously in vitro.HAP nanoparticles the entered cancer cytoplasm,and cell proliferation is stopped at G 1 phase of cell cycle,thus,cancer cells die directly.

Preparation, Characterization and Antibacterial Property of Cerium Substituted Hydroxyapatite Nanoparticles

Nanoparticles of hydroxyapatite （HAP） and cerium substituted hydroxyapatite （CeHAP） with the atomic ratio of Ce/[ Ca ＋ Ce] （xco） from 0 to 0.2 were prepared by sol-gel-supercritical fluid drying （SCFD） method. The nanoparticles were characterized by TEM, XRD, and FT-IR, and the effects of cerium on crystal structure, crystallinity, and particle shape were discussed. With the tests of bacterial inhibition zone and antibacterial ratio, the antibacterial property of HAP and CeHAP nanoparticles on Escherichia coli, Staphylococcus aureus, Lactobacillus were researched. Results showed that the nanoparticles of HAP and CeHAP could be made by sol-gel-SCFD, cerium could partially substitute for calcium and enter the structure of HAP. After substitution, the crystallinity, the IR wavenumbers of bonds in CeHAP decreased gradually with increase of cerium substitution, and the morphology of the nanoparticles changed from the short rod-shaped HAP to the needle-shaped CeHAP. The nanoparticles of HAP and CeHAP with Xco below 0.08 had antibacterial property only forcibly contacting with the test bacteria at the test concentration of 0.1 g · mi^-1, however, the Ce- HAP nanoparticles had antibacterial ability at that concentration no matter statically or dynamically contacting with the test bacteria when Xco was above 0.08, and the antibacterial ability gets better with the increase Of Xce, indicating that the antibacterial property was improved after calcium was partially substituted by cerium. The improved antibacterial effects of CeHAP nanoparticle on Lactobacillus showed its potential ability to anticaries.

Synthesis,Characterization and Antibacterial Property of Strontium Half and Totally Substituted Hydroxyapatite Nanoparticles

Nanoparticles of hydroxyapatite（HAP）, strontium half substituted hydroxyapatite （SrCaHAP） and strontium totally substituted hydroxyapatite （SrHAP） were prepared by sol-gel-supercritical fluid drying （SCFD） method. The nanoparticles were characterized by element content analysis, FT-IR, XRD and TEM, and the effects of strontium substitution on crystal structure, crystallinity, particle shape and antibacterial properties of the nanoparticles on Escherichia coli, Staphylococcus aureus, Lactobacillus were researched. Results show that strontium can half and totally substitute for calcium and enter the structure of apatite according to the initial atomic ratios of Sr/[Sr＋Ca] as 0.5, 1. The substitution decreases the IR wavenumbers of SrCaHAP and SrHAP, and changes the morphology of the nanoparticles from short rod shaped HAP to needle shaped SrCaHAP, and back to short rod shaped SrHAP. The crystallinity of HAP is higher than that of SrCaHAP, but is lower than that of SrHAP. Moreover, the antibacterial property of SrCaHAP and SrHAP are improved after the calcium is half and totally substituted by strontium.

Arginine functionalized hydroxyapatite nanoparticles and its bioactivity for gene delivery

In order to further improve the transfection efficiency of hydroxyapatite nanoparticle （HAp）, arginine functionalized hydroxyapatite （HAp/Arg） was synthesized by hydrothermal synthesis. The morphology, crystallite size and zeta potential of the HAp/Arg were characterized by transmission electron microscopy （TEM）, atomic force microscopy （AFM） and zeta potential analyzer. The loading and protecting properties of HAp/Arg to DNA were tested by electrophoresis. Its cytotoxicity was also measured in Hela cells and HAEC cells by MTT and LDH, and its transfection efficiency was examined by fluorescence microscope and flow cytometry. The results reveal that HAp/Arg is short rod-like and nano single crystal, the mean diameter is 50-90 nm and zeta potential is 35.8 mV at pH 7.4. HAp/Arg to DNA can be condensed by electrostatic effect and protect DNA against degradation in DNase I, and shows high transfection efficiency without cytotoxicity. These results suggest that HAp/Arg can be a promising alternative as a novel gene delivery system.

