Report of the 1st International Electronic Conference on Toxins (IECT2021), 16-31 January 2021.

The 1st International Electronic Conference on Toxins (IECT2021) was successfully held online by https://sciforum [...].

Proteomic Changes during the Dermal Toxicity Induced by Nemopilema nomurai Jellyfish Venom in HaCaT Human Keratinocyte.

Jellyfish venom is well known for its local skin toxicities and various lethal accidents. The main symptoms of local jellyfish envenomation include skin lesions, burning, prickling, stinging pain, red, brown, or purplish tracks on the skin, itching, and swelling, leading to dermonecrosis and scar formation. However, the molecular mechanism behind the action of jellyfish venom on human skin cells is rarely understood. In the present study, we have treated the human HaCaT keratinocyte with Nemopilema nomurai jellyfish venom (NnV) to study detailed mechanisms of actions behind the skin symptoms after jellyfish envenomation. Using two-dimensional gel electrophoresis (2-DE) and matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF/MS), cellular changes at proteome level were examined. The treatment of NnV resulted in the decrease of HaCaT cell viability in a concentration-dependent manner. Using NnV (at IC50), the proteome level alterations were determined at 12 h and 24 h after the venom treatment. Briefly, 70 protein spots with significant quantitative changes were picked from the gels for MALDI-TOF/MS. In total, 44 differentially abundant proteins were successfully identified, among which 19 proteins were increased, whereas 25 proteins were decreased in the abundance levels comparing with their respective control spots. DAPs involved in cell survival and development (e.g., Plasminogen, Vinculin, EMILIN-1, Basonuclin2, Focal adhesion kinase 1, FAM83B, Peroxisome proliferator-activated receptor-gamma co-activator 1-alpha) decreased their expression, whereas stress or immune response-related proteins (e.g., Toll-like receptor 4, Aminopeptidase N, MKL/Myocardin-like protein 1, hypoxia up-regulated protein 1, Heat shock protein 105 kDa, Ephrin type-A receptor 1, with some protease (or peptidase) enzymes) were up-regulated. In conclusion, the present findings may exhibit some possible key players during skin damage and suggest therapeutic strategies for preventing jellyfish envenomation.

Preparation and Neutralization Efficacy of Novel Jellyfish Antivenoms against Cyanea nozakii Toxins.

Jellyfish stings are a common issue globally, particularly in coastal areas in the summer. Victims can suffer pain, itching, swelling, shock, and even death. Usually, hot water, vinegar, or alumen is used to treat the normal symptoms of a jellyfish sting. However, a specific antivenom may be an effective treatment to deal with severe jellyfish stings. Cyanea nozakii often reach a diameter of 60 cm and are responsible for hundreds of thousands of stings per year in coastal Chinese waters. However, there has been no specific C. nozakii antivenom until now, and so the development of this antivenom is very important. Herein, we collected C. nozakii antisera from tentacle extract venom immunized rabbits and purified the immunoglobulin (IgG) fraction antivenom (AntiCnTXs). Subsequently, two complete procedures to produce a refined F(ab')2 type of antivenom (F(ab')2-AntiCnTXs) and Fab type of antivenom (Fab-AntiCnTXs) by multiple optimizations and purification were established. The neutralization efficacy of these three types of antivenoms was compared and analyzed in vitro and in vivo, and the results showed that all types of antibodies displayed some neutralization effect on the lethality of C. nozakii venom toxins, with the neutralization efficacy as follows: F(ab')2-AntiCnTXs ≥ AntiCnTXs > Fab-AntiCnTXs. This study describes the preparation of novel C. nozakii jellyfish antivenom preparations towards the goal of developing a new, effective treatment for jellyfish stings.

The Curious Case of the "Neurotoxic Skink": Scientific Literature Points to the Absence of Venom in Scincidae.

