RESUMO
BACKGROUND: Each year, 3,800 cases of snakebite envenomation are reported in Mexico, resulting in 35 fatalities. The only scientifically validated treatment for snakebites in Mexico is the use of antivenoms. Currently, two antivenoms are available in the market, with one in the developmental phase. These antivenoms, produced in horses, consist of F(ab')2 fragments generated using venoms from various species as immunogens. While previous studies primarily focused on neutralizing the venom of the Crotalus species, our study aims to assess the neutralization capacity of different antivenom batches against pit vipers from various genera in Mexico. METHODOLOGY: We conducted various biological and biochemical tests to characterize the venoms. Additionally, we performed neutralization tests using all three antivenoms to evaluate their effectiveness against lethal activity and their ability to neutralize proteolytic and fibrinogenolytic activities. RESULTS: Our results reveal significant differences in protein content and neutralizing capacity among different antivenoms and even between different batches of the same product. Notably, the venom of Crotalus atrox is poorly neutralized by all evaluated batches despite being the primary cause of envenomation in the country's northern region. Furthermore, even at the highest tested concentrations, no antivenom could neutralize the lethality of Metlapilcoatlus nummifer and Porthidium yucatanicum venoms. These findings highlight crucial areas for improving existing antivenoms and developing new products. CONCLUSION: Our research reveals variations in protein content and neutralizing potency among antivenoms, emphasizing the need for consistency in venom characteristics as immunogens. While Birmex neutralizes more LD50 per vial, Antivipmyn excels in specific neutralization. The inability of antivenoms to neutralize certain venoms, especially M. nummifer and P. yucatanicum, highlights crucial improvement opportunities, given the medical significance of these species.
Assuntos
Antivenenos , Testes de Neutralização , Antivenenos/farmacologia , Antivenenos/imunologia , Animais , México , Mordeduras de Serpentes/tratamento farmacológico , Mordeduras de Serpentes/imunologia , Viperidae , Crotalus , Venenos de Crotalídeos/imunologiaRESUMO
We used ultrasonography and radiography to assess the sexual organs and characterize the reproductive cycle of captive golden lancehead (Bothrops insularis) and Alcatrazes lancehead (B. alcatraz), two endangered island snake species in Brazil. We assessed 46- individuals of golden lancehead and 12 of Alcatrazes lancehead kept in captivity between 2014 and 2020. Follicular development was similar between species, but follicles in Alcatrazes lancehead were smaller than in the golden lanceheads. Female golden lanceheads produced 24 live young, seven stillborn and 73 undeveloped eggs. Parturition of live young occurred between midsummer (February) and early autumn and gestation averaged 8 months. Female Alcatrazes lanceheads produced four live young in midsummer, and one undeveloped egg in early autumn. Males and females of both species have seasonal and biennial reproductive cycles. Sperm storage in both sexes is essential to coordinate male and female cycles. The data obtained with golden lancehead and Alcatrazes lancehead in captivity, demonstrate a degree of conservatism, following data from other Bothrops.
Assuntos
Bothrops , Serpentes Peçonhentas , Viperidae , Humanos , Animais , Masculino , Feminino , Sêmen , Radiografia , Ultrassonografia/veterinária , Espécies em Perigo de ExtinçãoRESUMO
Serpentirhabdias mexicanus n. sp. (Nematoda: Rhabdiasidae) is described from the lung of the nauyaca viper Bothrops asper in Puebla State, central Mexico. This new species is the fifth of the genus described having onchia. Among the species included in this group, the new species is morphologically closest to S. viperidicus and S. atroxi. However, it differs from both species mainly by having only one excretory gland (compared to two present in S. viperidicus and S. atroxi). In addition, S. mexicanus n. sp. can be separated of S. viperidicus by tail length, shape of vulval lips, geographic distribution and host species and from S. atroxi by body length, number of papillae in the cephalic region, as well as the host species and geographic distribution. In the present study, we propose the new species based on morphological, host spectrum and genetic evidence. Phylogenetic analysis indicated Serpentirhabdias as a monophyletic group, with two subgroups that are congruent with the presence/absence of onchia in the esophagostome, host association and other relevant morphological characters.
