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

Abstract

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, La3+, Mn2+, Zn2+, Cu2+ and Fe2+, 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 (Kd) values as follows: La3+ 1.5 mM, Mn2+ 93.2 mM, Zn2+ 38.6 mM, Cu2+ 71.9 μM and Fe2+ 32.8 mM. The transparent non-selective pore blocker La3+ did not affect the absorbance of hemoglobin, while Mn2+ reduced it slightly. Other cations, including Zn2+, Cu2+ and Fe2+, greatly decreased the absorbance with Kd values of 35.9, 77.5 and 17.6 mM, respectively. After correction, the inhibitory Kd values were 1.4 mM, 45.8 mM, 128.5 μM and 53.1 mM for La3+, Zn2+, Cu2+ and Fe2+, respectively. Mn2+ did not inhibit TE-induced hemolysis. Moreover, the inhibitory extent at the maximal given dose of all cations except La3+ 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 La3+, 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.

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“…In brief, freshly collected blood samples from mice were immediately mixed with anticoagulant (1% heparin in PBS, pH 7.4) to prevent blood coagulation. For a pure suspension of erythrocytes, 1 mL of whole blood was made up to 20 mL in phosphate buffered saline (PBS, pH 7.4) and centrifuged at 1000× g for 10 min at 4 • C. The supernatant and buffy coats were then discarded by gentle aspiration, and the erythrocyte pellet was washed twice and suspended in the same buffer to a final concentration of 0.5% (v/v) [38]. Various concentrations of NnV (0.05, 0.2, 0.8, 3.2, 12.8, 51.2, and 204.8 µg/mL) were added to the erythrocyte suspension (100 µL, 0.…”

Raising Awareness on the Clinical and Forensic Aspects of Jellyfish Stings: A Worldwide Increasing Threat

“…Hemolysis is a common effect of several jellyfish venoms. Some venom components attain a hemolytic effect by altering cell permeability, causing ion currents, cell swelling, and osmotic lysis, while others degrade the phospholipids bilayer or form pores in the membrane [ 23 , 24 , 25 ].…”