In contrast to the clearly documented evolution of venom in many animal lineages, the origin of reptilian venom is highly debated. Historically, venom has been theorised to have evolved independently in snakes and lizards. However, some of the recent works have argued for the common origin of venom in "Toxicofera" reptiles, which include the order Serpentes (all snakes), and Anguimorpha and Iguania lizards. Nevertheless, in both these contrasting hypotheses, the lizards of the family Scincidae are considered to be harmless and devoid of toxic venoms. Interestingly, an unusual clinical case claiming neurotoxic envenoming by a scincid lizard was recently reported in Southern India. Considering its potentially significant medicolegal, conservation and evolutionary implications, we have summarised the scientific evidence that questions the validity of this clinical report. We argue that the symptoms documented in the patient are likely to have resulted from krait envenomation, which is far too frequent in these regions.

Centipede Venom Peptides Acting on Ion Channels.

Centipedes are among the oldest venomous arthropods that use their venom to subdue the prey. The major components of centipede venom are a variety of low-molecular-weight peptide toxins that have evolved to target voltage-gated ion channels to interfere with the central system of prey and produce pain or paralysis for efficient hunting. Peptide toxins usually contain several intramolecular disulfide bonds, which confer chemical, thermal and biological stability. In addition, centipede peptides generally have novel structures and high potency and specificity and therefore hold great promise both as diagnostic tools and in the treatment of human disease. Here, we review the centipede peptide toxins with reported effects on ion channels, including Nav, Kv, Cav and the nonselective cation channel polymodal transient receptor potential vanilloid 1 (TRPV1).

Selective Loss of Responsiveness to Exogenous but Not Endogenous Cyclic-Dinucleotides in Mice Expressing STING-R231H.

Stimulator of interferon genes (STING) plays a central role in innate immune responses to viral and intracellular bacterial infections, and cellular damage. STING is a cytosolic sensor of cyclic dinucleotides (CDNs) including those produced by pathogenic bacteria and those arising endogenously as products of the DNA sensor cGAS (e.g., 2'3' cGAMP). The two most common alternative allelic variants of STING in humans are STING-R71H-G230A-R293Q (STING-HAQ) and STING-R232H that are found in 20.4% and 13.7-17.6% of the population, respectively. To determine the biologic consequences of these genotypic variations, we generated knock-in mice containing the murine equivalents of each variant and studied their responsiveness to CDNs. Homozygous STING-HAQ (R71H-I229A-R292Q) and STING-R231H mice were found to be unresponsive to all exogenous CDNs tested (ci-di-GMP, ci-di-AMP, 3'3' cGAMP and Rp,Rp-CDA). Responses of homozygous STING-HAQ mice to endogenous 2'3' cGAMP was also greatly impaired. However, homozygous STING-R231H mice are fully responsive to 2'3' cGAMP. Analysis of heterozygous mice revealed reduced responsiveness to exogenous and endogenous CDNs in mice carrying a single copy of STING-HAQ, while STING-R231H heterozygous mice exhibit reduced responsiveness to exogenous but not endogenous CDNs. These findings confirm and extend previous reports by demonstrating differing impact of allelic variation of STING on the ability to sense and respond to exogenous vs. endogenous CDNs. Finally, the STING-R231H variant mouse represents a useful tool with which to examine the relative contributions of STING sensing of exogenous and endogenous CDNs in the context of bacterial infections and CDN-based cancer immunotherapeutics.

Expression of Ihh signaling pathway in condylar cartilage after bite-raising in adult rats.

Temporomandibular joint osteoarthritis (TMJOA) is a complex inflammatory condition with multiple factors and degenerative processes co-occurring. However, its pathogenesis remains uncertain. The purpose of the study was to observe the expression of Indian hedgehog (Ihh) signal related molecules in TMJOA induced by bite-raising and to study the effect and mechanism of Ihh signaling. Our research indicated that Ihh signaling pathway can be activated in condylar cartilage induced by bite-raising. The histological analysis showed TMJOA-like structural changes of condylar cartilage in experiment groups. Ihh, Smoothened (Smo), and Gli zinc finger transcription factors-1 (Gli-1) were activated in the experimental groups, and the expression levels increased significantly over time, whereas the sham control groups showed no fluctuation. Additionally, the expression levels of matrix metalloproteinase-13 (MMP-13) and cysteinyl aspartate specific proteinase-3 (Caspase-3) in the experiment groups increased in a time-dependent manner compared with the matched sham control groups. In conclusion, our results indicated that the Ihh signaling pathway may activate the occurrence of TMJOA by mediating the hypertrophy of chondrocytes, which may be an important regulatory mechanism and potential therapeutic target in the repair of condylar cartilage.

