BthTX-I, a phospholipase A2-like toxin, is inhibited by the plant cinnamic acid derivative: chlorogenic acid.

Snakebite is a significant health concern in tropical and subtropical regions, particularly in Africa, Asia, and Latin America, resulting in more than 2.7 million envenomations and an estimated one hundred thousand fatalities annually. The Bothrops genus is responsible for the majority of snakebite envenomings in Latin America and Caribbean countries. Accidents involving snakes from this genus are characterized by local symptoms that often lead to permanent sequelae and death. However, specific antivenoms exhibit limited effectiveness in inhibiting local tissue damage. Phospholipase A2-like (PLA2-like) toxins emerge as significant contributors to local myotoxicity in accidents involving Bothrops species. As a result, they represent a crucial target for prospective treatments. Some natural and synthetic compounds have shown the ability to reduce or abolish the myotoxic effects of PLA2-like proteins. In this study, we employed a combination approach involving myographic, morphological, biophysical and bioinformatic techniques to investigate the interaction between chlorogenic acid (CGA) and BthTX-I, a PLA2-like toxin. CGA provided a protection of 71.8% on muscle damage in a pre-incubation treatment. Microscale thermophoresis and circular dichroism experiments revealed that CGA interacted with the BthTX-I while preserving its secondary structure. CGA exhibited an affinity to the toxin that ranks among the highest observed for a natural compound. Bioinformatics simulations indicated that CGA inhibitor binds to the toxin's hydrophobic channel in a manner similar to other phenolic compounds previously investigated. These findings suggest that CGA interferes with the allosteric transition of the non-activated toxin, and the stability of the dimeric assembly of its activated state.

Neutralization of a bothropic PLA2-like protein by caftaric acid, a novel potent inhibitor of ophidian myotoxicity.

Envenoming by snakebite is an important global health issue that has received little attention, leading the World Health Organization to naming it as neglected tropical disease. Several snakebites present serious local symptoms manifested on victims that may not be efficiently neutralized by serum therapy. Phospholipase A2-like (PLA2-like) toxins are present in Viperidae venoms and are responsible for local myotoxic activity. Herein, we investigated the association between BthTX-I toxin and caftaric acid (CFT), a molecule present in plants. CFT neutralized neuromuscular blocking and muscle-damaging activities promoted by BthTX-I. Calorimetric and light-scattering assays demonstrated that CFT inhibitor interacted with dimeric BthTX-I. Bioinformatics simulations indicated that CFT inhibitor binds to the toxin's hydrophobic channel (HCh). According to the current myotoxic mechanism, three different regions of PLA2-like toxins have specific tasks: protein allosteric activation (HCh), membrane dockage (MDoS), and membrane rupture (MDiS). We propose CFT inhibitor interferes with the allosteric activation, which is related to the conformation change leading to the exposure/alignment of MDoS/MDiS region. This is the first report of a PLA2-like toxin fully inhibited by a compound that interacts only with its HCh region. Thus, CFT is a novel candidate to complement serum therapy and improve the treatment of snakebite.

Effects of AT1 receptor antagonism on interstitial and ultrastructural remodeling of heart in response to a hypercaloric diet.

Palatable hypercaloric feeding has been associated with angiotensin-II type 1 receptor (AT1R) stimulation and cardiac remodeling. This study analyzed whether AT1R antagonism attenuates cardiac remodeling in rats subjected to a palatable hypercaloric diet. Male Wistar-Kyoto rats were subjected to a commercial standard rat chow (CD) or a palatable hypercaloric diet (HD) for 35 weeks and then allocated into four groups: CD, CL, HD, and HL; L groups received losartan in drinking water (30 mg/kg/day) for 5 weeks. Body weight, adiposity, and glycemia were evaluated. The cardiovascular study included echocardiography, and myocardial morphometric and ultrastructural evaluation. Myocardial collagen isoforms Type I and III were analyzed by Western blot. Both HD and HL had higher adiposity than their respective controls. Cardiomyocyte cross-sectional-area (CD 285 ± 49; HD 344 ± 91; CL 327 ± 49; HL 303 ± 49 µm2 ) and interstitial collagen fractional area were significantly higher in HD than CD and unchanged by losartan. HD showed marked ultrastructural alterations such as myofilament loss, and severe mitochondrial swelling. CL presented higher Type I collagen expression when compared to CD and HL groups. The ultrastructural changes and type I collagen expression were attenuated by losartan in HL. Losartan attenuates systolic dysfunction and ultrastructural abnormalities without changing myocardial interstitial remodeling in rats subjected to a palatable hypercaloric diet.