Assuntos
Nematoides , Viperidae , Animais , Bothrops asper , México , Filogenia , Especificidade da EspécieRESUMO
Viperids of the genus Lachesis, also known as bushmasters, are capable of injecting great amounts of venom that cause severe envenomation incidents. Since phospholipases type A2 are mainly involved in edema and myonecrosis within the snakebite sites, in this work, the isolation, amino acid sequence and biochemical characterization of the first phospholipase type A2 from the venom of Lachesis acrochorda, named Lacro_PLA2, is described. Lacro_PLA2 is an acidic aspartic 49 calcium-dependent phospholipase A2 with 93% similarity to the L. stenophrys phospholipase. Lacro_PLA2 has a molecular mass of 13,969.7 Da and an experimental isoelectric point around 5.3. A combination of N-terminal Edman degradation and MS/MS spectrometry analyses revealed that Lacro_PLA2 contains 122 residues including 14 cysteines that form 7 disulfide bridges. A predicted 3D model shows a high resemblance to other viperid phospholipases. Nevertheless, immunochemical and phospholipase neutralization tests revealed a notorious level of immunorecognition of the isolated protein by two polyclonal antibodies from viperids from different genus, which suggest that Lacro_PLA2 resembles more to bothropic phospholipases. Lacro_PLA2 also showed significantly high edema activity when was injected into mice; so, it could be an alternative antigen in the development of antibodies against toxins of this group of viperids, seeking to improve commercial polyclonal antivenoms.
Assuntos
Crotalinae , Viperidae , Animais , Camundongos , Viperidae/metabolismo , Espectrometria de Massas em Tandem , Fosfolipases A2/química , Venenos de Víboras/toxicidade , Edema/induzido quimicamenteRESUMO
Most anti-inflammatory drugs currently adopted to treat chronic inflammatory joint diseases can alleviate symptoms but they do not lead to remission. Therefore, new and more efficient drugs are needed to block the course of joint inflammatory diseases. Animal venoms, rich in bioactive compounds, can contribute as valuable tools in this field of research. In this study, we first demonstrate the direct action of venoms on cells that constitute the articular joints. We established a platform consisting of cell-based assays to evaluate the release of cytokines (IL-6, IL-8, TNFα, IL-1ß, and IL-10) by human chondrocytes, synoviocytes and THP1 macrophages, as well as the release of neuropeptides (substance-P and ß-endorphin) by differentiated sensory neuron-like cells, 24 h after stimulation of cells with 21 animal venoms from snake and arthropod species, sourced from different taxonomic families and geographic origins. Results demonstrated that at non-cytotoxic concentrations, the venoms activate at varying degrees the secretion of inflammatory mediators involved in the pathology of articular diseases, such as IL-6, IL-8, and TNF-α by chondrocytes, synoviocytes, and macrophages and of substance P by neuron-like cells. Venoms of the Viperidae snake family were more inflammatory than those of the Elapidae family, while venoms of Arthropods were less inflammatory than snake venoms. Notably, some venoms also induced the release of the anti-inflammatory IL-10 by macrophages. However, the scorpion Buthus occitanus venom induced the release of IL-10 without increasing the release of inflammatory cytokines by macrophages. Since the cell types used in the experiments are crucial elements in joint inflammatory processes, the results of this work may guide future research on the activation of receptors and inflammatory signaling pathways by selected venoms in these particular cells, aiming at discovering new targets for therapeutic intervention.
Assuntos
Animais Peçonhentos , Venenos de Artrópodes , Artrópodes , Artropatias , Venenos de Escorpião , Escorpiões , Viperidae , Animais , Humanos , Interleucina-10 , Interleucina-6 , Interleucina-8 , Venenos de Serpentes/química , Citocinas , Fator de Necrose Tumoral alfa , Anti-InflamatóriosRESUMO
This study presents the first complete mitogenome of the Brazilian Atlantic bushmaster Lachesis with insights into snake evolution. The total length was 17,177 bp, consisting of 13 PCGs, 22 tRNAs, two rRNAs and a duplicate control region (CRs). Almost all genes were encoded by the heavy-strand, except for the ND6 gene and eight tRNAs (tRNA-Gln, Ala, Asn, Cys, Tyr, Ser[TGA anticodon], Glu, Pro). Only ATG, ATA, and ATC were starting codons for protein-coding sequences. Stop codons mainly were TAA, AGA, AGG, and TAG; whereas ND1, ND3, and CYTB terminated with incomplete stop codons. Phylogeny retrieved Lachesis within the Crotalinae as the sister group of Agkistrodon; and the Lachesis+Agkistrodon clade as the sister group of (Sistrurus+Crotalus)+Bothrops. The tree supports Crotalinae, Viperinae, and Azemiopinae in the Viperidae family, being sister taxa of Colubridae+(Elapidae+Psammophiidae). The mean genetic distance across 15 snake families and 57 nucleotide sequences was 0.37. The overall mean value of genetic distance across the Crotalinae was 0.23, with Lachesis muta exhibiting the shortest distance of 0.2 with Agkistrodon piscivorus, Protobothrops dabieshanensis and P. flavoviridis and the greatest 0.25 with Gloydius blomhoffii, Trimeresurus albolabris, S. miliarius, and Deinagkistrodon acutus. The complete Atlantic L. muta mitogenome presented herein is only the third annotated mitogenome from more than 430 described Brazilian snake species.
