Figure 1.
Schematic of a protrusion induced on an adherent living cell using optical tweezers.
The protrusion forms in response to application of a tensile force (F) on the cell surface in y direction by a laser trapped microsphere (bead) (drawn not to scale). A quadrant photodetector records the instantaneous displacement of the trapped microsphere from the trapping center for force measurements while the stage is driven in the negative y direction.
Figure 2.
Schematic of bead-cell contact system.
Dashed lines show initial cell-bead contact where there is no external force applied on the bead. Solid lines show the bead-cell system once the cell is moved away from the trapped bead by distance xpzt. Bead is displaced from the trapping center by xbead and the length of the protrusion is xpt.
Figure 3.
Cell protrusions as a viscoelastic structure.
(A) Standard linear solid (SLS) model of the protrusion. The model includes a Maxwell arm (viscous coefficient of η0 in series with spring with stiffness k0) in parallel with a spring with stiffness k1. (B) Two illustrative force-length profiles of protrusion elongation fitted with the SLS model (blue solid lines). The red circles show data for a cell with intact F-actin and normal membrane cholesterol content. Data in black circles are from cells with disrupted F-actin using Latrunculin-A, but with and normal membrane cholesterol level.
Figure 4.
An example force-time plot involving subsequent “pull” and “push” processes. The ∼1 second time interval between zero second and point A indicates bead-cell contact prior to the push-pull experiments beginning at A.
Figure 5.
Effect of membrane cholesterol content and cytoskeletal F-actin on maximum protrusion force.
Maximum protrusion force (Fmax) is defined as the force that results in separation of membrane from cytoskeleton.
Figure 6.
Effect of membrane cholesterol content and cytoskeletal F-actin on maximum protrusion length.
Maximum length of protrusion (lpt-max) is defined the length at which the membrane becomes separated from the cytoskeleton.
Figure 7.
Effect of membrane cholesterol content on: (A) stiffness parameter k0, and (B) stiffness parameter k1 for cells with intact or disrupted F-actin.
Figure 8.
Effect of membrane cholesterol content on the viscosity coefficient of the protrusion for cells with intact or disrupted F-actin. Asterisk (*) indicates a statistically significant difference with control; ** indicates a statistically significant difference between the two indicated data sets; and + indicates a statistically significant difference between intact F-actin and F-actin disrupted cells at the same cholesterol concentration.
Table 1.
Plasma membrane and protrusion stiffness and viscosity.
Figure 9.
Energy loss associated with protrusion hysteresis.
(A) Effect of membrane cholesterol content on the percent of energy loss associated with protrusion hysteresis for intact and disrupted F-actin cells. (B) Force-length plots indicating hysteresis in cells with intact and disrupted F-actin. lpar;C) Protrusion force during protrusion elongation to 3 µm (time interval D–E) followed by force relaxation after the protrusion reaches 3 µm, and maintained at that length (time interval E–F). The interval F–G displays the force as the protrusion is unloaded (pushed back). The inset shows the SLS-fit to the force relaxation during the time interval between the end of the pulling time and beginning of the pushing time (E–F). The general solution for the protrusion force relaxation under constant length is: , where xpt is the constant length of the protrusion (3 µm), and F0 is the force at the beginning of relaxation. The ∼7 seconds time interval between zero second and point D indicates bead-cell contact prior to the push-pull experiments beginning at D.