Engineered substrates incapable of induction of chondrogenic differentiation compared to the chondrocyte imprinted substrates.

It is well established that surface topography can affect cell functions. However, finding a reproducible and reliable method for regulating stem cell behavior is still under investigation. It has been shown that cell imprinted substrates contain micro- and nanoscale structures of the cell membrane that serve as hierarchical substrates, can successfully alter stem cell fate. This study investigated the effect of the overall cell shape by fabricating silicon wafers containing pit structure in the average size of spherical-like chondrocytes using photolithography technique. We also used chondrocyte cell line (C28/I2) with spindle-like shape to produce cell imprinted substrates. The effect of all substrates on the differentiation of adipose-derived mesenchymal stem cells (ADSCs) has been studied. The AFM and scanning electron microscopy images of the prepared substrates demonstrated that the desired shapes were successfully transferred to the substrates. Differentiation of ADSCs was investigated by immunostaining for mature chondrocyte marker, collagen II, and gene expression of collagen II, Sox9, and aggrecan markers. C28/I2 imprinted substrate could effectively enhanced chondrogenic differentiation compared to regular pit patterns on the wafer. It can be concluded that cell imprinted substrates can induce differentiation signals better than engineered lithographic substrates. The nanostructures on the cell-imprinted patterns play a crucial role in harnessing cell fate. Therefore, the patterns must include the nano-topographies to have reliable and reproducible engineered substrates.

The evaluation of prepared microstructure pattern by carbon-dioxide laser on zirconia-based ceramics for dental implant application: an in vitro study.

3 mol% yttria-stabilized zirconia ceramics have been gaining attention as promising restorative materials that are extensively used in dental implant applications. However, implant failure due to bacterial infection and its bioinert surface slow osseointegration in vivo, which are significant issues in clinical applications. In this work, surface modification was achieved using a continuous wave carbon dioxide laser at a wavelength of 10.6 µm in an air atmosphere. Changes in the surface characteristics were evaluated using X-ray diffraction (XRD), field-emission scanning electron microscopy (FESEM), atomic force microscopy (AFM), and 2D roughness and hardness tests. The bioactivity of the laser-treated samples was studied by examining their behavior when immersed in the SBF solution. The formation of the hydroxyapatite phase on the laser-treated sample was much more uniform than that of its untreated counterparts. The antibacterial properties of surface-treated zirconia ceramics against Streptococcus mutans and Escherichia coli bacteria were rigorously examined. These results indicate that the laser-induced nanoscale grooves significantly improved antibacterial activity by creating hydrophobic surfaces. The cellular response was evaluated for 7 days on microtextures on the zirconia surfaces and an untreated sample with MC3T3-E1 pre-osteoblast cell line cultured under basal conditions. Surface topography was revealed to improve the cellular response with increased metabolic activity compared to the untreated sample and showed modulation of cell morphology for the entire time. These results suggest that laser modification can be an appropriate non-contact method for designing nanoscale microtextures to improve the biological response and antibacterial behavior of zirconia ceramics in restorative dentistry.

Effect of Dioxane and N-Methyl-2-pyrrolidone as a Solvent on Biocompatibility and Degradation Performance of PLGA/nHA Scaffolds

