Mouse Strain– and Charge-Dependent Vessel Permeability of Nanoparticles at the Lower Size Limit

Remarkable advancement has been made in the application of nanoparticles (NPs) for cancer therapy. Although NPs have been favorably delivered into tumors by taking advantage of the enhanced permeation and retention (EPR) effect, several physiological barriers present within tumors tend to restrict the diffusion of NPs. To overcome this, one of the strategies is to design NPs that can reach lower size limits to improve tumor penetration without being rapidly cleared out by the body. Several attempts have been made to achieve this, such as selecting appropriate nanocarriers and modifying surface properties. While many studies focus on the optimal design of NPs, the influence of mouse strains on the effectiveness of NPs remains unknown. Therefore, this study aimed to assess whether the vascular permeability of NPs near the lower size limit differs among mouse strains. We found that the vessel permeability of dextran NPs was size-dependent and dextran NPs with a size below 15 nm exhibited leakage from postcapillary venules in all strains. Most importantly, the leakage rate of 8-nm fluorescein isothiocyanate dextran was significantly higher in the BALB/c mouse strain than in other strains. This strain dependence was not observed in slightly positive TRITC-dextran with comparable sizes. Our results indicate that the influence on mouse strains needs to be taken into account for the evaluation of NPs near the lower size limit

Synthesis and characterization of nanocrystalline Zn, Mg substituted hydroxyapatite

Hydroxyapatite (HA) is the most important bioceramic material for hard tissue replacement in human bodies. Recently, cation substituted HA has been a research focus in order to further enhance HA bioactivity and to adapt various application requirements. Magnesium and zinc are potentially the elements to partially substitute calcium in the HA for improving osteointegration, because Mg and Zn are the most important minor and trace elements respectively in human hard tissues. Magnesium depletion affects all stages of skeletal metabolism adversely, causing cessation of bone growth, decreasing osteoblastic and osteoclastic activity, osteopenia, and bone fragility. Zn can promote bone metabolism and growth, increase bone density and prevent bone loss. Mg, Zn substitutions in the crystal structure of HA are expected to have excellent biocompatibility and biological properties. Previous researches showed no direct evidence to prove that Zn/Mg successfully substituted partial Ca in the apatite lattice but not just absorbed on the apatite surface. The apatite structural changes after substitution even the maximum amount of Mg and Zn substitution for Ca are still in controversy. Nanocrystalline Mg, Zn substituted HA particles with various Mg and Zn substitution amount were synthesized by wet chemical method. Diffraction, spectroscopy, microscopy characterization techniques were employed to investigate the substituted apatites. Especially the X-ray diffraction Rietveld Refinement method was used to examine the effects of Mg and Zn substitution on crystal structure changes. Experimental results showed that Mg, Zn successfully substituted Ca in HA. The substitution limit was between 15~20 at. % Zn and different new phases presented over this limit. While Mg substitution was rather limited (less than 5 at.%)，various non apatite phases came out when the Mg/(Mg+Ca) atomic ratio was over 15 at. % in aqueous solution. Both Zn and Mg increased the resistance of HA crystallization. After substitution, the crystallite size and crystallinity decreased. Pure HA was well crystallized and in regular shape. With the substitution content increasing, crystallites were smaller, shorter, more irregular and formed serious agglomerates. Also both Mg and Zn substitution destabilized the apatite structure and reduced the decomposition temperature of HA. The lattice parameters a and c decreased up to 10at. % in solution and then started to slightly increase

