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In tissue engineering and regenerative medicine mesenchymal stem cells (MSC) are widely used to replace and restore the function of dysfunctional or missing tissue. Recent studies have shown significant enhancements of the in vivo healing process following dentofacial bone augmentation procedures employing stem cell-loaded xenografts. We conducted experimental and numerical investigations in perfusion flow bioreactor-xenograft-systems to identify flow conditions as well as bioreactor design features that allow for homogeneous MSC-distribution in Geistlich Bio- Oss Block xenografts. Pressure gradient - velocity characteristics and flow distributions were investigated experimentally and numerically for two bioreactor designs at steady-state flow conditions with Reynolds numbers (Re) ranging from 0.01 ≤ Re ≤ 0.32. Distilled water at 20°C with a dynamic viscosity of 1.002 mPa∙s and a density of 998 kg/m3 was used. The geometry of the xenograft utilized in three-dimensional computational fluid dynamics (CFD) simulation was obtained by means of micro-computed tomography (μCT) at an isotropic spatial resolution of 9.5 μm. The permeability values calculated from the experimental data are in good accordance with the numerical results. The investigations showed that the increase of the inflow- and outflow-area diameter, as well as the decrease of the volumetric flow rate, result in a decreasing heterogeneity of the flow distribution within the xenograft. The calculated wall shear stress rates in the three-dimensional (3D) scaffold range from 1∙10-12Pa ≤ τ ≤ 0.2 Pa. Experimentally validated CFD simulations introduced in this study provide an applicable tool to assess optimal flow conditions for homogeneous MSC distribution in bioreactor-xenograft-systems.

Background Regarding strategies improving the efficacy of stem cell transplantation in adipose tissue engineering, cell tracking might be useful. Here we report the in vivo tracking of adipose tissue derived stem cells (ASC) by means of nanoparticle labeling and magnetic resonance imaging (MRI). Here we report the in vivo tracking of adipose tissue derived stromal cells (ASC) by means of nanoparticle labeling and magnetic resonance imaging (MRI). Materials and methods Human ASC were amplified and labeled with two types of magnetic nanoparticles (MNP), BNF starch and nanomag®- D -spio. Adipose tissue constructs were fabricated by seeding collagen scaffolds with labeled and unlabeled ASCs. Constructs were implanted subcutaneously in the back of severe combined immunodeficient (SCID) mice (n =69, group 1: control with cells w/o label, group 2: BNF starch labeled cells, group 3: nanomag®- D -spio labeled cells). MRI scans were performed at 24 hours, four, twelve and 28 days and four months in a 7.1 T animal device. Explanted constructs were analyzed histomorphometrically. Results MRI scans showed high contrast of the labeled cells in t2-tse-sequence compared to unlabeled controls. Loss of volume of the implants was observed over time due to partial loss for transplanted cells without significant difference (level of significance p Additional Prussian blue stain showed iron in all implants. Significant differences between the three groups (significance level p Conclusion Both MNPs might be suitable for tracking of ASC in vivo and show long term stability over 4 months.

Objective There is a high demand for temporary wound dressings that improve wound healing and regeneration. Silicon (as SiO 2 ) has been shown to support the growth and collagen formation in biological systems. Methods A nanocomposite was made from PVP (polyvinylpyrrolidon), nano-sized silica aggregates and water and served for fabrication of a wet dressing material (SiO 2 –PVP gel, by cross-linking the gel) and a freeze-dried dressing material (SiO 2 –PVP fleece). Materials were characterized by SAXS, DSC, EDX and viscosity measurements. A 10 mm circular defect was set on both sides of the back of SKH1-hr mice (n = 40) and both dressing materials were compared with untreated controls. After 3, 6, 9, 12 and 15 days, the defect regions were explanted and evaluated by histomorphometric measurements and CD31-immunohistochemistry. Results The microstructure of the compound was composed of fiber like structures. SiO 2 nano-aggregates inside the composite remained stable and embedded in a rigid amorphous PVP fraction. In animal experiments, all groups showed a non-irritated defect closure after 9 days. EDX of SiO 2 –PVP gel and fleeces revealed SiO 2 –PVP diffusion into the wound. Wound contraction was significantly enhanced after treatment with SiO 2 –PVP gel followed by SiO 2 –PVP fleece compared to controls. Re-epithelialization was increased in SiO 2 –PVP treated wounds and the regenerated epidermis showed a well-differentiated layer structure compared to untreated controls. Conclusions The results indicate that silica diffuses from the dressing into the wound. Both dressings affect the wound healing. The SiO 2 -based wound dressing may counteract scarring and might be suitable as a temporary wound dressing.

Magnetic resonance imaging (MRI) is to be considered as an emerging detection technique for cell tracking experiments to evaluate the fate of transplanted progenitor cells and develop successful cell therapies for tissue engineering. Adipose tissue engineering using adipose tissue-derived progenitor cells has been advocated for the cure of soft tissue defects or for persistent soft tissue augmentation. Adipose tissue-derived progenitor cells were differentiated into the adipogenic lineage and labeled with two different types of magnetic iron oxide nanoparticles in varying concentrations which resulted in a concentration-dependent reduction of gene expression of adipogenic differentiation markers, adiponectin and fatty acid-binding protein 4 (FABP4), whereas the metabolic activity was not altered. As a result, only low nanoparticle concentrations for labeling were used for in vivo experiments. Cells were seeded onto collagen scaffolds and subcutaneously implanted into severe combined immunodeficient (SCID) mice. At 24 h as well as 28 days after implantation, MRI analyses were performed visualizing nanoparticle-labeled cells using T2-weighted sequences. The quantification of absolute volume of the scaffolds revealed a decrease of volume over time in all experimental groups. The distribution of nanoparticle-labeled cells within the scaffolds varied likewise over time.

