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Davos, Switzerland

Czekanska E.M.,AO Research Institute Davos
Methods in molecular biology (Clifton, N.J.)

The Alamar Blue assay is based on enzymatic reduction of indicator dye by viable cells and serves as an effective tool for assessing cell proliferation and as a screening technique. It can be applied in studies concentrating on animal, plant, yeast, and bacteria cells. Among the various methods for cell viability and cytotoxicity, it utilises all features of ideal and reliable test; it is one-step, sensitive, safe, non-toxic for cells, and cost-effective. Source

BACKGROUND:: Application of platelet-rich plasma (PRP) and stem cells has become of importance in regenerative medicine. Recent literature supports the use of PRP as a cell culture media supplement to stimulate proliferation of adipose-tissue derived mesenchymal stem cells (ASCs). The underlying mechanism of proliferation stimulation by PRP has not been investigated so far. METHODS:: ASCs were cultured in α-MEM supplemented with PRP or fetal calf serum (FCS). Cell proliferation was assessed by means of cell cycle kinetics using flow cytometric analyses after 48 hours. Differences in proteome expression of ASCs exposed to PRP or FCS were analysed using a reverse phase protein array (RPPA) to quantify 214 proteins. Complementary Ingenuity Pathways Analyses (IPA) and gene set enrichment analyses (GSEA) were performed using protein data, and confirmed by Western Blot. RESULTS:: A higher percentage of ASCs in the S-phase in the presence of PRP advocates the proliferation stimulation. IPA and GSEA confirm the involvement of our selected proteins in the process of cell growth and proliferation. IPA revealed a participation in the top-ranked canonical pathways PI3K/AKT, PTEN, ILK, and IGF-1. GSEA identified our protein set as being part of significantly regulated protein sets with the focus on cell cycle, metabolism, and the KEGG TGF-ß Signaling pathway. CONCLUSION:: The present study provides evidence that the application of PRP stimulates proliferation and induces a unique change in the proteomic profile of ASCs. The interpretation of altered expression of regulatory proteins represents a step forward towards achieving good manufacturing practice-compliant criteria for cell-based strategies. ©2016American Society of Plastic Surgeons Source

Wagner D.,AO Research Institute Davos
Journal of orthopaedic research : official publication of the Orthopaedic Research Society

The complex anatomy of the sacrum makes surgical fracture fixation challenging. We developed statistical models to investigate sacral anatomy with special regard to trans-sacral implant fixation. We used computed tomographies of 20 intact adult pelves to establish 3D statistical models: a surface model of the sacrum and the trans-sacral corridor S1, including principal component analysis (PCA), and an averaged gray value model of the sacrum given in Hounsfield Units. PCA demonstrated large variability in sacral anatomy markedly affecting the diameters of the trans-sacral corridors. The configuration of the sacral alae and the vertical position of the auricular surfaces were important determinants of the trans-sacral corridor dimension on level S1. The statistical model of trans-sacral corridor S1 including the adjacent parts of the iliac bones showed main variation in length; however, the diameter was the main criterion for the surgically available corridor. The averaged gray value model revealed a distinct pattern of bone mass distribution with lower density particularly in the sacral alae. These advanced 3D statistical models provide a thorough anatomical understanding demonstrating the impact of sacral anatomy on positioning trans-sacral implants. © 2014 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. Source

Sakai D.,Tokai University | Sakai D.,Collaborative Research Partner Annulus Fibrosus Repair Program | Grad S.,AO Research Institute Davos | Grad S.,Collaborative Research Partner Annulus Fibrosus Repair Program
Advanced Drug Delivery Reviews

The healthy intervertebral disc (IVD) fulfils the essential function of load absorption, while maintaining multi-axial flexibility of the spine. The interrelated tissues of the IVD, the annulus fibrosus, the nucleus pulposus, and the cartilaginous endplate, are characterised by their specific niche, implying avascularity, hypoxia, acidic environment, low nutrition, and low cellularity. Anabolic and catabolic factors balance a slow physiological turnover of extracellular matrix synthesis and breakdown. Deviations in mechanical load, nutrient supply, cellular activity, matrix composition and metabolism may initiate a cascade ultimately leading to tissue dehydration, fibrosis, nerve and vessel ingrowth, disc height loss and disc herniation. Spinal instability, inflammation and neural sensitisation are sources of back pain, a worldwide leading burden that is challenging to cure. In this review, advances in cell and molecular therapy, including mobilisation and activation of endogenous progenitor cells, progenitor cell homing, and targeted delivery of cells, genes, or bioactive factors are discussed. © 2014 Elsevier B.V. Source

Stoddart M.J.,AO Research Institute Davos
Methods in molecular biology (Clifton, N.J.)

The measurement of cell viability plays a fundamental role in all forms of cell culture. Sometimes it is the main purpose of the experiment, such as in toxicity assays. Alternatively, cell viability can be used to -correlate cell behaviour to cell number, providing a more accurate picture of, for example, anabolic -activity. There are wide arrays of cell viability methods which range from the most routine trypan blue dye exclusion assay to highly complex analysis of individual cells, such as using RAMAN microscopy. The cost, speed, and complexity of equipment required will all play a role in determining the assay used. This chapter aims to provide an overview of many of the assays available today. Source

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