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Made available in DSpace on 2022-04-29T08:33:17Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-10-30 Acute kidney injury pathogenesis in envenoming by snakes is multifactorial and involves immunologic reactions, hemodynamic disturbances, and direct nephrotoxicity. Sildenafil (SFC), a phosphodiesterase 5 inhibitor, has been reported to protect against pathological kidney changes. Objective: This study aimed to investigate the protective effect of sildenafil against Bothrops alternatus snake venom (BaV)-induced nephrotoxicity. Methods: Kidneys from Wistar rats (n = 6, weighing 260–300 g) were isolated and divided into four groups: (1) perfused with a modified Krebs-Henseleit solution (MKHS) containing 6 g% of bovine serum albumin; (2) administered 3 μg/mL SFC; (3) perfused with 3 μg/mL BaV; and (4) administered SFC + BaV, both at 3 μg/mL. Subsequently, the perfusion pressure (PP), renal vascular resistance (RVR), urinary flow (UF), glomerular filtration rate (GFR), and percentage of electrolyte tubular sodium and chloride transport (%TNa+, %TCl−, respectively) were evaluated. The cyclic guanosine monophosphate (cGMP) levels were analyzed in the perfusate, and the kidneys were removed to perform oxidative stress and histopathological analyses. Results: All renal parameters evaluated were reduced with BaV. In the SFC + BaV group, SFC restored PP to normal values and promoted a significant increase in %TNa+ and %TCl−. cGMP levels were increased in the SFC + BaV group. The oxidative stress biomarkers, malondialdehyde (MDA) and glutathione (GSH), were reduced by BaV. In the SFC + BaV group, a decrease in MDA without an increase in GSH was observed. These findings were confirmed by histological analysis, which showed improvement mainly in tubulis. Conclusion: Our data suggest the involvement of phosphodiesterase-5 and cGMP in BaV-induced nephrotoxicity since its effects were attenuated by the administration of SFC. Department of Physiology and Pharmacology School of Medicine Federal University of Ceara, Coronel Nunes de Melo St., 1127 Drug Research and Development Center (NPDM) Federal University of Ceara, Coronel Nunes de Melo St., 1000 Department of Dental Clinic School of Pharmacy Dentistry and Nursing Federal University of Ceara, Monsenhor Furtado St. Center for the Study of Venoms and Venomous Animals Fazenda Experimental Lageado São Paulo State University, José Barbosa de Barros St. 1780 Center for the Study of Venoms and Venomous Animals Fazenda Experimental Lageado São Paulo State University, José Barbosa de Barros St. 1780

Made available in DSpace on 2022-04-29T08:32:34Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-10-15 Snake venoms are substances mostly composed by proteins and peptides with high biological activity. Local and systemic effects culminate in clinical manifestations induced by these substances. Pain is the most uncomfortable condition, but it has not been well investigated. This review discusses Bothrops snakebite-induced nociception, highlighting molecules involved in the mediation of this process and perspectives in treatment of pain induced by Bothrops snake venoms (B. alternatus, B. asper, B. atrox, B. insularis, B. jararaca, B. pirajai, B. jararacussu, B. lanceolatus, B. leucurus, B. mattogrossensis, B. moojeni). We highlight, the understanding of the nociceptive signaling, especially in snakebite, enables more efficient treatment approaches. Finally, future perspectives for pain treatment concerning snakebite patients are discussed. Graduate Program in Tropical Diseases Botucatu Medical School (FMB) São Paulo State University Department of Physiology and Pharmacology and Drug Research and Development Center Federal University of Ceará Center of Mathematics Computing Sciences and Cognition Federal University of ABC Graduate Program in Tropical Diseases Botucatu Medical School (FMB) São Paulo State University

Graphic Abstract 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.

Snake venom is produced, transported and delivered by the sophisticated venom delivery system (VDS). When snakes bite, the venom travels from the venom gland through the venom duct into needle-like fangs that inject it into their prey. To counteract breakages, fangs are continuously replaced throughout life. Currently, the anatomy of the connection between the duct and the fang has not been described, and the mechanism by which the duct is reconnected to the replacement fang has not been identified. We examined the VDS in 3D in representative species from two families and one subfamily (Elapidae, Viperidae, Atractaspidinae) using contrast-enhanced microCT (diceCT), followed by dissection and histology. We observed that the venom duct bifurcates immediately anterior to the fangs so that both the original and replacement fangs are separately connected and functional in delivering venom. When a fang is absent, the canal leading to the empty position is temporarily closed. We found that elapid snakes have a crescent-shaped venom reservoir where venom likely pools before it enters the fang. These findings form the final piece of the puzzle of VDS anatomy in front-fanged venomous snakes. Additionally, they provide further evidence for independent evolution of the VDS in these three snake taxa.