Serum Metabolomics Analysis in Wasp Sting Patients.

To analyze the dynamic changes of serum metabolomics in wasp sting victims, we collected serum from 10 healthy volunteers and 10 patients who had been stung 3 hours, 24 hours, and 72 hours before sample collection. We analyzed the metabolomics by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) techniques and then performed enrichment analysis. A total of 838 metabolites were identified. Serum metabolomics analysis using MetaboAnalyst revealed 289 metabolites that were significantly different among patients in the 3-hour group versus healthy controls (P<0.001). Pathway analysis of those metabolites indicated that those metabolic sets were associated with sphingolipid metabolism. Based on the differences among the control, 3-hour, 24-hour, and 72-hour groups, we classified serum metabolites into different categories. The first and second categories included 297 and 280 metabolites that were significantly different in terms of concentration among healthy controls versus the participants whose sera were analyzed 3 hours, 24 hours, and 72 hours after wasp stings. Pathway analysis of those metabolites indicated that those metabolic sets were associated with thiamine metabolism. The third category included 269 significant metabolites. The fourth category included 28 significant metabolites. Pathway analysis of the metabolites in third and fourth categories indicated that those metabolic sets were associated with phenylalanine, tyrosine, and tryptophan biosynthesis. The fifth category included 31 metabolites, which were not significantly different between the control and 3-hour groups but were higher in concentration in the 24-hour and 72-hour groups. Pathway analysis of the fifth category of significant metabolites identified linoleic acid metabolism. In conclusion, multiple metabolic pathways are associated with wasp stings, and these might provide a basis for exploring mechanisms of wasp sting injury and potential targets for therapy.

Decreased expression of STING predicts poor prognosis in patients with gastric cancer.

STING (stimulator of interferon genes) has recently been found to play an important role in host defenses against virus and intracellular bacteria via the regulation of type-I IFN signaling and innate immunity. Chronic infection with Helicobacter pylori is identified as the strongest risk factor for gastric cancer. Thus, we aim to explore the function of STING signaling in the development of gastric cancer. Immunohistochemistry was used to detect STING expression in 217 gastric cancer patients who underwent surgical resection. STING protein expression was remarkably decreased in tumor tissues compared to non-tumor tissues, and low STING staining intensity was positively correlated with tumor size, tumor invasion depth, lymph mode metastasis, TNM stage, and reduced patients' survival. Multivariate analysis identified STING as an independent prognostic factor, which could improve the predictive accuracy for overall survival when incorporated into TNM staging system. In vitro studies revealed that knock-down of STING promoted colony formation, viability, migration and invasion of gastric cancer cells, and also led to a defect in cytosolic DNA sensing. Besides, chronic H. pylori infection up-regulated STING expression and activated STING signaling in mice. In conclusion, STING was proposed as a novel independent prognostic factor and potential immunotherapeutic target for gastric cancer.

Poison and alarm: the Asian hornet Vespa velutina uses sting venom volatiles as an alarm pheromone.

In colonial organisms, alarm pheromones can provide a key fitness advantage by enhancing colony defence and warning of danger. Learning which species use alarm pheromone and the key compounds involved therefore enhances our understanding of how this important signal has evolved. However, our knowledge of alarm pheromones is more limited in the social wasps and hornets compared with the social bees and ants. Vespa velutina is an economically important and widespread hornet predator that attacks honey bees and humans. This species is native to Asia and has now invaded Europe. Despite growing interest in V. velutina, it was unknown whether it possessed an alarm pheromone. We show that these hornets use sting venom as an alarm pheromone. Sting venom volatiles were strongly attractive to hornet workers and triggered attacks. Two major venom fractions, consisting of monoketones and diketones, also elicited attack. We used gas chromatography coupled to electroantennographic detection (GC-EAD) to isolate 13 known and 3 unknown aliphatic ketones and alcohols in venom that elicited conspicuous hornet antennal activity. Two of the unknown compounds may be an undecen-2-one and an undecene-2,10-dinone. Three major compounds (heptan-2-one, nonan-2-one and undecan-2-one) triggered attacks, but only nonan-2-one did so at biologically relevant levels (10 hornet equivalents). Nonan-2-one thus deserves particular attention. However, the key alarm releasers for V. velutina remain to be identified. Such identification will help to illuminate the evolution and function of alarm compounds in hornets.