Experimental and computational studies on carbonated apatite

The inorganic component of mineralized tissues (bone and teeth) is an analogue of carbonated apatite (CAp), it is therefore important to elucidate the crystal structure of CAp for the fundamental comprehension of bone regeneration in bodies and further for developing new artificial bones with desirable bioactivity and biocompatibility. However, the details of the crystal-chemical relationship between carbonate ions and apatite structure are not well known. Lack of single crystals of CAp with large enough size for direct structure analyses has kept the problem unresolved. This doctoral study aims to: 1) develop synthesis of biomimetic CAp nanocrystals; 2) Systematically study the carbonate effect on the structure, chemical composition, crystal size, morphology, and crystallinity of the apatite; 3) investigate the exact location and configuration of carbonate substitution in the apatite structure by ab initio simulation; 4) verify and refine the criterion to distinguish two types of CAp by infrared (IR) spectroscopy. CAp nanocrystals were successfully synthesized by controlled precipitation from solutions containing calcium, phosphate, and carbonate ions at pH = 11. In view of experimental evidence from XRD studies, Ca/P molar ratio changes and IR spectra difference, carbonate indeed substituted for partial phosphate in the precipitated CAp. Hydrothermal method was also developed to prepare CAp nanocrystals. The optimum parameters for synthesis of well crystalline CAp nanocrystals are pH = 10, C/P = 2/3, 200°C for 24 h. The structure of CAp has been investigated by ab initio simulations. The results show that type-A CAp are energetically preferred to type-B CAp. The most energetically favored configuration of type-A CAp had its carbonate triangular plane almost parallel to c-axis, making an angle of about 2° at z = 0.46. In the lowest energy structure of type-B CAp, the nearest Ca(2) ion was replaced by a sodium ion and the carbonate group was lying almost flat in b/c-plane of the apatite lattice, the normal to carbonate plane making an angle 88° to c-axis. Of all the models considered, mixed substitution type-AB where two carbonate ions replacing one phosphate group and one hydroxyl group was the most stable structure. The carbonate ions in the apatite lattice tend to be parallel to c-axis in both type-A and type-B sites. The lattice parameters a and b in type-A CAp unit cell expanded but the c parameter contracted. However, the most stable type-B CAp structure showed the opposite trend. The lattice parameter changes due to different substitution types may also be used as evidence to distinguish CAp. The bands at ~880 cm-1, ~1413 cm-1, and ~1450 cm- should not be used to identify CAp individually since they may result from carbonate absorption on apatite crystals. The IR signatures of carbonate environment in apatites are: v3 band at ~1465 cm-1 due to CO32- substituting for PO43- ; and v3 band at ~1546 cm-1 due to CO32- substituting for OH-. Based on these IR signatures, we can identify that the precipitated CAp is type-B substitution, the CAp prepared by hydroxyapatite reaction with CO2 at high temperature and hydrothermal method, and biological apatites are mixed type-AB substitution. This study is expected to contribute to synthesize CAp biomaterials with excellent biological properties and to help understanding the chemical and structural properties of mineralized tissues

Ab initio simulation on the crystal structure and elastic properties of carbonated apatite

Ab initio quantum mechanical (QM) calculations were employed to study the crystal structure and elastic properties of carbonated apatite (CAp). Two locations for the carbonate ion in the apatite lattice were considered: carbonate substituting for OH- ion (type-A), and for PO43- ion (type-B) with possible charge compensation mechanisms. A combined type-AB substitution (two carbonate ions replacing one phosphate group and one hydroxyl group, respectively) was also investigated. The results show that the most energetically stable substitution is type-AB, followed by type-A and then type-B. The most stable configuration of type-A has its carbonate triangular plane almost parallel to c-axis at z=0.46. The lowest energy configuration of type-B is that with a sodium ion substituting for a calcium ion for charge balance and the carbonate lying on the b/c-plane of apatite. Lattice parameter changes after carbonate substitution in hydroxyapatite (HA) agree with reported experimental results qualitatively: for type-A, lattice parameter a increases but c decreases; and for type-B, lattice parameter a decreases but c increases. Using the calculated CAp stable structures, we also calculated the elastic properties of CAp and compared them with those of HA and biological apatites. (C) 2013 Elsevier Ltd. All rights reserved

Synthesis and characterization of nano-crystalline calcium phosphates with EDTA-assisted hydrothermal method

This study aims to prepare the biologically important hydroxyapatite (HA), dicalcium phosphate dihydrate (DCPD) and dicalcium phosphate anhydrous (DCPA) phases in simple EDTA - assisted hydrothermal method. X-ray diffraction results showed that pure DCPD, DCPA and HA nano-crystals were obtained in the Ca-EDTA/PO4 solutions at 120 degrees C, 180 degrees C and 210 degrees C, respectively. Thermal gravinnetric analysis of the DCPD precipitates revealed that phase transformations of DCPD to DCPA and DCPA to HA occurred at 139 degrees C and 195 degrees C, respectively, which resulted in the different Ca-P phases during hydrothermal synthesis at different temperature ranges. (C) 2009 Elsevier Ltd. All rights reserved

Ab Initio Simulations on the Carbonated Apatite Structure

ab initio simulations were employed to investigate the crystal structure of carbonated apatite (CAp). Two possible sites for the carbonate ions in the apatite lattice were considered: carbonate substituting for OH- ion (type-A) and for PO43- ion (type-B). A combined type-AB substitution was also proposed and numerous possible charge compensation mechanisms were treated. The results show that the most stable type-A CAp had its carbonate triangular plane almost parallel to c-axis, making an angle of about 2 degrees at z = 0.46. In the most stable type-B CAp structure, the nearest Ca(2) ion was replaced by a sodium ion and the carbonate group was lying almost flat in b/c-plane. Of all the models considered, mixed substitution type-AB where two carbonate ions replacing one phosphate group and one hydroxyl group shows the most stable structure