Background: Solvent casting/particulate leaching is one of the most conventional methods for fabricating polymer/ceramic composite scaffolds. In this method, the solvent generally affects resulting scaffold properties, including porosity and degradation rate. Methods: Herein, composite scaffolds of PLGA (poly(lactide-co-glycolide))/ nano-hydroxyapatite (nHA) with different percentages of nHA (25, 35, and 45 wt. %) were prepared by the solvent casting/particle leaching combined with freeze drying. The effects of two different solvents, 1,4-dioxane (DIO) and N-methyl-2-pyrrolidone (NMP), on morphology, porosity, bioactivity, degradation rate, and biocompatibility of the resulting scaffolds were investigated. Results: The results revealed that increasing the nano-hydroxyapatite (nHA) percentages had no significant effect on the porosity and interconectivity of scaffolds (p > 0.05), whereas altering the solvent from DIO into NMP decreased the porosity from about 87% into 71%, respectively. Moreover, scaffolds of DIO illustrated the high results of cell proliferation compared to those of NMP; the cell viability of GD25 decreased from 85% to 65% for GN25. The findings also indicated that scaffolds prepared by NMP had a higher rate of losing weight in comparison to DIO. Adding nHA to PLGA had a significant effect on the bioactivity of scaffolds (p < 0.05), composite scaffolds with 45 wt % nHA had at least 30% more weight gain compared to the neat polymer scaffolds. Conclusion: The DIO scaffolds have higher rates of porosity, interconnectivity, bioactivity, and biocompatibility than NMP scaffolds due to its high evaporation rate.

Enhancing degradability, bioactivity, and osteocompatibility of poly (propylene fumarate) bone filler by incorporation of Mg-Ca-P nanoparticles.

As an alternative for polymethyl methacrylate, poly(propylene fumarate) (PPF) has been considered as injectable and biodegradable bone cement; however, its mechanical and biological properties need more attention. Hence, the current study aimed to develop the properties by compositing PPF with magnesium calcium phosphate (MCP) nano-powders. In this regard, the pure PPF was compared with PPF/MCP by evaluating their composition, mechanical properties, hydrophilicity, and biodegradability. Furthermore, their bioactivity in the simulated body fluid (SBF) and, via applying MG-63 cells, their cell interaction, including proliferation, adhesion, differentiation, and mineralization, were assessed. The addition of MCP improved compressive strength and elastic modulus of PPF, e.g., 10 wt% MCP increased them to 32.7 and 403 MPa, respectively. Also, hydrophilicity and biodegradation of PPF were enhanced in the presence of MCP; so that the highest hydrophilicity, 42% higher than PPF, was achieved at the presence of 20 wt% MCP. In this condition, after 21-day immersion in SBF, the surface of the sample was covered with a dense and continuous layer of hydroxyapatite. The composite proliferation, adhesion, differentiation, and mineralization of MG-63 cells improved in comparison to the pure PPF. Hence, controllable strength and biodegradation of the composite, along with its proved bioactivity and osteoconductivity, make PPF/MCP as a candidate for bone therapeutic application.

Poly(propylene fumarate)/magnesium calcium phosphate injectable bone composite: Effect of filler size and its weight fraction on mechanical properties.

This study aimed to produce a composite of poly(propylene fumarate)/magnesium calcium phosphate as a substitutional implant in the treatment of trabecular bone defects. So, the effect of magnesium calcium phosphate particle size, magnesium calcium phosphate:poly(propylene fumarate) weight ratio on compressive strength, Young's modulus, and toughness was assessed by considering effective fracture mechanisms. Micro-sized (∼30 µm) and nano-sized (∼50 nm) magnesium calcium phosphate particles were synthesized via emulsion precipitation and planetary milling methods, respectively, and added to poly(propylene fumarate) up to 20 wt.%. Compressive strength, Young's modulus, and toughness of the composites were measured by compressive test, and effective fracture mechanisms were evaluated by imaging fracture surface. In both micro- and nano-composites, the highest compressive strength was obtained by adding 10 wt.% magnesium calcium phosphate particles, and the enhancement in nano-composite was superior to micro-one. The micrographs of fracture surface revealed different mechanisms such as crack pinning, void plastic growth, and particle cleavage. According to the results, the produced composite can be considered as a candidate for substituting hard tissue.

Evaluation of setting time and compressive strength of a new bone cement precursor powder containing Mg-Na-Ca.

