Forced extension of delipidated red blood cell cytoskeleton with little indication of spectrin unfolding.
Force-extension curves obtained on intact human red blood cells (RBC) were compared with those of delipidated RBCs to assess the contribution of cytoskeletal flexibility to the extensibility of the intact membrane skeleton. The RBCs were first delipidated by treatment with phospholipase A₂; tensile properties of the exposed cytoskeletal structures were measured using an atomic force microscope (AFM). The AFM probes were modified either with the Band 3 specific lectin, concanavalin A, (Con A) or anti-F-actin antibody, to localize the point of interaction between the probe and the cytoskeleton. Extension of the spectrin-based cytoskeleton reached up to 2-3 μm with a force less than 70 pN without showing any force peaks before the final rupture of the adhesive bonds. Our interpretation of the result is that the spectrin-based network was slack enough to allow the observed degree of extension without unfolding the tetrameric spectrin molecules. The force-extension curves obtained either on Band 3-ankyrin loci or on junction nodes of the cytoskeleton were not significantly different. Experimental results were verified by computer simulation of pulling mechanics of a network model of the RBC cytoskeleton. Our experimental results are also in agreement with the theoretical prediction of Mirijanian and Voth [2008; Proc Natl Acad Sci USA 105:1204-1208].
Innovation Laboratory, Tokyo Institute of Technology, Yokohama, Japan.
Rehana
R
Afrin
Masato
M
Nakaji
Hiroshi
H
Sekiguchi
David
D
Lee
Kikuo
K
Kishimoto
Atsushi
A
Ikai
D000328:Adult; D001457:Anion Exchange Protein 1, Erythrocyte / Q000737:chemistry*; D003198:Computer Simulation; D003599:Cytoskeleton / Q000737:chemistry / Q000648:ultrastructure*; D004912:Erythrocytes / Q000737:chemistry / Q000648:ultrastructure*; D006801:Humans; D008297:Male; D018625:Microscopy, Atomic Force / Q000379:methods; D013049:Spectrin / Q000737:chemistry*
D001457:Anion Exchange Protein 1, Erythrocyte; D013049:Spectrin