Calcium phosphate bioceramics induce mineralization modulated by proteins

Proteins play an important role in the process of biomineralization, which is considered the critical process of new bone formation. The calcium phosphate (Ca-P) mineralization happened on hydroxyapatite (HA), beta-tricalcium phosphate (beta-TCP) and biphasic calcium phosphate (BCP) when proteins presented were investigated systematically. The results reveal that the presence of protein in the revised simulated body fluid (RSBF) did not alter the shape and crystal structure of the precipitated micro-crystals in the Ca-P layer formed on the three types of bioceramics. However, the morphology of the Ca-P precipitates was regulated but the structure of Ca-P crystal was unchanged in vivo. The presence of proteins always inhibits Ca-P mineralization in RSBF and the degree of inhibitory effect is concentration dependent. Furthermore, Protein presence can increase the possibility of HA precipitation in vitro and in vivo. The results obtained in this study can be helpful for better understanding the mechanism of biomineralization induced by the Ca-P bioceramics. (c) 2013 Elsevier B.V. All rights reserved

Infrared spectroscopic characterization of carbonated apatite: A combined experimental and computational study

A combined experimental and computational approach was employed to investigate the feasibility and effectiveness of characterizing carbonated apatite (CAp) by infrared (IR) spectroscopy. First, an experimental comparative study was conducted to identify characteristic IR vibrational bands of carbonate substitution in the apatite lattice. The IR spectra of pure hydroxyapatite (HA), carbonate adsorbed on the HA surface, a physical mixture of HA and sodium carbonate monohydrate, a physical mixture of HA and calcite, synthetic CAps prepared using three methods (precipitation method, hydrothermal route, and solid-gas reaction at high temperature) and biological apatites (human enamel, human cortical bone, and two animal bones) were compared. Then, the IR vibrational bands of carbonate in CAp were calculated with density functional theory. The experimental study identified characteristic IR bands of carbonate that cannot be generated from surface adsorption or physical mixtures and the results show that the bands at approximate to 880, 1413, and 1450 cm(-1) should not be used as characteristic bands of CAp since they could result from carbonate adsorbed on the apatite crystals surface or present as a separate phase. The combined experimental and computational study reveals that the carbonate v(3) bands at approximate to 1546 and 1465 cm(-1) are, respectively, the IR signature bands for type A CAp and type B CAp. (c) 2013 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 102A: 496-505, 2014

Characterization and structural analysis of zinc-substituted hydroxyapatites

The substitution of Zn in hydroxyapatite (HA) crystals was examined via comprehensive characterization techniques. Nanosized HA crystals were synthesized by the wet chemical method in aqueous solutions including various amounts of Zn ions. X-ray fluorescent spectroscopy was used to examine the amount of Zn in the HA precipitates. Scanning electron microscopy and high-resolution transmission electron microscopy were employed to examine the effects of Zn on the morphology and crystal size of the precipitates. Conventional powder X-ray diffraction and the Rietveld refinement method revealed the apatite lattice parameters and phase changes with the inclusion of Zn. The results indicated that Zn ions partially substituted for Ca ions in the apatite structure. They were not simply adsorbed on the apatite surface or in the amorphous phase. The precipitates maintained the apatite phase up to a Zn:(Zn + Ca) ratio of 15-20 mol.\% in the solution. Pure HA was well crystallized and the crystals had regular shapes, whereas the Zn-substituted apatite crystals became irregular and formed agglomerates. The lattice parameters, a and c, decreased at a Zn:(Zn + Ca) ratio of 10 mol.\%. (C) 2009 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved

Molecular dynamics simulation of protein effects on interfacial energy between HA surfaces and solutions

Proteins play an important role in apatite formation in physiological environments. The effects of proteins partially result from changing apatite interfacial energy, and in turn, promoting or inhibiting apatite precipitation in physiological environment. The interfacial energies of hydroxyapatite (HA) crystallographic planes in aqueous solutions containing proteins and calcium and phosphate ions were calculated by molecular dynamics simulations. It provided reference interfacial energies for comparative study of protein effects that both acidic Human serum albumin and basic Lysozyme reduces the interfacial energy of HA (001) and HA (100). The simulations provide information to achieve better understanding of protein effects on the biomineralization process at atomic level. (c) 2014 Elsevier B.V. All rights reserved