Taking the advantage of a novel magnesium phosphate precursor containing Na and Ca, the cementation rate of the cement, including only Mg/Mg-Na-Ca, was studied. Besides, two effective parameters, that is, calcination temperature, 650 °C and 800 °C, and powder-to-cement liquid ratio, 1 and 1.5 g/mL, were assessed. X-ray diffraction, scanning electron microscopy, ion chromatography, particle size analyser, Vicat needle and compression test were used to characterize the powders and obtained cements. The sample containing Mg-Na-Ca, calcined at 800 °C with powder-to-cement liquid ratio of 1.5, obtained the highest compressive strength, 20 MPa, but set fast. To control the kinetics of cementation, the powder containing Mg-Na-Ca calcined at 950 °C with powder-to-cement liquid ratio of 1.5 and 2 g/mL was assessed and the one with 2 g/mL set in 9 min possessing 22 MPa compressive strength was selected as optimal condition to be used as a candidate, injectable bone cement.

Incorporation of Nanoalumina Improves Mechanical Properties and Osteogenesis of Hydroxyapatite Bioceramics.

A handful of work focused on improving the intrinsic low mechanical properties of hydroxyapatite (HA) by various reinforcing agents. However, the big challenge regarding improving mechanical properties is maintaining bioactivity. To address this issue, we report fabrication of apatite-based composites by incorporation of alumina nanoparticles (n-Al2O3). Although numerous studies have used micron or submicron alumina for reinforcing hydroxyapatite, only few reports are available about the use of n-Al2O3. In this study, spark plasma sintering (SPS) method was utilized to develop HA-nAl2O3 dense bodies. Compared to the conventional sintering, decomposition of HA and formation of calcium aluminates phases are restricted using SPS. Moreover, n-Al2O3 acts as a bioactive agent while its conventional form is an inert bioceramics. The addition of n-Al2O3 resulted in 40% improvement in hardness along with a 110% increase in fracture toughness, while attaining nearly full dense bodies. The in vitro characterization of nanocomposite demonstrated improved bone-specific cell function markers as evidenced by cell attachment and proliferation, alkaline phosphatase activity, calcium and collagen detection and nitric oxide production. Specifically, gene expression analysis demonstrated that introduction of n-Al2O3 in HA matrix resulted in accelerated osteogenic differentiation of osteoblast and mesenchymal stem cells, as expression of Runx-2 and OSP showed 2.5 and 19.6 fold increase after 2 weeks (p < 0.05). Moreover, protein adsorption analysis showed enhanced adsorption of plasma proteins to HA-nAl2O3 sample compared to HA. These findings suggest that HA-nAl2O3 could be a prospective candidate for orthopedic applications due to its improved mechanical and osteogenic properties.

Doxorubicin loaded large-pore mesoporous hydroxyapatite coated superparamagnetic Fe3O4 nanoparticles for cancer treatment.

In the present study, a series of multifunctional drug delivery systems based on mesostructured hydroxyapatite coating and superparamagnetic nanoparticles with pH-responsive characters was prepared. The structure of each new synthesized nanoscale composite was fully characterized by XRD, FTIR, TEM, VSM and BET. The results showed a good ordered mesostructure having large pores, high pore volume, high surface area, and varied super paramagnetic properties. The mesoporous hydroxyapatite coated super paramagnetic Fe3O4 nanoparticles were applied as a drug delivery carrier loaded with doxorubicin (DOX) as a model drug. The storage/release properties of the developed nonocarriers in phosphate buffer saline (PBS) were studied in two certain pHs: pH=7.4 (the human blood pH) and pH=5.5 (pH of cancer cells). The large pores in the synthesized mesoporous acted as an excellent carrier for DOX molecules with a loading efficiency of ≈93% which is much higher than that of the conventional hydroxyapatite particles. When the pH of the release medium (PBS) was changed from 7.4 to 5.5, the drug release increased significantly from 10% of the adsorbed drug to about 70%. DOX-loaded mesostructure hydroxyapatite reduced the viability of SKBR3 and T47D cells by 54.7 and 57.3%, respectively, which were very similar to 56.8 and 60.4% reduction resulted from free DOX incubation. This new drug delivery system which benefits from both super paramagnetic properties and pH-responsive performances may serve as a suitable platform for developing new biocompatible drug carriers and could have a good potential use in targeted cancer therapy.

