Inhibition of Nematocyst Discharge from Pelagia noctiluca (Cnidaria: Scyphozoa)-Prevention Measures against Jellyfish Stings.

Pelagia noctiluca stings are common in Mediterranean coastal areas and, although the venom is non-lethal, they are painful. Due to its high toxicity and abundance, P. noctiluca is considered a target species for the focus of research on active ingredients to reduce the symptoms of its sting. To determine the effect of 31 substances and formulations on nematocyst discharge, we performed three tests: (1) screening of per se discharge activator solutions, (2) inhibitory test with nematocyst chemical stimulation (5% acetic acid) and (3) inhibitory test quantifying the hemolytic area. Ammonia, barium chloride, bleach, scented ammonia, carbonated cola, lemon juice, sodium chloride and papain triggered nematocyst discharge. All of them were ruled out as potential inhibitors. Butylene glycol showed a reduction in nematocyst discharge, while the formulations of 10% lidocaine in ethanol, 1.5% hydroxyacetophenone in distilled water + butylene glycol, and 3% Symsitive® in butylene glycol inhibited nematocyst discharge. These last results were subsequently correlated with a significant decrease in hemolytic area in the venom assays versus seawater, a neutral solution. The presented data represent a first step in research to develop preventive products for jellyfish stings while at the same time attempting to clarify some uncertainties about the role of various topical solutions in P. noctiluca first-aid protocols.

The architecture and operating mechanism of a cnidarian stinging organelle.

The stinging organelles of jellyfish, sea anemones, and other cnidarians, known as nematocysts, are remarkable cellular weapons used for both predation and defense. Nematocysts consist of a pressurized capsule containing a coiled harpoon-like thread. These structures are in turn built within specialized cells known as nematocytes. When triggered, the capsule explosively discharges, ejecting the coiled thread which punctures the target and rapidly elongates by turning inside out in a process called eversion. Due to the structural complexity of the thread and the extreme speed of discharge, the precise mechanics of nematocyst firing have remained elusive7. Here, using a combination of live and super-resolution imaging, 3D electron microscopy, and genetic perturbations, we define the step-by-step sequence of nematocyst operation in the model sea anemone Nematostella vectensis. This analysis reveals the complex biomechanical transformations underpinning the operating mechanism of nematocysts, one of nature's most exquisite biological micro-machines. Further, this study will provide insight into the form and function of related cnidarian organelles and serve as a template for the design of bioinspired microdevices.

Nematocyst types and venom effects of Aurelia aurita and Velella velella from the Mediterranean Sea.

Natural substances produced by venomous marine organisms are thought to be possible sources of useful compounds and new drugs having the potential to open new ways for pharmacology, nutrition and environmental applications. In this framework, cnidarians are very interesting being widely distributed and all are venomous organisms; so, a deep knowledge of their occurrence, morphology of venomous structures and of effects of venoms at cellular level is fundamental to evaluate the possible utilization of venomous compounds or extracts. In this research, the morphology and occurrence of nematocysts in two cnidarian species (Aurelia aurita, Velella velella), and the preliminary evaluation of the cytotoxicity of V. velella crude extract, of which cytotoxicity on cell cultures at present is unknown, were considered. The specimens were sampled in Güllük Bay, Southwestern coast of Turkey, and in the Gulf of Genova, Northwestern coast of Italy. Six nematocyst types (a-isorhiza, A-isorhiza, O-isorhiza, eurytele, polyspiras, birhopaloid) having different sizes, were observed in A. aurita, and two types (eurytele and stenotele) in V. velella. The crude extract from V. velella showed cytotoxic activity against cultured fibroblasts L929 at high doses, while inducing cell proliferation at low doses. The protein content in the extract increased remarkably after disruption of nematocysts.

Inhibitory Effect of Metalloproteinase Inhibitors on Skin Cell Inflammation Induced by Jellyfish Nemopilema nomurai Nematocyst Venom.

