Specific interaction between tetrandrine and Quillaja saponins in promoting permeabilization of plasma membrane in human leukemic HL-60 cells

AbstractSpontaneous Ni2+ entry (leak), measured as fluorescence quench in fura-2-loaded HL-60 cells at the excitation wavelength of 360 nm, was strongly inhibited by tetrandrine (TET, 100 μM), a Ca2+ antagonist of Chinese herbal origin. Exposure of the cells for 5 min to saponins from Quillaja saponaria (QS, 30 μg/ml), surfactants well known to permeabilize the plasma membrane by complexing with cholesterol, promoted Ni2+ entry without causing fura-2 leak-out. Unexpectedly, TET caused an immediate (within 2.5 min) augmentation of QS-promoted Ni2+ entry; and a 5-min treatment with both TET and QS resulted not only in an enhanced Ni2+ entry, but also a fura-2 leak-out. Ginseng saponins (100 μg/ml) alone or together with TET did not cause such a permeabilization. Permeabilization induced by 1–3 μM digitonin, another cholesterol-complexing glycoside, could not be enhanced by TET. TET did not affect permeabilization induced by Triton X-100 (0.01%), a detergent which non-specifically disrupts the hydrophobic interaction at the plasma membrane. TET also did not enhance Ni2+ entry triggered by ionomycin (0.35 μM) or SK&F 96365 (20 μM). Further, it did not augment Ni2+ entry when the plasma membrane fluidity was modulated by changes of temperature (27–47°C) or treatment with 5% ethanol. This QS-promoted Ni2+ entry could not be amplified by other lipophilic Ca2+ antagonists, such as diltiazem (100 μM) and verapamil (100 μM). The results hence indicate that TET enhanced Ni2+ entry (or permeabilization) elicited by QS treatment, but not other perturbations of the plasma membrane. We suggest that pore formation at the plasma membrane, a consequence of QS-cholesterol interaction, can be specifically enhanced by TET. Also, a comparative study of the effects of TET and its very close analogues, hernandezine and berbamine, reveals that the methoxyl group at the R2 position of TET appears to be crucial in enhancing QS-promoted Ni2+ entry