Investigation of the mechano-sensitivity of osteoblasts by combined atomic-force fluorescence microscopy and optical flow stress analysis

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Our goal is to analyse the reception of mechanical stimuli by bone cells (osteoblasts) which is a key process for the understanding of osteoporosis. We use a combination of an atomic force apparatus with a fluorescence microscope to investigate the reaction of cells onto application of well defined mechanical forces. Forces applied by the cells to the substrate, either self applied or forces transmitted through the cell by the AFM, are analysed by a type of finite element analysis of the substrate distortion. Any reactions of the cell upon mechanical stimulation involve variations of the free intracellular calcium concentration. Therefore, we detect changes in the intracellular free calcium concentration by fluorescent imaging as an unambiguous indicator for a cellular reaction, i.e. whether a cell reacts to a particular stimulus or not. Our central goals are to determine the optimum stimulation conditions (e.g. stimulation frequency) and to locate the mechanosensor in the cells. We will analyse the forces and energies required for stimulation and compare them to the tension forces of the cell. Moreover, we will investigate the origin of the calcium ions causing the intracellular free calcium increase after stimulation and analyse the communication of adjacent cells in order to contribute to the understanding of the whole complex mechanoreception process. We will perform comparative studies on different cell lines to work out the specific function of osteoblasts in the reception of mechanical stimuli.