Freely Suspended Actin Cortex Models on Arrays of Microfabricated Pillars

Abstract

Numerous chemomechanical processes of cells, such as pseudopod formation during cell locomotion [1] and the capping process preceding the engulfment of pathogens by macrophages during immunological responses, [2,3] are mediated by the actin-based cytoskeleton. In quiescent cells, the cytoskeleton consists of a partially crosslinked network of actin filaments forming a shell, several hundreds of nanometers thick, called the actin cortex. On the other hand, the activation of cells (for example, the endothelial cells lining the inner walls of blood vessels, by inflammation-mimicking agents such as thrombin) often leads to the formation of actin bundles coexisting within the random actin network. [2, 4] Several families of actin-manipulating proteins control the structure and viscoelastic properties of the actin cytoskeleton. These proteins include: 1) sequestering molecules which control the fraction of polymerized actin; 2) severing proteins which control the filament length; and 3) linker proteins mediating crosslinking between actin filaments and their coupling to membranes.Several studies of the structural reorganization of the actinbased cytoskeleton during pseudopod formation, [5] the centripetal contraction of endothelial cells by inflammation signalers, such as thrombin, [6] and the formation of focal contacts to stabilize cell adhesion [7, 8] have yielded some insight into the regulation of the actin cytoskeleton by biochemical signaling. Micromechanical studies of cell membranes have provided some information on the correlation between the viscoelastic behavior of cells and the structure of the actin cortex, [9] and on its role for the generation of forces. [10] of 700 mV in 0.2 M LiClO 4 0.1 M monomer (pyrrole or 1-methylpyrrole) acetonitrile solution.2) By consecutive square waves of potential (3 seconds at À 300 mV followed by 8 seconds at 700 mV and back to À 300 mV until established polymerization charge (oxidation minus reduction charge) was completed. Polythiophene and poly(3-methylthiophene) were generated in 0.2 M LiClO 4 0.1 M monomer (thiophene or 3-methylthiophene) acetonitrile solution by those consecutive potential steps.3) By potential cycling (polyaniline and poly(methylaniline) films were grown by potential cycling at 50 mVs À1 between À 100 and 900 mV in 0.2 M aniline 0.1 M H 2 SO 4 aqueous solutions, until the overall polymerization charge was completed.The thickness of the obtained films was around 0.22 mm, according to SEM observations of the dry films. After electrogeneration each polymer-coated electrode was rinsed with acetonitrile and transferred into the solution in which both electrochemical and electrochromic control would take place: 0.2 M lithium perchlorate acetonitrile solutions. There the electrodes were reduced at the compaction potential (E c ) for 1 min. Each of the studied polymers has a different value for its closing potential [22] (E s ) and the selected compaction potentials, À 600 mV for polythiophene, À 900 mV for poly(3-methylthiophene), À 1800 mV for p...

Microstructured Blood Vessel Surrogates Reveal Structural Tropism of Motile Malaria Parasites

“…The PDMS micropillar substrates were then made by photolithographic and replicate molding techniques essentially as described in. [7,39] To allow wetting of the hydrophobic pillar arrays they were first incubated with an alkaline solution containing 10% Extran (Merck) in water for 20 minutes and gently shaken. Following, the structures were washed three times for 10 minutes in distilled water.…”

“…[4,5] The latter, also called pillar arrays, are generated by fabricating cylindrical micropillars from the crosslinked polymer gel polydimethylsiloxane (PDMS) in defined geometries. [1,4,6,7] By coating the pillar tops with adhesive ligands and tuning pillar stiffness to cell forces, such arrays are commonly used measure cellular traction forces. [8,9] We have previously adapted pillar arrays for the investigation of small motile cells that can be placed between the pillars.…”

Mechanical Response Analysis and Power Generation by Single‐Cell Stretching

“…The cytoskeleton responds to both, stimulation by mechanical forces and biochemical signaling. [6,7] Cellular signaling may be induced, for instance, by surface recognition via intergrins, which causes cascades of biochemical reactions that lead to a precise regulation of the cytoskeleton structure and cell-adhesion forces. [8] A quantitative and precise determination of the forces generated by cells-in addition to the characterization of their viscoelastic behavior as a function of well-defined biochemical signals-and their description by physical models will contribute significantly to the understanding of the link between biomechanics and biochemistry.…”

Elucidating the molecular mechanisms underlying cellular response to biophysical cues using synthetic biology approaches

“…This approach can be extended to better understand the dynamics of cytoskeletal proteins such as actin filaments. [75][76][77] For instance, in studies using microfabricated epoxy pillars to quantify the elastic properties of individual actin filaments (Fig. 4A and 4B).…”