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

Spontaneous 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 mu M), a Ca2+ antagonist of Chinese herbal origin. Exposure of the cells for 5 min to saponins from Quillaja saponaria (QS, 30 mu g/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 mu g/ml) alone or together,with TET did not cause such a permeabilization. Permeabilization induced by 1-3 mu M digitonin, another cholesterol-complexing glycoside, could not be enhanced by TET. TET did nor 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 mu M) or SK&F 96365 (20 mu M). Further, it did not augment Ni2+ entry when the plasma membrane fluidity was modulated by changes of temperature (27-47 degrees C) or treatment with 5% ethanol. This QS-promoted Ni2+ entry could not be amplified by other lipophilic Ca2+ antagonists, such as diltiazem (100 mu M) and verapamil (100 mu M). 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 TBT and its very close analogues, hernandezine and berbamine, reveals that the methoxyl group at the R-2 position of TET appears to be crucial in enhancing QS-promoted Ni2+ entry.