The venom color variation of Crotalus durissus terrificus (Cdt) is attributed to the presence of the toxin L-amino acid oxidase (LAAO). During the venom milking routine of Instituto Butantan, we have noticed that most venoms of captive Cdt specimens show a yellowish color, while most venoms of wild specimens are white. Here we describe a comparative analysis of long-term captive (LTC) and recently wild-caught (RWC) Cdt, focusing on LAAO variation. For the identification of LAAO in individual venoms, four different approaches were employed: evaluation of the enzymatic activity, SDS-PAGE, Western blotting, and ELISA. In addition, mass spectrometry analysis was performed using pooled samples. Although some variation among these methodologies was observed, it was possible to notice that the presence of LAAO was significantly higher in the venom of LTC individuals. LAAO was identified in 60–80% LTC specimens and in only 10–12% of RWC specimens. Furthermore, this enzyme accounts for 5.6% of total venom proteins of LTC Cdt pooled venom, while it corresponds to only 0.7% of RWC Cdt pooled venom. These findings strongly suggest that captive maintenance increases the expression of LAAO in Cdt venom.

Viruses are associated with several human diseases that infect a large number of individuals, hence directly affecting global health and economy. Owing to the lack of efficient vaccines, antiviral therapy and emerging resistance strains, many viruses are considered as a potential threat to public health. Therefore, researches have been developed to identify new drug candidates for future treatments. Among them, antiviral research based on natural molecules is a promising approach. Phospholipases A2 (PLA2s) isolated from snake venom have shown significant antiviral activity against some viruses such as Dengue virus, Human Immunodeficiency virus, Hepatitis C virus and Yellow fever virus, and have emerged as an attractive alternative strategy for the development of novel antiviral therapy. Thus, this review provides an overview of remarkable findings involving PLA2s from snake venom that possess antiviral activity, and discusses the mechanisms of action mediated by PLA2s against different stages of virus replication cycle. Additionally, molecular docking simulations were performed by interacting between phospholipids from Dengue virus envelope and PLA2s from Bothrops asper snake venom. Studies on snake venom PLA2s highlight the potential use of these proteins for the development of broad-spectrum antiviral drugs.

Geographical variation of animal venom is common among venomous animals. This kind of intraspecific variation based on geographical location mainly concerned from envenomation cases and brought new problems in animal venom studies, including venom components regulatory mechanisms, differentiation of venom activities, and clinical treatment methods. At present, food is considered as the most related factor influencing venom development. Related research defined the variational venomous animal species by the comparison of venom components and activities in snakes, jellyfish, scorpions, cone snails, ants, parasitoid wasps, spiders and toads. In snake venom studies, researchers found that antivenom effectiveness was variated to different located venom samples. As described in some snake venom research, developing region-specific antivenom is the development trend. The difficulties of developing region-specific antivenom and theoretical solutions have been discussed. This review summarized biological studies of animal venom geographical variation by species, compared venom components and major biological activities of the vary venom from the same species, and listed the basic methods in comparing venom protein compositions and major toxicity differences to provide a comprehensive reference.

The aim of this study was to explore the activity of Naja naja oxiana venom on Leishmania tropica and its modes of action. Different fractions of Naja naja oxiana venom (NNOV) were prepared and characterized by high-performance liquid chromatography. The superior component, fraction k (FK) was selected. The activity of the fraction was assessed using advanced assays. Interleukin (IL)-12, TNF-α and iNOS gene expression as the indicators of Th1 significantly increased. In contrast, the level of IL-10, as the marker of T helper 2 substantially decreased (p

