Importance of jararhagin disintegrin-like and cysteine-rich domains in the early events of local inflammatory response

Jararhagin is a multi-domain SVMP from Bothrops jararaca venom comprising catalytic, disintegrin-like and cysteine-rich domains, which cause a local reaction manifested by hemorrhage, edema, cytokine release and inflammatory cell recruitment. In this study, the importance of disintegrin-like/cysteine-rich domains of jararhagin was addressed by analyzing the effects of jararhagin-C, which lacks the catalytic domain, in induction of leukocyte rolling and release of pro-inflammatory cytokines. Jararhagin-C was isolated from B. jararaca venom conserving the same ability of complete jararhagin molecule in inhibiting collagen-induced platelet-aggregation. Treatment of trans-illuminated cremaster muscle in vivo with jararhagin-C increased number of rolling leukocytes (approximately 250%) in post-capillary venules in all periods analyzed, without interfering with microvasculature haemodinamic, like vessel diameter, the erythrocyte speed or the blood flow rate. The release of pro-inflammatory cytokines TNF-α, IL-1β and IL-6 was significantly enhanced in the local of jararhagin-C injection, showing the maximum levels in periods between 2 and 4 h after treatment. Besides the action of jararhagin-C, the presence of the inactivated catalytic domain in o-phenanthrolin-treated jararhagin was related to a higher increase in the number of rolling leukocytes. Moreover, the levels of IL-6 and IL-1β induced by catalytically active jararhagin were higher than those induced by jararhagin-C. In conclusion, our findings suggest that the disintegrin-like/cysteine-rich domains of jararhagin are sufficient to locally activate the early events of an acute inflammatory response as leukocyte rolling and pro-inflammatory cytokine release and this action may add to the effect of catalysis, which enhances the primary cell activation.     

Inflammatory mediators generated at the site of inoculation of Loxosceles gaucho spider venom

Patients bitten by Loxosceles spiders generally manifest marked local inflammatory reaction and dermonecrosis. This report evaluated edema formation, leukocyte infiltration and release of inflammatory mediators at the injection site of Loxosceles gaucho venom. BALB/c mice were i.d. injected with venom and thereafter paws were disrupted and homogenized to obtain differential counts of migrated cells, as well to assay the levels of cytokines, chemokines and lipid mediators. Increased footpad thickness was detected as soon as 30 min after venom injection, and 24 h later was similar to that of the control group. Loxosceles venom mildly augmented the recruitment of leukocytes to the footpad in comparison with PBS-injected mice. Moreover, it stimulated the release of IL-6, MCP-1 and KC at 2 and 24 h after venom injection. In addition, higher levels of PGE₂ were detected 30 min after venom injection in comparison with control group. However, the venom failed to increase levels of IL-1β, TNF-α, TXB₂ and LTB₄. Our results demonstrate that L. gaucho venom evokes an early complex inflammatory reaction, stimulating the secretion of pro-inflammatory cytokines and lipid mediators (PGE₂), and recruiting leukocytes to the footpad which contribute to the local reaction induced by L. gaucho venom.     

Unusual profile of leukocyte recruitment in mice induced by a skin secretion of the tree frog Phyllomedusa hypochondrialis

Phyllomedusa hypochondrialis skin secretion can cause both systemic and local effects. In this study, we describe the pattern of local acute inflammatory response after P. hypochondrialis skin secretion injection. The inflammatory reaction in the mice footpad was analysed, including the leukocyte recruitment into local tissue from the peripheral blood, in a mouse model of tissue injury. We also investigated the release of the cytokines IL-1, IL-6 and TNF-α, chemokines KC and MCP-1 and the eicosanoids LTB 4 and PGE₂ in mice. The present findings support the ability of P. hypochondrialis skin secretion to induce local inflammation. In addition, these skin secretion components play a role in the initial rolling and slowing of recruited leukocytes and the transition from rolling to adhesion. Levels of the proinflammatory cytokine IL-6, chemokines KC and MCP-1 as well as the eicosanoid PGE₂ were significantly increased after injection of a skin secretion of P. hypochondrialis (0.6 μg/30 μl intraplantar), whereas no changes in other parameters were observed. Finally, the mechanisms involved in the local inflammatory process induced by P. hypochondrialis skin secretion is one of the questions of relevance related to the complex pathophysiology induced by this particular secretion and other toxins.     

