Enzymatic Analysis of Iranian Echis carinatus Venom Using Zymography

Snakebite is a common problem especially in tropical areas all over the world including Iran. Echis carinatus as one of the most dangerous Iranian snakes is spreading in this country excluding central and northwest provinces. In this study gelatinase and fibrinogenolytic properties as two disintegrating matrix metalloproteinase enzymes were evaluated by a strong clear halo between 56-72 kDa in addition to another band located 76-102 kDa for gelatinase and one major band around 38 kDa for fibrinogenolytic enzyme respectively. The electrophorectc profile of our venom demonstrated at least one protein band between 24-31 kDa like previous reports and another two bands between 52-76 kDa and below 17 kDa stemmed probably due to the effect of natural selection in one species. According to our results Razi institute antivenin could neutralize in-vitro effects of gelatinase enzyme comprehensively. The electrophoretic profile of Iranian commercial antivenom as the main intravenous treatment of envenomed patients showed impurities in addition to F (abʹ)2 weighing 96 kDa in SDS-PAGE analysis. It proposes more efforts for refinement to avoid short and long unwanted effects in envenomed patients

A biodistribution study of Hemiscorpius lepturus scorpion venom and available polyclonal antivenom in rats

The purpose of the present study was to investigate the biodistribution profile of the venom of Hemiscorpius lepturus, the most dangerous scorpion in Iran. Blood and tissue samples were taken at various predetermined intervals during a 400-minute period for the venom and a 360-minute period for the antivenom in rats. The radio-iodination was carried out using the chloramine-T method. The results showed that the descending order of venom uptake was skin, kidneys and intestine, respectively. The descending order of polyclonal antivenom uptake was kidneys, intestine, heart and lungs. The calculated pharmacokinetic parameters of the venom were Telimination half-life = 521.5 ± 12.6 minutes; Vd/F (apparent volume of distribution) = 14.9 ± 3.3 mL; clearance (CL/F, apparent total clearance of the drug from plasma) 0.02 ± 0.005 mL/minute and for the antivenom Telimination half-life = 113.7 ± 7.4 minutes; Vd/F = 13 ± 1.2 mL and CL/F 0.08 ± 0.01 mL/minute. The pharmacokinetics profile comparison of the venom with that of the antivenom shows that serotherapy may be more effective if administered within 2-4 hours following envenomation by H. lepturus

Cardiotoxic and Arrhythmogenic Effects of Hemiscorpius lepturus Scorpion Venom in Rats

Background: Envenomation by Hemiscorpius lepturus is not painful and the clinical manifestations include bloody urine due to hemoglobinuria or hematuria, dermonecrotic reactions,cardiac arrhythmia and in minority of cases acute renal failure which may lead to death following disseminated intravascular coagulation in infants. Cardiac effects of envenomation by H. lepturus venom including inotropic, chronotropic and arrhythmogenic properties are not studied as now in rat hearts with Langendorff apparatus. Methods: Rat hearts were allowed to equilibrate in its buffer and cardiotropic plus arrhythmogenic effects induced by injection of different doses of H. lepturus venom were detected and recorded by computer acquisition based data in Langendorff apparatus. The neutralizing effects of Razi Institute antivenom and autonomic drugs were assayed in parallel studies. Results: Hemiscorpius lepturus venom (25 μg/100 l) treatment caused a negative inotropic (65.4 ± 3.2 versus 110.2 ± 3.4) and chronotropic effects (186.3 ± 4.2 versus 302 ± 6.3) in comparison to normal saline. Arrhythmogenic aspects including bradycardia, QRS widening and ST depression were induced by venom injection. Pre venom treatment (20 min) of Razi Institute antivenom (10 μl) neutralized cardiotropic effects but post venom injection (15 min later) had no therapeutic role. Pre (10 min before) and post (15 min after) injection of adrenaline (10 μl) neneutralized cardiotropic effects while pre venom injection (20 min) of propanolol (10 μl) had aggravating effects. Conclusion: Our study paves the way for further in vivo investigation of cardiovascular effects of this venom for finding suitable treatments instead of its ordinary antivenom medication in cardiogenic shock induced by the veno