Preparation and application of hydroxyapatite(HA) nanoparticles/NR2B-siRNA complex

Hydroxyapatite(HA) nanoparticles were prepared by coprecipitation-hydrothermal synthesis and their exosyndrome was estimated via transmission electron microscopy. Agarose gel electrophoresis and ultraviolet spectrophotometry were used to evaluate the ability of HA to bind NR2B-siRNA at different pH values and at different HA-NR2B-siRNA ratios. And the stability of the complex in saline was also evaluated. The effect of HA/NR2B-siRNA complex on chronic inflammatory pain was evaluated in vivo in mice. Transmission electron microscopy(TEM) reveals that HA nanoparticles are thin strips or short rod in shape and the one-dimensional particle size of HA nanoparticles is 40-50 nm. Under the acid or neutral condition, the Zeta potential of HA is positive; nanoparticles can completely bind NR2B-siRNA when the HA:NR2B-siRNA ratio is at or larger than 35-1; while under the alkaline condition, the affinity of HA to NR2B-siRNA is rather weak. HA/NR2B-siRNA complex is not dissociated when being resuspended in saline. The nociception of the tonic phase induced by formalin is significantly reduced in the HA/NR2B-siRNA treated mice as compared with the controls. Therefore, HA may be a new siRNA nano-vector material.

Physical and Chemical Transformation of Hydroxyapatite Nanoparticles in Aqueous Sol after Preparation and in vitro

The co-precipitation method followed by ultrasound and heat treatment is a common way to prepare below 100 nm sized hydroxyapathe nanoparticles for biomedical studies and applications. The size and pH value of the obtained calcium phosphate nanoparticles in aqueous sol have a strong impact on the interactions with cells and tissue. The physical and chemical properties of material samples for in vitro and in vivo studies are often assumed to remain constant from the time after fabrication to the actual use. Only little attention is paid to eventual changes of the material over time or due to the different in vitro conditions. In this study, the physical and chemical transformation of calcium phosphate nanoparticles after preparation and in vitro was investigated. As the result showed, dispersed nano sized amorphons calcium phosphate precipitation as well as crystallized hydroxyapathe nanoparticles continue to crystallize even when kept at 4℃ leading to declining pH values and particle sizes. Due to the pH buffer in the medium the pH value of the cell culture remained stable after adding 20% nanoparticle sol in vitro. However, hydroxyapathe nanoparticles immediately became unstable in the presents of cell culture medium. The resulting loose agglomerations showed a size of above 500 nm.

Effects of Hydroxyapatite Nanoparticles on Apoptosis and Invasion of Human Renal Cell Carcinoma 786-0 Cells

Renal cell carcinoma is the most common cancer of the kidney, and resistant to traditional therapies. The aim of this study is to investigate the effects of hydroxyapatite nanoparticles on human renal cell carcinoma 786-0 cells. Cell proliferation was assessed with an 3-（4,5-dimethylthiazol-2-yl） 2,5-diphenyltetrazolium bromide（MTT） staining kit. The apoptosis assay was assessed with an FITC Annexin V Apoptosis Detection Kit. Caspase-3 and caspase-12 were detected by immunocytochemical staining and semi-quantitative RT-PCR. Cell wound healing assay was used to ensure cell motility. Matrigel invasion assay was analysed via transwell chambers. Our results showed that hydroxyapatite nanoparticles significantly reduced cell proliferation, invasion and induced apoptosis of 786-0 cells. The inhibiting action may have relation with up-regulated caspase-12, leading the cells to apoptosis. This study suggests that hydroxyapatite nanoparticles may be an effective and delivery system for renal cell carcinoma therapy.

Effect of hydroxyapatite nanoparticles on the growth and p53/c-Myc protein expression of implanted hepatic VX_2 tumor in rabbits by intravenous injection

AIM： To evaluate the effect of hydroxyapatite nano- particles （Nano HAP） by intravenous injection on the inhibition of implanted hepatic VX2 tumor growth in rabbits and cell p53/c-Myc protein expression. METHODS： 60 hepatic VX2 tumor-bearing rabbits was randomly divided into five groups. Nano HAP collosol 20 mglkg, 40 mg/kg, 5-FU solutions 20 mg/mL, mixed liquor of 5-FU solution 20 mg/mL and Nano HAP collosol 20 mg/kg were infused by vein, normal saline conducted as the control. The general state, weight, liver function and gross tumor volume were detected dynamically. The expression of p53 and c-Myc gene protein in tumor tissue was detected by immunohistochemistry methods. RESULTS： The growth of implanted hepatic VX2 tumors was significantly inhibited in all therapy groups, 3 wk after the injection, the tumor control rates in Nano HAP collosol groups were 25.5% and 32.5% respectively, and the gross tumor volumes were obviously less than that of control group. （24.81 ± 5.17 and 22.73 ± 4.23 vs 33.32 ± 5.26, P 〈 0.05）. The tumor control rate of 5-FU group was 43.7% （18.74 4± 4.40 vs 33.32 ± 5.26, P 〈 0.05）, but the general state of the animals after injection aggravated; and the adverse reaction in the drug combination group obviously decreased. Due to the effect of Nano HAP, the positive expression of tumor associated the mutated p53 and c-Myc in tumor tissue was decreased obviously compared with the control group. CONCLUSION： Nano HAP has evident inhibitory action on rabbit implanted hepatic VX2 tumor in vivo, which may be the result of decreasing the expression of the mutated p53 and c-myc, and drug combination can obviously decrease the adverse reaction of 5-FU.