Characterization of biological activity of Scatophagus argus venom

Scatophagus argus of the family Scatophagidae inflicts painful wounds in fishermen while handling it. The venom induces prominent local tissue damage characterized by pain, edema and necrosis. The pathogenesis of acute muscle damage in gastrocnemius muscle induced by S. argus venom was studied in mice. The inflammatory response induced by S. argus venom in the mice hind paw was studied measuring paw edema. Intramuscular injection of S. argus venom induced motoxicity. The effect of S. argus venom on the cellular components of inflammatory response was investigated. Venom from S. argus were quantitatively analyzed for enzymic and biochemical activity. The biochemical changes induced by the sublethal concentration of S. argus venom and histopathological studies of effect of venom on mice were carried out. Venom induced a rapid increment in serum creatine kinase (CK) and lactate dehydrogenase (LDH) showing the myotoxicity of venom. Concomitant with this a reduction of muscle CK and LDH activity was observed, where as no increment in muscle lactate was detected. Our findings showed that the edematic activity was dose dependent and remained significantly elevated over 48 h after injection. Administration of S. argus venom caused a significant cell accumulation of neutrophils in to peritoneal cavity as well as foot pad up to 24 h with maximal being at 4-6 h. The venom components analyzed showed the presence of phosphodiesterase, acid phosphatases, alkaline phosphatases, proteinase, and caseinolytic activity. SDS PAGE revealed the presence of major and minor protein bands between 6.5 and 68 kDa. The biochemical changes induced by the sublethal concentration of S. argus venom showed reversible changes in the hematological (blood cell count, hematocrit, hemoglobin, mean corpuscular volume, mean corpuscular hemoglobin and platelet count) parameters which were significantly altered at 6 and 24 h (GLM repeated measures p < 0.05). Serum enzymes such as AST, ALT, ACP, ALP, LDH and urea were altered significantly which in turn confirmed the damage of vital organ tissue.     

Isolation and characterization of ellagic acid derivatives isolated from Casearia sylvestris SW aqueous extract with anti-PLA2 activity

The Casearia sylvestris SW (Flacourtiaceae) is utilized in folk medicine (Brazil and all Latin American) to treat several pathologic processes as inflammation, cancer, microbial infection and snake bites. Studies showed that C. sylvestris aqueous extract can inhibit many toxic effects caused by snake venoms (or caused by phospholipase A₂ isolated) from different species, mainly of Bothrops genus. Inhibition of enzymatic and myotoxic activities, decrease of edema formation and increase of the survival rate of rats injected with lethal doses of bothropic venoms are some toxic effects inhibited by C. sylvestris. In this study, four ellagic acid derivatives from aqueous extracts of C. sylvestris were isolated, characterized, and tested against effects from both total venom and PLA₂ (Asp 49 BthTX-II) from the venom of Bothrops jararacussu. The isolated compounds were as follows: ellagic acid (A), 3′-O-methyl ellagic acid (B), 3,3′-di-O-methyl ellagic acid (C), 3-O-methyl-3′,4′-methylenedioxy ellagic acid (D). The inhibition constant values (Ki) for enzymatic activity, as well the IC₅₀ values found in the edematogenic and myotoxic activities, indicate that the ellagic acid is the best inhibitor of these activities, while compounds C and D are the substances with lowest capacity on inhibiting these same effects. Our results show that the presence of hydroxyls at position 3 or 3′ (compounds A and B) increases the capacity of these derivatives on inhibiting these toxic effects. However, the presence of methoxyl groups at position 3 or 3′ reduced, but did not completely inhibit the capacity of compounds C and D on inhibiting all the toxic effects studied.     

