Therapeutic outcome of quercetin nanoparticles on Cerastes cerastes venom-induced hepatorenal toxicity: a preclinical study.

Aim: The objective of this study was to investigate the therapeutic potential of quercetin (QT) and QT-loaded poly(lactic-co-glycolic acid) nanoparticles (QT-NPs) on Cerastes cerastes venom-mediated inflammation, redox imbalance, hepatorenal tissue damage and local hemorrhage. Methods: The developed QT-NPs were first submitted to physicochemical characterization and then evaluated in the 'challenge then treat' and 'preincubation' models of envenoming. Results: QT-NPs efficiently alleviated hepatorenal toxicity, inflammation and redox imbalance and significantly attenuated venom-induced local hemorrhage. Interestingly, QT-NPs were significantly more efficient than free QT at 24 h postenvenoming, pointing to the efficacy of this drug-delivery system. Conclusion: These findings highlight the therapeutic potential of QT-NPs on venom-induced toxicity and open up the avenue for their use in the management of snakebite envenoming.

Therapeutic Outcome of Anti-inflammatory and Antioxidative Medicines on the Dermonecrotic Activity of Cerastes cerastes Venom.

Envenomation by Cerastes cerastes often results in local dermonecrotic lesions. While immunotherapy is effective in reversing systemic symptoms, this strategy remains deficient in counteracting the extended dermonecrosis induced from the bite site. In this study, the therapeutic effect of pharmacological drugs on the dermonecrotic activity of the venom was investigated. Venom administration caused a marked dermonecrotic lesion with increased levels of oxidative stress biomarkers (MPO, EPO, NO, H2O2, MDA, protein carbonyl, and thiol levels). Antioxidant capacity was decreased, as evidenced by reduced catalase, glutathione, and selenium levels. Histopathological analysis of skin biopsies revealed necrotic lesions accompanied by hemorrhage and epidermis thickening. The efficiency of cyproheptadine (C), dexamethasone (D), and tetracycline (T), as a monotherapy or in association, were evaluated on the dermonecrotic activity of the venom. Most of the treatments (CD, CT, DT, and CDT) largely reduced tissue necrosis to, respectively, 84.29, 87.83, 83.77, and 82.71% and significantly decreased MPO and EPO activities and NO, H2O2, MDA, and protein carbonyl levels in skin tissue homogenates. CT and CDT associations significantly increased the antioxidant status as indicated by enhanced catalase, glutathione, and selenium levels. The second challenge of the pharmacological associations was more effective in improving the oxidative/antioxidative balance. Skin tissue sections from treated animals with CT or CDT revealed tissue structure close to that observed in control animals. Therefore, the synergistic action of all tested drugs on the major pathways of inflammation (phospholipases A2, metalloproteinases, and histamine) seems to be efficient to neutralize the necrotic activity of the venom.

Mast Cells Modulate the Immune Response and Redox Status of the Gastrointestinal Tract in Induced Venom Pathogenesis.

The pathogenesis of Androctonus autralis hector (Aah) scorpion venom involved cellular and molecular mechanisms resulting in multi-organ dysfunction. However, little is reported about the effects of venom on the gastrointestinal axis. Mast cells (MCs) are known to play a crucial role in modulating immune response of the gut. This study aims to investigate the involvement of this cell type in venom-induced gastric and intestinal disorders in a time course (3 and 24h). The obtained results revealed that Aah scorpion venom induced inflammatory cell infiltration as shown by the increase of the myeloperoxidase and eosinophil peroxidase activities. Overexpression of the c-kit receptor (CD117) severely imbalanced the redox status with depletion of antioxidant systemic accompanied by gastrointestinal tissue damage. Moreover, an increased level of lactate dehydrogenase in the serum was correlated with tissue injuries. Pharmacological inhibition of MCs targeting tyrosine kinase (TK) reduces the generation of reactive oxygen species and normalizes catalase, and gluthation S-transferase activities to their physiological levels. In addition, histopathological alterations were restored after pretreatment with c-kit receptor inhibitor associated with a considerable reduction of MC density. Interestingly, obtained results indicate that MCs might be involved in gastric modulation and intestinal inflammation through c-kit signaling following sub-cutaneous Aah venom injection.

