Biocompatibility of magnesium implants in primary human reaming debris-derived cells stem cells in vitro.

BACKGROUND: Use of magnesium for resorbable metal implants is a new concept in orthopaedic and dental medicine. The majority of studies on magnesium's biocompatibility in vitro have assessed the short-term effect of magnesium extract on cells. The aim of this study was to evaluate the influence of direct exposure to magnesium alloys on the bioactivity of primary human reaming debris-derived (HRD) cells. MATERIALS AND METHODS: Pure Mg, Mg2Ag, WE43 and Mg10Gd were tested for biocompatibility. The study consisted of assessment of cell viability by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test, evaluation of alkaline phosphatase (ALP) content, and study of cell morphology under light microscopy, scanning electron microscopy (SEM) and transmission electron microscopy (TEM), along with determination of calcification and pH changes induced by magnesium. RESULTS: The number of viable cells in the presence of Mg2Ag was high over the entire observation period. Inhibition of ALP content in osteogenic differentiating HRD was caused by pure Mg at day 14 and 28. All other magnesium alloys did not affect the ALP content. Exposure of HRD to magnesium increased the amount of lysosomes and endocytotic vesicles. Cellular attachment was generally the best for those crystals that formed on the surface of all materials. A decrease was observed in Ca(2+) in the medium from day 1 to day 14. CONCLUSIONS: In terms of cell morphology, cell viability and differentiation, cell density and the effect on the surrounding pH, Mg2Ag showed the most promising results. All magnesium materials induced calcification, which is beneficial for orthopaedic and dental applications.

Effects of dendritic core-shell glycoarchitectures on primary mesenchymal stem cells and osteoblasts obtained from different human donors.

The biological impact of novel nano-scaled drug delivery vehicles in highly topical therapies of bone diseases have to be investigated in vitro before starting in vivo trials. Highly desired features for these materials are a good cellular uptake, large transport capacity for drugs and a good bio-compatibility. Essentially the latter has to be addressed as first point on the agenda. We present a study on the biological interaction of maltose-modified poly(ethyleneimine) (PEI-Mal) on primary human mesenchymal stem cell, harvested from reaming debris (rdMSC) and osteoblasts obtained from four different male donors. PEI-Mal-nanoparticles with two different molecular weights of the PEI core (5000 g/mol for PEI-5k-Mal-B and 25,000 g/mol for PEI-25k-Mal-B) have been administered to both cell lines. As well dose as incubation-time dependent effects and interactions have been researched for concentrations between 1 µg/ml to 1 mg/ml and periods of 24 h up to 28 days. Studies conducted by different methods of microscopy as light microscopy, fluorescence microscopy, transmission-electron-microscopy and quantitative assays (LDH and DC-protein) indicate as well a good cellular uptake of the nanoparticles as a particle- and concentration-dependent impact on the cellular macro- and micro-structure of the rdMSC samples. In all experiments PEI-5k-Mal-B exhibits a superior biocompatibility compared to PEI-25k-Mal-B. At higher concentrations PEI-25k-Mal-B is toxic and induces a directly observable mitochondrial damage. The alkaline phosphatase assay (ALP), has been conducted to check on the possible influence of nanoparticles on the differentiation capabilities of rdMSC to osteoblasts. In addition the production of mineralized matrix has been shown by von-Kossa stained samples. No influence of the nanoparticles on the ALP per cell has been detected. Additionally, for all experiments, results are strongly influenced by a large donor-to-donor variability of the four different rdMSC samples. To summarize, while featuring a good cellular uptake, PEI-5k-Mal-B induces only minimal adverse effects and features clearly superior biocompatibility compared to the larger PEI-25k-Mal-B.

Regulation of acetylcholine receptors during differentiation of bone mesenchymal stem cells harvested from human reaming debris.

