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The Journal of Immunology, Vol 157, Issue 11 5070-5075, Copyright © 1996 by American Association of Immunologists
ARTICLES |
M Aepfelbacher, M Essler, E Huber, A Czech and PC Weber
Institute for Prevention of Cardiovascular Diseases, University of Munich, Germany.
Monocyte spreading is a sequential series of events including cell flattening, formation of new actin structures and focal adhesions, as well as development of cytoplasmic projections. To investigate the involvement of the GTP-binding protein Rho in spreading, we treated human blood monocytes or PMA-stimulated U937 or THP1 cells with Clostridium botulinum C3-transferase (C3), which ADP-ribosylates and inactivates Rho in intact cells. The C3 treatment caused 1) a four- to fivefold increase in the number of THP1 cells that spread on fibronectin within 24 h of PMA stimulation, 2) a greater area covered by the spread cells, and 3) accelerated and enhanced development of macrophage-like filopodial and pseudopodial projections. Similar results were obtained with PMA-stimulated U937 cells and human blood monocytes. Furthermore, cell staining revealed disorganization of subcortical actin in C3-treated THP1 cells, whereas circular actin formations at the substrate-attached part of the cells and vinculin- containing focal complexes/adhesions were unaffected. Finally, we found a decrease in membrane-associated RhoA in normal spreading THP1 cells, which suggests endogenous inactivation of Rho and might provide an explanation for the acceleration of spreading caused by the C3- transferase. In conclusion, these results indicate that active Rho is an important, negative regulator of human monocyte spreading by maintaining cell tension and cortical actin organization.
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