Bioactive Molecules Derived from Snake Venoms with Therapeutic Potential for the Treatment of Thrombo-Cardiovascular Disorders Associated with COVID-19.

As expected, several new variants of Severe Acute Respiratory Syndrome-CoronaVirus-2 (SARS-CoV-2) emerged and have been detected around the world throughout this Coronavirus Disease of 2019 (COVID-19) pandemic. Currently, there is no specific developed drug against COVID-19 and the challenge of developing effective antiviral strategies based on natural agents with different mechanisms of action becomes an urgent need and requires identification of genetic differences among variants. Such data is used to improve therapeutics to combat SARS-CoV-2 variants. Nature is known to offer many biotherapeutics from animal venoms, algae and plant that have been historically used in traditional medicine. Among these bioresources, snake venom displays many bioactivities of interest such as antiviral, antiplatelet, antithrombotic, anti-inflammatory, antimicrobial and antitumoral. COVID-19 is a viral respiratory sickness due to SARS-CoV-2 which induces thrombotic disorders due to cytokine storm, platelet hyperactivation and endothelial dysfunction. This review aims to: (1) present an overview on the infection, the developed thrombo-inflammatory responses and mechanisms of induced thrombosis of COVID-19 compared to other similar pathogenesis; (2) underline the role of natural compounds such as anticoagulant, antiplatelet and thrombolytic agents; (3) investigate the management of coagulopathy related to COVID-19 and provide insight on therapeutic such as venom compounds. We also summarize the updated advances on antiviral proteins and peptides derived from snake venoms that could weaken coagulopathy characterizing COVID-19.

Long-term antibody response and protective effect induced by attenuated scorpion toxins: Involvement of memory plasma cells.

In Algeria, Androctonus australis hector scorpion envenomation remains a major problem of public health because of non-efficient therapy. The development of safe vaccine against scorpion venom could be one key strategy for the envenomation prevention. The irradiation of venom by γ-rays develops suitable immunogens which produced effective antivenom and safe vaccine. In this study, we investigated the ability of the irradiated toxic fraction (γ-FtoxG50) to induce long-term memory humoral response in immunized animals (mice and rabbits), by involving the long-lived plasma cells to prevent efficiently the lethality of scorpion envenomation. For this purpose, an appropriate immunization schedule was established in mice and rabbits using three (3) similar doses of γ-FtoxG50 associated with Alum adjuvant. Obtained results indicate that the long-term immunogenicity of γ-FtoxG50 is able to induce the long-term memory humoral response with a high level of specific antibodies. The long-term persistence of antibody levels could depend on bone marrow memory plasma cells. These cells produce continuously antibodies without antigen stimulus. Furthermore, an enhanced memory response was obtained post-repeated envenomation with toxic native venom that leads to improved protection of animals. Together, pre-existing protective antibodies and the activation of memory B-cells could induce a rapid neutralization of scorpion toxins and long-term protection against scorpion envenomation.

"Computational and Functional Characterization of a Hemorrhagic Metalloproteinase Purified from Cerastes cerastes Venom".

