Cortical stiffness measurement in VSMCs.
(A) Controlled force pulses generated by magnetic tweezers displace aortic VSMC-adherent, RGD-coated superparamagnetic beads (2.8 µm) to measure the stiffness of the bead-focal adhesion-cortical cytoskeleton linkage (see Methods). The force F exerted on a bead depends on the induced magnetic moment in the bead m and the spatial gradient of the magnetic field B, which depends critically on the sharpness of the probe’s tip, characterized by its radius of curvature R. (B) Calibration curve relating the force exerted on a bead (d = 2.8 µm) to its distance from the MT tip (R = 40 µm) and the current through the electromagnet solenoid (I = 1.5 A). The gray box denotes the operating range used for the MT experiments, i.e. the distance that is set between the MT tip and a bead before pulling commences. (C) Mean cortical stiffness increases with LPA stimulation in a PP2-attenuated manner. Right: BSA beads, which do not bind integrins but adhere nonspecifically, do not register an increase in stiffness with LPA. *p<0.05, **p<0.01, n.s. – not significant, unpaired, two-tailed Student’s t-test, assuming unequal variances.