Role of intracellular free Ca(II) and Zn(II) in dexamethasone‐induced apoptosis and dexamethasone resistance in human leukemic CEM cell lines

Abstract

The levels of intracellular free Ca(II) and Zn(II) during dexamethasone (dex)-induced apoptosis in CEM cell lines were determined by 19F nuclear magnetic resonance (NMR), using the fluorinated intracellular chelator 1,2-bis-(2- amino-5-fluorophenoxy)ethane-N,N,N',N'-tetraacetic acid (5-FBAPTA). The effects of these divalent metal ions on growth rate and DNA degradation were evaluated. Measurements were done on one dex-sensitive (CEM-C7) and three different dex-resistant variants (CEM-C1, CEM-4R4, and CEM-ICR27). Dex caused a continuous increase in the Ca(II) level in dex-sensitive CEM-C7 cells, while in CEM-C1 cells dex caused an initial increase in the Ca(II) level which in approximately 36 h was restored to its normal value. The intracellular Ca(II) level in CEM-4R4 cells was not significantly affected by dex, while that of CEM-ICR27 cells decreased after dex incubation. Only the dex-sensitive CEM-C7 cells showed dex-induced DNA degradation. An intracellular free Zn(II) level of approximately 1 nM was measured for the dex-resistant CEM-C1 cells. No detectable level of intracellular Zn(II) was found in the other cell lines. Incubation with < 100 microM Zn(II) did not inhibit dex-induced apoptosis in CEM-C7 cells (e.g., DNA degradation). Treatment with approximately 250 microM Zn(II) caused significant decrease in growth rate in all cell lines and prevented dex-induced DNA degradation in CEM-C7 cells. A calibrated amount of Ca(II) ionophore (A23187), used to increase Ca(II) concentrations up to the dex-induced levels, did not induce DNA degradation in CEM-C7 or CEM-C1 cells. While elevation of intracellular Ca(II) by itself is not sufficient to initiate apoptosis in CEM-C7 cells, the results reported here suggest that Ca(II) is involved in the killing mechanism as a secondary factor. The combination of dex and ionophore caused significant DNA degradation in CEM-C1 cells, which normally showed resistance to each compound individually. The combination of dex and the Zn(II) chelator phenanthroline also caused extensive DNA degradation in the normally dex-resistant CEM-C1 cells, suggesting that Zn(II) plays a role in the dex resistance of these cells.

Measuring steady-state and dynamic endoplasmic reticulum and Golgi Zn 2+ with genetically encoded sensors

“…Numerous studies have reported measurement of cytosolic free Zn 2þ in a variety of cell types, including red blood cells (34), leukemic cells (35), splenocytes and thymocytes (36), PC-12 (6), , HEK 293 (5), INS-1 (832/13) (5), hepatocytes (37), and neurons (38), yielding estimates for resting free Zn 2þ concentrations from 5 to 1,000 pM. In this paper, we report estimates of free Zn 2þ in ER and Golgi, which are 0.9 and 0.6 pM, respectively, at least 100 times lower than our estimate of the cytosolic Zn 2þ concentration (80 pM) in HeLa cells (SI Appendix, Table S1).…”

Metal ionophores – An emerging class of anticancer drugs

“…Zinc, for example, can alter cellular signal transduction (1), DNA synthesis (2, 3), gene transcription (4,5), post transcriptional modifications (6), protein-protein interactions (7), and protein function (8,9). Increases or decreases in intracellular zinc have been associated with apoptotic cell death (10)(11)(12)(13). As the essential role of metals in cell biology is well recognized, there have been relatively few attempts to alter the biology of cancer cells by altering their metal content.…”

Calcium-dependent upregulation of E4BP4 expression correlates with glucocorticoid-evoked apoptosis of human leukemic CEM cells

“…In thymocytes and T-lymphoblastoid cells, an increase in intracellular calcium ([Ca 2+ ] i ) levels represents an early biochemical change linked to GC-evoked apoptosis [19]. A correlation between elevation of [Ca 2+ ] i and apoptosis has been reported in human leukemic CEM cells [20]. Several mechanisms for calcium-dependent apoptosis have been proposed, including activation of specific Ca 2+ -dependent nucleases, calmodulin (CaM)-mediated activation of calcineurin, and transcriptional changes.…”