Cytolytic and systemic toxic effects induced by the aqueous extract of the fire coral Millepora alcicornis collected in the Mexican Caribbean and detection of two types of cytolisins

Abstract

BackgroundMillepora alcicornis is a branching hydrocoral common throughout the Caribbean Sea. Like other members of this genus, this species is capable of inducing skin eruptions and blisters with severe pain after contact. In the present study, we investigated the toxicity of the M. alcicornis aqueous extract on several animal models. Considering that some cnidarian hemolysins have been associated to local tissue damage, since they also induce lysis of other cell types, we also made a partial characterization of the hemolytic activity of M. alcicornis aqueous extract. This information is important for understanding the defense mechanisms of the “fire corals”.MethodsThe effects of pH, temperature, and some divalent cations on the hemolytic activity of the extract were assayed, followed by a zymogram analysis to detect the cytolysins and determine their approximate molecular weight. The toxicity of the aqueous extract was assayed in mice, by intravenous administration, and histopathological changes on several tissues were analyzed by light microscopy. The toxicity of the extract was also tested in Artemia salina nauplii, and the damages caused on the crustaceans were analyzed by transmission and scanning electron microscopy.ResultsThe hemolytic activity of the hydrocoral extract was enhanced in the presence of Ca2+ (≥2 mM), Mg2+ (≥6 mM), and Ba2+ (≥0.1 mM); however, it was reduced in the presence of Cu2+ (≥0.1 mM), Zn2+ (≥6 mM), and EDTA (≥0.34 mM). Differences in the pH did not affect the hemolytic activity, but it was temperature-sensitive, since preincubation at ≥ 50 °C sharply reduced hemolysis. The zymogram showed the presence of two types of hemolysins: ~ 28–30 kDa proteins with phospholipase A2 activity and ~ 200 kDa proteins that do not elicit enzymatic activity. The aqueous extract of this cnidarian was lethal to mice (LD50 = 17 μg protein/g), and induced kidney, liver, and lung damages. Under denaturing conditions, the aqueous extract completely lost its toxic and hemolytic activities.ConclusionsThe results showed that the M. alcicornis aqueous extract contains two types of thermolabile hemolysins: proteins of approximately 28–30 kDa with PLA2 activity, while the others are larger proteins of approximately 200 kDa, which do not possess PLA2 activity. Those thermolabile cytolysins, which are stable to pH changes and whose activity is calcium dependent, are capable of inducing damage in lung, kidney and liver tissues, resulting in a slow death of mice. M. alcicornis cytolysins also provoke tissue dissociation in Artemia salina nauplii that might be attributed to pore forming mechanisms.

Heated Debates: Hot-Water Immersion or Ice Packs as First Aid for Cnidarian Envenomations?

“…There is abundant evidence from in vitro and in vivo studies which suggest that cndarian venoms from all classes in the phylum are heat labile (Table 4) [31,32,33,34,35,36,37,38,39,40,41,42,43,44,45]. Some have suggested that the mechanism by which heat provides pain relief is unrelated to inactivation of venom components.…”

Marine Natural Products from the Yucatan Peninsula

“…Out of the 66 marine natural products identified, 26 correspond to structures that were not previously reported. These 26 new chemical entities correspond to three aliphatic polyketides (1-3), one glycolipid (7), two diterpenes (15,16), two steroids (24,25), one triterpenoid saponin (40), and 17 conotoxins (49)(50)(51)(52)(53)(54)(55)(56)(57)(58)(59)(60)(61)(62)(63)(64)(65). Figure 15 displays the overall biogenetic distribution of the reported compounds.…”

“…The results suggested not only that these conotoxins can operate as nicotinic acetylcholine receptor inhibitors, but also that they bind to nicotinic acetylcholine receptors with a very high affinity, increasing their intrinsic cholinergic response, and making them excellent model tools for studying toxin-receptor interaction [30]. The fourth new peptide, conotoxin sr11a (56), with a molecular weight of 3650.77 Da, was reported in 2007, being the first I-conotoxin that was isolated from the Western Atlantic. This peptide produces a stiffening of body, limbs, and tail when intracranially injected into mice [31].…”

Evaluation of Cyanea capillata Sting Management Protocols Using Ex Vivo and In Vitro Envenomation Models

“…Indeed recent research has shown that a crude venom extract from Pelagia noctiluca (another scyphozoan jellyfish) also exhibits a loss of potency at temperatures higher than 40 °C [ 48 , 49 ]). Similarly, biochemical studies have shown marked 45 °C heat related loss of activity in vitro [ 50 , 51 , 52 , 53 , 54 , 55 , 56 , 57 , 58 , 59 , 60 , 61 , 62 , 63 , 64 ] which may reflect thermal unfolding or aggregation. Additional research is needed to determine exactly why heat has this direct, negative effect on venom activity; there may be less evolutionary pressure for heat tolerance among cnidarian venom proteins than mammalian proteins.…”