Antiangiogenic effects of phospholipase A2 Lys49 BnSP-7 from Bothrops pauloensis snake venom on endothelial cells: An in vitro and ex vivo approach

Antiangiogenic strategies are promising tools for cancer treatment and several other disorders. In this sense, phospholipases A₂ (PLA₂s) from snake venom have been described to possess antiangiogenic properties. In this study, we evaluated both in vitro and ex vivo antiangiogenic effects induced by BnSP-7, a Lys49 PLA₂ isolated from Bothrops pauloensis snake venom. BnSP-7 was able to inhibit endothelial cell (HUVEC) proliferation, which was indeed confirmed by a modulation of cell cycle progression. Interestingly, BnSP-7 also inhibited the adhesion and migration of HUVECs and blocked in vitro angiogenesis in a VEGF-dependent manner, an important proangiogenic factor. Finally, BnSP-7 was capable of inhibiting sprouting angiogenic process through an ex vivo aortic ring assay. Taken together, these results indicate that BnSP-7 has potent in vitro and ex vivo antiangiogenic effect.     

Biochemical and biological characterization of a PLA2 from crotoxin complex of Crotalus durissus cumanensis

A new PLA₂ (Cdcum6) from crotoxin complex of Colombian Crotalus durissus cumanensis rattlesnake was purified using molecular exclusion chromatography and RP-HPLC. The molecular mass of Cdcum6 was determined by SDS-PAGE ∼14 KDa and confirmed by MALDI-TOF (14321.98 Da). The enzyme showed K_m 6.0 mM, V_max 3.44 nmol/min, optimum pH was 8.0 and temperature was between 30 and 45 °C, and it had a strict requirement of Ca²⁺ for its activity. The N-terminal sequence of PLA₂ was SLVQF EKMIK EVAGK NGVPWY. Comparison of amino acid sequence data with other PLA₂ from South American Crotalus durissus rattlesnakes showed that Cdcum6 shares the highest sequence identity with Cdr13 an isoform PLA₂ from Crotalus durissus ruruima, nevertheless, Cdcum6 showed high content of basic and hydrophobic amino acids. In mice, Cdcum6 presented higher LD₅₀ than crotoxin complex from C. d. cumanensis. Additionally, Cdcum6 induced a conspicuous local myotoxic effect and moderate footpad edema; in vitro, it was antigoagulant in doses as low as 0.5 μg/ml, and it was not cytotoxic on myoblast but Cdcum6 was able to lyse myotubes.     

Agglutinin isolated from the red marine alga Hypnea cervicornis J. Agardh reduces inflammatory hypernociception: Involvement of nitric oxide

Hypnea cervicornis agglutinin (HCA), a lectin isolated from the red marine alga has been previously shown to have an antinociceptive effect. In the present study in rats, mechanisms of action of HCA were addressed regarding mechanical hypernociception induced by carrageenan, ovalbumin (as antigen), and also by prostaglandin E₂ in rats. The lectin administered intravenously inhibited carrageenan- and antigen-induced hypernociception at 1, 3, 5 and 7 h. This inhibitory effect was completely prevented when lectin was combined with mucin, demonstrating the role of carbohydrate-binding sites. The inhibition of inflammatory hypernociception by HCA was associated with the prevention of neutrophil recruitment to the plantar tissue of rats but was not associated with the inhibition of the release of pro-hypernociceptive cytokines (TNF-α, IL-1β and CINC-1). HCA also blocked mechanical hypernociception induced by PGE₂, which was prevented by the administration of nitric oxide synthase inhibitors. These results were corroborated by the increased circulating levels of NO metabolites following HCA treatment. These findings suggest that the anti-hypernociceptive effects of HCA are not associated with the inhibition of pro-inflammatory cytokine production. However, these effects seem to involve the inhibition of neutrophil migration and also the increase in NO production.     