Jellyfish envenomations result in extensive dermatological symptoms, clinically named as jellyfish dermatitis, which can seriously affect the daily activities and physical health of people. Inflammatory response accompanies the whole process of jellyfish dermatitis and the complexity of jellyfish venom components makes it difficult to treat jellyfish dermatitis symptoms effectively. Moreover, inhibiting inflammation is essential for the treatment of jellyfish stings and exploring the main components of jellyfish venom that cause inflammation is an urgent research area. In this study, the inhibitory effects of matrix metalloproteinase (MMP) inhibitors for venom-induced inflammation were explored at a cellular level. The expression of the three inflammatory factors, IL-6, TNF-α and MCP-1 in two skin cell lines, human keratinocyte cells (HaCaT) and human embryonic skin fibroblasts cells (CCC-ESF-1), at the cellular level, after treatment with the inhibitors of jellyfish Nemopilema nomurai (N. nomurai) nematocyst venom (NnNV-I), were determined. The results showed that inhibitors of MMP can significantly reduce the toxic effects of jellyfish Nemopilema nomurai nematocyst venom (NnNV) to skin cells. The expression levels of the three inflammatory factors IL-6, MCP-1, and TNF-α in the cells were also significantly decreased, indicating that MMPs in jellyfish venom are probably vital factors leading to jellyfish dermatitis. This study is beneficial in the prevention and treatment of jellyfish stings.

Crude venom from nematocysts of Pelagia noctiluca (Cnidaria: Scyphozoa) elicits a sodium conductance in the plasma membrane of mammalian cells.

Cnidarians may negatively impact human activities and public health but concomitantly their venom represents a rich source of bioactive substances. Pelagia noctiluca is the most venomous and abundant jellyfish of the Mediterranean Sea and possesses a venom with hemolytic and cytolytic activity for which the mechanism is largely unknown. Here we show that exposure of mammalian cells to crude venom from the nematocysts of P. noctiluca profoundly alters the ion conductance of the plasma membrane, therefore affecting homeostatic functions such as the regulation and maintenance of cellular volume. Venom-treated cells exhibited a large, inwardly rectifying current mainly due to permeation of Na+ and Cl-, sensitive to amiloride and completely abrogated following harsh thermal treatment of crude venom extract. Curiously, the plasma membrane conductance of Ca2+ and K+ was not affected. Current-inducing activity was also observed following delivery of venom to the cytosolic side of the plasma membrane, consistent with a pore-forming mechanism. Venom-induced NaCl influx followed by water and consequent cell swelling most likely underlie the hemolytic and cytolytic activity of P. noctiluca venom. The present study underscores unique properties of P. noctiluca venom and provides essential information for a possible use of its active compounds and treatment of envenomation.

The nematocysts venom of Chrysaora helvola Brandt leads to apoptosis-like cell death accompanied by uncoupling of oxidative phosphorylation.

The present work investigated the effects of the nematocysts venom (NV) from the Chrysaora helvola Brandt (C. helvola) jellyfish on the human nasopharyngeal carcinoma cell line, CNE-2. The medium lethal concentration (LC50), quantified by MTT assays, was 1.7 ± 0.53 µg/mL (n = 5). An atypical apoptosis-like cell death was confirmed by LDH release assay and Annexin V-FITC/PI staining-based flow cytometry. Interestingly, activation of caspase-4 other than caspase-3, -8, -9 and -1 was observed. Moreover, the NV stimuli caused a time-dependent loss of mitochondrial membrane potential (ΔΨm) as was an intracellular ROS burst. These results indicated that there was uncoupling of oxidative phosphorylation (UOP). An examination of the intracellular pH value by a pH-sensitive fluorescent probe, BCECF, suggested that the UOP was due to the time-dependent increase in the intracellular pH. This is the first report that jellyfish venom can induce UOP.

Crude Venom from Nematocysts of the Jellyfish Pelagia noctiluca as a Tool to Study Cell Physiology.