Serological, pathological and scintigraphic assessment of Hemiscorpius lepturus effects on renal dysfunction in rats

Objective(s): Hemiscorpius lepturus is one of the dangerous scorpions of Iran leading to acute kidney injury (AKI) especially in infants. The purpose of this animal study was to compare the serological, pathological and scintigraphic data to quickly predict the occurrence of this disorder.Materials and Methods: In two groups of animals, each contained five rats, H. lepturus venom (1200 µg/Kg) were injected intravenously via the tail vein. At three hours and one week later, 99m Tc-DMSA (3 mCi) was intravenously injected and renal scintigraphy was performed after an hour. Moreover, plasma levels of creatinine, sodium, potassium, and blood urea nitrogen (BUN) were measured. At the end of the study, renal tissues were excised and prepared to perform pathological evaluation after Hematoxylin and Eosin staining.Results: All serological indices were remained unchanged compared to control. A large number of glomerular fibrin thrombi with entrapped red blood cells and simplified tubular epithelium in dilated and ectatic tubules were observed in high power field (×100) four hours after envenomation, which reduced significantly one week later. In our scintigraphic study, there was a statistically significant difference (

Cloning of Metalloproteinase 17 Genes from Oriental Giant Jellyfish Nemopilema nomurai (Scyphozoa: Rhizostomeae)

We previously demonstrated that Nemopilema nomurai jellyfish venom metalloproteinases (JVMPs) play a key role in the toxicities induced by N. nomurai venom (NnV), including dermotoxicity, cytotoxicity, and lethality. In this study, we identified two full-length JVMP cDNA and genomic DNA sequences: JVMP17-1 and JVMP17-2. The full-length cDNA of JVMP17-1 and 17-2 contains 1614 and 1578 nucleotides (nt) that encode 536 and 525 amino acids, respectively. Putative peptidoglycan (PG) binding, zinc-dependent metalloproteinase, and hemopexin domains were identified. BLAST analysis of JVMP17-1 showed 42, 41, 37, and 37% identity with Hydra vulgaris, Acropora digitifera, Megachile rotundata, and Apis mellifera venom metalloproteinases, respectively. JVMP17-2 shared 38 and 36% identity with H. vulgaris and A. digitifera, respectively. Alignment results of JVMP17-1 and 17-2 with other metalloproteinases suggest that the PG domain, the tissue inhibitor of metalloproteinase (TIMP)-binding surfaces, active sites, and metal (ion)-binding sites are highly conserved. The present study reports the gene cloning of metalloproteinase enzymes from jellyfish species for the first time. We hope these results can expand our knowledge of metalloproteinase components and their roles in the pathogenesis of jellyfish envenomation

The toxinology of sea snakes: A systematic review

Background: Sea snakes belong to Hydrophiidae family mainly are found in tropical and subtropical waters of the world including the Persian Gulf. Their highly lethal venoms are more potent than snakes with terrestrial origin and contain complex mixtures of organic and inorganic bioactive substances, such as enzyme, and non-enzymatic proteins. There were limited studies on the venoms and toxins of sea snake; hence, the aim of this systematic review was to investigate the toxinological dimensions of sea snakes. Materials and Methods: In order to investigate of the toxins of sea snakes,  the “Hydrophis schistosus toxin", "Hydrophis cyanocinctus toxin", " Hydrophis lapemoides toxin", "Hydrophis spiralis toxin", and "Lapemis curtus toxin " terms were searched separately, in "PubMed database", in 10/08/2016  which obtained 32, 9, 2, 2 and 4 papers, respectively. The "Hydrophis gracilis toxin" term had no any results.  For the “Hydrophis gracilis" term, two studies were obtained. The first one related to the sea snakes phylogenetic characteristics and the second one shared with other search results and well-connected with the issue. Some papers were similar in different searches. Of these, those studies were selected that had direct relevance to the subject  . Results: The main isolated toxins from different sea snakes venoms included three-finger toxin (3FTx (short chain neurotoxins isoforms: AAL54893, AAL54892, ABN54806; long chain neurotoxins isoforms: AAL54894, AAL54895, P68416 , ABN54805), pelamitoxin (P62388), phospholipase A2 (both the basic and acidic PLA2), two phospholipases A2 (PLA2-H1 and H2), cysteine-rich secretory protein (CRISP), snake venom Zn2+-metalloproteinase (SVMP), L-amino acid oxidase (LAAO), 5′-nucleotidase, Hydrophitoxins a, b and c, Hydrophis ornatus a, Hydrophis lapemoides a, PDGF and α- neurotoxins of rSN311, rSN316 and rSN285. Each toxin and protein family presents a wide range of pharmacological activities. Some of these neurotoxins were linked to acetylcholine receptors in the neuromuscular junctions. These toxins showed protease (gelatinase and caseinase) activities, and/or they produce the myonecrosis and biochemical and histopathological changes. Conclusion: There is scant variability in the venom composition in the same and different species of sea snakes. Our study revealed that there is a rather simple venom profile with an affinity towards a lethal mixture of high abundance of neurotoxins and PLA2s, and lower amounts of toxins such as CRISP, SVMP and LAAO

