Deshan Zhou

Cytoplasmic elongation and rupture in megakaryoblastic leukemia cells via activation of adhesion and motility by staurosporine on fibronectin-bound substratum

Human megakaryoblastic leukemia Meg‐01 cells were attached to fibronectin (FN)‐coated substratum, on which remarkable spreading and cytoplasmic elongation was induced by treatment with a protein kinase inhibitor, staurosporine (stp). This effect was inhibited by RGDS and was also not seen on FN‐lacking substratum. The extended cytoplasm had swollen terminals and nodes, which contained GpIIb and β‐thromboglobulin, occasionally included α granules, and tended to form particles (2–5 μm) after rupture of the narrowed cytoplasm. Among other protein kinase modulators tested, only K252a promoted the elongation, while calphostin, herbimycin, TPA, and calyculin suppressed it. The cells began to migrate soon after addition of stp, with attachment to the substratum held at some sites during the migration. This tethered movement seemed to cause the cytoplasmic elongation and the rupture into particles. The elongation was retarded by pretreating the cells with cytochalasin A and Clostridium C3 toxin but not with demecolcine. Actin microfilaments in the stp‐treated Meg‐01 cells accumulated in the filopodia and periphery of the extended cytoplasm, in which vinculin was colocalized as adhesion plaques. The microtubules were longitudinally oriented through the cytoplasmic extension and showed no ring profile in the nodes and particles. Thus, stp in the presence of FN appears to stimulate reorganization of actin‐based cytoskeleton and formation of focal contacts in Meg‐01 cells. This leads to the activation of cell adhesion and motility, and then cytoplasmic elongation and rupture into particles. J. Cell. Physiol. 179:179–192, 1999.

Acceleration of dna damage-induced apoptosis in leukemia cells by interfering with actin system

Abstract The apoptosis induced by actinomycin D (AD) in human mega-karyoblastic leukemia CMK-7 cell line is accelerated by microtubule poisons such as colcemid. We previously reported that the cell fragmentation in this apoptosis was suppressed by cytochalasin D (CD). 1 Here, we report that the AD-induced apoptosis is also accelerated by CD. The caspase-3 activation and DNA cleavage reached the maximum at least 10 hr earlier in the presence of CD than without this actin polymerization inhibitor. Decrease in organelle membrane potential, cytochrome c release from mitochondria, and cleavage of procaspase-9 to its active form began to appear prior to the caspase-3 activation. Apaf-1 was detected in the cytosol. These results suggest that the autocatalytic activation of caspase-9 starts the usual caspase cascade, and that the determinant, cytochrome c liberation, is promoted by interfering with the actin system. The cell surface F-actin disappeared by the AD-CD treatment and instead broad actin fibers appeared in the nucleus. These fibers were remote from the condensed chromatins, while F-actin in the cells undergoing apoptosis without CD accumulated around the nuclear fragments. The acceleration of AD-induced apoptosis by CD was found in U937 cells as well. Apoptosis induced by other DNA damaging agents (CDDP and MMC) was also accelerated by CD. The apoptosis by AD and CD was counteracted by antioxidants such as α-tocopherol and luteolin. Thus, the DNA damage-induced apoptosis is affected by disruption of the actin system, and probably, reactive oxygen species are involved in the process. 1 Exp. Hematol. , 27, Suppl. 1, 51 (1999).