Marine animals represent a source of novel bioactive compounds considered as a good research model, whose mechanism of action is intriguing and still under debate. Among stinging animals, Cnidarians differentiated highly specialized cells, termed nematocytes, containing a capsule fluid with toxins and an inverted tubule, synergistically responsible for mechanisms of defence and predation. Such compounds include proteins and secondary metabolites with toxic action. With the aim of better elucidating the effects of Cnidarian venom upon cell targets, this short review reports on the current knowledge about the toxicological activity of venom extracted from nematocysts of the jellyfish Pelagia noctiluca, whose notable blooming is well known in the Strait of Messina (Italy). The effects on cultured cells, from both mammals and invertebrates, and erythrocytes are here being considered. What is known about the biological activity of Pelagia noctiluca crude venom accounts for a powerful biological activity at different levels, suggesting that cell damage may be due to a pore formation mechanism on cell membrane target leading to osmotic lysis, and /or to oxidative stress events. In this light, the study of venom activity may contribute to: i) validate suitable biological assays for venom testing; ii) elucidate cell function features; iii) understand the pathophysiology of envenoming.

Cell death in relation to DNA damage after exposure to the jellyfish Pelagia noctiluca nematocysts.

Studies on the toxicity of Mediterranean jellyfish have gained attention owing to their weak toxic properties. Our research has been mainly performed on the Scyphomedusae. Pelagia noctiluca is a scyphozoan jellyfish which causes a danger to sea bathers and fishery damages in the Mediterranean Sea. To check whether the cytotoxicity of Pelagia noctiluca nematocysts was associated to DNA lesions, we have looked for DNA fragmentation by means of the Comet and chromosome aberration assays. To specify cell death pathway, we have investigated caspase-3 activation. Our results have shown that nematocysts reduced cell viability and induced DNA fragmentation in a concentration-dependent manner with a maximum effect at 150 000 nematocysts mL(-1). The high percentage of chromosome aberrations also emphasized the genotoxic character of Pelagia noctiluca nematocysts in Vero cells. This fragmentation was correlated to apoptosis induction which was confirmed by caspase-3 activation. In conclusion, the present report has suggested that Pelagia noctiluca nematocysts were able to promote apoptosis in Vero cells and therefore may be useful in cancer therapy.

Sea water acidification affects osmotic swelling, regulatory volume decrease and discharge in nematocytes of the jellyfish Pelagia noctiluca.

BACKGROUND: Increased acidification/PCO2 of sea water is a threat to the environment and affects the homeostasis of marine animals. In this study, the effect of sea water pH changes on the osmotic phase (OP), regulatory volume decrease (RVD) and discharge of the jellyfish Pelagia noctiluca (Cnidaria, Scyphozoa) nematocytes, collected from the Strait of Messina (Italy), was assessed. METHODS: Isolated nematocytes, suspended in artificial sea water (ASW) with pH 7.65, 6.5 and 4.5, were exposed to hyposmotic ASW of the same pH values and their osmotic response and RVD measured optically in a special flow through chamber. Nematocyte discharge was analyzed in situ in ASW at all three pH values. RESULTS: At normal pH (7.65), nematocytes subjected to hyposmotic shock first expanded osmotically and then regulated their cell volume within 15 min. Exposure to hyposmotic ASW pH 6.5 and 4.5 compromised the OP and reduced or totally abrogated the ensuing RVD, respectively. Acidic pH also significantly reduced the nematocyte discharge response. CONCLUSION: Data indicate that the homeostasis and function of Cnidarians may be altered by environmental changes such as sea water acidification, thereby validating their use as novel bioindicators for the quality of the marine environment.

Sulphate and chloride-dependent potassium transport in human erythrocytes are affected by crude venom from nematocysts of the jellyfish Pelagia noctiluca.