Assuntos
Crotalinae , Genoma Mitocondrial , Viperidae , Humanos , Animais , Crotalinae/genética , Viperidae/genética , Genoma Mitocondrial/genética , Códon de Terminação , BrasilRESUMO
OBJECTIVE.: To evaluate the in vitro inhibitory activity of Plantago major "llantén" and Piper aduncum "matico" extracts on phospholipase A2 (PLA2) from the venom of the snake Lachesis muta muta. MATERIALS AND METHODS.: We carried out an explanatory study with experimental design. Leaves of P. major and P. aduncum were collected in the province of Huarochirí in Lima, Peru. Then, we prepared alcoholic extracts diluted in distilled water and conducted phytochemical assays, quantification of phenols and flavonoids, thin layer chromatography (TLC) on cellulose and enzymatic activity with PLA2. The ability to inhibit PLA2 with the extracts under study and their fractions was analyzed. The Kruskal Wallis test and Bonferroni multiple comparisons were used during statistical analysis. RESULTS.: Phenols, flavonoids and tannins were qualitatively identified in both P. major and P. aduncum; in addition, P. aduncum presented saponins. The inhibition of PLA2 activity of the venom by the total extract of P. major was 45.3%, and its fractions showed the following inhibition values: 31.1% for LLF-1, 66.3% for LLF-2 and 65.5% for LLF-3. The inhibition values for the total extract of P. aduncum were 86.9%, and its fractions showed the following inhibition rates: 34.3% for MF-1, 67.1% for MF-2 and 54.9% for MF-3. Statistical analysis showed significant differences in the inhibition of PLA2 (p=0.009) by the extracts. CONCLUSION.: The tests demonstrated an association between the anti-inflammatory effect of the extracts and PLA2 inhibition.
OBJETIVO.: Evaluar la actividad inhibitoria in vitro de los extractos de Plantago major «llantén¼ y Piper aduncum «matico¼ sobre Fosfolipasa A2 (PLA2) del veneno de la serpiente Lachesis muta muta. MATERIALES Y MÉTODOS.: Esta investigación fue de tipo explicativa con diseño experimental. Se recolectaron hojas de P. major y P. aduncum en la provincia de Huarochirí en Lima, Perú. Se prepararon extractos alcohólicos diluidos en agua destilada y se realizaron los ensayos fitoquímicos, la cuantificación de fenoles y flavonoides, la cromatografía de capa fina (CCF) en celulosa y la actividad enzimática con PLA2. Se analizó la capacidad de inhibir la PLA2 con los extractos en estudio y sus fracciones. Para el análisis estadístico se utilizó la prueba de Kruskal Wallis y comparaciones múltiples de Bonferroni. RESULTADOS.: Tanto en P. major como en P. aduncum se identificó cualitativamente la presencia de fenoles, flavonoides y taninos; además, P. aduncum presentó saponinas. La inhibición de la actividad de la PLA2 del veneno por el extracto total de P. major fue del 45,3%, y sus fracciones mostraron valores de inhibición: LLF-1 con 31,1%, LLF-2 con 66,3% y LLF-3 con 65,5%. En P. aduncum, los valores de inhibición para el extracto total fueron de 86,9%, y sus fracciones presentaron inhibiciones: MF-1 con 34,3%, MF-2 con 67,1% y MF-3 con 54,9%. El análisis estadístico demostró diferencias significativas en la inhibición de la PLA2 (p=0,009) por los extractos. CONCLUSIÓN.: Los ensayos realizados demostraron una asociación entre el efecto antiinflamatorio de los extractos y la inhibición de la PLA2.