A Comparative Analysis of Saffron and Methylprednisolone on Bleomycin-Induced Pulmonary Fibrosis in Rats

Background: The purpose of this study was to compare the effects of saffron and methylprednisolone on bleomycin-induced pulmonary fibrosis in rats. Methods: This study was conducted in Bushehr, southern Iran in 2017.The animals were divided into four groups of five rats each. Three groups were injected with a single intratracheal dose of bleomycin (5 mg/kg). The fourth group was administered with normal saline at the same volume (200 µl). Saffron extract dissolved in water was given to one group (100 mg /body weight) orally while intraperitoneal injection of methylprednisolone (2.5 mg/kg) injected to another one for 16 days. The rats were sacrificed 28 days following surgery and their right and left lungs were removed and washed for measuring lung indices, myeloperoxidase activities and finally histopathological examination. Results: Injection of bleomycin caused decrement of body weight aggravated by intraperitoneal methylprednisolone treatment. Lung indices were increased in the bleomycin-treated group compared with the control, while methylprednisolone, unlike saffron, had no preventive effects on it. Both saffron and methylprednisolone treatment prevented the increase in lung myeloperoxidase as a destructive enzyme. In addition, excessive collagen deposition and thickening of alveolar septa were significantly prevented with saffron treatment as compared to methylprednisolone injection following hematoxylin and eosin staining. Conclusion: Saffron with established antioxidant properties could prevent some detrimental effects in bleomycin-induced pulmonary fibrosis even more than methylprednisolone injection known as a standard therapy in this murine model. More investigations must be carried out to examine the beneficial or harmful effects of this remedy

Hemodynamic Changes in Experimentally Envenomed Anaesthetized Rats by Intravenous Injection of Hemiscorpius lepturus Venom

Background: We investigated the hemodynamic changes (Inotropic, chronotropic and arrhythmogenic) in intrave­nously envenomed anesthetized rats with Hemiscorpius lepturus venom. The neutralizing potencies of different drugs and commercial antivenom were assessed simultaneously. Methods: Different doses of the crude venom (100, 200 and 400μg/rat) were injected during five minutes via the femoral vein and cardiovascular changes were recorded in rats in Razi Institute Corporation, Karaj, Iran in 2017. The drugs (Atropine, lidocaine, propranolol and prazosin) were injected before the venom for determination of the coun­teracting effects. Different volumes (100, 500 and 1000µl) of the antivenom were pre envenomed to neutralize cardi­ovascular changes. Results: Temporary hypertension and bradycardia with no arrhythmogenic effects were depicted within twenty minutes. There was a difference in arterial pressure between the venom (400μg/rat) and the vehicle at 8 minutes (114.68±5.1mmHg versus 70.2±4.3mmHg). Elevation of the mean arterial pressure was inhibited by propranolol (2 mg/kg) and neutralized by prazosin (1mg/kg) while lidocaine (4mg/kg) and atropine (1mg/kg) had no effects. Pre­medication with Iranian commercial antivenom (1000μl) produced surprisingly temporary hypertension compared to the vehicle (140.84±4.5 versus 84.3±3.2). It had no neutralizing properties on blood pressure variation before the venom injection. Volume-expanded hypertension phenomenon was ruled out in a parallel study. Conclusion: This venom has vasoconstrictive effects in rats probably due to the presence of norepinephrine like ma­terials in its content or liberated from adrenal gland inhibited by prazosin premedication. The neutralizing effects of antivenom on venom-induced hypertension are questionable