Snakebite is a neglected medical emergency causing fatalities and long-term disabilities throughout the world, especially in tropical countries. The effectiveness of therapy against snakebite is reliant on the unambiguous identification of bitten species of snake followed by immediate administration of venom-specific monovalent antivenom. However, this is a challenging task and therefore, over the several years scientists are constantly trying to address this issue by developing species-specific snake venom diagnostic kits as an alternative to classical methods of snake identification in clinics. Recently quite a few modern tools and techniques have been deployed for the development of simple, inexpensive, rapid, specific, and sensitive snake venom detection kits. However, despite these efforts a lone snakebite diagnostic kit is available until now which is a severe concern for efficacious snakebite therapy. In this article, we have reviewed the key issues pertaining to the rapid diagnosis of snake envenomation, tools and techniques developed and/or invented particularly over the past 40 years for the detection of snakebite as well as quantity of venom in the body fluids and/or tissues of victims. To overcome the practical constraints against the successful commercialization of these diagnostic kits, much more intensive studies for their improvement in terms of efficacy, affordability, storage stability, and usability, in addition to standardization of techniques for use in clinics are required to fulfil the objectives of the user-friendliness and commercial viability of snakebite diagnostic kits particularly in the rural and underdeveloped areas of tropical countries showing the maximum incidence of snakebite.

C-type lectins (CTL) and CTL-like proteins (snaclecs) are important toxins found in snake venom which can disrupt hemostasis by binding platelet membrane glycoproteins. Traditional identification of these toxins usually relies on an “activity-directed fractionation” approach which is very arduous. Here, we report a new method for rapid screening of these proteins in snake venom. Methods A conserved and immunogenic peptide found in svCTLs (CTL and snaclecs) was identified by sequence alignment using DNAStar software. The peptide was de novo synthesized and conjugated to keyhole limpet hemocyanin (KLH). Rabbit antibodies were generated against the peptide by classical immunization. Deinagkistrodon acutus venom was separated by two-dimensional electrophoresis (2DE) followed by Western blot and CTLs immunodetected using the isolated polyclonal antibody. The same svCTL spots on a parallel 2DE gel were isolated and analyzed by MALDI-TOF-MS. Results A highly conserved peptide with the sequence “KTWDDAEKFCTEQ” was identified as a common epitope in svCTLs. The polyclonal antibody against the 13aa-peptide was successfully prepared and purified. Its usefulness to detect svCTLs in D. acutus venom was tested by 2DE-WB and we determined that it positively identified all known D. acutus venom CTLs. Conclusions Immunodetection with antibodies against KTWDDAEKFCTEQ is an efficient strategy to identify novel svCTLs in the context of a complex proteome.

Cervical cancer is the fourth most common cancer worldwide in women. Apoptosis reactivation has become the main strategy for decreasing cancer proliferation. There is a need to extend the search for new drugs to implement more effective and less toxic strategies for cervical cancer treatment. Research has been carried out to find new drugs that have minimal side effects and that focus on the tumor microenvironment, particularly in the induction of cellular apoptosis and cell migration and the inhibition of angiogenesis. Potent toxins from snake venoms have shown potential as sources for the synthesis of new drugs with such characteristics. The present work aimed to describe cervical cancer characteristics, associated risk factors, current treatments and to highlight the effects of toxins isolated from the venom of snakes of the Viperidae family on cervical cancer cell lines.

Melanoma is a deadly type of skin cancer that is particularly difficult to treat owing to its resistance to radiation therapy. Here, we attempted to determine the key proteins responsible for melanoma radioresistance, with the aim of improving disease response to radiation therapy. Two melanoma cell lines, SK-Mel5 and SK-Mel28, with different radiosensitivities were analysed via RNA-Seq (Quant-Seq) and target proteins with higher abundance in the more radioresistant cell line, SK-Mel28, identified. Among these proteins, integrin αvβ3, a well-known molecule in cell adhesion, was selected for analysis. Treatment of SK-Mel28 cells with cilengitide, an integrin αvβ3 inhibitor, as well as γ-irradiation resulted in more significant cell death than γ-irradiation alone. In addition, Akt, a downstream signal transducer of integrin αvβ3, showed high basic activation in SK-Mel28 and was significantly decreased upon co-treatment with cilengitide and γ-irradiation. MK-2206, an Akt inhibitor, exerted similar effects on the SK-Mel28 cell line following γ-irradiation. Our results collectively demonstrate that the integrin αvβ3-Akt signalling pathway contributes to radioresistance in SK-Mel28 cells, which may be manipulated to improve therapeutic options for melanoma.

