Sodium Salicylate-induced Apoptosis In HL-60 Cells And Its Partial Reversing Effect On Multidrug Resistance In HL-60/VCR Cells

Objective Multidrug resistance(MDR) is the biological basis of refractory leukemia. Searching for an efficient reversing agent of MDR is always important in the clinical research of leukemia. Sodium salicylate(Na-Sal), as a sort of classical nonsteroidal anti-inflamatory drugs(NSAID), rescently showed strong apoptosis-inducing activity against some tumor cells. Because the unability of apoptosis is one of the main mechanisms of MDR in leukemia, we took HL-60 and HL-60/VCR cells as targets to probe into the proapoptotic activity of Na-Sal and its partial reversing effect upon MDR of leukemia. Methods The cytotoxicity of Na-Sal and chemotherapeutical drugs to HL-60 and HL-60/VCR cells was measured by MTT assay; the morphologic changes of target cells were observed by means of light and electronic microscope; the phosphatidylserine exposure was identified with the help of Annexin-V binding assay; the DNA fragmentation was determined by means of agarose gel electrophoresis; the changes of the concentration of intracellular free Ca2+ was analyzed with the help of confocal laser scanning microscope (CLSM); the aneuploid DNA and the concentrationof intracellular daunorubicin(DNR) were analyzed through flow cytometry. Result Na-Sal was able to inhibit the growth of HL-60 cells in a dose- and time-dependent manner. After treatment of HL-60 cells with Na-Sal in a concentration of Smmol-L'^ICso) for lOh 16h, the target cells were characterized morphologically according to condensation of the nuclear chromatin, shrinkage and formation of apoptotic bodies; the phosphatidylserine exposure was highly increased compared with control (0.24 versus 0.03); the DNA from target cells showed typical aneuploid peak by flow cytometry and characteristic "ladder" pattern in agarose gel electrophoresis; and, meanwhile, the concentration of intracellular Ca2+ in target cells was progressively elevated. After exposure of HL-60/VCR cells with non-cytotoxic dose of Na-Sal(lmmol-L"'), the sensitivity of target cells to chemotherapeutical agents was remarkably increased , indicating that the MDR of HL-60/VCR cells was partly reversed. The reversal times varied from 1.01 to 3.26 with a relative reversal efficiency reaching 0.52 % to 73.62 % .With the help of flow cytometry, we found no increase of intracellular concentration of daunorubicin in HL-60/VCR cells during Na-Sal-mediated reverse of MDR. Conclusion (1) Na-Sal is potent in inducing apoptosis of HL-60 cells, which appears to be closely associated with the increase in the concentration of intracellular free Ca2+; (2) Na-Sal can partly reverse MDR of HL-60/VCR cells without changes of the concentration of intracellular DNR, showing that Na-Sal-mediated reverse of MDR in HL-60/VCR cells appears not to affect the pump action of glycoprotein 170(gpl70).