Saffron Protection against Bleomycin-Induced Pulmonary Fibrosis in Rats

Background: Bleomycin-induced lung fibrosis has been accepted as an animal model for fibrosis in rats. The aim of this study was to evaluate the effects of saffron aqueous extract on this disorder paving the way for more investigation in treating idiopathic pulmonary fibrosis in human. Methods: Male Wistar rats (250–300 gr) were instilled a single dose of bleomycin (5 mg/kg) via intratracheal tube (n=6) in 2015. Sham group received normal saline. Saffron aqueous extract (50 mg/kg and 100 mg/kg) were given orally in two different treated groups with bleomycin for 28 days. Lung Indices was calculated at the end of this experiment. Lung segments fixed in 10% formaldehyde were used for pathological preparation with Hematoxylin & Eosin and trichrome staining. Results: The body weight was decreased and lung Indices increased in bleomycin group (P<0.5). Bleomycin administration increased myeloperoxidase, malondialdehyde and finally TNF-α in lung tissue homogenates (P<0.05) compared with sham group. The fibrotic process and thickening of alveolar septa in treated rats with bleomycin were increased by H&E and Masson Trichrome staining. Saffron treatment (50 and 100 mg/kg) attenuated the increase in MDA (264.43±10.4 nmol/g by the higher dose versus 378.4±18.1nmol/g), MPO (0.19±0.03 and 0.13± 0.04 IU/ml versus 0.39.2±0.05 IU/ml) and TNF-α level (18.42±3.7 ng/ml and14.31±3.6 ng /ml versus 35.32±4.2) in lung homogenates compared to bleomycin group (P<0.05). It decreased collagen accumulation and alveolar destructive patterns in pulmonary fibrosis. Conclusion: This study introduces saffron as novel anti-fibrotic agent against bleomycin-induced fibrosis due to histological examinations and preventive effects on destructive enzyme release in rats

Cloning of Metalloproteinase 17 Genes from Oriental Giant Jellyfish <i>Nemopilema nomurai</i> (Scyphozoa: Rhizostomeae)

We previously demonstrated that Nemopilema nomurai jellyfish venom metalloproteinases (JVMPs) play a key role in the toxicities induced by N. nomurai venom (NnV), including dermotoxicity, cytotoxicity, and lethality. In this study, we identified two full-length JVMP cDNA and genomic DNA sequences: JVMP17-1 and JVMP17-2. The full-length cDNA of JVMP17-1 and 17-2 contains 1614 and 1578 nucleotides (nt) that encode 536 and 525 amino acids, respectively. Putative peptidoglycan (PG) binding, zinc-dependent metalloproteinase, and hemopexin domains were identified. BLAST analysis of JVMP17-1 showed 42, 41, 37, and 37% identity with Hydra vulgaris, Acropora digitifera, Megachile rotundata, and Apis mellifera venom metalloproteinases, respectively. JVMP17-2 shared 38 and 36% identity with H. vulgaris and A. digitifera, respectively. Alignment results of JVMP17-1 and 17-2 with other metalloproteinases suggest that the PG domain, the tissue inhibitor of metalloproteinase (TIMP)-binding surfaces, active sites, and metal (ion)-binding sites are highly conserved. The present study reports the gene cloning of metalloproteinase enzymes from jellyfish species for the first time. We hope these results can expand our knowledge of metalloproteinase components and their roles in the pathogenesis of jellyfish envenomation