The g and F contents in megakaryocyte cell lines after stimulation with phorbol myristate acetate

Megakaryocyte polyploidization responds to platelet demand and results from the lack of cytoplasmic separation while the nucleus keeps dividing. In order to investigate the role of actin in the megakaryocyte polyploidization, phorbol myristate acetate (PMA, 5 x 10^-9 M), a differentiation marker known to induce megakaryocyte polyploidization, was added to human megakaryocyte cell lines (DAMI and HEL) and G, F and total actins were estimated by DNase I inhibition. After four days of culture in the presence of PMA, G actin contents in pg per 10⁶ cells were 13.0 pg ± 2.8 ard 1.0 pg ± 0.1 for unstimulated DAMI and HEL cells. F actin contents per 10⁶ cells were 5.8 pg ± 1.5 and 0.1 pg ± 0.0 for DAMI and HEL cells. Addition of PMA for four days to culture significantly increased G actin contents (235% and 268% of controls) and F actin contents (234% and 394%), for DAMI and HEL cell lines, respectively (p < 0.05 by t-test). In contrast, G/F actin ratio was not affected (p < 0.05 by t-test) by PMA. DAMI cells from each ploidy classes were then sorted on an ELITE Coulter and assayed for actin content. While total actin, G actin and F actin per cell increased in polyploid cells cultured with PMA, there was a reduction in G, F and total actin contents per diploid equivalent when cells became polyploid. In conclusion, megakaryocyte polyploidization of these cell lines is not related to an unbalance between G and F actins but would be rather due to a defect in total actin production that could lead to a prevention of the formation of the construction ring in telophase.