Assuntos
Venenos de Crotalídeos , Plantago , Viperidae , Animais , Fosfolipases A2 , Flavonoides , FenóisRESUMO
The skull of vipers is a highly kinetic anatomical structure involved in envenomating and consuming of prey. Morphological knowledge about the viperid skull is based on studies on some groups of species, but information on its variation within the whole family and its functional morphology is still scarce. In this study, we aimed to explore variation in skull morphology among species of the three subfamilies of Viperidae, and test whether that variation correlates with macrohabitat and diet. We performed quantitative analyses of the viperid skull based on broad taxonomic sampling and two methodological approaches: linear and geometric morphometrics. The results of both approaches showed that much of the variation lies in differences of shape and relative size of the premaxilla, the nasals, the frontals, and the parietals. The results indicated that phylogeny and size influence the shape of the skull, but we also found evidence of morphological differentiation between arboreal and terrestrial species and in species with mammal specialist diet. Our findings imply that, besides evolutionary allometry and phylogenetic signal, demands of particular diets coupled with use of certain habitats have in part shaped morphological evolution of the viperid skull.
Assuntos
Viperidae , Animais , Filogenia , Viperidae/anatomia & histologia , Crânio/anatomia & histologia , Cabeça , Ecossistema , Evolução Biológica , MamíferosRESUMO
In Colombia, there are two species of bushmaster snakes, Lachesis acrochorda, which is distributed mainly in the west of the country (in the Choco region), and Lachesis muta in the southeast (in the Amazon and Orinoquia region), whose presence has been reduced due to the destruction of their habitats. Captive maintenance is challenging, making it difficult to obtain their venom for study and antivenom manufacturing. They are the largest vipers in the world. The occurrence of human envenomation is quite rare, but when it occurs, it is associated with high mortality. Bushmaster venom is necrotizing, hemorrhagic, myotoxic, hemolytic, and cardiovascular depressant. Due to the presence of bradycardia, hypotension, emesis, and diarrhea in some patients (Lachesis syndrome), the possibility of a vagal or cholinergic effect is raised. The treatment of envenomation is hindered by the scarcity of antivenom and the need to use high doses. A review of the most relevant biological and medical aspects of bushmaster snakes is presented, mainly for those occurring in Colombia, to facilitate their recognition and raise awareness about the need for special attention to improve their conservation and advance scientific knowledge, in particular, about their venom.
Assuntos
Venenos de Crotalídeos , Viperidae , Animais , Humanos , Antivenenos/uso terapêutico , ColômbiaRESUMO
Bitis arietans is a medically important snake found in Sub-Saharan Africa. The envenomation is characterized by local and systemic effects, and the lack of antivenoms aggravates the treatment. This study aimed to identify venom toxins and develop antitoxins. The F2 fraction obtained from Bitis arietans venom (BaV) demonstrated the presence of several proteins in its composition, including metalloproteases. Titration assays carried out together with the immunization of mice demonstrated the development of anti-F2 fraction antibodies by the animals. The determination of the affinity of antibodies against different Bitis venoms was evaluated, revealing that only BaV had peptides recognized by anti-F2 fraction antibodies. In vivo analyses demonstrated the hemorrhagic capacity of the venom and the effectiveness of the antibodies in inhibiting up to 80% of the hemorrhage and 0% of the lethality caused by BaV. Together, the data indicate: (1) the prevalence of proteins that influence hemostasis and envenomation; (2) the effectiveness of antibodies in inhibiting specific activities of BaV; and (3) isolation and characterization of toxins can become crucial steps in the development of new alternative treatments. Thus, the results obtained help in understanding the envenoming mechanism and may be useful for the study of new complementary therapies.