Objective Snake envenomation has been poorly studied in developing countries. ‘Early morning neuroparalytic syndrome’ (EMNS), the classical clinical constellation caused by krait bites, refers to nighttime, indoor bites where nonspecific symptoms progress to neuroparalysis. Literature regarding EMNS in children is scarce. This study was planned to describe the clinical profile, intensive care needs and predictors of outcome in children with EMNS. Methods It is a retrospective study of children below 12 years admitted with a clinical diagnosis of snake envenomation to the pediatric intensive care unit (PICU) of a tertiary care hospital in North India. Patient records were reviewed from the electronic patient database manager. Comparison was made between the EMNS group and the non-EMNS group and between survivors and nonsurvivors within the EMNS group. Results Of the 111 children with snake envenomation, 76 had neuroparalysis (68%) and 51 had EMNS. In the EMNS cohort, 37 (72.5%) belonged to rural areas, 46 (90.2%) had indoor bites and 39 (76.5%) were witnessed. Patients with EMNS were more likely to have absent fang marks, hypoxemia at admission, bulbar palsy and need for PICU admission. Mortality rate was 13.7% in EMNS; predictors included younger age, presence of ptosis, cardiac arrest at admission and nonavailability of PICU bed (univariable analysis) but none of them independently predicted mortality. Conclusion Younger age, presence of ptosis, cardiac arrest at admission and nonavailability of intensive care beds increase the risk of mortality in children with EMNS. Timely recognition and respiratory support may reduce mortality in these children.

Snake venoms are primarily composed of proteins and peptides, and these toxins have developed high selectivity to their biological targets. This makes venoms interesting for exploration into protein evolution and structure–function relationships. A single venom protein superfamily can exhibit a variety of pharmacological effects; these variations in activity originate from differences in functional sites, domains, posttranslational modifications, and the formations of toxin complexes. In this review, we discuss examples of how the major venom protein superfamilies have diversified, as well as how newer technologies in the omics fields, such as genomics, transcriptomics, and proteomics, can be used to characterize both known and unknown toxins.Because toxins are bioactive molecules with a rich diversity of activities, they can be useful as therapeutic and diagnostic agents, and successful examples of toxin applications in these areas are also reviewed. With the current rapid pace of technology, snake venom research and its applications will only continue to expand.

BACKGROUND AND PURPOSE: Animal toxins have contributed significantly to our understanding of the neurobiology of receptors and ion channels. We studied the venom of the coral snake Micrurus fulvius fulvius and identified and characterized the structure and pharmacology of a new homodimeric neurotoxin, fulditoxin, that exhibited novel pharmacology at nicotinic ACh receptors (nAChRs). EXPERIMENTAL APPROACH: Fulditoxin was isolated by chromatography, chemically synthesized, its structure determined by X‐ray crystallography, and its pharmacological actions on nAChRs characterized by organ bath assays and two‐electrode voltage clamp electrophysiology. KEY RESULTS: Fulditoxin's distinct 1.95‐Å quaternary structure revealed two short‐chain three‐finger α‐neurotoxins (α‐3FNTxs) non‐covalently bound by hydrophobic interactions and an ability to bind metal and form tetrameric complexes, not reported previously for three‐finger proteins. Although fulditoxin lacked all conserved amino acids canonically important for inhibiting nAChRs, it produced postsynaptic neuromuscular blockade of chick muscle at nanomolar concentrations, comparable to the prototypical α‐bungarotoxin. This neuromuscular blockade was completely reversible, which is unusual for snake α‐3FNTxs. Fulditoxin, therefore, interacts with nAChRs by utilizing a different pharmacophore. Unlike short‐chain α‐3FNTxs that bind only to muscle nAChRs, fulditoxin utilizes dimerization to expand its pharmacological targets to include human neuronal α4β2, α7, and α3β2 nAChRs which it blocked with IC(50) values of 1.8, 7, and 12 μM respectively. CONCLUSIONS AND IMPLICATIONS: Based on its distinct quaternary structure and unusual pharmacology, we named this new class of dimeric Micrurus neurotoxins represented by fulditoxin as Σ‐neurotoxins, which offers greater insight into understanding the interactions between nAChRs and peptide antagonists.