The effect of aqueous media on the mechanical properties of fluorapatite-mullite glass-ceramics.

OBJECTIVES: To verify the effects of alternating thermal changes in aqueous media and chemical composition on mechanical properties of apatite-mullite glass-ceramics and to investigate concentration of ions eluted from glass-ceramics in aqueous media. MATERIALS AND METHODS: The glass compositions were from SiO2Al2O3P2O5CaOTiO2BaOZrO2CaF2 system. Glass-ceramics were prepared by heat-treating at 1100°C for 3h samples alternately immersed in water at 5 and 60°C. The 3-point bending strength (n=10) were determined using 3×4×25mm/bar and a universal testing machine, at a cross-head speed of 0.1mm/min. Vickers micro hardness were evaluated by applying a total of 15-20 indentations under a 100g load for 30s. Concentrations of ions eluted from glass-ceramics immersed in 60±5°C double distilled water were determined by ion chromatography. The toxicity of glass-ceramics was assessed by seeding the osteosarcoma cells (MG63) on powder for different days and their cell proliferation assessment was investigated by MTT assay. The data were analyzed using one way analysis of variance and the means were compared by Tukey's test (5% significance level). RESULTS: The highest flexural strength and hardness values after thermal changes belonged to TiO2 and ZrO2 containing glass-ceramics which contained lower amount of released ions. BaO containing glass-ceramic and sample with extra amount of silica showed the highest amount of reduction in their mechanical strength values. These additives enhanced the concentration of eluted ions in aqueous media. MTT results showed that glass-ceramics were almost equivalent concerning their in-vitro biological behavior. SIGNIFICANCE: Thermal changes and chemical compositions had significant effects on flexural strength and Vickers micro-hardness values.

Enhancement of mechanical properties of experimental composite by Fuller's earth nanofibers for cervical restoration.

In this research, we studied improvement of mechanical properties of dimethacrylate-silica based dental composites by addition of Fuller's Earth (FE) clay. Three composites were made as base compounds consisting of 68, 58, and 48 wt % resin and 31, 41, and 51 wt % silica, respectively. Afterward, the composites were modified by adding FE. Mechanical properties including flexural strength, flexural modulus, work-of-fracture, fracture toughness, and microhardness were measured. Clay particles and fracture surface of composites consisting of 51 wt % silica (with and without FE) were examined by SEM. Measured results showed that flexural strength, work-of-fracture, flexural modulus, and microhardness of all composites increased by including FE nanofibers. Fracture toughness except for composite including 51 wt % silica had similar variations. It seems that locating FE nanofibers in weak resin region among silica particles leads to strengthening mechanisms, such as bridging and crack deflection, which cause improvement in mechanical properties.

Hydrothermal synthesis and characterization of hydroxyapatite and fluorhydroxyapatite nano-size powders.

Pure hydroxyapatite (HAp) and fluoride-containing apatite powders (FHAp) were synthesized using a hydrothermal method. The powders were assessed by x-ray diffraction (XRD), Fourier transform infrared (FTIR), scanning electron microscope (SEM) and F-selective electrode. X-ray diffraction results revealed the formation of single phase apatite structure for all the compositions synthesized in this work. However, the addition of a fluoride ion led to a systematic shift in the (3 0 0) peak of the XRD pattern as well as modifications in the FTIR spectra. It was found that the efficiency of fluoride ion incorporation decreased with the increase in the fluoride ion content. Fluorine incorporation efficiency was around 60% for most of the FHAp samples prepared in the current study. Smaller and less agglomerated particles were obtained by fluorine substitution. The bioactivity of the powder samples with different fluoride contents was compared by performing cell proliferation, alkaline phosphatase (ALP) and Alizarin red staining assays. Human osteoblast cells were used to assess the cellular responses to the powder samples in this study. Results demonstrated a strong dependence of different cell activities on the level of fluoridation.

