Shin-Woo Ha

Bioactive Silica Nanoparticles Promote Osteoblast Differentiation through Stimulation of Autophagy and Direct Association with LC3 and p62

We recently identified an engineered bioactive silica-based nanoparticle formulation (designated herein as NP1) that stimulates in vitro differentiation and mineralization of osteoblasts, the cells responsible for bone formation, and increases bone mineral density in young mice in vivo. The results demonstrate that these nanoparticles have intrinsic biological activity; however, the intracellular fate and a complete understanding of the mechanism(s) involved remains to be elucidated. Here we investigated the cellular mechanism(s) by which NP1 stimulates differentiation and mineralization of osteoblasts. We show that NP1 enters the cells through a caveolae-mediated endocytosis followed by stimulation of the mitogen activated protein kinase ERK1/2 (p44/p42). Our findings further revealed that NP1 stimulates autophagy including the processing of LC3β-I to LC3β-II, a key protein involved in autophagosome formation, which is dependent on ERK1/2 signaling. Using a variant of NP1 with cobalt ferrite magnetic metal core (NP1-MNP) to pull down associated proteins, we found direct binding of LC3β and p62, two key proteins involved in autophagosome formation, with silica nanoparticles. Interestingly, NP1 specifically interacts with the active and autophagosome associated form of LC3β (LC3β-II). Taken together, the stimulation of autophagy and associated signaling suggests a cellular mechanism for the stimulatory effects of silica nanoparticles on osteoblast differentiation and mineralization.

Fabrication of triacetylcellulose-SiO2 nanocomposites by surface modification of silica nanoparticles.

We have successfully fabricated triacetylcellulose (TAC) polymer-silica nanocomposite films having up to 40 wt % of incorporated silica nanoparticles by deliberately designing a surface ligand that has a structure similar to that of polymer repeating units and effectively modifying the surface of silica nanoparticles through chemical bonding. Cross-sectional TEM analysis reveals no significant aggregation in all TAC-silica nanocomposite films. Thermal analysis results suggested that TAC-silica nanocomposites had higher T(g) and T(c) values as compared to pure TAC, and the increase in T(g) and T(c) was affected by the silica content. The transparency of all the nanocomposite films was over 80% in the visible range, confirming the excellent compatibility of nanoparticles with TAC. In this study, we enhance the interaction between nanoparticles and polymer matrices by modifying the surface of nanoparticles with a ligand that has a structure similar to that of polymer repeating units. It is expected that this method can be applied to other polymer systems to develop useful nanocomposites.

Excellent Photostability of Phosphorescent Nanoparticles and Their Application as a Color Converter in Light Emitting Diodes

The phosphorescent Ir(III) complexes were modified by allylation and consecutive hydrosilylation, and covalently incorporated into the silica nanoparticles by hydrolysis and condensation reaction with TEOS. These nanoparticles showed an excellent photochemical and thermal stability, and a very high luminescent efficiency due to the blocking of O(2) quenching and suppression of energy transfer through the amorphous silica solid solution. The limited mobility of complexes in the silica matrix also resulted in a decrease in the vibration relaxation and restrained the nonradiative decay. It is expected that these photostable and very efficient phosphorescent nanoparticles can be used in various fields ranging from nanobiotechnology to nanoengineering materials, where long-term stability with the high luminescent efficiency is required. As an example of the use of excellent photostability, a preliminary test result in which they are used as a color converter in a light emitting diode (LED) is also discussed.

Impact of Phosphorus-Based Food Additives on Bone and Mineral Metabolism

CONTEXT Phosphorus-based food additives can substantially increase total phosphorus intake per day, but the effect of these additives on endocrine factors regulating bone and mineral metabolism is unclear. OBJECTIVE This study aimed to examine the effect of phosphorus additives on markers of bone and mineral metabolism. Design and Setting, and Participants: This was a feeding study of 10 healthy individuals fed a diet providing ∼1000 mg of phosphorus/d using foods known to be free of phosphorus additives for 1 week (low-additive diet), immediately followed by a diet containing identical food items; however, the foods contained phosphorus additives (additive-enhanced diet). Parallel studies were conducted in animals fed low- (0.2%) and high- (1.8%) phosphorus diets for 5 or 15 weeks. MAIN OUTCOME MEASURES The changes in markers of mineral metabolism after each diet period were measured. RESULTS Participants were 32 ± 8 years old, 30% male, and 70% black. The measured phosphorus content of the additive-enhanced diet was 606 ± 125 mg higher than the low-additive diet (P < .001). After 1 week of the low-additive diet, consuming the additive-enhanced diet for 1 week significantly increased circulating fibroblast growth factor 23 (FGF23), osteopontin, and osteocalcin concentrations by 23, 10, and 11%, respectively, and decreased mean sclerostin concentrations (P < .05 for all). Similarly, high-phosphorus diets in mice significantly increased blood FGF23, osteopontin and osteocalcin, lowered sclerostin, and decreased bone mineral density (P < .05 for all). CONCLUSIONS The enhanced phosphorus content of processed foods can disturb bone and mineral metabolism in humans. The results of the animal studies suggest that this may compromise bone health.