Assuntos
Mordeduras de Serpentes , Viperidae , Camundongos , Animais , Viperidae/metabolismo , Venenos de Serpentes/metabolismo , Antivenenos , Metaloproteases/metabolismo , Hemorragia , Imunoglobulina G/metabolismoRESUMO
Lachesis acrochorda envenomation has a lethality rate of approximately 90%. Despite its high lethality, little is known about its local and systemic effects and its relationship with its protein content. Thus, to increase knowledge of L. acrochorda snake venom from the Southwestern ecoregion of Colombia, we developed a proteomic analysis using a "bottom-up shotgun proteomic profiling" approach. Besides, we evaluated toxinological properties and compared the effects with the Bothrops asper snake venom activities. The RP-HPLC profile showed similarities with the L. acrochorda snake venom from the Northwestern ecoregion of Colombia. However, the results displayed differences in the protein families identified, probably due to the proteomic identification strategy. The in vitro and in vivo tests showed a L. acrochorda snake venom with Phospholipase A2 and metalloproteinase activities related to myotoxic, edematic, and hemorrhagic effects. Nevertheless, the L. acrochorda snake venom displayed a low lethality despite a large amount of inoculated venom. This investigation's results will help us improve the knowledge about the relationship between the clinical manifestations of L. acrochorda envenomation and the venom protein content.
Assuntos
Proteômica , Viperidae , Animais , Humanos , Colômbia , Venenos de Serpentes , HemorragiaRESUMO
Crotalines (pitvipers) in the Americas are distributed from southern Canada to southern Argentina, and are represented by 13 genera and 163 species that constitute a monophyletic group. Their phylogenetic relationships have been assessed mostly based on DNA sequences, while morphological data have scarcely been used for phylogenetic inquiry. We present a total-evidence phylogeny of New World pitvipers, the most taxon/character comprehensive phylogeny to date. Our analysis includes all genera, morphological data from external morphology, cranial osteology and hemipenial morphology, and DNA sequences from mitochondrial and nuclear genes. We performed analyses with parsimony as an optimality criterion, using different schemes for character weighting. We evaluated the contribution of the different sources of characters to the phylogeny through analyses of reduced datasets and calculation of weighted homoplasy and retention indexes. We performed a morphological character analysis to identify synapomorphies for the main clades. In terms of biogeography, our results support a single colonization event of the Americas by pitvipers, and a cladogenetic event into a Neotropical clade and a North American/Neotropical clade. The results also shed light on the previously unstable position of some taxa, although they could not sufficiently resolve the position of Bothrops lojanus, which may lead to the paraphyly of either Bothrops or Bothrocophias. The morphological character analyses demonstrated that an important phylogenetic signal is contained in characters related to head scalation, the jaws and the dorsum of the skull, and allowed us to detect morphological convergences in external morphology associated with arboreality.
Assuntos
Bothrops , Crotalinae , Viperidae , Animais , Filogenia , Viperidae/genética , Crotalinae/genética , Evolução Biológica , Sequência de Bases , Bothrops/genéticaRESUMO
In the Brazilian Amazon, envenomations by lancehead pit vipers prevail across the region, while bushmaster (Lachesis muta) envenomations are rarely confirmed. Here, we described a moderate snakebite, diagnosed as a lancehead pit viper envenomation upon admission and treated with four vials of Bothrops antivenom. Blood remained unclottable for 4 days of hospitalization. On day 4, after admission, the patient presented pictures of the perpetrating snake to the hospital staff, which was identified as a Lachesis muta specimen. After administering 10 vials of Lachesis antivenom, blood became clottable 12 hours after treatment. The patient was discharged without complaints.
Assuntos
Bothrops , Venenos de Crotalídeos , Mordeduras de Serpentes , Viperidae , Animais , Antivenenos/uso terapêutico , Brasil , Mordeduras de Serpentes/complicações , Mordeduras de Serpentes/terapiaRESUMO
BACKGROUND: Snakebite envenomation exerts a heavy toll in sub-Saharan Africa. The design and production of effective polyspecific antivenoms for this region demand a better understanding of the immunological characteristics of the different venoms from the most medically important snakes, to select the most appropriate venom combinations for generating antivenoms of wide neutralizing scope. Bitis spp. and Echis spp. represent the most important viperid snake genera in Africa. METHODOLOGY/PRINCIPAL FINDINGS: Eight rabbit-derived monospecific antisera were raised against the venoms of four species of Bitis spp. and four species of Echis spp. The effects of immunization in the rabbits were assessed, as well as the development of antibody titers, as judged by immunochemical assays and neutralization of lethal, hemorrhagic, and in vitro coagulant effects. At the end of immunizations, local and pulmonary hemorrhage, together with slight increments in the plasma activity of creatine kinase (CK), were observed owing to the action of hemorrhagic and myotoxic venom components. Immunologic analyses revealed a considerable extent of cross-reactivity of monospecific antisera against heterologous venoms within each genus, although some antisera provided a more extensive cross-reactivity than others. The venoms that generated antisera with the broadest coverage were those of Bitis gabonica and B. rhinoceros within Bitis spp. and Echis leucogaster within Echis spp. CONCLUSIONS/SIGNIFICANCE: The methodology followed in this study provides a rational basis for the selection of the best combination of venoms for generating antivenoms of high cross-reactivity against viperid venoms in sub-Saharan Africa. Results suggest that the venoms of B. gabonica, B. rhinoceros, and E. leucogaster generate antisera with the broadest cross-reactivity within their genera. These experimental results in rabbits need to be translated to large animals used in antivenom production to assess whether these predictions are reproduced in horses or sheep.
