Cortical stiffness of keratinocytes measured by lateral indentation with optical tweezers

Abstract

Keratin intermediate filaments are the principal structural element of epithelial cells. Their importance in providing bulk cellular stiffness is well recognized, but their role in the mechanics of cell cortex is less understood. In this study, we therefore compared the cortical stiffness of three keratinocyte lines: primary wild type cells (NHEK2), immortalized wild type cells (NEB1) and immortalized mutant cells (KEB7). The cortical stiffness was measured by lateral indentation of cells with AOD-steered optical tweezers without employing any moving mechanical elements. The method was validated on fixed cells and Cytochalasin-D treated cells to ensure that the observed variations in stiffness within a single cell line were not a consequence of low measurement precision. The measurements of the cortical stiffness showed that primary wild type cells were significantly stiffer than immortalized wild type cells, which was also detected in previous studies of bulk elasticity. In addition, a small difference between the mutant and the wild type cells was detected, showing that mutation of keratin impacts also the cell cortex. Thus, our results indicate that the role of keratins in cortical stiffness is not negligible and call for further investigation of the mechanical interactions between keratins and elements of the cell cortex.

Antimicrobial peptide nisin induces spherical distribution of macropinocytosis-like cytokeratin 5 and cytokeratin 17 following immediate derangement of the cell membrane

“…After culturing for 2 days, cells were treated with a drug for 30 minutes or 24 hours. Either 93 μg/ml nisin or 93 μg/ml nisin with 10 μM cytochalasin D was added to the culture medium, as previously reported [ 8 , 40 , 41 ]. Cells grown in a medium containing 0.0015% acetic acid was used as the control.…”

A mathematical model for the dependence of keratin aggregate formation on the quantity of mutant keratin expressed in EGFP-K14 R125P keratinocytes

“…In the present study, we looked further into the topic of keratin aggregates formation and to the possible mechanisms involved. We addressed this by observing the effect that different ratios between mutant and WT keratin proteins expressed in cultured keratinocytes may have on aggregate formation in vitro , and how keratin aggregate formation affects the mechanical properties of cells at the cell cortex [ 14 ]. This is an important question, as although the vast majority of EBS cases are inherited in a dominant negative fashion, the actual ratio between WT and mutant keratin protein expressed in cells is not known, and this in turn may also affect the patient’s phenotype.…”