Structural and functional aspects of snake venoms metalloproteinases (SVMPs) have been extensively studied due to their role in envenomation. However, in the detection of certain coagulation disorders these biomolecules have been used and applied for the production of new thrombolytic drugs. CcMP-II, a SVMP-II metalloproteinase with a hemorrhagic activity, isolated from the venom of Cerastes cerastes, its sequence of 472 amino acids was identified. Predicted 3D structure showed an arrangement of CcMP-II into two distinct domains: i) a metalloproteinase domain where the zinc-binding motif is found (HXXGHNLGIDH) in addition to the sequence Cys-Ile-Met (CIM) at the Met-turn and ii) disintegrin-like domain containing RGD motif. CcMP-II inhibits platelet aggregation induced by collagen in a dose-dependent manner with an IC50 value estimated of 0.11 nM. This proteinase inhibits also aggregation of platelet stimulated by collagen even if the metal chelating agents (EDTA and 1, 10-phenontroline) are present suggesting that anti-aggregating effect is not due to its metalloproteinase domain, but to its disintegrin-like domain. Capillary pathological modifications caused by CcMP-II following intramuscular injection have as well been examined in mice. The key morphological alterations of the capillary vessels were clearly apparent from the ultrastructural study. The CcMP-II can play a key function in the pathogenesis of disorders that occurs following envenomation of Cerastes cerastes. The three-dimensional model of CcMP-II was built to explain structure-function relationships in ADAM/ADAMTs, a family of proteins having significant therapeutic potential. In order to explain structure-function relationships in ADAM / ADAMT, a family of proteins with considerable therapeutic potential, the three-dimensional model of CcMP-II was constructed.

Immunomodulatory and protective effects of interleukin-4 on the neuropathological alterations induced by a potassium channel blocker.

The pathophysiology of neurological diseases related to potassium-channel dysfunction such as epilepsy is increasingly linked to immune system modulation. However, there are limited reports of which interleukin-4 (IL-4) can act on the neuroinflammatory response after seizure. Hence, we evaluated the effect of IL-4 in murine model of neuroexcitotoxcity using kaliotoxin (KTx), a potassium-channel blocker. Results showed that IL-4 treatment can significantly reduce the neuronal death induced by KTx. Probably by decreasing mitochondria swelling, reversing oxidative damage and enhancing Bcl-2 expression. Furthermore, IL-4 treatment significantly reduced TNF-α expression and enhanced GFAP and IL-10 expressions in the brain. IL-4 can be neuroprotective in epileptogenesis.

Development and evaluation of polymeric nanoparticles as a delivery system for snake envenoming prevention.

Polymer-based nanoparticles have become an appealing carrier for improving vaccine delivery efficiency. In this study, we investigated an interesting approach based on PLGA nanoparticles encapsulating Cerastes cerastes venom as an intranasal vaccine delivery system for snake envenomation prevention. Particles were synthesized by double emulsion solvent evaporation method and characterized for their size, morphology, distribution, and venom-nanoparticles interactions. An immunization trial was performed in mice by the intranasal route to evaluate the immune response, the reactogenicity, and the protective effect of this nanovaccine. The physicochemical and structural characteristics of Cc-loaded PLGA NPs revealed that the particles exhibited a spherical shape with a diameter of 370 nm, and a negatively charged surface with a zeta potential value of 19,9 mV. The immunization with Cc-PLGA NPs can induce a systemic innate and humoral immune response and confers protection against Cerastes cerastes venom (Cc) over than 6 LD50 with a cross-protection against Vipera lebetina venom (Vl) over than 5 LD50. Nano-encapsulation of Cc venom reduced its toxicity and the induced tissue alterations. Our results confirm that the nano-formulation Cc-PLGA NPs is a potent adjuvant system that improve the humoral immune response and provide protection against high lethal doses of viper venoms.

Isolation and Characterization of CD39-like Phosphodiesterase (Cc-PDE) from Cerastes cerastes Venom: Molecular Inhibitory Mechanism of Antiaggregation and Anticoagulation.