Pharmacological characterization of cnidarian extracts from the Caribbean Sea: evaluation of anti-snake venom and antitumor properties

Background: Cnidarians produce toxins, which are composed of different polypeptides that induce pharmacological effects of biotechnological interest, such as antitumor, antiophidic and anti-clotting activities. This study aimed to evaluate toxicological activities and potential as antitumor and antiophidic agents contained in total extracts from five cnidarians: Millepora alcicornis, Stichodactyla helianthus, Plexaura homomalla, Bartholomea annulata and Condylactis gigantea (total and body wall). Methods: The cnidarian extracts were evaluated by electrophoresis and for their phospholipase, proteolytic, hemorrhagic, coagulant, fibrinogenolytic, neuromuscular blocking, muscle-damaging, edema-inducing and cytotoxic activities. Results: All cnidarian extracts showed indirect hemolytic activity, but only S. helianthus induced direct hemolysis and neurotoxic effect. However, the hydrolysis of NBD-PC, a PLA2 substrate, was presented only by the C gigantea (body wall) and S. helianthus. The extracts from P. homomalla and S. helianthus induced edema, while only C gigantea and S. helianthus showed intensified myotoxic activity. The proteolytic activity upon casein and fibrinogen was presented mainly by B. annulata extract and all were unable to induce hemorrhage or fibrinogen coagulation. Cnidarian extracts were able to neutralize clotting induced by Bothrops jararacussu snake venom, except M. alcicornis. All cnidarian extracts were able to inhibit hemorrhagic activity induced by Bothrops moojeni venom. Only the C. gigantea (body wall) inhibited thrombin-induced coagulation. All cnidarian extracts showed antitumor effect against Jurkat cells, of which C. gigantea (body wall) and S. helianthus were the most active; however, only C. gigantea (body wall) and M. alcicornis were active against B16F10 cells...

Structural Basis for the Inhibition of a Phospholipase A2-Like Toxin by Caffeic and Aristolochic Acids.

One of the main challenges in toxicology today is to develop therapeutic alternatives for the treatment of snake venom injuries that are not efficiently neutralized by conventional serum therapy. Venom phospholipases A2 (PLA2s) and PLA2-like proteins play a fundamental role in skeletal muscle necrosis, which can result in permanent sequelae and disability. This leads to economic and social problems, especially in developing countries. In this work, we performed structural and functional studies with Piratoxin-I, a Lys49-PLA2 from Bothropspirajai venom, complexed with two compounds present in several plants used in folk medicine against snakebites. These ligands partially neutralized the myotoxic activity of PrTX-I towards binding on the two independent sites of interaction between Lys49-PLA2 and muscle membrane. Our results corroborate the previously proposed mechanism of action of PLA2s-like and provide insights for the design of structure-based inhibitors that could prevent the permanent injuries caused by these proteins in snakebite victims.

Extensive impact of saturated fatty acids on metabolic and cardiovascular profile in rats with diet-induced obesity: a canonical analysis.

