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B Lomonte

Publications and source records attributed to B Lomonte.

At least 73 records · Page 4Linked to original sources
PubMed1992

Individual expression patterns of myotoxin isoforms in the venom of the snake Bothrops asper.

1. In order to investigate the distribution of myotoxin isoforms in the snake Bothrops asper, venoms from individual specimens were analyzed by a cathodic electrophoretic system for basic proteins under native conditions. 2. The electrophoretic system resolved at least five bands with slight differences in mobility, corresponding to the fastest migrating proteins in the venom. The identity of the bands was confirmed by immunoblotting, using a rabbit anti-myotoxin serum. 3. There were clear differences in the individual patterns of myotoxin isoform expression, both in specimens from the Atlantic and Pacific regions of Costa Rica. Some individuals possessed all five variants. 4. In agreement with previous reports, the venoms of ten newborn (less than 10 days of age) specimens completely lacked myotoxin bands, indicating an ontogenetic regulation in the expression of these toxins in B. asper. 5. One of the bands, corresponding to the lysine-49 phospholipase myotoxin II, was the only isoform present in all individuals studied, suggesting a possible selective pressure for the conservation of this type of protein in the venom of B. asper.

Blotting, Western↗
PubMed1992

Immunoenzymatic quantitation of antibodies to Bothrops asper myotoxins after polyvalent antivenom administration in mice.

1. Two quantitative enzyme-immunoassays (EIA) for Bothrops asper myotoxin and anti-myotoxin antibodies, respectively, were utilized to study their in vivo distribution in mice (Swiss, 18 to 20 g). 2. After polyvalent antivenom (0.4 ml) administration by the iv route, there was an immediate peak in plasma anti-myotoxin antibodies which declined rapidly during the first hour, and then decreased more gradually. Anti-myotoxin antibodies were detected in muscular tissue (gastrocnemius) following iv injection of antivenom. After im injection of antivenom (0.4 ml), a slow and steady increase in plasma anti-myotoxin levels was observed, with a peak at 24 h. 3. Mice that received antivenom (0.4 ml) by the iv or im route 15 min after im injection of B. asper venom (100 micrograms) had lower levels of plasma anti-myotoxin antibodies than controls injected with antivenom only, suggesting that at least a fraction of the antibodies combines with myotoxins in vivo. 4. Myotoxin was not detected in plasma at any time after venom injection by the im (100 micrograms) or ip (40 micrograms) route. Following iv injection of 50 micrograms of purified myotoxin II, all plasma samples were also negative, at a detection limit of 10 ng/ml. 5. It was demonstrated that myotoxin II binds to mouse erythrocytes in vitro, a fact that could partially explain its rapid in vivo disappearance from plasma. 6. The present results on the distribution of anti-myotoxin antibodies in vivo are in agreement with previous experimental studies reporting the poor neutralization of myotoxicity induced by B. asper venom when antivenom is injected im, in comparison to i.v. injection.

Animals↗
PubMed1991

The effect of myotoxins isolated from Bothrops snake venoms on multilamellar liposomes: relationship to phospholipase A2, anticoagulant and myotoxic activities.

The effect of four myotoxins isolated from Bothrops snake venoms on the release of peroxidase trapped in large multilamellar liposomes was studied and correlated to their phospholipase A2, myotoxic and anticoagulant activities. The four myotoxins affected negatively-charged liposomes in a dose-dependent way, having no effect on positively-charged liposomes. Conditions that inhibited phospholipase A2 activity, i.e., substitution of calcium by EDTA, reduced liposome-disrupting activity of Bothrops asper myotoxin I and Bothrops atrox myotoxin, both of which have high phospholipase A2 activity, but did not affect the action of B. asper myotoxin II and Bothrops moojeni myotoxin II, which have extremely low phospholipase A2 activity. However, all myotoxins disrupted to some extent negatively-charged liposomes under conditions where phospholipase A2 activity was abolished. Since these toxins behave as amphiphilic proteins in charge-shift electrophoresis, it is suggested that membrane-disorganization is at least partially due to a non-enzymatic penetration and alteration of bilayers. There was no strict correlation between liposome-disrupting activity and myotoxicity in vivo. Thus, although both effects probably depend on the toxins' ability to disturb membranes, it is likely that variation in complexity between skeletal muscle plasma membrane and liposome bilayers are the basis for this difference. The anticoagulant effect seems to depend on the ability of the toxins to enzymatically degrade phospholipids, since only B. asper myotoxin I and B. atrox myotoxin prolonged the plasma recalcification time.

Animals↗
PubMed1991

Myotoxin II from Bothrops asper (Terciopelo) venom is a lysine-49 phospholipase A2.