The Middle East and Northern Africa, collectively known as the MENA region, are inhabited by a plethora of venomous animals that cause up to 420,000 bites and stings each year. To understand the resultant health burden and the key variables affecting it, this review describes the epidemiology of snake, scorpion, and spider envenomings primarily based on heterogenous hospital data in the MENA region and the pathologies associated with their venoms. In addition, we discuss the venom composition and the key medically relevant toxins of these venomous animals, and, finally, the antivenoms that are currently in use to counteract them. Unlike Asia and sub-Saharan Africa, scorpion stings are significantly more common (approximately 350,000 cases/year) than snakebites (approximately 70,000 cases/year) and present the most significant contributor to the overall health burden of envenomings, with spider bites being negligible. However, this review also indicates that there is a substantial lack of high-quality envenoming data available for the MENA region, rendering many of these estimates speculative. Our understanding of the venoms and the toxins they contain is also incomplete, but already presents clear trends. For instance, the majority of snake venoms contain snake venom metalloproteinases, while sodium channel–binding toxins and potassium channel–binding toxins are the scorpion toxins that cause most health-related challenges. There also currently exist a plethora of antivenoms, yet only few are clinically validated, and their high cost and limited availability present a substantial health challenge. Yet, some of the insights presented in this review might help direct future research and policy efforts toward the appropriate prioritization of efforts and aid the development of future therapeutic solutions, such as next-generation antivenoms.

As said by former United Nations Secretary-General Kofi Annan, “Snakebite is the most important tropical disease you’ve never heard of.” Listed as a priority neglected tropical disease by the World Health Organization, snakebite envenoming (SBE) kills in excess of 125,000 people per year. However, due to the complexity and overlap of snake venom compositions, few reliable venom diagnostic methods for genus-/species-specific identification, which is crucial for successful SBE therapy, are available. Here, we develop a strategy to select and prepare genus-specific snake venom antibodies, which allows rapid and efficient clinical diagnosis of snakebite. Multi-omics approaches are used to choose candidate antigens from snake venoms and identify genus-specific antigenic epitope peptide fragments (GSAEPs) with ideal immunogenicity, specificity, and spatial accessibility. Double-antibody sandwich ELISA kit was established by matching a polyclonal antibody against a natural antigen and a monoclonal antibody that was prepared by natural protein as antigen and can specifically target the GSAEPs. The kit shows the ability to accurately identify venoms from similar genera of Trimeresurus and Protobothrops with a detection limit of 6.25 ng/ml on the snake venoms and a little cross-reaction, thus proving high feasibility and applicability.

Hemorrhage induced by snake venom metalloproteinases (SVMPs) is a complex phenomenon that involves capillary disruption and blood extravasation. HF3 (hemorrhagic factor 3) is an extremely hemorrhagic SVMP of Bothrops jararaca venom. Studies using proteomic approaches revealed targets of HF3 among intracellular and extracellular proteins. However, the role of the cleavage of plasma proteins in the context of the hemorrhage remains not fully understood. The main goal of this study was to analyze the degradome of HF3 in human plasma. For this purpose, approaches for the depletion of the most abundant proteins, and for the enrichment of low abundant proteins of human plasma, were used to minimize the dynamic range of protein concentration, in order to assess the proteolytic activity of HF3 on a wide spectrum of proteins, and to detect the degradation products using mass spectrometry-based untargeted peptidomics. The results revealed the hydrolysis products generated by HF3 and allowed the identification of cleavage sites. A total of 61 plasma proteins were identified as cleaved by HF3. Some of these proteins corroborate previous studies, and others are new HF3 targets, including proteins of the coagulation cascade, of the complement system, proteins acting on the modulation of inflammation, and plasma proteinase inhibitors. Overall, the data indicate that HF3 escapes inhibition and sculpts the plasma proteome by degrading key proteins and generating peptides that may act synergistically in the hemorrhagic process.