Effect of Tb/Mg doping on composition and physical properties of hydroxyapatite nanoparticles for gene vector application

Transfection efficiency of hydroxyapatite nanoparticles(HAnps)is relative to the particle size,morphology,surface charge,surface modifier and so on.This study prepared HAnps with doped Tb/Mg by hydrothermal synthesis method(HTSM)and investigated the effects of different Tb/Mg contents on the morphology,particle size,surface charge,composition and cellular endocytosis of HAnps.The results showed that Mg-HAnps possessed better dispersion ability than Tb-HAnps.With increasing doping content of Tb/Mg-HAnps,the granularity of Tb-HAnps increased,while that of Mg-HAnps declined.Both particle size and zeta potential of Mg-HAnps were lower than those of Tb-HAnps.7.5%Mg-doping HAnps presented relatively uniform slender rod morphology with average size of30nm,while10%Mg-doping HAnps were prone to agglomeration.Moreover,Mg-HAnps-GFP(green fluorescent protein)endocytosed by MG63cells was dotted in the perinuclear region,while Tb-HAnps were more likely to aggregate.In conclusion,as gene vectors,Mg-HAnps showed enhanced properties compared to Tb-HAnps.

Effects of the Interaction between Hydroxyapatite Nanoparticles and Hepatoma Cells

To gain a better understanding of the anticancer effects of hydroxyapatite （HAP） nanoparticles in vivo and in vitro, the effects of the interaction of HAP nanoparticles with hepatoma cells were explored. HAP nanoparticles were prepared by homogeneous precipitation and characterized by laser particle analysis and transmission electron microscopy （TEM）. HAP nanoparticles were observed to be uniformly distributed, with rod-like shapes and diameters in the range of 42.1-87.1 nm. Overnight attached, suspended, and proliferating Bel-7402 cells were incubated with HAP nanoparticles. Inverted microscopy observation revealed that HAP nanoparticles with a cell membrane showed good adsorption. TEM demonstrated that HAP nanoparticles were present on the surface of cells, continuously taken up by cells through endocytosis, and transported in vesicles close to the nucleus. Fluorescence microscopy showed that the concentrations of intracellular Ca2＋ labeled with Fluo-3 calcium fluorescent probe were significantly enhanced. In addition, inverted microscopy observation revealed that suspended cells treated with HAP nanoparticles did not adhere to the culture bottle, resulting in cell death. After the overnight attached cells were treated with HAP nanoparticles for 96 h with increasing doses of HAP nanoparticles, inverted microscopy observation revealed that cell proliferation was slowed and ceU-ceU adhesion was weakened. Feulgen staining and image analysis indicated that the nuclear DNA content of the cells was markedly reduced, and argyrophilic nucleolar organizer region （AgNOR） staining and image analysis indicated that the number of AgNORs was significantly decreased. Therefore, hepatoma cells brought about the adsorption, uptake, transport and degradation of HAP nanoparticles. In addition, HAP nanoparticles affected hepatoma cells with regard to cell-cell adhesion, cell and extracellular matrix adhesion, and DNA and protein synthesis; thus inhibiting cell proliferation. This understanding of the effects of interaction between HAP nanoparticles and hepatoma cells is useful for further study of the anticancer mechanisms of HAP nanoparticles.