Animal toxins and renal ion transport: Another dimension in tropical nephrology.

Renal vascular and tubular ion channels and transporters involved in toxin injury are reviewed. Vascular ion channels modulated by animal toxins, which result in haemodynamic alterations and changes in blood pressure, include ENaC/Degenerin/ASIC, ATP sensitive K channels (KATP ), Ca activated K channels (Kca) and voltage gated Ca channels, mostly L-type. Renal tubular Na channels and K channels are also targeted by animal toxins. NHE3 and ENaC are two important targets. NCC and NKCC may be involved indirectly by vasoactive mediators induced by inflammation. Most renal tubular K channels including voltage gated K channels (Kv1), KATP , ROMK1, BK and SK are blocked by scorpion toxins. Few are inhibited by bee, wasp and spider venoms. Due to small envenoming, incomplete block and several compensatory mechanisms in renal tubules, serum electrolyte charges are not apparent. Changes in serum electrolytes are observed in injury by large amount of venom when several channels or transporters are targeted. Envenomings by scorpions and bees are examples of toxins targeting multiple ion channels and transporters.

A pain-inducing centipede toxin targets the heat activation machinery of nociceptor TRPV1.

The capsaicin receptor TRPV1 ion channel is a polymodal nociceptor that responds to heat with exquisite sensitivity through an unknown mechanism. Here we report the identification of a novel toxin, RhTx, from the venom of the Chinese red-headed centipede that potently activates TRPV1 to produce excruciating pain. RhTx is a 27-amino-acid small peptide that forms a compact polarized molecule with very rapid binding kinetics and high affinity for TRPV1. We show that RhTx targets the channel's heat activation machinery to cause powerful heat activation at body temperature. The RhTx-TRPV1 interaction is mediated by the toxin's highly charged C terminus, which associates tightly to the charge-rich outer pore region of the channel where it can directly interact with the pore helix and turret. These findings demonstrate that RhTx binding to the outer pore can induce TRPV1 heat activation, therefore providing crucial new structural information on the heat activation machinery.

Continuous drug release by sea anemone Nematostella vectensis stinging microcapsules.

Transdermal delivery is an attractive option for drug delivery. Nevertheless, the skin is a tough barrier and only a limited number of drugs can be delivered through it. The most difficult to deliver are hydrophilic drugs. The stinging mechanism of the cnidarians is a sophisticated injection system consisting of microcapsular nematocysts, which utilize built-in high osmotic pressures to inject a submicron tubule that penetrates and delivers their contents to the prey. Here we show, for the first time, that the nematocysts of the starlet sea anemone Nematostella vectensis can be isolated and incorporated into a topical formulation for continuous drug delivery. We demonstrate quantitative delivery of nicotinamide and lidocaine hydrochloride as a function of microcapsular dose or drug exposure. We also show how the released submicron tubules can be exploited as a skin penetration enhancer prior to and independently of drug application. The microcapsules are non-irritant and may offer an attractive alternative for hydrophilic transdermal drug delivery.

Rules and mechanisms of punishment learning in honey bees: the aversive conditioning of the sting extension response.

Honeybees constitute established model organisms for the study of appetitive learning and memory. In recent years, the establishment of the technique of olfactory conditioning of the sting extension response (SER) has yielded new insights into the rules and mechanisms of aversive learning in insects. In olfactory SER conditioning, a harnessed bee learns to associate an olfactory stimulus as the conditioned stimulus with the noxious stimulation of an electric shock as the unconditioned stimulus. Here, we review the multiple aspects of honeybee aversive learning that have been uncovered using Pavlovian conditioning of the SER. From its behavioral principles and sensory variants to its cellular bases and implications for understanding social organization, we present the latest advancements in the study of punishment learning in bees and discuss its perspectives in order to define future research avenues and necessary improvements. The studies presented here underline the importance of studying honeybee learning not only from an appetitive but also from an aversive perspective, in order to uncover behavioral and cellular mechanisms of individual and social plasticity.