African spitting cobras are unique among cobras for their potent anticoagulant venom activity arising from strong inhibition of Factor Xa. This anticoagulant effect is exerted by venom phospholipase A2 (Group I PLA2) toxins whose activity contributes to the lethality of these species. This anticoagulant toxicity is particularly problematic as it is not neutralized by current antivenoms. Previous work demonstrated this trait for Naja mossambica, N. nigricincta, N. nigricollis, and N. pallida. The present work builds upon previous research by testing across the full taxonomical range of African spitting cobras, demonstrating that N. ashei, N. katiensis, and N. nubiae are also potently anticoagulant through the inhibition of Factor Xa, and therefore the amplification of potent anticoagulant activity occurred at the base of the African spitting cobra radiation. Previous work demonstrated that the enzyme-inhibitor varespladib was able to neutralize this toxic action for N. mossambica, N. nigricincta, N. nigricollis, and N. pallida venoms. The current work demonstrates that varespladib was also able to neutralize N. ashei, N. katiensis, and N. nubiae. Thus varespladib is shown to have broad utility across the full range of African spitting cobras. In addition, we examined the cross-reactivity of the metalloprotease inhibitor prinomastat, which had been previously intriguingly indicated as being capable of neutralizing viperid venom PLA2 (Group II PLA2). In this study prinomastat inhibited the FXa-inhibiting PLA2 toxins of all the African spitting cobras at the same concentration at which it has been shown to inhibit metalloproteases, and thus was comparably effective in its cross-reactivity. In addition we showed that the metalloprotease-inhibitor marimastat was also able to cross-neutralize PLA2 but less effectively than prinomastat. Due to logistical (cold-chain requirement) and efficacy (cross-reactivity across snake species) limitations of traditional antivenoms, particularly in developing countries where snakebite is most common, these small molecule inhibitors (SMIs) might hold great promise as initial, field-based, treatments for snakebite envenoming as well as addressing fundamental limitations of antivenom in the clinical setting where certain toxin effects are unneutralized.

Snakebite envenomation (SBE) is an important public health issue that is now receiving renewed attention following its reclassification as a Neglected Tropical Disease (NTD). Most incidences occur in rural areas of resource-limited countries, as such, timely and appropriate medical care for SBE is often inaccessible. The administration of anti-snake venom serum (ASV) is the only effective definitive treatment of SBE, but treatment failure to available ASVs is not uncommon. Emerging evidence highlights the potential of small-molecule compounds as inhibitors against toxins of snake venom. This presents an encouraging prospect to develop an alternative therapeutic option for the treatment SBE, that may be amenable for use at the point of care in resource-constraint settings. In view of the pivotal role of natural products in modern drug discovery programmes, there is considerable interest in ethno-pharmacological mining of medicinal plants and plant-derived medicinal compounds toward developing novel snake venom-neutralising therapeutics. In this review, we compile a collection of medicinal plants used in the treatment of SBE in West Africa and highlight their promise as potential botanical drugs or as sources of novel small-molecule compounds for the treatment of SBE. The challenges that must be surmounted to bring this to fruition including the need for (sub) regional collaboration have been discussed.

Snake envenomation is a severe problem in Sri Lanka (SL) and Indian polyvalent antivenom (PAV) is mostly used for treating snakebite albeit due to geographical variation in venom composition, Indian PAV shows poor efficacy in neutralizing the lethality and toxicity of venom from the same species of snakes in SL. Therefore, the quality and in vivo venom neutralization potency of a country-specific PAV produced against the venom of the five most medically important snakes of SL (Daboia russelii, Echis carinatus, Hypnale hypnale, Naja naja, Bungarus caeruleus) was assessed. LC-MS/MS analysis of two batches of PAV showed the presence of 88.7–97.2% IgG and traces of other plasma proteins. The tested PAVs contained minor amounts of undigested IgG and F(ab′)2 aggregates, showed complement activation, were devoid of IgE, endotoxin, and content of preservative was below the threshold level. Immunological cross-reactivity and in vitro neutralization of enzymatic activities, pharmacological properties demonstrated superior efficacy of SL PAV compared to Indian PAV against SL snake venoms. The in vivo neutralization study showed that the tested PAVs are potent to neutralize the lethality and venom-induced toxicity of SL snake venoms. Therefore, our study suggests that introduction of SL-specific PAV will improve snakebite management in SL.

