Affiliations: Medical Engineering Division, Department of Engineering, Queen Mary University of London, London, UK | Graduate School of Engineering, Tohoku University, Japan | Graduate School of Systems Life Sciences, Kyusyu University, Japan
Note: [] Address for correspondence: Dr. Martin Knight, Medical Engineering Division, Department of Engineering, Queen Mary University of London, Mile End Rd., London, E1 4NS, UK. Tel.: +44 207 882 5512; E-mail: [email protected].
Abstract: The present study utilised pipette aspiration and simultaneous confocal microscopy to test the hypothesis that chondrocyte deformation is associated with distortion of intracellular organelles and activation of calcium signalling. Aspiration pressure was applied to isolated articular chondrocytes in increments of 2 cm of water every 60 seconds up to a maximum of 10 cm of water. At each pressure increment, confocal microscopy was used to visualise the mitochondria and nucleus labelled with JC-1 and Syto-16, respectively. To investigate intracellular calcium signalling, separate cells were labelled with Fluo 4, rapidly aspirated to 5 cm of water and then imaged for 5 minutes at a tare pressure of 0.1 cm of water. Partial cell aspiration was associated with distortion of the mitochondrial network, elongation of the nucleus and movement towards the pipette mouth. Treatment with cytochalasin D or nocodazole produced an increase in cell aspiration indicating that both the actin microfilaments and microtubules provide mechanical integrity to the cell. When the data was normalised to account for the increased cell deformation, both actin microfilaments and microtubules were shown to be necessary for strain transfer to the intracellular organelles. Mitochondria and nucleus deformation may both be involved in chondrocyte mechanotransduction as well as cellular and intracellular mechanics. In addition, pipette aspiration induced intracellular calcium signalling which may also form part of a mechanotransduction pathway. Alternatively calcium mobilisation may serve to modify actin polymerisation, thereby changing cell mechanics and membrane rigidity in order to facilitate localised cell deformation. These findings have important implications for our understanding of cell mechanics and mechanotransduction as well as interpretation and modelling of pipette aspiration data.