BACKGROUND: Cerastes cerastes venom contains several bioactive proteins with inhibitory potential of platelet aggregation and blood coagulation. OBJECTIVE: The current study deals with purification, characterization and determination of structural properties of Cc-PDE, the first phosphodiesterase from Cerastes cerastes venom. MATERIAL AND METHODS: The purification process consists of three successive chromatographies including G75-Sephadex size exclusion, DEAE exchange chromatography and affinity using Sildenafil as a main PDEs' specific inhibitor. The amino acid sequence of purified Cc-PDE was determined by liquid chromatography coupled off line to MALDI-TOF/TOF. Modeling and structural features were obtained using several bioinformatics tools. In vivo and in vitro antiplatelet aggregation and anticoagulant assays were performed. RESULTS: Cc-PDE (73 506.42 Da) is a 654-residue single polypeptide with 1-22 signal peptide and it is characterized by the presence of predominant basic amino acids suitable to alkaline pI (8.17). Cc-PDE structure is composed of ß-strands (17%) and α-helices (24%) and it shares a high identity with homologous snake venom PDEs. Cc-PDE hydrolyzes both Bis-p-nitrophenyl phosphate (Km = 2.60 ± 0.95 mM, Vmax = 0.017 ± 0.002569 µmol.min-1) and p-nitrophenyl phosphate (Km = 7.13 mM ± 0.04490 mM, Vmax = 0.053 ±0.012 µmol.min-1). Cc-PDE prevents ADP- and ATP-induced platelet aggregation by hydrolyzing ADP and ATP, reducing surface P-selectin expression and attenuating platelet function. In addition, Cc-PDE inhibits coagulation factors involved in the intrinsic pathway demonstrated by a significant prolongation of activated partial thromboplastin time and in vivo long-lasting anticoagulation. CONCLUSION: The obtained results revealed that Cc-PDE may have a therapeutic potential and could be a remedy for thromboembolic diseases as an alternative of anticoagulant and antiplatelet aggregation chemical origins.

Valorisation potentielle des biomolécules issues des venins de scorpions contre la COVID-19 / Potential role of bioactive molecules from scorpion venoms against COVID-19

Les produits d?origine naturelle sont considérés comme une source précieuse de substances médicamenteuses et constituent une grande diversité structurelle et fonctionnelle. Les venins, de par les propriétés de leurs composés représentent pour l?animal venimeux un moyen de défense contre des prédateurs et d?immobilisation de leurs proies. Ces venins sont responsables des effets néfastes induits lors d?une envenimation en induisant une stimulation du système nerveux autonome par des peptides toxiques, une libération massive des neurotransmetteurs et une réponse inflammatoire systémique excessive. Ces processus sont à l?origine des dommages tissulaires et d?une défaillance multiviscérale. Par ailleurs, certains des composants des venins représentent un potentiel pharmacologique d?intérêt dans le développement de nouveaux médicaments pouvant être utilisés dans plusieurs pathologies telles que la COVID-19. La COVID-19 est une maladie infectieuse transmissible. Lors d?une infection, le virus SARS-CoV-2 induit une réaction immunitaire inadaptée associée à un orage cytokinique et à une coagulopathie, responsables d?un syndrome de détresse respiratoire dans les cas les plus graves. La pandémie de la COVID-19 a causé à travers le monde et ce depuis fin 2019, une crise sanitaire sans précédent et d'énormes pertes économiques. Dans cette revue bibliographique, les principales avancées scientifiques concernant essentiellement la réponse immunitaire et les mécanismes immunopathologiques induits lors d?une infection avec le SARS-CoV-2 seront décrites. Le rôle bénéfique des biomolécules isolées à partir des venins sera également rapporté pour une éventuelle utilisation de nouvelles biomolécules contre la COVID-19.

