Preparation and functional study of pH-sensitive amorphous calcium phosphate nanocarriers

Recently,multifunctional nanoparticles have shown great prospects in cancer treatment,which have the ability to simultaneously deliver the drug,image and target tumor cells.In this paper,we designed a luminescent nanoparticles platform based on hydrothermal hyaluronic acid/amorphous calcium phosphate(HA-FCNs/ACP)with multifunctional properties for drug delivery,bio-imaging,and targeting treatment.HA-FCNs/ACP shows an ability to load curcumin(Cur)with pH-sensitive responsive drug release behavior and excellent biocompatibility.HA-FCNs/ACP dispersed in the cytoplasm through the overexpressed CD44 receptor that is actively targeted into human lung cancer cells(A549 cells).Meanwhile,the viability of A549 cells was significantly inhibited in vitro.The prepared HA-FCNs and HA-FCNs/ACP both exhibit excellent targeted bioimaging performance on cancer cells.Hence,the as-prepared nanoparticles have promising applications in treating tumor disease.

Effect of Fluoride on the Ion-association of Calcium Phosphate and Crystallization of Hydroxyapatite

Using a titration setup to accurately control the reaction conditions and in situ monitor the reaction,we showed that fluoride exhibited negligible effects on the ion association process of calcium and phosphate and the formation of ACP nanospheres in a buffer solution with constant ionic strength.However,the stability of ACP increased with increasing fluoride concentration,which was ascribed to the inhibitory effect of fluoride on the aggregation of ACP nanospheres and the nucleation of nanocrystals on the surface of ACP nanospheres.Furthermore,fluoride could inhibit the lateral growth of HAP nanosheets and promote the formation of rod-like crystals.These results further improve our understanding of the crystallization pathway of HAP crystals and the regulatory effects of fluoride.

Redispersible and stable amorphous calcium phosphate nanoparticles functionalized by an organic bisphosphate

Although much effort has been focused on the preparation of stable amorphous calcium phosphate （ACP） nanoparticles in aqueous solution, the redispersibility and long-term stability of ACP nanoparticles in aqueous solution remains an unresolved problem. In this work, stable colloidal ACPs were prepared by using an organic bisphosphonate （BP） as a sterically hindered agent in aqueous solution. The harvested calcium phosphate nanoparticles were characterized by inductively coupled plasma atomic emission spectrometry （ICP-AES）, Fourier transform infrared （FTIR）, X-ray diffraction （XRD）, dynamic light scattering （DLS） and transmission electron microscopy （TEM）. ICP-AES, FTIR and XRD results suggested the particles were ACP. DLS and TEM results indicated that the size of the ACP nanoparticles were in the range of 60 nm with a spherical morphology. The resulting calcium phosphate nanoparticles retained its amorphous nature in aqueous solution for at least 6 months at room temperature due to the stabilizing effect of the organic bisphosphonate. Moreover, the surface of the ACP nanoparticles adsorbed with the organic bisphosphate used showed good redispersibility and high colloid stability both in organic and aqueous solutions.

Clinical evaluation of remineralization potential of casein phosphopeptide amorphous calcium phosphate nanocomplexes for enamel decalcification in orthodontics

Background Enamel decalcification in orthodontics is a concern for dentists and methods to remineralize these lesions are the focus of intense research. The aim of this study was to evaluate the remineralizing effect of casein phosphopeptide amorphous calcium phosphate （CPP-ACP） nanocomplexes on enamel decalcification in orthodontics. Methods Twenty orthodontic patients with decalcified enamel lesions during fixed orthodontic therapy were recruited to this study as test group and twenty orthodontic patients with the similar condition as control group. GC Tooth Mousse, the main component of which is CPP-ACP, was used by each patient of test group every night after tooth-brushing for six months. For control group, each patient was asked to brush teeth with toothpaste containing 1100 parts per million （ppm） of fluoride twice a day. Standardized intraoral images were taken for all patients and the extent of enamel decalcification was evaluated before and after treatment over this study period. Measurements were statistically compared by t test. Results After using CPP-ACP for six months, the enamel decalcification index （EDI） of all patients had decreased; the mean EDI before using CPP-ACP was 0.191＋0.025 and that after using CPP-ACP was 0.183＋0.023, the difference was significant （t=5.169, P 〈0.01）. For control group, the mean EDI before treatment was 0.188±0.037 and that after treatment was 0.187±0.046, the difference was not significant （t=1.711, P 〉0.05）. Conclusion CPP-ACP can effectively improve the demineralized enamel lesions during orthodontic treatment, so it has some remineralization potential for enamel decalcification in orthodontics.