Effects of acute in vitro exposure of murine precision-cut lung slices to gaseous nitrogen dioxide and ozone in an air–liquid interface (ALI) culture

The aim of this study was to establish an air–liquid interface (ALI) culture of precision-cut lung slices (PCLS) for direct exposure of lung cells to gaseous contaminants. Nitrogen dioxide (NO₂) and ozone (O₃) were selected as model gas compounds. Acute pro-inflammatory and toxic effects of NO₂ and O₃ on live lung tissue were investigated. Murine PCLS were exposed to different flow rates (3–30 mL/min) of synthetic air, O₃ (3.5–8.5 ppm), or NO₂ (1–80 ppm). Tissue survived ex vivo in ALI culture and resisted exposure to NO₂ (1–10 ppm) and O₃ (3.5–8.5 ppm) for 1 h. Longer exposure to NO₂ resulted in a clear loss of viability, whereas exposure to O₃ was less effective. Exposure to NO₂ dose-dependently induced release of the pro-inflammatory IL-1α (40%), whereas RANTES, IL-12, and eotaxin remained unchanged. Early secretion of IL-1α (80%), RANTES (>800%), MIP-1β (44%), and MCP-1 (60%) was already detected after 1 h of exposure to O₃. The obtained data showed that direct exposure to O₃ and NO₂ induced cytotoxicity and pro-inflammatory responses in PCLS with ALI culture. This provides a model that more closely resembles in vivo exposure of airborne contaminants, and thus should be appropriate for toxicity testing.     

A unified algorithm for predicting partition coefficients for PBPK modeling of drugs and environmental chemicals

The algorithms in the literature focusing to predict tissue:blood PC (P_tb) for environmental chemicals and tissue:plasma PC based on total (K_p) or unbound concentration (K_pu) for drugs differ in their consideration of binding to hemoglobin, plasma proteins and charged phospholipids. The objective of the present study was to develop a unified algorithm such that P_tb, K_p and K_pu for both drugs and environmental chemicals could be predicted. The development of the unified algorithm was accomplished by integrating all mechanistic algorithms previously published to compute the PCs. Furthermore, the algorithm was structured in such a way as to facilitate predictions of the distribution of organic compounds at the macro (i.e. whole tissue) and micro (i.e. cells and fluids) levels. The resulting unified algorithm was applied to compute the rat P_tb, K_p or K_pu of muscle (n = 174), liver (n = 139) and adipose tissue (n = 141) for acidic, neutral, zwitterionic and basic drugs as well as ketones, acetate esters, alcohols, aliphatic hydrocarbons, aromatic hydrocarbons and ethers. The unified algorithm reproduced adequately the values predicted previously by the published algorithms for a total of 142 drugs and chemicals. The sensitivity analysis demonstrated the relative importance of the various compound properties reflective of specific mechanistic determinants relevant to prediction of PC values of drugs and environmental chemicals. Overall, the present unified algorithm uniquely facilitates the computation of macro and micro level PCs for developing organ and cellular-level PBPK models for both chemicals and drugs.     

Purification, characterization and biological activities of the l-amino acid oxidase from Bungarus fasciatus snake venom

l-Amino acid oxidases (LAAOs) are widely distributed in snake venoms, which contribute to the toxicity of venoms. However, LAAO from Bungarus fasciatus (B. fasciatus) snake venom has not been isolated previously. In the present study, LAAO from B. fasciatus snake venom was purified by SP-Sepharose HP anion exchange chromatography followed by Heparin-Sepharose FF affinity chromatography procedure and the purified enzyme was named BF-LAAO. BF-LAAO presented an estimated molecular weight of 55 kDa in SDS-PAGE and an apparent molecular weight of 70 kDa in size-exclusion chromatography suggesting that BF-LAAO is a monomeric protein. Kinetics studies showed that BF-LAAO was very active against l-Tyr, l-Asp, l-Phe, l-Glu, l-Trp, l-His, l-Gln, l-Ile, l-Met, l-Leu and moderately active against l-Lys, l-Arg, l-Ala and l-Asn. BF-LAAO exhibited a cytotoxic effect on A549 cells and caused up to 41.2% apoptosis of A549 cells following 12 h incubation period. In the mouse peritoneum, BF-LAAO provoked a marked increase in the number of neutrophils, lymphocytes and macrophages following injection. It also induced rabbit platelet aggregation in a dose-dependent manner. At 3 h following injection, BF-LAAO elicited severe inflammation in the gastrocnemius muscles of mice, but failed to induce significant organ damage. In conclusion, the cytotoxic and proinflammatory activities of BF-LAAO could be the main cause of the local inflammation, which helps us to understand the pathogenesis of snakebite.     