Context affects nestmate recognition errors in honey bees and stingless bees.

Nestmate recognition studies, where a discriminator first recognises and then behaviourally discriminates (accepts/rejects) another individual, have used a variety of methodologies and contexts. This is potentially problematic because recognition errors in discrimination behaviour are predicted to be context-dependent. Here we compare the recognition decisions (accept/reject) of discriminators in two eusocial bees, Apis mellifera and Tetragonisca angustula, under different contexts. These contexts include natural guards at the hive entrance (control); natural guards held in plastic test arenas away from the hive entrance that vary either in the presence or absence of colony odour or the presence or absence of an additional nestmate discriminator; and, for the honey bee, the inside of the nest. For both honey bee and stingless bee guards, total recognition errors of behavioural discrimination made by guards (% nestmates rejected + % non-nestmates accepted) are much lower at the colony entrance (honey bee: 30.9%; stingless bee: 33.3%) than in the test arenas (honey bee: 60-86%; stingless bee: 61-81%; P<0.001 for both). Within the test arenas, the presence of colony odour specifically reduced the total recognition errors in honey bees, although this reduction still fell short of bringing error levels down to what was found at the colony entrance. Lastly, in honey bees, the data show that the in-nest collective behavioural discrimination by ca. 30 workers that contact an intruder is insufficient to achieve error-free recognition and is not as effective as the discrimination by guards at the entrance. Overall, these data demonstrate that context is a significant factor in a discriminators' ability to make appropriate recognition decisions, and should be considered when designing recognition study methodologies.

The importance of early bedside echocardiography in children with scorpion envenomation.

Scorpion sting may cause myocardial injury and heart failure (HF). Clinical signs of failure may develop several hours or even days after the sting, while electrocardiography (ECG) and blood examination soon after the sting may be normal. We sought to examine whether normal echocardiographic (echo) examination performed shortly after hospital arrival would exclude subsequent HF. We also sought to check if blood troponin and natriuretic peptide values measured shortly after arrival may predict or exclude subsequent HF. Natriuretic peptide activities have not been measured in scorpion sting victims. We also wanted to check if HF occurs in envenomated young infants. In a 3-year prospective study we looked at the demographic, clinical, laboratory, ECG, and echo data of all patients with general envenomation who arrived at the emergency department (ED) after scorpion sting. Clinical, laboratory, ECG, and echo results on arrival and 24 h after arrival were checked and compared between groups of patients with normal and abnormal echo on arrival. We then looked for differences in clinical course, therapy, and outcome between groups. The study included 98 children aged 80 days to 19 years (median 53.1 months), 25 were below the age of 2 years. Envenomation by the "yellow scorpion"Leiurus quinquestriatus was suspected in 74 cases. Median time between sting and ED arrival was 80 min. Echo was performed on arrival in 93 of the 98 patients, (in 5 occasions it was not performed or not recorded) 74 were normal and 19 were abnormal. Abnormal echo included hypokinesia and low fractional shortening and ejection fraction of the left ventricle. Clinical signs, abnormal ECG, and laboratory results were not discriminative between groups on arrival. Mean troponin T was higher in patients with abnormal echo, but within normal range in 13 of the 19 patients with abnormal echo and above normal in 2 of the 74 patients with normal echo-missing sensitivity and specificity. Mean N-terminal pro B-type natriuretic peptide was above normal in both groups but within normal range in 5 patients with abnormal echo and above normal range in 24 patients with normal echo-missing sensitivity and specificity. None of the patients with normal echo had subsequent HF and none of the children younger than 2 years of age had HF. All patients survived the intoxication and were discharged home without sequel. We conclude that early echo examination is an important procedure. In our study, normal examination excluded subsequent HF. Abnormal examination accelerated cardiac therapy which might have contributed to our favorable outcome. HF did not occur in infants younger than two years of age.

Biochemistry of envenomation.