A basic, dimeric myotoxic protein, myotoxin II, purified from Bothrops asper venom has a similar molecular weight and is immunologically cross-reactive with antibodies raised to previously isolated B. asper phospholipases A2, except that it shows only 0.1% of the phospholipase activity against L-alpha-phosphatidylcholine in the presence of Triton X-100. Its 121 amino acid sequence, determined by automated Edman degradation, clearly identifies it as a Lys-49 phospholipase A2. Key amino acid differences between myotoxin II and phospholipase active proteins in the Ca2(+)-binding loop region, include Lys for Asp-49, Asn for Tyr-28, and Leu for Gly-32. The latter substitution has not previously been seen in Lys-49 proteins. Other substitutions near the amino terminus (Leu for Phe-5 and Gln for several different amino acids at position 11) may prove useful for identifying other Lys-49 proteins in viperid and crotalid venoms. Myotoxin II shows greater sequence identity with other Lys-49 proteins from different snake venoms (Agkistrodon piscivorus piscivorus, Bothrops atrox, and Trimeresurus flavoviridis) than with another phospholipase A2 active Asp-49 molecule isolated from the same B. asper venom. This work demonstrates that phospholipase activity per se, is not required in phospholipase molecules for either myotoxicity or edema inducing activities.

Amino Acid Sequence↗
PubMed1991

Quantitation by enzyme-immunoassay of antibodies against Bothrops myotoxins in four commercially-available antivenoms.

An enzyme-immunoassay (EIA) for the quantitation of antibodies against myotoxins present in the venoms of Bothrops asper (Costa Rica), B. atrox (Colombia) and B. moojeni (Brazil), was developed. This EIA was utilized for the evaluation of four antivenoms produced in Mexico (Laboratorios Myn; MYN), Costa Rica (Instituto Clodomiro Picado; ICP), Colombia (Instituto Nacional de Salud; INS) and Brazil (Instituto Butantan; IB). Antivenoms ICP, IB and INS showed a higher titer of antibodies against the three myotoxins tested, with only slight differences between them, depending on the antigen utilized. In contrast, MYN antivenom had very low levels of antibodies to the three myotoxins. Seventeen batches of ICP antivenom were analyzed by EIA, using B. asper myotoxin II as antigen. Although all batches had high anti-myotoxin titers, these varied significantly. Batches produced after 1988 had, in general, higher titers than older (1986-1987) ones. Antivenom stored for one year at 37 degrees C had a slight, but significant (P less than 0.002) decrease of anti-myotoxin activity, compared to antivenom stored at 4 degrees C or 23 degrees C. No correlation was found between anti-myotoxin concentration and lethality-neutralizing ability of the ICP antivenom.

Animals↗
PubMed1991

Neutralization of coral snake Micrurus nigrocinctus venom by a monovalent antivenom.

1. The neutralizing ability of a monovalent anti-Micrurus nigrocinctus (coral snake) antivenom produced in Costa Rica was tested against the lethal, myotoxic and phospholipase A2 activities of homologous venom. In addition, immunodiffusion and Western blot analyses were performed. 2. In experiments where venom and antivenom were incubated prior to the test, antivenom was effective in neutralizing lethal, myotoxic and phospholipase A2 activities, with Effective Doses 50% of 2700 microliters antivenom/mg venom, 1840 microliters antivenom/mg venom, and 3630 microliters antivenom/mg venom, respectively. 3. When coral snake antivenom was administered at different times after coral snake venom injection, neutralization of lethality was achieved when antivenom was injected i.v. immediately and 15 min after venom. In contrast, lethality was not reduced when antivenom was administered by the im route. Only partial neutralization of myotoxicity was observed even when antivenom was injected i.v. immediately after envenomation. 4. Immunodiffusion and immunoblot analyses demonstrated the presence of antibodies in antivenom against several, but not all, venom components.

Animals↗
PubMed1990

Isolation of a galactose-binding lectin from the venom of the snake Bothrops godmani (Godmann's pit viper).

A galactose-binding lectin, isolated from the venom of B. godmani by affinity chromatography, is an acidic protein (pI 4.9) with a subunit mol. wt of about 14,000, occurring mostly as a disulfide-linked dimer of 28,000. A small proportion of lectin appears as a monomer and as a tetramer. The lectin agglutinates erythrocytes from mice, rabbit, cow and human (all ABO types, either Rh positive or negative), but does not agglutinate horse, sheep, goat and snake (Oxybelis aeneus, Colubridae) erythrocytes. The agglutinating activity is inhibited by 1 mM EDTA. The lectin is devoid of lethal, hemorrhagic, myotoxic, proteolytic and phospholipase A2 activities. It is not mitogenic for human peripheral blood mononuclear cells. The only effect observed was a moderate induction of edema in the footpad of mice, with a minimal edema-forming dose of 22 micrograms. This effect developed rapidly, and was significantly inhibited by i.p. administration of cyproheptadine, a histamine and serotonin antagonist, before injection of the lectin. Despite the edema-forming activity observed, the low concentration of lectin in crude venom, together with its relatively low potency, suggest that this lectin is not a key component in the development of edema following envenomations by B. godmani.