Natural products are considered as an excellent source of novel drugs and continue to provide greater structural and functional diversity. Venoms, by the properties of their compounds, are used by venomous animal for their defense against predators and to immobilize the prey. These venoms are responsible for the harmful effects induced during envenomation by inducing stimulation of the autonomic nervous system by toxic peptides, massive release of neurotransmitters and an excessive systemic inflammatory response. These processes cause tissue damage and multiple organ failure. However, some of the components of the venoms represent a valuable pharmacological potential in the development of new drugs that can be used in several pathologies such as COVID-19. The infectious, COVID-19 disease, caused by SARS-CoV-2, can lead to an inappropriate immune response, consequently causing cytokine storm and coagulopathy, responsible for respiratory distress syndrome in the most severe cases. Since its onset, the COVID-19 pandemic has spread over the world; it has not only impacted human health, but also the individual behavior, the functioning of society and the economy of all countries. This review provides an overview of the current evidence regarding immune response and the immune-pathological mechanisms involved during infection with SARS-CoV-2. The beneficial role of biomolecules isolated from venoms will be also presented as potential candidate molecules against COVID-19.

Isolation and Functional Identification of an Antiplatelet RGD-Containing Disintegrin from Cerastes cerastes Venom.

The current report focuses on purification, structural and functional characterization of Cerastategrin from Cerastes cerastes venom, a novel basic disintegrin (pI 8.36) with 128 amino acid residues and a molecular weight of 13 835.25 Da measured by MALDI-MSMS. The 3D structure of Cerastategrin is organized as α-helix (13%), ß-strand (15%) and disordered structure (30%) and presents homologies with several snake venom disintegrins. Structural modeling shows that Cerastategrin presents an RGD motif that connects specifically to integrin receptors. Cerastategrin exhibits the inhibition of ADP induced platelets with an IC50 of 0.88 µg/mL and shows in vivo long stable anticoagulation effect 24 h post-injection of increasing doses ranging from 0.2 to 1 mg/kg, therefore, Cerastategrin maintained irreversibly the blood incoagulable. Moreover, Cerastategrin decreases the amount of bounded αIIbß3 and reduced significantly the quantity of externalized P-Selectin. Cerastategrin acts as a molecule targeting specifically the receptor αIIbß3; therefore, it behaves as a potent platelet activation inhibitor. As a new peptide with promising pharmacological properties, Cerastategrin could have a potential therapeutical effect in the vascular pathologies and may be a new effective treatment approach.

Involvement of the Endothelin Receptor Type A in the Cardiovascular Inflammatory Response Following Scorpion Envenomation.

Elevated levels of endothelin-1 (ET-1) were recorded in sera of scorpion sting patients. However, no studies focused on the mechanism of ET-1 involvement in the pathogenesis of scorpion envenomation, particularly in the cardiovascular system which is seriously affected in severe cases of scorpion stings. Inflammation induced by Androctonus australis hector (Aah) scorpion venom in the heart together with the aorta was studied in mice pretreated with a specific endothelin A receptor (ETA-R) inhibitor. ETA-R inhibition resulted in the attenuation of the high amounts of cytokine (tumor necrosis factor alpha (TNF-α) and interleukin-17 (IL-17)) recorded in the sera of envenomed mice. The recovery of the oxidative stress marker balance and matrix metalloproteinase (MMP) expression were also observed, concomitantly with the reduction of tissular neutrophil infiltration. Additionally, the cardiac and the aortic tissue alterations, and the metabolic enzymes (creatine kinase (CK) and muscle-brain isoform creatine kinase (CK-MB)) overspread into sera were significantly attenuated. Obtained results suggest the implication of endothelin throughout its ETA receptors in the inflammatory response observed in the cardiovascular components during scorpion envenomation. Further knowledge is needed to better understand the implication of the endothelin axis and to improve the therapeutic management of severe scorpion sting cases.

Immune-toxicity effects of scorpion venom on the hypothalamic pituitary adrenal axis during rest and activity phases in a rodent model.