BACKGROUND: It has been reported that biologically active compounds extracted from Cnidaria venom may induce damage by oxidative stress. Erythrocytes are constantly exposed to oxidative stresses, which can contribute to sulphydril (SH-) group oxidation and cell membrane deformability accompanied with activation of K-Cl co-transport and inhibition of anion transport. In this regard, Band 3 protein is responsible for mediating the electroneutral exchange of chloride (Cl(-)) for bicarbonate (HCO3(-)), particularly in erythrocytes, where it is the most abundant membrane protein. The aim of this study was to elucidate the effect of crude venom extracted from Pelagia noctiluca nematocysts on Band 3 -mediated anion transport in human erythrocytes. METHODS: Erythrocytes were tested for SO4(2-) uptake, K(+) efflux, glutathione (GSH) levels and concentration of SH- groups. RESULTS: The rate constant of SO4(2-) uptake decreased progressively to 58% of control with increasing venom doses, and showed a 28% decrease after 2 mM NEM treatment. These effects can be explained by oxidative stress, which was reflected by decreased GSH levels in venom-treated erythrocytes. Hence, the decreased efficiency of anion transport may be due to changes in Band 3 structure caused by SH-group oxidation and reduced GSH concentration. In addition, an increased Cl(-)-dependent K(+) efflux was observed in venom-treated erythrocytes. CONCLUSION: Our results suggest that crude venom from Pelagia noctiluca alters cell membrane transport in human erythrocytes.

Is cell death induced by nematocysts extract of medusa Pelagia noctiluca related to oxidative stress?

Pelagia noctiluca, a jellyfish widely distributed in the Mediterranean waters, especially in coastal areas of Tunisia, has garnered attention because of its stinging capacity and the resulting public health hazard. Crude extracts of P. noctiluca nematocysts have been tested for their cytotoxicity on Vero cells. Our results clearly showed that nematocysts induced cell mortality in a dose- and time-dependent manner. A cytoprotective effect against cell mortality was obtained when Vero cells were treated with Vitamin E. This process was further confirmed by the generation of reactive oxygen species (ROS) and the induction of Hsp 70 and 27 protein expressions. Thus, our findings suggested that oxidative stress is involved in the toxicity of pelagia nematocysts and may therefore constitute the major mechanism of this medusa nematocysts toxicity.

Analgesic and antibutyrylcholinestrasic activities of the venom prepared from the Mediterranean jellyfish Pelagia noctiluca (Forsskal, 1775).

BACKGROUND: Toxins derived from jellyfishes have been exploited as a model for the development of new drug promising applications to treat neurodegenerative diseases. The present work is aimed to evaluate the acute toxicity of crude venom of Pelagia noctiluca and then to screen the analgesic and antibutyrylcholinestrasic (anti-BuChE) activities of the crude venom and its fractions. METHODS: Sephadex G75 gel was used to separate crude venom of Pelagia noctiluca, which led to some fractions. In addition, in vivo analgesic and in vitro plasma antibutyrylcholinestrasic activities were carried out with Pelagia crude venom and its fractions respectively. RESULTS: The crude venom and its fractions displayed analgesic and anti-BuChE activities at different doses without inducing acute toxicity. Fraction 2 possesses the highest analgesic and antibutyrylcholinestrasic properties. The crude venom and fraction 1 had shown to possess less significant inhibitory activity against analgesic and antibutyrylcholinestrasic models. CONCLUSIONS: Based on this study, the crude venom of Pelagia noctiluca is found to be a useful tool for probing pharmacological activity. The purification and the determination of chemical structures of compounds of active fractions of the venom are under investigation.

Application of nanoLC-MS/MS to the shotgun proteomic analysis of the nematocyst proteins from jellyfish Stomolophus meleagris.

The nematocyst proteins of jellyfish Stomolophus meleagris, a complicated mixture, contain many important bioactive molecules. In present study, to gain comprehensive insight into the protein component and search some novel bioactive molecules in the nematocyst proteins, shotgun proteomic analysis of the nematocyst proteins was carried out by nano liquid chromatography tandem mass spectrometry (nanoLC-MS/MS) for the first time. Digested peptides of the nematocyst proteins were analyzed by nanoLC-MS/MS and all MS/MS spectra were then automatically searched by the SEQUEST program. A total of 181 proteins had been identified, with the molecular weight ranging from 5268.06 to 843,487.57 and the pI from 4.49 to 11.39. Bioinformatic analysis was also applied to better understand the identified proteins. In the gene ontology (GO) annotation, all the identified proteins were classified into 13, 9 and 7 groups according to biological process, cellular component and molecular function, respectively. Pathways analysis of the identified proteins was conducted with 33 corresponding pathways found. On the basis of pathways analysis, we also constructed the gene network to analyze the relationship of those genes each other, which contained enzyme-enzyme relation, protein-protein interaction and gene expression interaction.

