The mechanical properties of a cell, as determined by its actin cytoskeleton, are important for nanoneedle insertion into a living cell

Abstract

Previously, we reported that a nanoneedle of 200 nm diameter manipulated by an atomic force microscope apparatus could be inserted into a living cell. The insertion probabilities varied according to cell type. However, the nanoneedle was never successfully inserted into artificial liposomes. In the current study, we found that the stress fibers and actin filaments comprising the plasmalemmal undercoat are important, determining factors as to whether a nanoneedle can be successfully inserted into a cell. Depolymerization of microtubules increased both the number of stress fibers and insertion efficiency in NRK cells. These results indicate that the insertion efficiency of a nanoneedle (200 nm in diameter) into a cell with a smaller actin meshwork in its plasmalemmal undercoat is enhanced and the formation of stress fibers obviously contributes to this incremental enhancement. These facts are not only important as technical information to improve the efficiency of cell manipulation but also as observations of the mechanical properties of the native cell cortex.

Penetration of Cell Membranes and Synthetic Lipid Bilayers by Nanoprobes

“…AFM has been used to study the penetration of synthetic lipid bilayers (26)(27)(28)(36)(37)(38)(39)(40)(41)(42)(43)(44)(45) and live cells (9)(10)(11)(12)24,29,46,47) by nanoprobes, but the reported membrane penetration forces range from <1 nN to >20 nN (12,29), and the variety of probe types, lipids, and other experimental variables make comparisons across studies difficult.…”

“…To separate lipid bilayer penetration from other force-relaxation events, we used the breakthrough signatures in synthetic lipid stacks as a template for detecting penetration events in cells. Because synthetic and cellular bilayers differ considerably in their structure and composition, we also chose an intermediate system for comparison: cells whose cytoskeleton was chemically disrupted by cytochalasin D (10). Such cells have intact cell surfaces and native membrane composition, but no underlying cortical cytoskeleton to provide rigidity to the plasma membrane.…”

“…Nanofabricated devices, including nanostraws (1-3), nanowires (4-7), nanoneedles (8)(9)(10)(11)(12), and nanoelectrodes (13)(14)(15)(16)(17)(18)(19)(20)(21)(22), are increasingly being investigated as tools for cellular studies, but these structures do not readily insert through the cell membrane (2)(3)(4)(5)13,23), and assessing when (or whether) penetration has occurred is difficult due to the nanoscale features of the probe-membrane interface. To design cell-penetrating nanoprobes, a systematic approach is needed to describe nanostructure-membrane interactions at relevant temporal and spatial scales, particularly the processes of nanoprobe insertion through (3,(8)(9)(10)(11)(12)(13)15,18,24,25) or fusion with (14,16,19,(26)(27)(28) the plasma membrane.…”

Nanoscale Imaging and Mechanical Analysis of Fc Receptor-Mediated Macrophage Phagocytosis against Cancer Cells

“…52 Actin cytoskeleton determines the mechanical properties of a cell. 53 When macrophage phagocytosis was activated, a rapid accumulation of F-actin and associated proteins in the periphagosomal region occurred. 8 This phenomenon can be clearly seen in Figure 7A,B.…”

Mechanical Criterion for the Rupture of a Cell Membrane under Compression

“…This also is in reasonable agreement with our results, especially taking into account that incorporating peptides into lipid bilayers (hence getting a model somewhat closer to a cell membrane) makes them easier to break, according to micropipette aspiration (44) and AFM indentation (42) measurements. Kagiwada et al (45) reported that a typical force of 3 nN is required to insert a nanoneedle of 200 nm in diameter into a cell. Here, the contact area is critical when estimating a compressive force: although using Eq.…”