Atomic force microscopy of microvillous cell surface dynamics at fixed and living alveolar type II cells.
Scanning probe techniques enable direct imaging of morphology changes associated with cellular processes at life specimen. Here, glutaraldehyde-fixed and living alveolar type II (ATII) cells were investigated by atomic force microscopy (AFM), and the obtained topographical data were correlated with results obtained by scanning electron microscopy (SEM) and confocal microscopy (CM). We show that low-force contact mode AFM at glutaraldehyde-fixed cells provides complementary results to SEM and CM. Both AFM and SEM images reveal fine structures at the surface of fixed cells, which indicate microvilli protrusions. If ATII cells were treated with Ca(2+) channel modulators known to induce massive endocytosis, changes of the cell surface topography became evident by the depletion of microvilli. Low force contact mode AFM imaging at fixed ATII cells revealed a significant reduction of the surface roughness for capsazepine and 2-aminoethoxydiphenyl-borate (CPZ/2-APB)-treated cells compared to untreated control cells (Rc of 99.7 ± 6.8 nm vs. Rc of 71.9 ± 4.6 nm for N = 22), which was confirmed via SEM studies. CM of microvilli marker protein Ezrin revealed a cytoplasmic localization of Ezrin in CPZ/2-APB-treated cells, whereas a submembranous Ezrin localization was observed in control cells. Furthermore, in situ AFM investigations at living ATII cells using low force contact mode imaging revealed an apparent decrease in cell height of 17% during stimulation experiments. We conclude that a dynamic reorganization of the microvillous cell surface occurs in ATII cells at conditions of stimulated endocytosis.
Institute of Analytical and Bioanalytical Chemistry, University of Ulm, Germany.
Elena
E
Hecht
Shariq M
SM
Usmani
Susanne
S
Albrecht
Oliver H
OH
Wittekindt
Paul
P
Dietl
Boris
B
Mizaikoff
Christine
C
Kranz
D056809:Alveolar Epithelial Cells / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure*; D000818:Animals; D001896:Boron Compounds; D002118:Calcium; D002211:Capsaicin / Q000031:analogs & derivatives; D002462:Cell Membrane / Q000378:metabolism* / Q000648:ultrastructure*; D002478:Cells, Cultured; D003598:Cytoskeletal Proteins / Q000276:immunology / Q000378:metabolism; D004705:Endocytosis; D005976:Glutaral; D021621:Imaging, Three-Dimensional; D008168:Lung / Q000166:cytology; D008297:Male; D018625:Microscopy, Atomic Force / Q000379:methods*; D018613:Microscopy, Confocal; D008855:Microscopy, Electron, Scanning / Q000379:methods; D008871:Microvilli / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D000099149:Ezrin
D001896:Boron Compounds; D002118:Calcium; D002211:Capsaicin; D003598:Cytoskeletal Proteins; D005976:Glutaral; D000099149:Ezrin; C109986:2-aminoethoxydiphenyl borate; C071423:capsazepine