Acetylcholine (ACh) is an important signaling molecule in non-neuronal systems where it is involved in regulation of viability, proliferation, differentiation and migration of mesenchymal stem cells (MSC) that are capable to differentiate into osteoblasts, chondrocytes and adipocytes. Patients with the systemic disease osteoporosis show altered MSC properties, reduced bone formation and mineral density followed by increased bone fragility and high fracture incidence. Here we asked whether nicotinic and muscarinic acetylcholine receptors (AChR) are expressed in osteoblasts, adipocytes and chondrocytes differentiated from bone MSC extracted from human reaming debris (RDMSC) that was harvested during surgery of long bone diaphyseal fractures. Using RT-PCR, AChR were detected in RDMSC, osteoblasts, chondrocytes and adipocytes of male and female bone-healthy and of female osteoporotic donors. An up-regulation in multiplicity and occurrence of AChR subtypes was found in female compared to male donors and in osteoblast of male donors compared to adipocytes. Real-time RT-PCR analysis resulted in a significant increase in relative expression of nAChR α9 in chondrocytes compared to adipocytes of healthy female donors. The nAChR subunit α10 was significantly up-regulated in osteoblasts of healthy compared to osteoporotic donors as well as the mAChR M5 that is additionally decreased in osteoporotic osteoblasts compared to MSC and chondrocytes of osteoporotic donors. In summary, the gene expression of AChR during differentiation of RDMSC and its regulation in cells of osteoporotic donors lead to the assumption that AChR signaling is involved in bone formation and might be utilized to stimulate bone remodeling processes.

Enhanced osteogenesis on titanium implants by UVB photofunctionalization of hydrothermally grown TiO2 coatings.

Even though Ti-based implants are the most used materials for hard tissue replacement, they may present lack of osseointegration on the long term, due to their inertness. Hydrothermal treatment (HT) is a useful technique for the synthesis of firmly attached, highly crystalline coatings made of anatase titanium dioxide (TiO2), providing favorable nanoroughness and higher exposed surface area, as well as greater hydrophilicity, compared to the native amorphous oxide on pristine titanium. The hydrophilicity drops even more by photofunctionalization of the nanostructured TiO2-anatase coatings under UV light. Human mesenchymal stem cells exhibited a good response to the combination of the positive surface characteristics, especially in respect to the UVB pre-irradiation. The results showed that the cells were not harmed in terms of viability; even more, they were encouraged to differentiate in osteoblasts and to become osteogenically active, as confirmed by the calcium ion uptake and the formation of well-mineralized, bone-like nodule structures. In addition, the enrichment of hydroxyl groups on the HT-surfaces by UVB photofunctionalization accelerated the cell differentiation process and greatly improved the osteogenesis in comparison with the nonirradiated samples. The optimal surface characteristics of the HT-anatase coatings as well as the high potentiality of the photo-induced hydrophilicity, which was reached during a relatively short pre-irradiation time (5 h) with UVB light, can be correlated with better osseointegration ability in vivo; among the samples, the superior biological behavior of the roughest and most hydrophilic HT coating makes it a good candidate for further studies and applications.

Biomimetic PEG-catecholates for stabile antifouling coatings on metal surfaces: applications on TiO2 and stainless steel.

Trimeric catecholates have been designed for the stable immobilization of effector molecules on metal surfaces. The design of these catecholates followed a biomimetic approach and was inspired by natural multivalent metal binders, such as mussel adhesion proteins (MAPs) and siderophores. Three catecholates have been conjugated to central scaffolds based on adamantyl or trisalkylmethyl core structures. The resulting triscatecholates have been immobilized on TiO2 and stainless steel. In a proof of concept study we have demonstrated the high stability of the resulting nanolayers at neutral and slightly acidic pH. Furthermore, polyethylene glycol (PEG) conjugates of our triscatecholates have been synthesized and were immobilized on TiO2 and stainless steel. The PEG coated surfaces showed excellent antifouling properties upon exposure to human blood and bacteria as demonstrated by fluorescence microscopy, ellipsometry and a bacterial assay with Staphylococcus epidermidis. In addition, our PEG-triscatecholates showed no cytotoxicity against bone-marrow stem cells on TiO2.