Intervention effects of five cations and their correction on hemolytic activity of tentacle extract from the jellyfish Cyanea capillata

Cations have generally been reported to prevent jellyfish venom-induced hemolysis through multiple mechanisms by spectrophotometry. Little attention has been paid to the potential interaction between cations and hemoglobin, potentially influencing the antagonistic effect of cations. Here, we explored the effects of five reported cations, La(3+), Mn(2+), Zn(2+), Cu(2+) and Fe(2+), on a hemolytic test system and the absorbance of hemoglobin, which was further used to measure their effects on the hemolysis of tentacle extract (TE) from the jellyfish Cyanea capillata. All the cations displayed significant dose-dependent inhibitory effects on TE-induced hemolysis with various dissociation equilibrium constant (K(d)) values as follows: La(3+) 1.5 mM, Mn(2+) 93.2 mM, Zn(2+) 38.6 mM, Cu(2+) 71.9 μM and Fe(2+) 32.8 mM. The transparent non-selective pore blocker La(3+) did not affect the absorbance of hemoglobin, while Mn(2+) reduced it slightly. Other cations, including Zn(2+), Cu(2+) and Fe(2+), greatly decreased the absorbance with K(d) values of 35.9, 77.5 and 17.6 mM, respectively. After correction, the inhibitory K(d) values were 1.4 mM, 45.8 mM, 128.5 μM and 53.1 mM for La(3+), Zn(2+), Cu(2+) and Fe(2+), respectively. Mn(2+) did not inhibit TE-induced hemolysis. Moreover, the inhibitory extent at the maximal given dose of all cations except La(3+) was also diminished. These corrected results from spectrophotometry were further confirmed by direct erythrocyte counting under microscopy. Our results indicate that the cations, except for La(3+), can interfere with the absorbance of hemoglobin, which should be corrected when their inhibitory effects on hemolysis by jellyfish venoms are examined. The variation in the inhibitory effects of cations suggests that the hemolysis by jellyfish venom is mainly attributed to the formation of non-selective cation pore complexes over other potential mechanisms, such as phospholipases A2 (PLA2), polypeptides, protease and oxidation. Blocking the pore-forming complexes may be a primary strategy to improve the in vivo damage and mortality from jellyfish stings due to hemolytic toxicity.