Neuroprotective effects of Schisandrin B against transient focal cerebral ischemia in Sprague–Dawley rats

Fruits of Schisandra have been traditionally used in East Asia for the treatment of dyspnea, cough, dysentery, insomnia, tonic-clonic seizures, and amnesia. Schisandrin B, a dibenzocyclooctadiene derivative isolated from Fructus Schisandrae, has been shown to produce antioxidant effect on rodent liver and heart. In the present study, we investigated the neuroprotective effects of Schisandrin B, a constituent drug of the fruit of Schisandra, against focal cerebral ischemia in rats. Schisandrin B (10, 30 mg/kg, i.p.) was twice administered 30 min before the onset of ischemia and 2 h after reperfusion. Schisandrin B 10 and 30 mg/kg treated groups showed infarct volumes reduced by 25.7% and 53.4%, respectively, 2 h after occlusion. Also, Schisandrin B treated animal treatment abrogated protein expression of TNF-α and IL-1β and degradation of MMP-2 and MMP-9 in ischemic hemispheres. These results suggest that Schisandrin B treatment provides a neuroprotective effect to rats after transient focal cerebral ischemia by inhibiting inflammation and by protecting against metalloproteinase degradation.     

Local inflammatory response induced by scorpionfish Scorpaena plumieri venom in mice

The Scorpaena plumieri fish venom induces a severe pain and edema, observed both clinically and experimentally. In order to understand more about the envenomation syndrome, the present study characterized experimentally the local acute inflammatory response induced by S. plumieri venom (SpV) in a mouse model of tissue injury. Our results demonstrated that the local inflammatory response provoked after 2 h of SpV injection in footpad of mice is characterized by release of pivotal pro-inflammatory mediators (TNF, IL-6 and MCP-1). These mediators could be associated with histopathological changes observed into paw tissue, characterized by cellular infiltration, mainly neutrophils. Additionally, an investigation of edema formation pathways involved in inflammatory response was performed. SpV-induced edema was reduced significantly by previous administration of aprotinin or icatibant (HOE-140). However, the pre-treatment with diclofenac sodium and promethazine had less effect on this response. These results demonstrate that the kallikrein-kinin system (KKS) plays a major role in the edema formation. Despite the whole venom hydrolyzed the kallikrein synthetic substrate S-2302 (Pro-Phe-Arg-pNA), its main pro-inflammatory fraction was devoid of kininogenase activity. Our results demonstrate that SpV evokes a complex inflammatory reaction stimulating a secretion of TNF, IL-6, MCP-1 and leukocytes recruitment at the site of venom injection. In addition provide clear evidence of the involvement of the KKS in inflammatory response induced by S. plumieri venom.     

Bitis arietans Snake Venom and Kn-Ba,a Snake Venom Serine Protease,Induce the Production of Inflammatory Mediators in THP-1 Macrophages

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-1β 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.     

Experimental pathology of local tissue damage induced by Bothrops asper snake venom

Envenomations by Bothrops asper are often associated with complex and severe local pathological manifestations, including edema, blistering, dermonecrosis, myonecrosis and hemorrhage. The pathogenesis of these alterations has been investigated at the experimental level. These effects are mostly the consequence of the direct action of zinc-dependent metalloproteinases (SVMPs) and myotoxic phospholipases A₂ (PLA₂s). SVMPs induce hemorrhage, blistering, dermonecrosis and general extracellular matrix degradation, whereas PLA₂s induce myonecrosis and also affect lymphatic vessels. In addition, the prominent vascular alterations leading to hemorrhage and edema may contribute to ischemia and further tissue necrosis. The mechanisms of action of SVMPs and PLA₂s are discussed in detail in this review. Venom-induced tissue damage plays also a role in promoting bacterial infection. A prominent inflammatory reaction develops as a consequence of these local pathological alterations, with the synthesis and release of abundant mediators, resulting in edema and pain. However, whether inflammatory cells and mediators contribute to further tissue damage is not clear at present. Muscle tissue regeneration after venom-induced pathological effects is often impaired, thus resulting in permanent tissue loss and dysfunction. SVMP-induced microvessel damage is likely to be responsible of this poor regenerative outcome. Antivenoms are only partially effective in the neutralization of B. asper-induced local effects, and the search for novel toxin inhibitors represents a potential avenue for improving the treatment of this serious aspect of snakebite envenomation.     