Scorpion venom is a complex mixture of peptides and proteins, rich in toxins. Its toxicological effects are related to central disruptions and autonomic disturbances, organ failure, as well as an excessive systemic inflammatory response. Since the role of the hypothalamic pituitary adrenal (HPA) axis is central in the neuroendocrine-immunological axis, the purpose of this study was, therefore, to examine the immunotoxic effect of Androctonus australis hector (Aah) venom on HPA-axis in synchronised-mice model. Taking into account the circadian activity of the HPA-axis, the variations of adrenocorticotropic hormone and corticosterone plasma levels, oxidative stress as well as inflammatory markers in cerebral, hypothalamic and adrenal tissue homogenates were investigated during the rest and activity phases of animals. Histopathology study was also performed. Results showed that Aah venom activated the HPA axis. This response seems to be dependent on time of envenomation, as a higher hormone levels were more operative during the active phase than in the rest phase when compared to time-matched control. The local toxicity-effects following Aah envenomation revealed an imbalance in oxidative stress with a higher antioxidant defences in darkness hypothalamic and cerebral tissues. Furthermore, there were significantly higher levels in vascular permeability in hypothalamic and cerebral tissues accompanied by a concomitant increase in immune-cell infiltration and/or activation as shown by expression of CD68 and myeloperoxidase activity during the active phase compared with the rest phase. Overall results suggested that Aah venom had a toxic impact on different HPA-axis areas and the effect varies according to the time of envenomation.

Chitosan nanoparticles as a delivery platform for neurotoxin II from Androctonus australis hector scorpion venom: Assessment of toxicity and immunogenicity.

In recent years, biodegradable polymers based nanoparticles received high interest for the development of vaccine delivery vehicles. In this study, chitosan nanoparticles encapsulating Aah II toxin (AahII-CNPs) isolated from Androctonus australis hector venom, were investigated as vaccine delivery system. Particles obtained by ionotropic gelation were characterized for their size, surface charge, morphology and toxin release profile from Aah II-CNPs. Toxin-nanoparticles interactions were assessed by Fourier Transform Infrared Spectrometry and X-Ray Diffraction. An immunization protocol was designed in mice to investigate anti-toxin immunity and the protective status induced by different Aah II immune formulations. Unloaded chitosan nanoparticles presenting a spherical shape and smooth surface, were characterized by a size of 185 nm, a dispersion index (PDI) of 0.257 and a zeta potential of +34.6 mV. Aah II toxin was successfully entrapped into chitosan nanoparticles as revealed by FTIR and XRD data. Entrapment efficiency (EE) and Loading capacity (LC) were respectively of 96.66 and 33.5%. Aah II-CNPs had a diameter of 208 nm, a PDI of 0.23 and a zeta potential of +30 mV. Encapsulation of Aah II reduced its toxicity and protected mice until 10 LD50. Mice were immunized via a dual prime-boost scheme. Nanoentrapped Aah II immunogen elicited systemic innate and humoral immune responses as well as local spleen parenchyma hyperplasic alterations. Aah II-CNPs immunized mice withstood high lethal doses of native Aah II, one-month post-boost inoculation. This study provided encouraging and promising results for the development of preventive therapies against scorpion envenoming mainly for the populations at-risk.

Involvement of Toll-like Receptor 4 in Neutrophil-Mediated Inflammation, Oxidative Stress and Tissue Damage Induced by Scorpion Venom.

Systemic inflammatory response and generation of oxidative stress are known to contribute to scorpion venom-induced tissue damage. TLR receptors might represent a link between oxidative stress and inflammation; we therefore investigated whether or not TLR4 is involved in venom-induced immunopathology. The obtained results showed that pharmacological targeting of TLR4 with the selective inhibitor TAK-242 (Resatorvid) prevents the inflammatory response induced by subcutaneous administration of Androctonus australis hector (Aah) venom, as revealed by a significant decrease of neutrophil cell count in peripheral blood associated with significant decline of neutrophil degranulation and sequestration to the lung, liver, and kidney tissues. Moreover, TAK-242 administration inhibited nitrite levels increase in serum, malondialdehyde (MDA), and protein carbonyl tissue contents concomitantly with a significant increase of catalase activity and reduced glutathione (GSH) level in tissue homogenates. Furthermore, venom-induced increases in serum levels of organ dysfunction markers (lactate deshydrogenase, aminotransferase ALT and AST, creatinine and urea) were also significantly suppressed by pre-treatment with TLR4 inhibitor, concordantly with a remarkable improvement in the histological features in lung and liver tissues. The results of the present study indicate the potential role of TLR4 in venom-induced immunopathology and show the in vivo requirement of TLR4 signaling in mediating venom-induced tissue damage.