Assuntos
Viperidae , África Subsaariana , Animais , Antivenenos , Hemorragia , Cavalos , Soros Imunes , Coelhos , Ovinos , Venenos de Serpentes , SerpentesRESUMO
BACKGROUND: Snakebites are a major problem in developing countries owing to their high morbidity rates, severity, and sequelae. In Brazil, most cases of envenomation are caused by Bothrops and Lachesis snakes. The present study aimed to evaluate variables associated with death, systemic complications, and amputations in victims of envenomation due to Bothrops or Lachesis snake. METHODS: An analytical epidemiological study was performed with data from the Notifiable Diseases Information System (Sistema de Informação de Agravos de Notificação [SINAN]) relating to the Brazilian Amazon for the period 2010-2015. A hierarchical Poisson regression analysis was performed with three aspects, namely, individual characteristics, sociodemographic characteristics, and clinical conditions. RESULTS: The following associations were observed: i) advanced age (>65 years), sepsis, renal failure, and hemorrhagic manifestations were related to two outcomes - death due to snakebite and death from all causes; ii) more advanced age (≥46 years), time to health care longer than 6 hours, renal and hemorrhagic manifestations, and region of occurrence being rural areas were associated with systemic complications; and iii) children (up to 12 years old) were associated with amputations. CONCLUSIONS: Knowledge of the characteristics associated with severe outcomes in snakebites may help identify patients who will require more intensive care or longer follow-up and may provide the physician with counseling rationale for their possible prognosis.
Assuntos
Bothrops , Mordeduras de Serpentes , Viperidae , Idoso , Animais , Antivenenos , Brasil/epidemiologia , Criança , Hemorragia , Humanos , Pessoa de Meia-Idade , Mordeduras de Serpentes/complicações , Mordeduras de Serpentes/epidemiologiaRESUMO
Viperid snakebite envenoming is often characterized by a venom-induced consumption coagulopathy due to the procoagulant effect of venom components, resulting in the alteration of clotting laboratory tests. There is a growing trend to use rotational thromboelastometry in the assessment of clotting disturbances in a variety of pathologies, although its use in experimental models of envenoming has been limited. An in vivo murine model was implemented to assess the coagulopathy induced by three Central American viperid venoms which have different mechanisms of action on clotting factors, i.e., Bothrops asper, Crotalus simus and Bothriechis lateralis. Venom was injected by the intravenous route and blood samples were collected at 1, 3, 5 and 24 h after envenoming. Coagulopathy was assessed by standard clotting tests and by routine rotational thromboelastometric parameters. In addition, the changes in platelet number were followed. B. asper and C. simus venoms induced coagulopathy and thrombocytopenia 1 h after injection, followed by a slow recovery at 3, 5 and 24 h, although the majority of clotting parameters were still significantly affected by 3 and 5 h, and were corrected by 24 h. In general, a similar time-course of alterations was observed for standard clotting tests and most rotational thromboelastomeric assays. However, some thromboelastometric parameters, especially those related to Fibtem, showed more drastic alterations than standard tests and remained altered even at 24 h in some cases. This is likely related to the low fibrinogen concentration observed at most time intervals. B. lateralis venom did not induce a consumption coagulopathy, although it caused a marked thrombocytopenia.