Isolation and functional characterization of a new acidic PLA2 Ba SpII RP4 of the Bothrops alternatus snake venom from Argentina

An acidic protein with phospholipase A₂ activity was purified to homogeneity from the venom of the Northeast Argentinian viperid Bothrops alternatus by two chromatographic steps: a conventional gel filtration on Sephadex G-75 and reversed phase on C18 HPLC column.A molecular mass of 14185.48 Da was determined by mass spectrometry, displaying a homodimer conformation. The kinetic assay demonstrated a catalytically active phospholipase A₂ in correspondence with Asp49 PLA₂ group. The enzyme designated Ba SpII RP4 contains an amino acid composition of 121 residues and a calculated theoretical pI value of 4.88. Amino acid sequence alignments with other Bothrops PLA₂ revealed a high degree of homology sequence (90-56%). Ba SpII RP4 did not show myotoxic activity upon muscular fibers at doses up to 100 μg i.m. route injection or lethal response when it was i.p. injected at the hightest dose of 200 μg. This toxin generates slight biological activities like paw edema inflammation and a delay in the clotting time, although Ba SpII RP4 exhibited catalytic activity. The primary amino acid sequence, determined a quadruple-time of flight (Q-TOF) hybrid mass spectrometer Q-TOF Ultima from Micromass (Manchester, UK) equipped with a nano Zspray source operating in a positive ion mode and tandem mass spectrum, an ESI/MS mass spectrum (TOF MS mode) “de novo amino acid sequencing”, also provides more database about the small group of the non-myotoxic PLA₂s isolated up to the present.     

Expression of proliferative and inflammatory markers in a full-thickness human skin equivalent following exposure to the model sulfur mustard vesicant, 2-chloroethyl ethyl sulfide

Sulfur mustard is a potent vesicant that induces inflammation, edema and blistering following dermal exposure. To assess molecular mechanisms mediating these responses, we analyzed the effects of the model sulfur mustard vesicant, 2-chloroethyl ethyl sulfide, on EpiDerm-FT™, a commercially available full-thickness human skin equivalent. CEES (100-1000 μM) caused a concentration-dependent increase in pyknotic nuclei and vacuolization in basal keratinocytes; at high concentrations (300-1000 μM), CEES also disrupted keratin filament architecture in the stratum corneum. This was associated with time-dependent increases in expression of proliferating cell nuclear antigen, a marker of cell proliferation, and poly(ADP-ribose) polymerase (PARP) and phosphorylated histone H2AX, markers of DNA damage. Concentration- and time-dependent increases in mRNA and protein expression of eicosanoid biosynthetic enzymes including COX-2, 5-lipoxygenase, microsomal PGE₂ synthases, leukotriene (LT) A₄ hydrolase and LTC₄ synthase were observed in CEES-treated skin equivalents, as well as in antioxidant enzymes, glutathione S-transferases A1-2 (GSTA1-2), GSTA3 and GSTA4. These data demonstrate that CEES induces rapid cellular damage, cytotoxicity and inflammation in full-thickness skin equivalents. These effects are similar to human responses to vesicants in vivo and suggest that the full thickness skin equivalent is a useful in vitro model to characterize the biological effects of mustards and to develop potential therapeutics.     