Aim: We aimed to analyze the suitability of nanoparticles (M4E) for safe human mesenchymal stem cell (hMSC) labeling and determined cell labeling maintenance in 2D and 3D culture. Materials & methods: We investigated cell–particle interaction and the particles’ impact on cell viability, growth and proliferation. We analyzed cell labeling maintenance in 2D and 3D culture invasively and noninvasively. Results: M4E do not affect cell viability, growth and proliferation and do not cause chromosomal aberrations. Cell labeling maintenance is up to five-times higher in 3D conditions compared with 2D culture. Conclusion: M4E allow safe hMSC labeling and noninvasive identification. Our hMSC-loaded, 3D tissue-engineered construct could serve as a graft for regenerative therapies, in which M4E-labeled hMSCs can migrate to their target.

The control of mesenchymal stem cells (MSC) by physical cues is of great interest in regenerative medicine. Because integrin receptors function as mechanotransducers, we applied drag forces to β1 integrins on the apical surface of adherent human MSC. In addition to mechanical forces, the technique we used involved also the exposure of the cells to an inhomogeneous magnetic field. In order to assess the influence of the substrate on cell adhesion, cells were cultured on plain tissue culture polystyrene (TCP) or on coated well plates, which allowed only adhesion to embedded fibronectin or RGD peptides. We found that the expression of collagen I, which is involved in osteogenesis, and VEGF, a factor which stimulates angiogenesis, increased as a result of short-term mechanical integrin stress. Whereas, collagen I expression was stimulated by mechanical forces when the cells were cultured on fibronectin and RGD peptides but not on TCP, VEGF expression was enhanced by physical stimulation on TCP. The study further revealed that magnetic forces enhanced Sox 9 expression, a marker of chondrogenesis, and reduced the expression of ALP. Concerning the intracellular mechanisms involved, we found that the expression of VEGF induced by physical forces depended on Akt activation. Together, the results implicate that biological functions of MSC can be stimulated by integrin-mediated mechanical forces and a magnetic field. However, the responses of cells depend strongly on the substrate to which they adhere and on the cross-talk between integrin-mediated signals and soluble factors.

Designing of implant surfaces using a suitable ligand for cell adhesion to stimulate specific biological responses of stem cells will boost the application of regenerative implants. For example, materials that facilitate rapid and guided migration of stem cells would promote tissue regeneration. When seeded on fibronectin (FN) that was homogeneously immmobilized to NCO-sP(EO-stat-PO), which otherwise prevents protein binding and cell adhesion, human mesenchymal stem cells (MSC) revealed a faster migration, increased spreading and a more rapid organization of different cellular components for cell adhesion on fibronectin than on a glass surface. To further explore, how a structural organization of FN controls the behavior of MSC, adhesive lines of FN with varying width between 10 mu m and 80 mu m and spacings between 5 mu m and 20 mu m that did not allow cell adhesion were generated. In dependance on both line width and gaps, cells formed adjacent cell contacts, were individually organized in lines, or bridged the lines. With decreasing sizes of FN lines, speed and directionality of cell migration increased, which correlated with organization of the actin cytoskeleton, size and shape of the nuclei as well as of focal adhesions. Together, defined FN lines and gaps enabled a fine tuning of the structural organization of cellular components and migration. Microstructured adhesive substrates can mimic the extracellular matrix in vivo and stimulate cellular mechanisms which play a role in tissue regeneration.

Magnetic resonance imaging (MRI) using measurement of the transverse relaxation time (R2*) is to be considered as a promising approach for cell tracking experiments to evaluate the fate of transplanted progenitor cells and develop successful cell therapies for tissue engineering. While the relationship between core composition of nanoparticles and their MRI properties is well studied, little is known about possible effects on progenitor cells. This in vitro study aims at comparing two magnetic iron oxide nanoparticle types, single vs. multi-core nanoparticles, regarding their physico-chemical characteristics, effects on cellular behavior of adipose tissue-derived stem cells (ASC) like differentiation and proliferation as well as their detection and quantification by means of MRI. Quantification of both nanoparticle types revealed a linear correlation between labeling concentration and R2* values. However, according to core composition, different levels of labeling concentrations were needed to achieve comparable R2* values. Cell viability was not altered for all labeling concentrations, whereas the proliferation rate increased with increasing labeling concentrations. Likewise, deposition of lipid droplets as well as matrix calcification revealed to be highly dose-dependent particularly regarding multi-core nanoparticle-labeled cells. Synthesis of cartilage matrix proteins and mRNA expression of collagen type II was also highly dependent on nanoparticle labeling. In general, the differentiation potential was decreased with increasing labeling concentrations. This in vitro study provides the proof of principle for further in vivo tracking experiments of progenitor cells using nanoparticles with different core compositions but also provides striking evidence that combined testing of biological and MRI properties is advisable as improved MRI properties of multi-core nanoparticles may result in altered cell functions.