Assuntos
Transtornos da Coagulação Sanguínea , Venenos de Crotalídeos , Coagulação Intravascular Disseminada , Mordeduras de Serpentes , Viperidae , Animais , Antivenenos/farmacologia , Transtornos da Coagulação Sanguínea/etiologia , Testes de Coagulação Sanguínea , Venenos de Crotalídeos/toxicidade , Camundongos , Mordeduras de Serpentes/complicações , Venenos de Serpentes/toxicidade , TromboelastografiaRESUMO
Bothorps atrox is responsible for most of the ophidism cases in Perú. As part of the envenoming, myotoxicity is one of the most recurrent and destructive effects. In this study, a myotoxin, named BaMtx, was purified from B. atrox venom to elucidate its biological, immunological, and molecular characteristics. BaMtx was purified using CM-Sephadex-C-25 ion-exchange resin and SDS-PAGE analysis showed a unique protein band of 13 kDa or 24 kDa under reducing or non-reducing conditions, respectively. cDNA sequence codified a 122-aa mature protein with high homology with other Lys49-PLA2s; modeled structure showed a N-terminal helix, a ß-wing region, and a C-terminal random coil. This protein has a poor phospholipase A2 enzymatic activity. BaMtx has myotoxic (DMM = 12.30 ± 0.95 µg) and edema-forming (DEM = 26.00 ± 1.15 µg) activities. Rabbit immunization with purified enzyme produced anti-BaMtx antibodies that reduced 50.28 ± 10.15% of myotoxic activity and showed significant cross-reactivity against B. brazili and B pictus venoms. On the other hand, BaMtx exhibits mild anti-proliferative and anti-migratory effects on breast cancer cells, affecting the ROS and NADH levels, which may reduce mitochondrial respiration. These results contribute to the understanding of B. atrox Lys49-PLA2 effects and establish the anticancer potential de BaMtx.
Assuntos
Bothrops , Venenos de Crotalídeos , Viperidae , Sequência de Aminoácidos , Animais , Bothrops/metabolismo , Miotoxicidade , Peru , Fosfolipases A2/química , Coelhos , Viperidae/metabolismoRESUMO
SUMMARY: Myodural bridges (MDB) are anatomical connections between the suboccipital muscles and the cervical dura mater which pass through both the atlanto-occipital and the atlanto-axial interspaces in mammals. In our previous studies, we found that the MDB exists in seven terrestrial mammal species, two marine mammal species, two reptilian species, and one bird species. A recent study suggested that given the "ubiquity" of myodural bridges in terrestrial vertebrates, the MDB may also exist in snakes. Specifically, we focused on the Gloydius shedaoensis, a species of Agkistrodon (pit viper snake) that is only found on Shedao Island, which is in the southeastern sea of Dalian City in China. Six head and neck cadaveric specimens of Gloydius shedaoensis were examined. Three specimens were used for anatomical dissection and the remaining three cadaveric specimens were utilized for histological analysis. The present study confirmed the existence of the MDB in the Gloydius shedaoensis. The snake's spinalis muscles originated from the posterior edge of the supraoccipital bones and the dorsal facet of the exocciput, and then extended on both sides of the spinous processes of the spine, merging with the semispinalis muscles. On the ventral aspect of this muscular complex, it gave off fibers of the MDB. These MDB fibers twisted around the posterior margin of the exocciput and then passed through the atlanto-occipital interspace, finally terminating on the dura mater. We observed that the MDB also existed in all of the snakes' intervertebral joints. These same histological findings were also observed in the Gloydius brevicaudus, which was used as a control specimen for the Gloydius shedaoensis. In snakes the spinal canal is longer than that observed in most other animals. Considering the unique locomotive style of snakes, our findings contribute to support the hypothesis that the MDB could modulate cerebrospinal fluid (CSF) pulsations.