BACKGROUND: Although hypercaloric interventions are associated with nutritional, endocrine, metabolic, and cardiovascular disorders in obesity experiments, a rational distinction between the effects of excess adiposity and the individual roles of dietary macronutrients in relation to these disturbances has not previously been studied. This investigation analyzed the correlation between ingested macronutrients (including sucrose and saturated and unsaturated fatty acids) plus body adiposity and metabolic, hormonal, and cardiovascular effects in rats with diet-induced obesity. METHODS: Normotensive Wistar-Kyoto rats were submitted to Control (CD; 3.2 Kcal/g) and Hypercaloric (HD; 4.6 Kcal/g) diets for 20 weeks followed by nutritional evaluation involving body weight and adiposity measurement. Metabolic and hormonal parameters included glycemia, insulin, insulin resistance, and leptin. Cardiovascular analysis included systolic blood pressure profile, echocardiography, morphometric study of myocardial morphology, and myosin heavy chain (MHC) protein expression. Canonical correlation analysis was used to evaluate the relationships between dietary macronutrients plus adiposity and metabolic, hormonal, and cardiovascular parameters. RESULTS: Although final group body weights did not differ, HD presented higher adiposity than CD. Diet induced hyperglycemia while insulin and leptin levels remained unchanged. In a cardiovascular context, systolic blood pressure increased with time only in HD. Additionally, in vivo echocardiography revealed cardiac hypertrophy and improved systolic performance in HD compared to CD; and while cardiomyocyte size was unchanged by diet, nuclear volume and collagen interstitial fraction both increased in HD. Also HD exhibited higher relative ß-MHC content and ß/α-MHC ratio than their Control counterparts. Importantly, body adiposity was weakly associated with cardiovascular effects, as saturated fatty acid intake was directly associated with most cardiac remodeling measurements while unsaturated lipid consumption was inversely correlated with these effects. CONCLUSION: Hypercaloric diet was associated with glycemic metabolism and systolic blood pressure disorders and cardiac remodeling. These effects directly and inversely correlated with saturated and unsaturated lipid consumption, respectively.

Expression of genes related to quality of Longissimus dorsi muscle meat in Nellore (Bos indicus) and Canchim (5/8 Bos taurus × 3/8 Bos indicus) cattle.

This study was performed to compare CAPN1, CAPN2, CAST, TG, DGAT1 and LEP gene expressions and correlate them with meat quality traits in two genetic groups (Nellore and Canchim) in order to assess their expression profile and use their expression profile as genetic markers. We analyzed 30 young bulls (1year old), 15 of each genetic group. Samples of the Longissimus dorsi muscle were collected for analysis of: total lipids (TL) and meat tenderness measured as Warner-Bratzler shear force (SF) and myofibrillar fragmentation (MFI) at day of slaughter and 7days of aging. Gene expression profiles were obtained via RT-qPCR. TL and MFI showed differences between breeds, higher MFI in Canchim and higher TL in Nellore. Calpains showed no differential expression between groups, as did DGAT1, TG, and LEP. CAST was expressed more in the Nellore cattle. The only significant within-breed correlation (0.79) between gene expression and meat traits was found for DGAT1 and MFI in Canchim breed. Although the number of animals used in this study was small, the results indicate that the increased expression of CAST in Nellore may reflect tougher meat, but the lack of correlations with the meat traits indicates it is not a promising genetic marker.

Morfologia e histoenzimologia do músculo extensor longo do halux em ratos senis / Morphologic and histochemistry of extensor hallucis longus muscle of Wistar senile rat

O músculo extensor longo do halux em ratos senis apresentou alteraçöes nas fibras musculares e tecido conjuntivo. O grau de alteraçäo morfológica variou entre os animais, sendo mais acentuado nas idades mais avançadas. Algumas fibras apresentaram área reduzida, angulaçäo acentudada e contornos irregulares; aparência de necrose; divisäo longitudinal splitting; além do aumento do tecido conjuntivo entre as fibras musculares. Observou-se uma diminuiçäo na área das fibras FG, comparando-se com a área das fibras FG de animais jovens