Isolation, identification and characterization of a novel antioxidant protein from the nematocyst of the jellyfish Stomolophus meleagris.

Reactive oxygen species (ROS) can damage the lipids, proteins and DNA when produced excessively in cells. Here, we describe the isolation and identification of a novel antioxidant protein named SmP90 from the nematocyst of jellyfish Stomolophus meleagris by 50% ammonium sulfate precipitation and gel filtration chromatography, superdex75. HPLC and SDS-PAGE analysis revealed >95% purity of SmP90 with apparent molecular weight of 90 kDa, approximately. The identification of SmP90 was confirmed by both N-terminal amino acids sequencing, with the sequences of NLDTPYCFYSGDYGG, and peptide mass fingerprint (PMF) analysis by MALDI-TOF-MS. However, no known protein had been completely matched in the database, which indicated that SmP90 might be a novel protein. The antioxidant assay result showed that it had strong superoxide anion radical-scavenging activity with the half-scavenging concentration (EC(50)) of about 16 µg/mL. Therefore, the present study is the first time to demonstrate a high efficient method of isolating a novel antioxidant protein from the nematocyst of jellyfish S. meleagris.

Cytotoxicity of the venom from the nematocysts of jellyfish Cyanea nozakii Kishinouye.

In this article, the cytotoxicity of the venom from the nematocysts of jellyfish Cyanea nozakii Kishinouye on human hepatoma cells (Bel-7402, SMMC-7721) and human colon cancer cells (H630) was investigated first. Of the three kinds of cells, the venom had the strongest cytotoxicity on H630 cells with the 50% lethal concentration (IC(50)) of 5.1 µg/ml. However, the IC(50) on Bel-7402 and SMMC-7721 was approximate 17.9 and 24.3 µg/ml, respectively. The cytotoxicity of the venom was affected by pH, temperature and storage conditions. At the pH 4.5-8.5, the venom displayed obvious cytotoxicity and the percentage of survival was about 50%. When pre-incubated at temperatures over 60°C for as short as 10 min, the percentage of survival sharply improved from 4.6% up to 80%. The venom was stored in a more stable condition at -80°C and in lyophilized state compared to other storage conditions used in this study. Lactate dehydrogenase release assay performed on H630 cells indicated that exposure to the venom could result in damage to the cell membrane.

Partial characterization of the hemolytic activity of the nematocyst venom from the jellyfish Cyanea nozakii Kishinouye.

Using a recently developed technique to extract jellyfish venom from nematocysts, the present study investigated the hemolytic activity of Cyanea nozakii Kishinouye nematocyst venom on chicken erythrocytes. Venom extract caused a significant concentration-dependent hemolytic effect. The extract could retain its activity at -80 degrees C but was unstable when kept at 4 degrees C and -20 degrees C for 2 days. The hemolytic activity was inhibited by heating within the range of 37-100 degrees C. The extract was active over a pH range of 5.0-8.63 and the pH optima for the extract was 7.8. Incubation of the venom with sphingomyelin specially inhibited hemolytic activity by up to 70%. Cu(2+) and Mn(2+) greatly reduced the hemolytic activity while Mg(2+), Sr(2+) and Ba(2+) produced a relatively low inhibiting effect on the hemolytic activity. Treatment with Ca(2+) induced a concentration-dependent increase in the hemolytic activity. In the presence of 5 mM EDTA, all the hemolytic activity was lost, however, the venom containing 1.5 mM EDTA was stable in the long-term storage. PLA(2) activity was also found in the nematocyst venom of C. nozakii. These characteristics provide us a fundamental knowledge in the C. nozakii nematocyst venom which would benefit future research.