Myotoxicity induced by Cerastes cerastes venom: Beneficial effect of heparin in skeletal muscle tissue regeneration.

Myonecrosis is a relevant tissue damage induced by snakes of Viperidae family often leading to permanent tissue and function loss and even amputation. The aim of this study was to evaluate the effect of heparin on skeletal muscle tissue regeneration after Cerastes cerastes envenomation. Mice received either the venom (1 LD50) by i.m. route, or the venom followed, by heparin administration by i.v. route at 15 min and 4 h. Obtained results showed that Cerastes cerastes venom induced deep tissue structure alterations, characterized mainly by edema, hemorrhage, myonecrosis and inflammation. Myotoxicity was correlated with increased CK levels in sera, concomitant with their decrease in muscle tissue homogenates. Muscle wet weight was restored within 2 weeks after heparin treatment and 28 days in the envenomed group. Heparin treatment significantly decreased MPO activity, suggesting an anti-inflammatory effect. NO, HGF, VEGF and G-CSF levels were increased after heparin administration. These mitogenic factors constitute potent stimuli for satellite and endothelial cells improving, thus, muscle regeneration. This study showed that muscle tissue recovery was significantly enhanced after heparin treatment. Heparin use seems to be a promising therapeutic approach after viper envenomation.

Purification and characterization of a thrombin-like enzyme isolated from Vipera lebetina venom: its interaction with platelet receptor.

: Snake venoms contain various molecules that can be used as tools in the diagnosis and in the treatment of hemostatic disorders. This study reports the isolation and functional characterization of a new thrombin-like enzyme and its role in the modulation of platelet aggregation and coagulation. The molecule was purified by gel filtration, anion exchange chromatography and reverse-phase-HPLC on C8 column; its molecular weight was determined. Natural and synthetic substrates were used to evaluate its enzymatic activities. The fibrinogenolytic activity was tested electrophoretically and by reverse-phase-HPLC on C18 column. Otherwise, the effect on blood coagulation and deficient plasma factors were also evaluated. The mechanism by which a thrombin-like enzyme VLCV (thrombin-like enzyme)-induced platelet aggregation was explored in presence of ticlopidin, clopidogrel and aspirin. VLCV (45 kDa) isolated from Vipera lebetina as a thrombin-like enzyme seems to be able to modulate platelet function. This enzyme showed an amidolytic activity by hydrolyzing the chromogenic-specific substrate of thrombin and the α-chain of fibrinogen. It is also able to clot human plasma and the deficient human plasma in factor X, suggesting that it is involved in the intrinsic and common pathways. The aggregating effect of VLCV is more sensitive to ticlopidine than to the clopidogrel suggesting the involvement of ADP/P2Y12/PI3K pathway. VLCV seems to be able to promote human platelet aggregation suggesting an interaction between P2Y12 and PAR1. Due to its ability to replace the missing factor X and its proaggregating activity, VLCV could be used as molecular tool to better understand the hemostasis mechanism.

Cerebrospinal inflammatory response following scorpion envenomation: role of histamine H1 and H3 receptors.

