Effects of Plasma Membrane Cholesterol Level and Cytoskeleton F-Actin on Cell Protrusion Mechanics

Abstract

Protrusions are deformations that form at the surface of living cells during biological activities such as cell migration. Using combined optical tweezers and fluorescent microscopy, we quantified the mechanical properties of protrusions in adherent human embryonic kidney cells in response to application of an external force at the cell surface. The mechanical properties of protrusions were analyzed by obtaining the associated force-length plots during protrusion formation, and force relaxation at constant length. Protrusion mechanics were interpretable by a standard linear solid (Kelvin) model, consisting of two stiffness parameters, k 0 and k 1 (with k 0>k 1), and a viscous coefficient. While both stiffness parameters contribute to the time-dependant mechanical behavior of the protrusions, k 0 and k 1 in particular dominated the early and late stages of the protrusion formation and elongation process, respectively. Lowering the membrane cholesterol content by 25% increased the k 0 stiffness by 74%, and shortened the protrusion length by almost half. Enhancement of membrane cholesterol content by nearly two-fold increased the protrusion length by 30%, and decreased the k 0 stiffness by nearly two-and-half-fold as compared with control cells. Cytoskeleton integrity was found to make a major contribution to protrusion mechanics as evidenced by the effects of F-actin disruption on the resulting mechanical parameters. Viscoelastic behavior of protrusions was further characterized by hysteresis and force relaxation after formation. The results of this study elucidate the coordination of plasma membrane composition and cytoskeleton during protrusion formation.

Understanding the role of cholesterol in cellular biomechanics and regulation of vesicular trafficking: The power of imaging

“…These results indicated that interaction of the actin cortical cytoskeleton was stronger with membranes containing lower amount of cholesterol. A follow up work from the same group, now also using cells treated with latrunculin, confirmed previous results showing that cholesterol disruption alters membrane interaction with the actin cytoskeleton leading to increased cell stiffness [57]. These results also confirmed that both plasma membrane and cytoskeleton contribute to the viscoelastic behavior observed for the membrane tethers.…”

“…Additionally and most important, they showed that the effects of plasma membrane itself in mechanical properties of cells are relatively minor in comparison with the effects of the cytoskeleton. On the other hand, changes in plasma membrane content, as in the case of cholesterol, might induce changes in the cytoskeleton and therefore indirectly contribute to cellular mechanical properties [57]. These results were corroborated by a work from our group, showing that the levels of the plasma membrane cholesterol content influence cell rigidity [51].…”

3,4-dihydroxyphenylacetic acid, a microbiota-derived metabolite of quercetin, protects against pancreatic β-cells dysfunction induced by high cholesterol

“…Cholesterol: methyl-β-cyclodextrin complex (Sigma catalog no. C4951), a "water-soluble cholesterol" containing 47 mg of cholesterol/g solid according to Certificate of Analysis (molar ratio, 1:6 cholesterol/ methyl-β-cyclodextrin) was used to deliver cholesterol to the cells, as previously described [9,32,33]. Considering that methyl-β-cyclodextrin induces mitochondrial cholesterol depletion from cell membranes at very high concentration ($ 2% or 5 mM) [32,34], a ten times lower concentration of this compound in our experiments.…”

Impact of lipid rafts on theT-cell-receptor and peptide-major-histocompatibility-complex interactions under different measurement conditions

“…Thus the raft-disrupting agent, MβCD, plays a negative regulation role in the 2D receptor-ligand interactions from this perspective. However, more studies reveal that the effect of cholesterol depletion on the membrane bending rigidity is not universal but rather depends on the cell type and lipid architecture [59][60][61][62]. Therefore, the potential contributions of MβCD to the receptor-ligand interactions may be specific to the type of cells and lipids.…”