Alina Grzanka1, Dariusz Grzanka. 1. Department of Clinical Pathomorphology, L. Rydygier Medical University, Bydgoszcz, Poland. grzanka@ab-byd.edu.pl
Abstract
BACKGROUND: In the last decade there has been increasing interest in cytoskeletal research in relation to important process of cell biology. However, after decades of debate many issues remain unsettled. The aim of our study was to estimate the distribution of tubulin in etoposide-induced HL-60 cells and address the question of whether the reorganization of tubulin may be involved in apoptosis. MATERIAL/ METHODS: HL-60 cells were incubated with etoposide (0.02-200 mM/L) for 72 hours. Tubulin was estimated by the indirect immunofluorescence method. TRITC-labeled goat anti rabbit IgG was used to label tubulin. RESULTS: Significant changes occurred in cell morphology of and tubulin organization at 200 mM/L etoposide: large cells, brightly labeled for tubulin, in cytoplasm. At 200 uM/L etoposide, we also observed an increase in apoptotic cells with bright staining for tubulin at the site of apoptotic body formations. Cells with features of apoptosis were frequently seen also at 20 mM/L etoposide. The exposure of cells to 0.02, 0.2, or 2 mM/L etoposide did not result in any visible changes in morphology or tubulin organization. Cell size, the number of viable cells, and the percentage of apoptotic cells correlated significantly with the dose of etoposide (p<0.001). CONCLUSIONS: These results indicate that etoposide promotes changes in the distribution of tubulin in HL-60 cells, depending on the dose, and that the reorganization of tubulin seems to be involved in apoptosis
BACKGROUND: In the last decade there has been increasing interest in cytoskeletal research in relation to important process of cell biology. However, after decades of debate many issues remain unsettled. The aim of our study was to estimate the distribution of tubulin in etoposide-induced HL-60 cells and address the question of whether the reorganization of tubulin may be involved in apoptosis. MATERIAL/ METHODS: HL-60 cells were incubated with etoposide (0.02-200 mM/L) for 72 hours. Tubulin was estimated by the indirect immunofluorescence method. TRITC-labeled goat anti rabbit IgG was used to label tubulin. RESULTS: Significant changes occurred in cell morphology of and tubulin organization at 200 mM/L etoposide: large cells, brightly labeled for tubulin, in cytoplasm. At 200 uM/L etoposide, we also observed an increase in apoptotic cells with bright staining for tubulin at the site of apoptotic body formations. Cells with features of apoptosis were frequently seen also at 20 mM/L etoposide. The exposure of cells to 0.02, 0.2, or 2 mM/L etoposide did not result in any visible changes in morphology or tubulin organization. Cell size, the number of viable cells, and the percentage of apoptotic cells correlated significantly with the dose of etoposide (p<0.001). CONCLUSIONS: These results indicate that etoposide promotes changes in the distribution of tubulin in HL-60 cells, depending on the dose, and that the reorganization of tubulin seems to be involved in apoptosis