Anne Neumann

BMP2-loaded nanoporous silica nanoparticles promote osteogenic differentiation of human mesenchymal stem cells

Nanoporous (or mesoporous) silica materials have been extensively investigated as a novel biomaterial in the last few years, especially with regard to bone tissue engineering. Due to their high specific surface areas, capability for chemical modification, and their large pore volumes, nanoporous silica nanoparticles (NPSNPs) are of tremendous interest as drug delivery systems. In this study, we describe the immobilization of the bone growth factor BMP2 on NPSNPs in biologically relevant amounts, using coupling via an aminosilane linker. The amount of BMP2 loaded onto NPSNPs was determined using two different methods. The biocompatibility of NPSNPs was tested with different cell lines upon exposure to different particle concentrations. Whereas standard cell types (HepG2, NIH3T3) are relatively insensitive against NPSNPs, adMSC cells (adipose-derived human mesenchymal stem cells) show a somewhat reduced viability. adMSC cells were also used to test for the osteoinductive effect of the BMP2-carrying NPSNPs. Histological staining reveals the osteogenic differentiation of the human mesenchymal stem cells. Even BMP-free amino-modified NPSNPs show a certain osteoinductive effect, which is however significantly stronger with immobilized BMP2 and can furthermore be enhanced by supplemental dexamethasone. We therefore suggest that NPSNPs carrying BMP2 are suitable for application in bone tissue engineering, e.g. in the construction of scaffolds.

Stem Cell Differentiation Depending on Different Surfaces

Mesenchymal stem cells and 3D biomaterials are a potent assembly in tissue engineering. Today, a sizable number of biomaterials has been characterized for special tissue engineering applications. However, diverse material properties, such as soft or hard biomaterials, have a specific influence on cell behavior. Not only the cell attachment and proliferation, but also differentiation is controlled by the microenvironment. Material characteristics such as pore size, stiffness, roughness, and geometry affect not only the cell attachment and proliferation, but also the differentiation behavior of mesenchymal stem cells. Optimization of these features might enable direct differentiation without adjustment of the culture medium by applying expensive growth or differentiation factors. Future aspects include the design of multilayered biomaterials, where each zone fulfills a distinct function. Moreover, the embedding of growth and differentiation factors into the matrix with a controlled release rate might be advantageous to direct differentiation.

Potential for Osteogenic and Chondrogenic Differentiation of MSC

The introduction of mesenchymal stem cells (MSC) into the field of tissue engineering for bone and cartilage repair is a promising development, since these cells can be expanded ex vivo to clinically relevant numbers and, after expansion, retain their ability to differentiate into different cell lineages. Mesenchymal stem cells isolated from various tissues have been intensively studied and characterized by many research groups. To obtain functionally active differentiated tissue, tissue engineered constructs are cultivated in vitro statically or dynamically in bioreactors under controlled conditions. These conditions include special cell culture media, addition of signalling molecules, various physical and chemical factors and the application of different mechanical stimuli. Oxygen concentration in the culture environment is also a significant factor which influences MSC proliferation, stemness and differentiation capacity. Knowledge of the different aspects which affect MSC differentiation in vivo and in vitro will help researchers to achieve directed cell fate without the addition of differentiation agents in concentrations above the physiological range.

Adipose-derived stem cells cultivated on electrospun L-lactide/glycolide copolymer fleece and gelatin hydrogels under flow conditions - aiming physiological reality in hypodermis tissue engineering

INTRODUCTION Generation of adipose tissue for burn patients that suffer from an irreversible loss of the hypodermis is still one of the most complex challenges in tissue engineering. Electrospun materials with their micro- and nanostructures are already well established for their use as extracellular matrix substitutes. Gelatin is widely used in tissue engineering to gain thickness and volume. Under conventional static cultivation methods the supply of nutrients and transport of toxic metabolites is controlled by diffusion and therefore highly dependent on size and porosity of the biomaterial. A widely used method in order to overcome these limitations is the medium perfusion of 3D biomaterial-cell-constructs. In this study we combined perfusion bioreactor cultivation techniques with electrospun poly(l-lactide-co-glycolide) (P(LLG)) and gelatin hydrogels together with adipose-derived stem cells (ASCs) for a new approach in soft tissue engineering. METHODS ASCs were seeded on P(LLG) scaffolds and in gelatin hydrogels and cultivated for 24 hours under static conditions. Thereafter, biomaterials were cultivated under static conditions or in a bioreactor system for three, nine or twelve days with a medium flow of 0.3ml/min. Viability, morphology and differentiation of cells was monitored. RESULTS ASCs seeded on P(LLG) scaffolds had a physiological morphology and good viability and were able to migrate from one electrospun scaffold to another under flow conditions but not migrate through the mesh. Differentiated ASCs showed lipid droplet formations after 21 days. Cells in hydrogels were viable but showed rounded morphology. Under flow conditions, morphology of cells was more diffuse. DISCUSSION ASCs could be cultivated on P(LLG) scaffolds and in gelatin hydrogels under flow conditions and showed good cell viability as well as the potential to differentiate. These results should be a next step to a physiological three-dimensional construct for soft tissue engineering and regeneration.

Nanoporous silica nanoparticles as biomaterials: evaluation of different strategies for the functionalization with polysialic acid by step-by-step cytocompatibility testing.

Nanoporous silica materials have become a prominent novel class of biomaterials which are typically applied as nanoparticles or thin films. Their large surface area combined with the rich surface chemistry of amorphous silica affords the possibility to equip this material with variable functionalities, also with several different ones on the same particle or coating. Although many studies have shown that nanoporous silica is apparently non-toxic and basically biocompatible, any surface modification may change the surface properties considerably and, therefore, the modified materials should be checked for their biocompatibility at every step. Here we report on different silane-based functionalization strategies, firstly a conventional succinic anhydride-based linker system and, secondly, copper-catalyzed click chemistry, to bind polysialic acid, a polysaccharide important in neurogenesis, onto nanoporous silica nanoparticles (NPSNPs) of MCM-41 type. At each of the different modification steps, the materials are characterized by cell culture experiments. The results show that polysialic acid can be immobilized on the surface of NPSNPs by using different strategies. The cell culture experiments show that the kind of surface immobilization has a strong influence on the toxicity of the material versus the cells. Whereas most modifications appear inoffensive, NPSNPs modified by click reactions are toxic, probably due to residues of the Cu catalyst used in these reactions.

Combined effect of grain refinement and surface modification of pure titanium on the attachment of mesenchymal stem cells and osteoblast-like SaOS-2 cells

Surface modification is an important step in production of medical implants. Surface roughening creates additional surface area to enhance the bonding between the implant and the bone. Recent research provided a means to alter the microstructure of titanium by severe plastic deformation (SPD) in order to increase its strength, and thereby reduce the size of the implants (specifically, their diameter). The purpose of the present study was to examine the effect of bulk microstructure of commercially pure titanium with coarse-grained (CG) and ultrafine-grained (UFG) bulk structure on the surface state of these materials after surface modification by sand blasting and acid etching (SLA). It was shown that SLA-modified surface characteristics, in particular, roughness, chemistry, and wettability, were affected by prior SPD processing. Additionally, biocompatibility of UFG titanium was examined using osteosarcoma cell line SaOS-2 and primary human adipose-derived mesenchymal stem cell (adMSC) cultures. Enhanced cell viability as well as increased matrix mineralization during osteogenic differentiation of MSCs on the surface of ultrafine-grained titanium was shown.