Actin reorganization during proplatelet formation.

The distribution of polymerized MK actin was investigated after cytocentrifugation by staining with rhodamine-conjugated phalloidin (red) and examining by fluorescent microscopy. (A) In the absence of plasma, polymerized actin is evenly distributed throughout the cells, and there is no proplatelet formation as determined from visual inspection of undisturbed cultures before cytocentrifugation. (B) In plasma-containing cultures that promote proplatelet formation, there are F-actin aggregates in the cells (arrows). These aggregates are not nuclei, as demonstrated by DAPI staining of cells in the same field to show nuclear DNA in blue (E). (C) F-actin staining in proplatelet-forming MKs. MKs grown in 10% plasma were fixed in culture with 2% paraformaldehyde to preserve proplatelet processes before cytospin preparation. At 48 hours, F-actin aggregations can be found in proplatelet-forming MKs (arrows). (D) PKC down-modulation by prolonged incubation of PMA prevented actin reorganization in plasma-containing culture, but did not reduce the amount of F-actin. (F) Nuclear DNA staining of cells in panel C was revealed by DAPI staining.