Morphologies of hydroxyapatite nanoparticles adjusted by organic additives in hydrothermal synthesis

Properties of hydroxyapatite （HA, Ca10（PO4）6（OH）2）, including bioactivity, biocompatibility, solubility and adsorption could be tailored over wide ranges by the control of particle composition, particle size and morphology. In order to satisfy various applications, well-crystallized pure HA nanoparticles were synthesized at moderate temperatures by hydrotherrnal synthesis, and HA nanoparticles with different lengths were obtained by adding organic additives. X-ray diffractometry （XRD） and Fourier transform infrared （FTIR） spectrometry were used to characterize these nanoparticles, and the morphologies of the HA particles were observed by transmission electron microscopy （TEM）. The results demonstrate that shorter rod-like HA particles can be prepared by adding cetyltrimethylammonium bromide （CTAB）, as the additive of CTAB can block the HA crystal growth along with c-axis. And whisker HA particles are obtained by adding ethylenediamine tetraacetic acid （EDTA）, since EDTA may have effect on the dissolution-repreeipitation process of HA.

Room temperature gas sensor based on tube-like hydroxyapatite modified with gold nanoparticles

The main goal of this work is to explore the possibility of using Au-modified hydroxyapatite(HA) as a potential sensor material. Tube-like HA structure was fabricated with the aid of a Nafion N-117 cation exchange membrane and gold(Au) nanoparticles were added by a hydrothermal method. The morphology, structure and composition were characterized by scanning electron microscopy(SEM), transmission electron microscopy(TEM), X-ray diffraction(XRD), and X-ray photoelectron spectroscopy(XPS). The gas sensing properties were also investigated. Results show that Au nanoparticles are dispersed into the HA powder, which is tube-like, with rough inner and outer surfaces. Compared with pure HA, Au-modified HA exhibits improved sensing properties for NH_3. 5%(mass fraction) Au-modified HA shows the highest response with relatively short response/recovery time. The response is up to 79.2% when the corresponding sensor is exposed to 200×10^(-6) NH_3 at room temperature, and the response time and recovery time are 20 s and 25 s, respectively. For lower concentration, like 50×10^(-6), the response is still up to 70.8%. Good selectivity and repeatability are also observed. The sensing mechanism of high response and selectivity for NH_3 gas was also discussed. These results suggest that Au-HA composite is a promising material for NH_3 sensors operating at room temperature.

Poly(Vinyl Alcohol)/Collagen/Hydroxyapatite Nanoparticles Hybrid System Containing Yttrium-90 as a Potential Agent to Treat Osteosarcoma

Osteosarcoma is the most common primary malignant bone tumors, affecting mostly children, adolescents and young adults. This is an aggressive tumor that results in a high mortality rate and poor prognosis. Due to the low sensitivity of osteosarcoma to ionizing radiation, such treatment is not used very often and it can be recommended only to postsurgical therapy. As an alternative therapy, functionalized nanomaterials allow their accumulation in tumor tissues due to their unique properties, making them good agents to act as stable carriers for radionuclides. In this work, mesoporous hydroxyapatite nanoparticles were synthesized and the functionalization process with poly(vinyl alcohol) and collagen was investigated. The samples were characterized by X-ray diffraction (XRD), N2 adsorption, elemental analysis (CHN), Fourier Transform Infrared Spectroscopy (FTIR), Nitrogen Adsorption, Transmission Electron Microscopy (TEM), and Thermal Analysis. Also, the yttrium incorporation potential and its release kinetics in the hydroxyapatite matrix were evaluated to study the capacity of this system to treat osteosarcomas. The results indicate that this material has a promisor potential to treat this kind of tumor.

Apoptosis of Cancer Cells Induced by HAP Nanoparticles

To confirm apoptosis is one of the hepatoma cells death pathways after HAP nanoparticles absorption,hepatoma cells were collected for ultrathin sections preparation and examined under a transmission electron microscope（TEM）after 1 h incubation with HAP nanoparticle.Apoptosis was detected by TUNEL technique.After absorption.some vacuoles with membrane containing HAP nanoparticles were found in cytoplasma.The nuclear enrelope shrinked.and some area pullulated from nucleus.The karyotin became pycnosis and assembled at the edge.An apoptosis body was found.and the data of IOD and numbers of the positive apoptosic signals in nuclear area of slides could illustrate much more apoptosis in the HAP group than those in the control group（P〈0.001）.The experimental results indicate that the HAP nanoparticles can induce cancer cells apoptosis.

Hot pressing of hydroxyapatite(HA)-Ti material and stability of HA

Hot pressing of hydroxyapatite(HA) Ti system material and the stability of HA component were investigated to supply the foundation of optimizing sintering procedure of HA Ti functionally graded material(FGM). The results show that the HA powders used have excellent thermal stability and no decomposition is observed at 1 300 ℃. The existence of Ti can promote the dehydration and decomposition of HA. However, no new compounds form between HA and Ti. By selecting sintering parameters properly, ideal HA Ti material can be acquired. The relative densities of the mixtures of HA and Ti are always lower than those of pure HA or Ti, which may be caused by the decomposition of HA in the mixtures.