Bio-active engineered 50 nm silica nanoparticles with bone anabolic activity: therapeutic index, effective concentration, and cytotoxicity profile in vitro.

Silica-based nanomaterials are generally considered to be excellent candidates for therapeutic applications particularly related to skeletal metabolism however the current data surrounding the safety of silica based nanomaterials is conflicting. This may be due to differences in size, shape, incorporation of composite materials, surface properties, as well as the presence of contaminants following synthesis. In this study we performed extensive in vitro safety profiling of ∼ 50 nm spherical silica nanoparticles with OH-terminated or Polyethylene Glycol decorated surface, with and without a magnetic core, and synthesized by the Stöber method. Nineteen different cell lines representing all major organ types were used to investigate an in vitro lethal concentration (LC) and results revealed little toxicity in any cell type analyzed. To calculate an in vitro therapeutic index we quantified the effective concentration at 50% response (EC50) for nanoparticle-stimulated mineral deposition activity using primary bone marrow stromal cells (BMSCs). The EC50 for BMSCs was not substantially altered by surface or magnetic core. The calculated Inhibitory concentration 50% (IC50) for pre-osteoclasts was similar to the osteoblastic cells. These results demonstrate the pharmacological potential of certain silica-based nanomaterial formulations for use in treating bone diseases based on a favorable in vitro therapeutic index.

Dental and Skeletal Applications of Silica-Based Nanomaterials

The unique combination of semistructured extracellular matrix, biomechanical properties, and active remodeling makes dentin and bone unique tissues, particularly suited to nanomaterials. Silica has long been used for dental applications because of its physical and optical properties as well as compatibility in composites; however, the emergence of nanotechnology has provided new opportunities to package and deliver certain elements at the nanoscale, with the intent of enhancing biological effects or properties. Recent studies have suggested that engineered silica nanoparticles possess beneficial properties that endow them with enhanced physical and mechanical properties for dental applications as well as therapeutic properties for bone. In this chapter, we will discuss the specific physicochemical properties of silica-based nanomaterials including synthesis methods, size, shape, surface properties, and biocompatibility in the context of both mechanical properties and potential biological applications to living cells relevant to both dentition and the skeleton.

Nano-hydroxyapatite stimulation of gene expression requires Fgf receptor, phosphate transporter, and Erk1/2 signaling.

Hydroxyapatite (HAp) is critical to health both as the main structural material of the skeleton and storage material of calcium and phosphate. Nanosized HAp (nHAp) is naturally produced by mineralizing cells during bone formation and remodeling and is the main constituent of the skeleton. As such, HAp is currently being investigated as a therapeutic biomaterial for orthopedic and dental purposes. Recent studies have suggested that extracellular nHAp can influence osteoblast lineage commitment and cell function through changes in gene expression; however, the mechanisms remain to be elucidated. Here, the cellular and molecular mechanism by which rod-shaped nHAp (10 × 100 nm) stimulates gene expression in preosteoblast bone marrow stromal cells was investigated. Electron microscopy detected a rapid and stable interaction of nHAp with the cell membrane, which correlated with a strong stimulation of the Erk1/2 signaling pathway. Results also identified the requirement of the Fgf receptor signaling and phosphate-transporters for nHAp regulated gene expression whereas a calcium-sensing receptor inhibitor had no effect. Collectively, the study uncovers novel signaling pathways and cellular events specifically stimulated by and required for the cellular response to free extracellular HAp. The results provide insight into the osteoblastic response to HAp relevant to functional mineralization and pathological calcification and could be used in the development of biomaterials for orthopedic purposes.

Synthesis of pH stable, blue light-emitting diode-excited, fluorescent silica nanoparticles and effects on cell behavior

To date, delivery of light-emitting diode (LED)-activated compounds to cells and tissue remains a challenge. Silica-based materials possess good biocompatibility and have advantages of control of size and shape. Fluorescent silica nanoparticles (NPs) have been synthesized and used for applications such as cell tracking and tumor identification. Here, we report the synthesis and optimization of fluorescent silica NPs, which incorporate a naphthalimide dye with triethoxysilanes that are excited by the blue LED wavelength (LEDex NPs). The NPs can be imaged in the 420–470 nm wavelength, demonstrate a high quantum yield, are stable in a range of pH, and are taken into the cells. Therefore, these NPs represent a novel imaging technology for biomedical applications.