Venoms and toxins are of significant interest due to their ability to cause a wide range of pathophysiological conditions that can potentially result in death. Despite their wide distribution among plants and animals, the biochemical pathways associated with these pathogenic agents remain largely unexplored. Impoverished and underdeveloped regions appear especially susceptible to increased incidence and severity due to poor socioeconomic conditions and lack of appropriate medical treatment infrastructure. To facilitate better management and treatment of envenomation victims, it is essential that the biochemical mechanisms of their action be elucidated. This review aims to characterize downstream envenomation mechanisms by addressing the major neuro-, cardio-, and hemotoxins as well as ion-channel toxins. Because of their use in folk and traditional medicine, the biochemistry behind venom therapy and possible implications on conventional medicine will also be addressed.

Receptor-targeting mechanisms of pain-causing toxins: How ow?

Venoms often target vital processes to cause paralysis or death, but many types of venom also elicit notoriously intense pain. While these pain-producing effects can result as a byproduct of generalized tissue trauma, there are now multiple examples of venom-derived toxins that target somatosensory nerve terminals in order to activate nociceptive (pain-sensing) neural pathways. Intriguingly, investigation of the venom components that are responsible for evoking pain has revealed novel roles and/or configurations of well-studied toxin motifs. This review serves to highlight pain-producing toxins that target the capsaicin receptor, TRPV1, or members of the acid-sensing ion channel family, and to discuss the utility of venom-derived multivalent and multimeric complexes.

Myotoxicity and nephrotoxicity by Micrurus venoms in experimental envenomation.

Micrurus venoms are essentially neurotoxic but other activities, such as myotoxicity, may be apparent under experimental conditions. Although this myotoxicity has been occasionally reported, there are no studies addressing it systematically across the genus, particularly in its relationship to other systemic manifestations such as renal impairment. The lethal potency of Micrurus fulvius, Micrurus nigrocinctus, Micrurus surinamensis, Micrurus altirostris, Micrurus balyocoriphus and Micrurus pyrrhocryptus venoms determined by us were in the range described for the genus and all venoms exhibited phospholipase activity, albeit at significantly different levels. Intramuscular venom injection caused variable local inflammation-edema; myotoxicity (as determined by plasma creatine kinase levels and histopathology) was apparent only in those venoms with highest phospholipase activity, namely M. fulvius, M. nigrocinctus and M. pyrrhocryptus. Kidneys of animals injected with these strongly myotoxic venoms showed lesions consisting in extensive tubular necrosis with nuclear fragmentation, destruction of the brush border, rupture of basal membrane and epithelial exfoliation of tubular cells, granular cast and thickening of tubules. The histological characteristics of the lesions suggest an important role for indirect glomerular damage by myoglobin deposits. Phospholipase and myotoxic activities did not correlate significantly to the lethal potency; renal lesions were, however, evident only in those venoms that caused extensive muscular damage. Although kidney lesions have not been described in clinical cases of Micrurus envenomation, the potential for nephrotoxicity of some of these venoms should be considered in the overall toxicological picture, at least in experimental conditions.

Salivary glands as the source of plasma brain-derived neurotrophic factor in stressed rats engaged in biting behavior.

Brain-derived neurotrophic factor (BDNF) is crucial for the survival and differentiation of the central and peripheral nervous systems. Recently, BDNF has been reported to exert broader biological activity on non-neural cells. A previous study examined the effect of immobilization stress on BDNF and its receptor tyrosine receptor kinase B in male rat submandibular glands. In the present study, we found that the rat submandibular gland is the major source of plasma BDNF during acute immobilization stress. Biting modulates the mRNA and protein levels of BDNF in the rat hippocampus, so we also investigated whether the plasma BDNF concentration is influenced by biting. Two hours of acute immobilization stress significantly increased the amount of BDNF mRNA within the rat submandibular glands. Moreover, allowing biting behavior for the second half of the 2-h stress exposure significantly increased the amount of salivary gland BDNF mRNA relative to stress alone. Similar results were found with plasma BDNF concentrations under the same conditions. We confirmed that biting during stress attenuates the increases in plasma adrenocorticotropic hormone and corticosterone concentrations, but this was not dependent on the submandibular glands. Increased BDNF, mRNA and protein expressions were observed in salivary duct cells as a result of immobilization stress and biting behavior, as demonstrated by real-time polymerase chain reaction, immunohistochemistry, western blotting, and enzyme-linked immunosorbent assay. Taken together, the findings indicate that the submandibular glands evidently contribute to the increase in plasma BDNF upon biting.