Effect of Hydroxyapatitc Nanoparticles on K562 Cells in vitro

Stable and single-dispersed hydroxyapatite （HAP） nanoparticles were synthesized with ultrasonic-assisted method. HAP nanoparticles were characterized by dynamic light scattering, XRD （X-ray diffraction） and TEM （Transmission Electron Microscopy）. The effect of HAP nanoparticles on the K562 human myelogenous leukemia cell line was investigated by MTT assay and cell count test, and the mechanism was studied through the changes of cell cycle and ultrastructure. The results showed that HAP nanoparticles inhibited the proliferation of K562 cells dramatically in vitro. HAP nanoparticles entered the cytoplasm of K562 cells and the cells were arrested at G/M phase, thus, the cells died directly.

Preparation and Characterization of Nanosized Hydroxyapatite Particles in AOT Inverse Microemulsion

Nanosized particles of hydroxyapatite (HAP) were synthesized by reacting Ca(H_2PO_4)_2·H_2O solution complex with equimolar Ca(OH)_2 saturated solution in sodium bis(2-ethylhexyl) sulfosuccinate (AOT)/isooctane water-in-oil microemulsion.The formation of microemulsion strongly depended on water content w (w=[H_2O]/[AOT] molar ratio) and concentration of surfactant and cosurfactant (1-octanol).By the variety of conductivity with w and the partial ternary phase diagram derived from a series of demarcation points,we set the basic component of microemulsions:[AOT]=0.1M(mol/dm^3),[1-octanol]=0.1M and w=3-9.Dynamic light scattering (DLS),UV-visible absorbance,TEM analysis and X-ray diffraction were used to characterize the microemulsion,formation of particles and resulting HAP particles.At low water content(w<9),the water pool radius of the droplet in the Ca(H_2PO_4)_2·H_2O microemulsion lineally depended on w.The size of final HAP particles was strongly affected by water content w and reactant concentration.With increasing water content w from 3 to 9,the size of HAP particles increased from 10-20nm to 40-50 nm at reactant concentration [Ca(H_2PO_4)2·H_2O]=12×10^(-3) mol/dm^3.The resulting HAP particles were poorly crystallized and spherical in morphology.

Size Depending Separation of HAP:Eu Nanoparticles in Dispersed Sol

The panicle size has a strong impact on the interactions between nanoparticles and cells. However, the synthesis process of nanoparticles limits the range of achievable average panicle sizes. When biocompatible hydroxyapatite nanoparticles （HAP） are doped with the luminescent rare earth elemeat Europium （Eu）, the panicle size becomes larger compared to pure HAP. Hence, a panicle size reduction is necessary to achieve similar experimental conditions when stbstituting pure HAP with luminescent HAP ： Eu nanoparticles to investigate particlecell-interactions in cell culture experiments. While the sedimentation process of particles in liquids and gels has been well described in literature, the separation of particles in dispersed colloids has not been studied, yet. In this study, the size depending separation and particle size redaction of a homogeneous dispersed nanoparticle sol by gravity and centrifugation were investigated. As the results showed, shorter time of centrifugation at higher speed can reduce the average particle size compared to the decline of the panicle concentation in the upper sol layer most efficiently. This ceatrifugation method has some similarity to the overspeeding technique which is commonly used to lower the transient time to reach the equilibrium of sedimentation.

Comparative Study on Lattice Parameters of HAP Nanoparticles with Those of HAP Whiskers

Hydroxyapatite whiskers are used as reinforcement for biomaterials because of their needlelike morphology and strong strength of single crystal. HAP nanoparitcles are used in drug delivery system, protein separation and anticancer drug besides their implant applications. Many new properties arise from nanoeffects while nanoeffects come from specialty of nanostructure. The Ca（NO3）2·4H2O and （NH4）2HPO4 were used as starting reagents for preparation of HAP whiskers and nanoparticles. The whiskers are 100-200 μm long while nanoparticles are 30-80 nm in size. XRD is applied to characterize the lattice parameter difference of whisker and nanoparticle. Compared with those of HAP whiskers, the a-axis and b-axis of HAP nanocrystals are shortened while the c-axis is elongated. That makes nanoparticles capable of their non-needle-like morphology, higher reaction activity and special bioeffects.