IntroductionMechanosensing 

Forces in Cell Motility and Mechanical Sensing of Cells

We will investigate lamellipodia extension of motile cells as an essential initial step in cell motility. As a first cell model we will focus on the neuronal growth cone. The ability of a neuron's growth cone to migrate actively through tissue is essential to nerve regeneration. We will focus on the forces generated by the leading edge (lamellipodium) of a nerve's growth cone and its sensing of mechanical stresses. Our novel AFM-based microrheology technique is perfectly suited to measure these forces and to trigger mechanotransduction events. To determine to what extent the forces generated by the leading edge of motile cells are driven by thermal-ratchet-like, polymerization-based processes, or are caused by molecular motors, we will use our new technique to compare PC12 cells with normal and with down-regulated myosin activity, as well as PC12 cells whose actin polymerization and depolymerization rates are slowed down. Indeed, our preliminary results on the lamellipodia extension of cells show strong evidence for a thermal ratchet-like force-generating mechanism. Furthermore, we have recently found that the leading edge of the growth cone retracts in response to quick mechanical stimuli at a threshold deforming stress of about 300 Pa and subsequently grows in another random direction. The low threshold relative to the mechanical strength of the growth cone (3000 Pa) indicates that these cells cannot push past soft obstacles. This may be a sign that growth cones can only generate low protrusion forces. The observed mechanosensing at the growth cone is caused by stress-induced Ca²⁺-channels. The Ca²⁺-influx activates gelsolin and depolymerizes the actin cytoskeleton, which results in retraction. We also investigate how variations in the elasticity of the lamellipodium impact cell migration. Our preliminary measurements show a clear decrease in the Young's modulus from the leading edge of normal fibroblasts towards their main cell body. In malignantly transformed fibroblasts, the Young's modulus at the leading edge is distinctively lower and a weak decrease of the Young's modulus towards the cell body is observed since the elastic strength at the cell body is comparable to the value of normal cells. In a Boyden chamber assay, the transformed cells exhibit a drastically increased ability to migrate, and in agreement with this finding they also showed an increased speed of lamellipodia extension. We will now extend this study to different metastatic breast cancer cell lines to see whether a higher elastic strength of the lamellipodium reduces the metastatic potential.