Comparative study of cytotoxicity of detonation nanodiamond particles with an osteosarcoma cell line and primary mesenchymal stem cells.

Recently, nanodiamonds (NDs) have attracted great interest due to their unique physical and chemical properties that could be used in various biological applications. However, depending on the origin, NDs often contain different impurities which may affect cellular functions and viability. Therefore, before their biomedical application, the cytotoxicity of newly produced NDs should be assessed. In the present study, we have evaluated cytotoxicity of four types of ND particles with two cell models: a human osteosarcoma cell line, MG-63, and primary rat mesenchymal stem cells (rMSCs). Detonation-generated nanodiamond (DND) particles were purified with different acid oxidizers and impurities' content was determined by elemental analysis. The particles size distribution was measured revealing that the DND particles have an average size in the range of 51-233 nm. Cytotoxicity was assessed by optical microscopy and proliferation assay after 72 hours exposure of the cells to nanoparticles. We observed cell-specific and material-specific toxicity for all tested particles. Primary stem cells demonstrated higher sensitivity to DND particles than osteosarcoma cells. The most toxic were the DND particles with the smallest grain size and slight content of non-diamond carbon, while DNDs with higher grain size and free from impurities had no significant influence on cell proliferation and morphology. In addition, the smaller DND particles were found to form large aggregates mainly during incubation with rMSCs. These results demonstrate the role of the purification method on the properties of DND particles and their cytotoxicity as well as the importance of cell types used for evaluation of the nanomaterials.

Bioactivity of polycrystalline silicon layers.

After oxygen, silicon is the second most abundant element in the environment and is present as an impurity in most materials. The widespread occurrence of siliceous biominerals as structural elements in lower plants and animals suggests that Si plays a role in the production and maintenance of connective tissue in higher organisms. It has been shown that the presence of Si is necessary in bones, cartilage and in the formation of connective tissue, as well as in some important metabolic processes. In this work, polycrystalline silicon layers are tested in terms of bioactivity, i.e., their ability to induce hydroxyapatite formation from simulated body fluid. Hydroxyapatite is a biologically compatible material with chemical similarity to the inorganic part of bones and teeth. Polycrystalline silicon layers are obtained by aluminum induced crystallization of Al and amorphous Si thin films deposited sequentially on glass substrates by radio-frequency magnetron sputtering and subsequently annealed in different atmospheres. The hydroxyapatite formation is induced by applying a method of laser-liquid-solid interaction. The method consists of irradiating the samples with laser light while immersed in a solution that is supersaturated with respect to Ca and P. As a result, heterogeneous porous sponge-like carbonate-containing hydroxyapatite is grown on the polysilicon surfaces. Crystals that are spherical in shape, containing Ca, P and O, Na, Cl, Mg, Al, Si and S, as well as well-faceted NaCl crystals are embedded in the hydroxyapatite layer. Enhancement of the hydroxyapatite growth and increased crystallinity is observed due to the applied laser-liquid-solid interaction.

Calcium phosphate formation on plasma immersion ion implanted low density polyethylene and polytetrafluorethylene surfaces.

The flexible structure of polymers has enabled them to be useful in a wide variety of medical applications due to the possibility to tailor their properties to suit desired applications. For a long time, there has been an increasing interest in utilizing polymers as matrices for calcium phosphate-based composites with applications in hard tissue implants. On the other side, polymers with application as heart valves, urea catheters and artificial vessels present a case where the formation of minerals (namely calcification) should be avoided. The modification of polymer surfaces by various ion beam treatments for reducing the calcification, as for example plasma immersion ion implantation (PIII), is well known and has a long time effect. This work is part of a wider investigation of the ability of plasma immersion ion implanted polymers to induce calcium phosphate formation from an aqueous solution resembling the human blood plasma. In the experiment described in this paper, topographical and chemical changes were inserted on the surfaces of two conventional polymers (low density polyethylene and polytetrafluorethylene) by PIII with nitrogen ions, and under conditions mimicking the natural mineral formation processes. The effect of the plasma modification on the calcium phosphate nucleation and growth from the aqueous solution was ambiguous. We suppose that the complex combination of surface characteristics influenced the ability of the plasma treated polymer films to induce the formation of a calcium phosphate layer.

Hydroxyapatite grown on a native extracellular matrix: initial interactions with human fibroblasts.

Proteins are known to modulate the physical properties of minerals, and thus we anticipate that they will strongly influence the structure and the biological properties of biomimetically prepared carbonate-containing hydroxyapatite. This study was designed to learn more about the main morphological characteristics of hydroxyapatite layer grown on different substrates coated with an extracellular matrix, a biological matrix that was produced by cultured osteoblast-like cells. The hydroxyapatite growth was carried out in a simulated body fluid, a solution that resembles the human blood plasma. It was found that the extracellular matrix may serve as a template for the mineralization of biomimetic hydroxyapatite on the surface of materials like stainless steel, silicon, and silica glass, leading to the formation of a homogeneous layer. The latter was consisting of nanometer-sized hydroxyapatite crystals grouped in particles with regular sphere shape and with a significantly higher average diameter in comparison to samples without extracellular matrix coating. Subsequent in vitro studies with living fibroblasts showed that the cellular behavior depended on the type of underlying substrate used for the hydroxyapatite growth, as well as on the immersion time of the samples in the simulated body fluid. Increasing the thickness of the hydroxyapatite layer altered visibly the cellular response, and the fibroblasts developed stellate morphology on the samples with a hydroxyapatite-extracellular matrix coating. Preadsorption with fibronectin significantly improved the initial cell adhesion and spreading to all surfaces. Thus, such an approach may contribute to the development of surfaces with better tissue compatibility.

White light scanning interferometry adapted for large-area optical analysis of thick and rough hydroxyapatite layers.

Understanding the mechanisms of biomineralization continues to be an important area of research in physics, chemistry, materials science, medicine, and dentistry due to its importance in the formation of bones, teeth, cartilage, etc. Stimulated by these fascinating natural examples, as well as by certain others such as shells and corals, attempts are being made to develop synthetic, biomimetic nanocomposites by simulating the basic principles of biomineralization. We have grown bio-like hydroxyapatite layers in vitro on substrates of stainless steel, silicon, and silica glass by using a biomimetic approach (i.e., immersion in a supersaturated simulated body fluid). Hydroxyapatite is one of the most common natural biomaterials and an important structural component of bones and teeth. Metal substrates are of interest for hard tissue implants, while semiconductors and glasses are under investigation for their use as biosensors. Using classical techniques such as stylus profiling, atomic force microscopy (AFM), and scanning and transmission electron microscopy (SEM and TEM), it was found difficult, ambiguous, destructive, or time-consuming to measure the topography, thickness, and profile of the grown heterogeneous, thick, and rough hydroxyapatite layers. On the other hand, coherence probe microscopy based on white light scanning interferometry and image processing provides rapid, contactless measurements of surface roughness and does not need any sample preparation. The results obtained have shown a typical layer thickness of up to 20 microm and an average root-mean-square (rms) roughness of about 4 mum. The hydroxyapatite investigated in this work presents nonetheless a challenge for this technique because of its semi-translucency, high surface roughness, and the presence of cavities formed throughout its volume. This results in a variable quality of fringe pattern, ranging from classical fringes (on a smooth surface) to complex fringes displaying properties of white light speckle (on a rough surface), together with multiple fringe signals along the optical axis in the presence of buried layer interfaces, which in certain configurations affect the axial and lateral precision of the measurement. In this paper we present the latest results for optimizing the measurement conditions in order to reduce such errors and to provide additional useful information concerning the layer.