BACKGROUND: The mechanism of the inflammatory process induced by scorpion venom in the cerebrospinal tissues has not yet been completely elucidated. Therefore, we aimed to investigate the role of histamine through its H1 and H3 receptors in this process. METHODS: Histamine H1 and H3 receptor antagonists, Hydroxyzine (10 mg/kg) and Betaserc (20 mg/kg), respectively, were administered by intraperitoneal route to mice 1 h before subcutaneous envenomation with a subletal dose (0.5 mg/kg) of Androctonus australis hector venom. Cerebrospinal inflammation response was assessed 24 h after envenomation by evaluating the vascular permeability changes, inflammatory cell infiltration, oxidative/nitrosative stress marker levels (hydrogen peroxide, nitric oxide, malondialdehyde, glutathione and catalase) and by histological examination of cerebrospinal tissue. RESULTS: Envenomed mice displayed an installation of an inflammatory response marked by increased vascular permeability (76% and 68% in brain and spinal cord, respectively, in comparison to controls), inflammatory cell infiltration, increased pro-oxidant levels and decreased anti-oxidant markers (p  < 0.05 to p  < 0.001). Scorpion venom also induced structural changes in brain and spinal cord tissues. Hydroxyzine seemed to be more efficient than Betaserc in the prevention of the induced cerebrospinal inflammation response, as evidenced by the decreased vascular permeability, inflammatory cell infiltration, pro-oxidant levels, increased anti-oxidant defense (p  < 0.05 to p  < 0.001) and a reduction of the anatomo-pathological alterations. CONCLUSION: The results showed that the histamine H1 receptor is more involved in the induced central nervous system inflammatory response during scorpion envenomation.

Serotherapy against Voltage-Gated Sodium Channel-Targeting αToxins from Androctonus Scorpion Venom.

Because of their venom lethality towards mammals, scorpions of the Androctonus genus are considered a critical threat to human health in North Africa. Several decades of exploration have led to a comprehensive inventory of their venom components at chemical, pharmacological, and immunological levels. Typically, these venoms contain selective and high affinity ligands for the voltage-gated sodium (Nav) and potassium (Kv) channels that dictate cellular excitability. In the well-studied Androctonusaustralis and Androctonusmauretanicus venoms, almost all the lethality in mammals is due to the so-called α-toxins. These peptides commonly delay the fast inactivation process of Nav channels, which leads to increased sodium entry and a subsequent cell membrane depolarization. Markedly, their neutralization by specific antisera has been shown to completely inhibit the venom's lethal activity, because they are not only the most abundant venom peptide but also the most fatal. However, the structural and antigenic polymorphisms in the α-toxin family pose challenges to the design of efficient serotherapies. In this review, we discuss past and present accomplishments to improve serotherapy against Androctonus scorpion stings.

Differential effect of Androctonus australis hector venom components on macrophage KV channels: electrophysiological characterization.

Neurotoxins of scorpion venoms modulate ion channels. Voltage-gated potassium (KV) channels regulate the membrane potential and are involved in the activation and proliferation of immune cells. Macrophages are key components of the inflammatory response induced by scorpion venom. The present study was undertaken to investigate the effect of Androctonus australis hector (Aah) venom on KV channels in murine resident peritoneal macrophages. The cytotoxicity of the venom was assessed using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) -based assay and electrophysiological recordings were performed using the whole-cell patch clamp technique. High doses of Aah venom (50, 125, 250 and 500 µg/ml) significantly decreased cell viability, while concentrations of 0.1-25 µg/ml were not cytotoxic towards peritoneal macrophages. Electrophysiological data revealed a differential block of KV current between resting and LPS-activated macrophages. Aah venom significantly reduced KV current amplitude by 62.5 ± 4.78% (n = 8, p < 0.05), reduced the use-dependent decay of the current, decreased the degree of inactivation and decelerated the inactivation process of KV current in LPS-activated macrophages. Unlike cloned KV1.5 channels, Aah venom exerted a similar blocking effect on KV1.3 compared to KV current in LPS-activated macrophages, along with a hyperpolarizing shift in the voltage dependence of KV1.3 inactivation, indicating a direct mechanism of current inhibition by targeting KV1.3 subunits. The obtained results, demonstrating that Aah venom differentially targets KV channels in macrophages, suggest differential outcomes for their inhibitions, and that further investigations of scorpion venom immunomodulatory potential are required.