Made available in DSpace on 2022-04-28T19:41:27Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-08-31 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Snakebite envenoming is the cause of an ongoing health crisis in several regions of the world, particularly in tropical and neotropical countries. This scenario creates an urgent necessity for new practical solutions to address the limitations of current therapies. The current study investigated the isolation, phytochemical characterization, and myotoxicity inhibition mechanism of gallic acid (GA), a myotoxin inhibitor obtained from Anacardium humile. The identification and isolation of GA was achieved by employing analytical chromatographic separation, which exhibited a compound with retention time and nuclear magnetic resonance spectra compatible with GA's commercial standard and data from the literature. GA alone was able to inhibit the myotoxic activity induced by the crude venom of Bothrops jararacussu and its two main myotoxins, BthTX-I and BthTX-II. Circular dichroism (CD), fluorescence spectroscopy (FS), dynamic light scattering (DLS), and interaction studies by molecular docking suggested that GA forms a complex with BthTX-I and II. Surface plasmon resonance (SPR) kinetics assays showed that GA has a high affinity for BthTX-I with a KD of 9.146 × 10−7 M. Taken together, the two-state reaction mode of GA binding to BthTX-I, and CD, FS and DLS assays, suggest that GA is able to induce oligomerization and secondary structure changes for BthTX-I and -II. GA and other tannins have been shown to be effective inhibitors of snake venoms' toxic effects, and herein we demonstrated GA's ability to bind to and inhibit a snake venom PLA2, thus proposing a new mechanism of PLA2 inhibition, and presenting more evidence of GA's potential as an antivenom compound. Instituto de Genética e Bioquímica Universidade Federal de Uberlândia UFU Departamento de Biofísica e Farmacologia Instituto de Biociências Universidade Estadual Paulista UNESP Laboratório de Biotecnologia de Proteínas e Compostos Bioativos LABIOPROT Centro de Estudos de Biomoléculas Aplicadas à Saúde CEBio Laboratório de Imunologia Celular Aplicada à Saúde Fundação Oswaldo Cruz FIOCRUZ Unidade Rondônia Instituto Brasileiro do Meio Ambiente e dos Recursos Naturais Renováveis IBAMA Departamento de Química Universidade Federal de Lavras UFLA Instituto Federal Goiano IF Goiano Faculty of Chemical Sciences University of Cuenca LAQV/Requimte Faculty of Sciences University of Porto Centro Universitário São Lucas UniSL Instituto Nacional de Ciência e Tecnologia em Epidemiologia da Amazônia Ocidental (INCT-EpiAmO) Departamento de Biofísica e Farmacologia Instituto de Biociências Universidade Estadual Paulista UNESP

The phospholipases A2 (PLA2) superfamily encompasses enzymes commonly found in mammalian tissues and snake venom. Many of these enzymes have unique tissue distribution, function, and substrate specificity suggesting distinct biological roles. In the past, much of the research on secretory PLA2s has analyzed their roles in inflammation, anti-bacterial actions, and atherosclerosis. In recent studies utilizing a variety of mouse models, pancreatic islets, and clinical trials, a role for many of these enzymes in the control of metabolism and insulin action has been revealed. In this review, this research, and the unique contributions of the PLA2 enzymes in insulin resistance and metabolism.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent responsible for the Coronavirus Disease-2019 (COVID-19) pandemic, has infected over 185 million individuals across 200 countries since December 2019 resulting in 4.0 million deaths. While COVID-19 is primarily associated with respiratory illnesses, an increasing number of clinical reports indicate that severely ill patients often develop thrombotic complications that are associated with increased mortality. As a consequence, treatment strategies that target COVID-associated thrombosis are of utmost clinical importance. An array of pharmacologically active compounds from natural products exhibit effects on blood coagulation pathways, and have generated interest for their potential therapeutic applications towards thrombotic diseases. In particular, a number of snake venom compounds exhibit high specificity on different blood coagulation factors and represent excellent tools that could be utilized to treat thrombosis. The aim of this review is to provide a brief summary of the current understanding of COVID-19 associated thrombosis, and highlight several snake venom compounds that could be utilized as antithrombotic agents to target this disease.

Snake venoms have evolved primarily to immobilize and kill prey, and consequently, they contain some of the most potent natural toxins. Part of that armory is a range of hemotoxic components that affect every area of hemostasis, which we have harnessed to great effect in the study and diagnosis of hemostatic disorders. The most widely used are those that affect coagulation, such as thrombin-like enzymes unaffected by heparin and direct thrombin inhibitors, which can help confirm or dispute their presence in plasma. The liquid gold of coagulation activators is Russell's viper venom, since it contains activators of factor X and factor V. It is used in a range of clotting-based assays, such as assessment of factor X and factor V deficiencies, protein C and protein S deficiencies, activated protein C resistance, and probably the most important test for lupus anticoagulants, the dilute Russell's viper venom time. Activators of prothrombin, such as oscutarin C from Coastal Taipan venom and ecarin from saw-scaled viper venom, are employed in prothrombin activity assays and lupus anticoagulant detection, and ecarin has a valuable role in quantitative assays of direct thrombin inhibitors. Snake venoms affecting primary hemostasis include botrocetin from the jararaca, which can be used to assay von Willebrand factor activity, and convulxin from the cascavel, which can be used to detect deficiency of the platelet collagen receptor, glycoprotein VI. This article takes the reader to every area of the diagnostic hemostasis laboratory to appreciate the myriad applications of snake venoms available in diagnostic practice.