Enhanced integrin-mediated human osteoblastic adhesion to porous amorphous calcium phosphate/poly（L-lactic acid） composite

Background The initial osteoblastic adhesion to materials characterizes the first phase of cell-material interactions and influences all the events leading to the formation of new bone. In a previous work, we developed a novel amorphous calcium phosphate （ACP）/poly（L-lactic acid） （PLLA） material that demonstrated morphologic variations in its microstructure. The aim of this study was to investigate the initial interaction between this material and osteoblastic cells. Cellular attachment and the corresponding signal transduction pathways were investigated. Methods A porous ACP/PLLA composite and PLLA scaffold （as a control） were incubated in fetal bovine serum （FBS） containing phosphate-buffered saline （PBS）, and the protein adsorption was determined. Osteoblastic MG63 cells were seeded on the materials and cultured for 1, 4, 8, or 24 hours. Cell attachment was evaluated using the MTS method. Cell morphology was examined using scanning electron microscopy （SEM）. The expression levels of the genes encoding integrin subunits αl, α5, αv, β1, focal adhesion kinase （FAK）, and glyceraldehyde-3-phosphate dehydrogenase （GAPDH） were determined using real-time reverse transcription polymerase chain reaction （RT-PCR）. Results The ACP/PLLA material significantly increased the protein adsorption by 6.4-fold at 1 hour and 2.4-fold at 24 hours, compared with the pure PLLA scaffold. The attachment of osteoblastic cells to the ACP/PLLA was significantly higher than that on the PLLA scaffold. The SEM observation revealed a polygonal spread shape of cells on the ACP/ PLLA, with the filopodia adhered to the scaffold surface. In contrast, the cells on the PLLA scaffold exhibited a spherical or polygonal morphology. Additionally, real-time RT-PCR showed that the genes encoding the integrin subunits αl, αv, β1, and FAK were expressed at higher levels on the ACP/PLLA composite. Conclusions The ACP/PLLA composite promoted protein adsorption and osteoblastic adhesion. The enhanced cell adhesion may be mediated by the binding of integrin subunits αl, αv, and β1, and subsequently may be regulated through the FAK signal transduction pathways.

Stereolithography printing of bone scaffolds using biofunctional calcium phosphate nanoparticles

Calcium phosphate(CaP)has been widely used for bone defect repair due to good biocompatibility and osteoconductivity.Additive manufacture of calcium phosphate bioceramics with tailored architectures and improved mechanical properties has recently attracted great attention.Herein,calcium phosphate nanoparticles with the size of~89-164 nm were synthesized by the hydrothermal treatment of amorphous calcium phosphate(ACP)precursors at 180°C for 24 h.Biofunctional elements including Mg,Sr and Zn have been doped into these calcium phosphate nanoparticles.Our results revealed that Mg^(2+)ions played critical roles in formation of whitlockite-type calcium phosphate(not hydroxyapatite)from ACP precursors.Moreover,gyroid scaffolds with bionic triply periodic minimal surface structures were fabricated using stereolithography printing of these calcium phosphate nanoparticles,which are likely used as biofunctional scaffolds for bone repair.

Biomineralization Precursor Carrier System Based on Carboxyl- Functionalized Large Pore Mesoporous Silica Nanoparticles

Bone and teeth are derived from intrafibrillarly mineralized collagen fibrils as the second level of hierarchy.According to polymer-induced liquid-precursor process,using amorphous calcium phosphate precursor(ACP)is able to achieve intrafibrillar mineralization in the case of bone biomineral in vitro.Therefore,ACP precursors might be blended with any osteoconductive scaffold as a promising bone formation supplement for in-situ remineralization of collagens in bone.In this study,mesoporous silica nanoparticles with carboxyl-functionalized groups and ultra large-pores have been synthesized and used for the delivery of liquid like biomimetic precursors(ACP).The precursor delivery capacity of the nanoparticles was verified by the precursor release profile and successful mineralization of 2D and 3D collagen models.The nanoparticles could be completely degraded in 60 days and exhibited good biocompatibility as well.The successful translational strategy for biomineralization precursors showed that biomineralization precursor laden ultra large pore mesoporous silica possessed the potential as a versatile supplement in demineralized bone formation through the induction of intrafibrillar collagen mineralization.

