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Contents IV 3.1.

Contents IV 3.1. Characterisation of hMSCs ............................................................................18 3.1.1. Osteogenic differentiation .......................................................................18 3.1.2. Adipogenic differentiation........................................................................22 3.1.3. Surface receptor expression ...................................................................24 3.2. Biological cell characteristics..........................................................................25 3.2.1. Cumulative population doubling ..............................................................25 3.2.2. Metabolic activity.....................................................................................27 3.2.3. Cell morphology ......................................................................................28 3.2.4. Cell volume measurement ......................................................................32 3.2.5. Time-laps analysis of cell behaviour .......................................................36 3.2.6. Integrin profile .........................................................................................38 4. Discussion........................................................................................ 39 5. Conclusions and Outlook ............................................................... 46 6. References ....................................................................................... 47 7. Supplementary data ........................................................................ 56 8. Appendix .......................................................................................... 58 List of abbreviations............................................................................... 61 List of figures ......................................................................................... 63 List of tables........................................................................................... 66 Acknowledgments.................................................................................. 67

Introduction 1 1. Introduction The first mammalian stem cells were described in the early 1960s 1,2 . Since then, a variety of totipotent, pluripotent or multipotent stem cells were discovered 3 . The most seldom stem cells are the totipotent cells which only appear during the first few cell division after fertilisation of an oocyte by a spermatozoon. These rare cells can differentiate into all cell types of an adult organism and any cell type of extraembryonic membranes (EEM). Pluripotent cells in contrast lost the capability to differentiate into EEM cell types. The proceeding cellular differentiation and maturation during embryonic development causes a specialisation of the existing stem cells. These multipotent stem cells can still differentiate into various cellular lineages, such as the hematopoietic and mesenchymal lineage 4 and are the only present stem cells in adult vertebrates. Severe traumata with great loss of tissues require a global body reaction with interplay of cell proliferation, cell migration, cytokines and growth factors. Healing of such injuries is very demanding for modern medicine. Great expectations are on the regeneration potential of adult stem cells, i.e. for the treatment of large bone defects. Up to date, the most common treatment of large segmental bone defects is the replacement with autologous corticocancellous bone grafts 5 . This technique has an eligible clinical outcome, but also has serious drawbacks such as increased surgical procedures, postoperative morbidity, limited availability and high costs for the socio-economic system 6 . A possible alternative is tissue engineering by using adult hMSCs. 1.1. Mesenchymal stem cells and their clinical application In the last decades, modern medicine has made great steps to treat various diseases and defects. Nevertheless, there are still untreatable diseases such as Parkinson’s disease, corneal blindness, sarcomas or large bone defects. This loss or failure of adult organs is one of the most challenging problems in human health care 7 . A back door out of this misery might be the emerging field of Tissue Engineering 8,9 .

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