Fabrication and characterization of poly(D,L-lactide-co-glycolide)/hydroxyapatite nanocomposite scaffolds for bone tissue regeneration.

Conventional methods in fabrication of scaffolds based on polymer/bioceramic composites frequently make use of solution casting then particle leaching. The residues of common organic solvents can get trapped in this technique hence provide safety concerns on final scaffold. In this study, N-methyl pyrrolidone was used as an approved solvent for parenteral pharmaceutical products especially implants with acceptable toxicological profile. A combined freeze drying and solvent casting methods were adopted for complete removal of the solvent from final scaffold structure. Biodegradable scaffolds based on poly (D,L-lactide-co-glycolide) and different percentages of nanohydroxyapatite (25, 35, and, 45% w/w) were characterized thoroughly regarding porosity, pore distribution as well as their bioactivity and biocompatibility. The results showed 70-80% porosity with a size distribution in the range of 50-200 mum for different conditions. Bioactivity of the scaffolds was directly dependent on the bioceramic content in the samples according to the results. Composites and neat samples showed the same cytocompatibility profile. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res, 2010.

Investigation of mechanical properties of experimental Bis-GMA/TEGDMA dental composite resins containing various mass fractions of silica nanoparticles.

PURPOSE: Mechanical properties of dental composite resins need to be improved in order to enhance their performance for applications in direct restorations. Application of nanoparticles in this field is a recent development. The aim of this study was to investigate the mechanical properties of experimental composites containing various mass fractions of silica nanoparticles. MATERIALS AND METHODS: Experimental composites were composed of a visible-light-curing monomer mixture (70 wt% Bis-GMA and 30 wt% TEGDMA) and silica nanoparticles of a size ranging from 20 nm to 50 nm modified with gamma-methacryloxy propyl trimethoxy silane (gamma-MPS) as reinforcing filler. The composites were classified into four groups according to their filler mass fractions ranging from 20% to 50%. Following the same preparation procedure, a conventional composite was also fabricated consisting of a mass percentage of 60% silica fillers having particle sizes ranging from 10 microm to 40 microm in the same organic matrix, which served as control. Ten specimens were prepared of each experimental group and also of the control. Fracture toughness was measured using single-edge notched bend (SENB) specimens. Specimen fracture surfaces were mounted on aluminum stubs with carbon cement, sputter-coated with gold and examined under scanning electron microscopy (SEM). Flexural strength was evaluated through a standard three-point bending test and Vickers microhardness test was performed to investigate the hardness of the samples. RESULTS: Filler mass fraction had a significant effect on composite properties. Fracture toughness, flexural strength, and hardness of composites at filler mass fraction of 40% of silica nanoparticles were (mean +/- SD) 1.43 +/- 0.08 MPa.m(1/2), 149.74 +/- 8.14 MPa, and 62.12 +/- 3.07 VHN, respectively; relevant values for composites at 50% mass fraction of silica nanoparticles were 1.38 +/- 0.07 MPa.m(1/2), 122.83 +/- 6.13 MPa, and 70.69 +/- 3.67 VHN, respectively, all of which were significantly higher than 1.07 +/- 0.06 MPa.m(1/2), 104.61 +/- 8.73 MPa, and 52.14 +/- 4.02 VHN of the control, respectively (Tukey's multiple comparison test; family confidence coefficient = 0.95). Measured values for composites at 20% mass fraction of silica nanoparticles were 0.94 +/- 0.06 MPa.m(1/2), 103.41 +/- 7.62 MPa, and 42.87 +/- 2.61 VHN, respectively; relevant values for composites at 30% mass fraction of silica nanoparticles were 1.16 +/- 0.07 MPa.m(1/2), 127.91 +/- 7.05 MPa, and 51.78 +/- 3.41 VHN, respectively. CONCLUSIONS: Reinforcement of dental composite resins with silica nanoparticles resulted in a significant increase in the evaluated mechanical properties in comparison with the conventional composite. The filler mass fraction played a critical role in determining the composite's mechanical properties.