Promoting effect of a calcium-responsive self-assemblyβ-sheet peptide on collagen intrafibrillar mineralization

Recently,a de novo synthetic calcium-responsive self-assemblyβ-sheet peptide ID8(Ile-Asp-Ile-Asp-Ile-Asp-Ile-Asp)has been developed to serve as the template inducing hydroxyapatite nucleation.The aim of this study was to evaluate the effect of ID8 on intrafibrillar mineralization of collagen making full use of its self-assembly ability.The mineralization experiments were carried out in vitro on both bare TypeⅠcollagen and fully demineralized dentin samples.The calcium-responsive self-assembly of ID8 was revealed by circular dichroism spectrum,8-anilino-1-naphthalenesulfonic acid ammonium salt hydrate assay,attenuated total reflection Fourier transform infrared spectrum(ATR-FTIR)and transmission electron microscope(TEM).Polyacrylic acid(450 kDa)with a concentration of 100μg ml^(-1)was selected as the nucleation inhibitor based on the determination of turbidimetry and TEM with selected area electron diffraction(TEM-SAED).The results showed that collagen intrafibrillar mineralization was significantly promoted with the pretreatment of self-assembly ID8 detected by TEM-SAED,SEM,X-ray diffraction and ATRFTIR.The pretreatment of collagen utilizing self-assembly ID8 not only enhanced intermolecular hydrogen bonding but also contributed to calcium retention inside collagen and significantly increased the hydrophilicity of collagen.These results indicated that peptides with self-assembly properties like ID8 are expected to be potential tools for biomimetic mineralization of collagen.

Epigallocatechin-3-gallate/mineralization precursors co-delivery hollow mesoporous nanosystem for synergistic manipulation of dentin exposure

As a global public health focus,oral health plays a vital role in facilitating overall health.Defected teeth characterized by exposure of dentin generally increase the risk of aggravating oral diseases.The exposed dentinal tubules provide channels for irritants and bacterial invasion,leading to dentin hypersensitivity and even pulp inflammation.Cariogenic bacterial adhesion and biofilm formation on dentin are responsible for tooth demineralization and caries.It remains a clinical challenge to achieve the integration of tubule occlusion,collagen mineralization,and antibiofilm functions for managing exposed dentin.To address this issue,an epigallocatechin-3-gallate(EGCG)and poly(allylamine)-stabilized amorphous calcium phosphate(PAH-ACP)co-delivery hollow mesoporous silica(HMS)nanosystem(E/PA@HMS)was herein developed.The application of E/PA@HMS effectively occluded the dentinal tubules with acid-and abrasion-resistant stability and inhibited the biofilm formation of Streptococcus mutans.Intrafibrillar mineralization of collagen fibrils and remineralization of demineralized dentin were induced by E/PA@HMS.The odontogenic differentiation and mineralization of dental pulp cells with high biocompatibility were also promoted.Animal experiments showed that E/PA@HMS durably sealed the tubules and inhibited biofilm growth up to 14 days.Thus,the development of the E/PA@HMS nanosystem provides promising benefits for protecting exposed dentin through the coordinated manipulation of dentin caries and hypersensitivity.

Biocompatible Poly(ε-caprolactone)-based Shape-memory Polyurethane Composite Scaffold with Bone-induced Activity

3D porous scaffold could provide suitable bone-like structure for cell adhesion and proliferation;however,surgical suffering from large volume implantation is a great challenge for patients.In this study,a shape programmable porous poly(ε-caprolactone)(PCL)-based polyurethane scaffold with memory effect was synthesized via gas foaming method,using Citrate modified Amorphous calcium Phosphate(CAP)as bioactive factor.The bending experiments indicated that the scaffolds achieved excellent shape-memory effect,which could be influenced by particle weight content.In vitro mineralization results suggested that the deposition of hydroxyapatite was promoted by scaffolds.Additionally,cell assay showed that composite scaffolds presented good cell toxicity and osteogenicity by the differentiation of rat Mesenchymal Stem Cells(rMSCs)into the osteogenic lineage.In the model of rat cranial implantation,the reparative tissue covered the defect site and bone-like structure deposited on the scaffold due to the formation of new bones.In summary,the porous smart shape-memory composite scaffolds could be a potential candidate in future distinctive bone repair applications.