Arsenic-induced myocardial injury: Protective role of Corchorus olitorius leaves

Groundwater arsenic contamination in Bangladesh and its adjoining part of West Bengal (India) is reported to be the biggest arsenic calamity in the world in terms of the affected population. Tossa jute, Corchorus olitorius is a popular crop of this arsenic prone population. The present study was undertaken to evaluate the protective effect of aqueous extract of C. olitorius leaves (AECO) against sodium arsenite (NaAsO₂) induced cardiotoxicity in experimental rats. The animals exposed to NaAsO₂ (10 mg/kg, p.o.) for 10 days exhibited a significant inhibition (p < 0.01) of superoxide dismutase, catalase, glutathione-S-transferase, glutathione peroxidase, glutathione reductase and reduced glutathione level in myocardial tissues of rats. In addition, it significantly increased (p < 0.01) oxidized glutathione, malondialdehyde and protein carbonyl content in myocardial tissue. Treatment with AECO (50 and 100 mg/kg, p.o.) for 15 days prior to NaAsO₂-intoxication significantly protected cardiac tissue against arsenic-induced oxidative impairment. In addition, AECO pretreatment significantly prevented NaAsO₂ induced hyperlipidemia, cardiac arsenic content and DNA fragmentation in experimental rats. Histological studies of myocardial tissue supported the protective activity of the AECO. The results concluded that the treatment with AECO prior to arsenic intoxication has significant protecting effect against arsenic-induced myocardial injury.     

Toxicity of Jatropha curcas phorbol esters in mice

Phorbol esters are the main toxins in Jatropha curcas seed and oil. The aim of this study was to assess the acute toxicity of phorbol esters given by intragastric administration and to determine the LD₅₀ for Swiss Hauschka mice. The LD₅₀ and 95% confidence limits for male mice were 27.34 mg/kg body mass and 24.90–29.89 mg/kg body mass; and the LD₅ and LD₉₅ were 18.87 and 39.62 mg/kg body mass, respectively. The regression equations between the probits of mortalities (Y) and the log of doses (D) was Y = −9.67 + 10.21 log (D). Histopathological studies on the organs from the dead mice showed: (1) no significant abnormal changes in the organs at the lowest dose (21.26 mg/kg body mass) studied, (2) prominent lesions mainly found in lung and kidney, with diffused haemorrhages in lung, and glomerular sclerosis and atrophy in kidney at doses ?32.40 mg/kg body mass, and (3) multiple abruption of cardiac muscle fibres and anachromasis of cortical neurons at the highest dose of 36.00 mg/kg body mass. The results obtained would aid in developing safety measures for the Jatropha based biofuel industry and in exploiting the pharmaceutical and agricultural applications of phorbol esters.     

Natural phospholipase A2 myotoxin inhibitor proteins from snakes, mammals and plants

A renewed interest in the phenomenon of inter- and intra-species resistance towards the toxicity of snake venoms, coupled with the search for new strategies for treatment of snake envenomations, has prompted the discovery of proteins which neutralize the major toxic components of these venoms. Among these emerging groups of proteins are inhibitors of toxic phospholipases A₂ (PLA₂s), many of which exhibit a wide range of toxic effects including muscle-tissue damage, neurotoxicity, and inflammation. These proteins have been isolated from both venomous and non-venomous snakes, mammals, and most recently from medicinal plant extracts. The snake blood-derived inhibitors have been grouped into three major classes, α, β, and γ, based on common structural motifs found in other proteins with diverse physiological properties. In mammals, DM64, an anti-myotoxic protein isolated from opossum serum, belongs to the immunoglobulin super gene family and is homologous to human α₁B-glycoprotein and DM43, a metalloproteinase inhibitor from the same organism. In plants, a short note is made of WSG, a newly described anti-toxic-PLA₂ glycoprotein isolated from Withania somnifera (Ashwaganda), a medicinal plant whose aqueous extracts neutralize the PLA₂ activity of the Naja naja venom. The implications of these new groups of PLA₂ toxin inhibitors in the context of our current understanding of snake biology as well as in the development of novel therapeutic reagents in the treatment of snake envenomations worldwide are discussed.