RESUMEN: Los puentes miodurales (MDB) son conexiones anatómicas entre los músculos suboccipitales y la duramadre cervical que pasan a través de los espacios intermedios atlanto-occipital y atlanto-axial en los mamíferos. En nuestros estudios anteriores, encontramos que el MDB existe en siete especies de mamíferos terrestres, dos especies de mamíferos marinos, dos especies de reptiles y una especie de ave. Un estudio reciente sugirió que dada la "ubicuidad" de los puentes miodurales en los vertebrados terrestres, el MDB también puede existir en las serpientes. Específicamente, nos enfocamos en Gloydius shedaoensis, una especie de Agkistrodon (serpiente víbora) que solo se encuentra en la isla Shedao, en el mar sureste de la ciudad de Dalian en China. Se examinaron seis especímenes cadavéricos de cabeza y cuello de Gloydius shedaoensis. Se utilizaron tres especímenes para la disección anatómica y los tres especímenes cadavéricos restantes se utilizaron para el análisis histológico. El presente estudio confirmó la existencia del MDB en Gloydius shedaoensis. Los músculos espinosos de la serpiente se originaron en el margen posterior de los huesos supraoccipital y la cara dorsal del exoccipucio, y luego se extendieron a ambos lados de los procesos espinosas de la columna vertebral, fusionándose con los músculos semiespinosos. En la cara ventral de este complejo muscular se desprendían fibras del MDB. Estas fibras MDB se ubican alrededor del margen posterior del exoccipucio y luego atraviesan el interespacio atlanto-occipital, terminando finalmente en la duramadre. Observamos que el MDB también existía en todas las articulaciones intervertebrales de las serpientes. Estos mismos hallazgos histológicos también se observaron en Gloydius brevicaudus, que se utilizó como muestra de control para Gloydius shedaoensis. En las serpientes, el canal espinal es más largo que el observado en la mayoría de los otros animales. Teniendo en cuenta el estilo único locomotor de las serpientes, nuestros hallazgos contribuyen a respaldar la hipótesis de que el MDB podría modular las pulsaciones del líquido cerebroespinal.
Assuntos
Animais , Líquido Cefalorraquidiano/fisiologia , Viperidae/anatomia & histologia , Tecido Conjuntivo , Dura-Máter/anatomia & histologia , Crotalinae , Anatomia ComparadaRESUMO
Oral tolerance is defined as a specific suppression of cellular and humoral immune responses to a particular antigen through prior oral administration of an antigen. It has unique immunological importance since it is a natural and continuous event driven by external antigens. It is characterized by low levels of IgG in the serum of animals after immunization with the antigen. There is no report of induction of oral tolerance to Bothrops jararaca venom. Here, we induced oral tolerance to B. jararaca venom in BALB/c mice and evaluated the specific tolerance and cross-reactivity with the toxins of other Bothrops species after immunization with the snake venoms adsorbed to/encapsulated in nanostructured SBA-15 silica. Animals that received a high dose of B. jararaca venom (1.8 mg) orally responded by showing antibody titers similar to those of immunized animals. On the other hand, mice tolerized orally with three doses of 1 µg of B. jararaca venom showed low antibody titers. In animals that received a low dose of B. jararaca venom and were immunized with B. atrox or B. jararacussu venom, tolerance was null or only partial. Immunoblot analysis against the venom of different Bothrops species provided details about the main tolerogenic epitopes and clearly showed a difference compared to antiserum of immunized animals.
Assuntos
Reações Cruzadas/imunologia , Venenos de Crotalídeos/imunologia , Tolerância Imunológica , Administração Oral , Animais , Anticorpos/sangue , Bothrops , Venenos de Crotalídeos/administração & dosagem , Feminino , Camundongos Endogâmicos BALB C , Nanoestruturas , Dióxido de Silício/química , Especificidade da Espécie , Venenos de Víboras/imunologia , ViperidaeRESUMO
Bitis arietans is a snake of medical importance found throughout sub-Saharan Africa and in savannas and pastures of Morocco and western Arabia. The effects of its venom are characterized by local and systemic alterations, such as inflammation and cardiovascular and hemostatic disturbances, which can lead to victims' death or permanent disability. To better characterize the inflammatory process induced by this snake's venom, the participation of eicosanoids and PAF (platelet- activating factor) in this response were demonstrated in a previous study. In addition, edema and early increased vascular permeability followed by an accumulation of polymorphonuclear (PMN) cells in the peritoneal cavity were accompanied by the production of the eicosanoids LTB4, LTC4, TXB2, and PGE2, and local and systemic production of IL-6 and MCP-1. In this context, the present study focused on the identification of inflammatory mediators produced by human macrophages derived from THP-1 cells in response to Bitis arietans venom (BaV), and Kn-Ba, a serine protease purified from this venom. Here, we show that Kn-Ba, and even the less intensive BaV, induced the production of the cytokine TNF and the chemokines RANTES and IL-8. Only Kn-Ba was able to induce the production of IL-6, MCP-1, and IP-10, whereas PGE2 was produced only in response to BaV. Finally, the release of IL-1ß in culture supernatants suggests the activation of the inflammasomes by the venom of Bitis arietans and by Kn-Ba, which will be investigated in more detail in future studies.