It has been suggested that pemphigus antibodies (PA) react with the surface molecules on keratinocytes, and induce the production and release of proteases resulting in acantholysis. If this is the case, the immunoreactions on the cell surface may send signals to the interior of the cell across the membrane. The present study was carried out to determine whether or not cytoskeletons [microtubules (MT) and keratin-intermediate filaments (KIF)] respond to PA-immunoreactions in cultured human keratinocytes, by indirect immunofluorescence microscopy using anti-keratin and anti-alpha-tubulin antibodies. During incubation for 30 min to 72 h in a PA-containing medium with a normal concentration of Ca2+ (1.2 mM), no changes in MT or KIF organization were detected. Alterations in the organization of these filaments were observed 96 h after addition of PA. When cells grown in a normal medium for 5-7 days were transferred to a medium containing PA and a low level of Ca2+ (0.07-0.14 mM) the reorganization of KIFs and MTs occurred after 1 h incubation. However, no reorganization of the cytoskeletons was detected in the absence of cell detachment. These observations suggest that the pemphigus antibody-induced reorganization of MTs and KIFs does not precede acantholysis and is probably secondary to it, but is not a direct transmembrane response. The present study also showed that immunofluorescence microscopy of KIFs may be one of the most sensitive methods for detecting early cell-to-cell dissociation in cultured keratinocytes.
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