Supplemental Digital Content is available in the text., Objective: The Australian snake venom ptFV (Pseudonaja textilis venom-derived factor V) variant retains cofactor function despite APC (activated protein C)-dependent proteolysis. Here, we aimed to unravel the mechanistic principles by determining the role of the absent Arg306 cleavage site that is required for the inactivation of FVa (mammalian factor Va). Approach and Results: Our findings show that in contrast to human FVa, APC-catalyzed proteolysis of ptFVa at Arg306 and Lys507 does not abrogate ptFVa cofactor function. Remarkably, the structural integrity of APC-proteolyzed ptFVa is maintained indicating that stable noncovalent interactions prevent A2-domain dissociation. Using Molecular Dynamics simulations, we uncovered key regions located in the A1 and A2 domain that may be at the basis of this remarkable characteristic. Conclusions: Taken together, we report a completely novel role for uniquely adapted regions in ptFVa that prevent A2 domain dissociation. As such, these results challenge our current understanding by which strict regulatory mechanisms control FVa activity.

Background Snakebite is a neglected tropical disease that causes high global rates of mortality and morbidity. Although snakebite can cause a variety of pathologies in victims, haemotoxic effects are particularly common and are typically characterised by haemorrhage and/or venom-induced consumption coagulopathy. Antivenoms are the mainstay therapeutic for treating the toxic effects of snakebite, but despite saving thousands of lives annually, these therapies are associated with limited cross-snake species efficacy due to venom variation, which ultimately restricts their therapeutic utility to particular geographical regions. Methodology/Principal findings In this study we explored the feasibility of generating globally effective pathology-specific antivenoms to counteract the haemotoxic signs of snakebite envenoming. Two different immunogen mixtures, consisting of seven and twelve haemotoxic venoms sourced from geographically diverse and/or medically important snakes, were used to raise ovine polyclonal antibodies, prior to characterisation of their immunological binding characteristics and in vitro neutralisation profiles against each of the venoms. Despite variability of the immunogen mixtures, both experimental antivenoms exhibited broadly comparable in vitro venom binding and neutralisation profiles against the individual venom immunogens in immunological and functional assays. However, in vivo assessments using a murine preclinical model of antivenom efficacy revealed substantial differences in venom neutralisation. The experimental antivenom generated from the seven venom immunogen mixture outperformed the comparator, by providing protective effects against venom lethality caused by seven of the eight geographically diverse venoms tested, including three distinct venoms that were not used as immunogens to generate this antivenom. These findings suggest that a core set of venom immunogens may be sufficient to stimulate antibodies capable of broadly neutralising a geographically diverse array of haemotoxic snake venoms, and that adding additional venom immunogens may impact negatively on the dose efficacy of the resulting antivenom. Conclusions/Significance Although selection of appropriate immunogens that encapsulate venom toxin diversity without diluting antivenom potency remains challenging and further optimisation is required, the findings from this pilot study suggest that the generation of pathology-specific antivenoms with global utility is likely to feasible, thereby highlighting their promise as future modular treatments for the world’s tropical snakebite victims., Author summary Snakebite is a major public health concern that causes extensive death and disability in the tropical world. Variation in venom composition causes major challenges for snakebite therapy by typically restricting antivenom efficacy to only the few snake species that each product was designed against, meaning that many geographically restricted therapeutics are manufactured for different parts of the world. In this study, we explored the feasibility of generating a global pathology-specific antivenom by using two different sets of haemotoxic venoms as immunogens, one including seven venoms and the other twelve venoms, to generate ovine polyclonal antibodies. Our in vitro immunological cross-reactivity and venom functional-neutralisation findings demonstrated broadly comparable venom recognition and inhibition against the various venom immunogens, irrespective of whether they were included in the immunising mixture. However, assessments of murine in vivo protection against venom-induced lethality revealed that the antivenom generated with fewer immunogens outperformed the comparator, and provided broader preclinical efficacy, even against some venoms not used to raise the antibodies. These findings suggest that careful selection of venoms used as immunogens is crucial for maintaining antivenom potency, and that a small subset of venoms may be capable of generating an antivenom with potential global efficacy against a specific pathology commonly observed after snakebite.