Animals↗
PubMed1990

Effect of storage temperature on the stability of the liquid polyvalent antivenom produced in Costa Rica.

The effect of storage temperature on the stability of the liquid polyvalent (crotaline) antivenom produced at the Instituto Clodomiro Picado, Costa Rica, was studied during a twelve-month period. The following parameters were evaluated: neutralizing potency against lethal activity of Bothrops asper venom; protein and phenol concentrations; pH; turbidity; safety; and sterility. Analyses were performed each month on different samples of a batch, stored at 4, 23, 30 and 37 degrees C. No significant (P greater than 0.1) variations occurred in potency, protein and phenol concentrations, pH, sterility or safety, at any of the storage temperatures during the study period. However, visual inspection revealed a moderate increase in turbidity of the samples stored at 23, 30 and 37 degrees C, at nine, four and three months, respectively. Culture of samples excluded the possibility of microbial contamination of the product leading to turbidity. Chromatographic and electrophoretic analyses demonstrated that turbidity was caused by the formation of heterogeneous protein aggregates of high molecular weight. Present results support the conclusion that, although storage temperature (up to 37 degrees C for twelve months) does not alter antivenom potency, it significantly influences the formation of protein aggregates. This phenomenon can be prevented by recommending the storage of antivenom at refrigeration temperature.

Animals↗
PubMed1990

Equine antibodies to Bothrops asper myotoxin II: isolation from polyvalent antivenom and neutralizing ability.

Equine antibodies to B. asper myotoxin II were isolated from polyvalent antivenom by affinity chromatography. Purified antibodies were among the most acidic serum immunoglobulins, migrating between the beta- and alpha 2-globulin regions by zone electrophoresis. This might be related to the fact that myotoxin II is a very basic antigen. At an antibody/toxin molar ratio of two or higher, myotoxic effect of myotoxin II was neutralized completely. Antibodies also neutralized myotoxic activity of crude venom in mice, reducing the effect by about 75% when a proportion of 16 mg antibodies/mg venom was tested. Despite the lack of phospholipase A2 activity of myotoxin II, antibodies were able to neutralize this enzymatic activity of myotoxin I, a previously described isoform. This finding is in agreement with the notion that myotoxin II is a phospholipase A2-analog devoid of this enzyme activity.

Animals↗
PubMed1990

Ontogenetic changes in the venom of the snake Lachesis muta stenophrys (bushmaster) from Costa Rica.

A comparative study was performed on some enzymatic and toxic activities of venoms collected from newborn, one-year old, two-years old and adult (more than five-years old) specimens of Lachesis muta stenophrys. There was an increase in lethal, hemorrhagic, edema-forming, myotoxic, proteolytic and phospholipase A2 activities of venoms as snakes aged. The venom of newborn specimens was almost devoid of toxicity. On the other hand, venom from newborn specimens showed the highest coagulant effect of human plasma. Electrophoretic and immunochemical results demonstrated conspicuous differences between venoms of different ages. Observations on the feeding behavior indicated that specimens of L. muta of different ages displayed a similar pattern, characterized by rapid strike and bite, holding the prey until they stopped their movements and swallowing them afterwards. It is concluded that venom of newborn L. muta has very low toxic and proteolytic activities and that it undergoes conspicuous changes during the first year of life.

Aging↗
PubMed1990

Isolation of basic myotoxins from Bothrops moojeni and Bothrops atrox snake venoms.

Three myotoxins, one from the venom of Bothrops atrox and two from the venom of B. moojeni, were isolated by ion-exchange chromatography on CM-Sephadex C-25. The three toxins are basic proteins with an estimated mol. wt of about 13,500, and similar amino acid compositions. When injected into the gastrocnemius muscle of mice, the three toxins induce drastic myonecrosis of rapid onset, as judged by histological observation and quantitation of plasma creatine kinase levels. B. atrox myotoxin also has phosphlipase A2 and anticoagulant activities, whereas B. moojeni myotoxins I and II lack these effects. The three toxins are antigenically similar to each other, and to previously isolated myotoxins I and II from the venom of B. asper, when tested by gel immunodiffusion against rabbit antiserum to B. asper myotoxin I. Two monoclonal antibodies against B. asper myotoxins were tested against the newly purified proteins. MAb-3 recognizes all of them, whereas MAb-4 recognizes only B. atrox myotoxin, by enzyme-immunoassay. B. atrox and B. moojeni myotoxins can be tentatively classified within a group of myotoxins having phospholipase A2 structure present in Bothrops venoms.

Amino Acids↗
PubMed1990

South American snake venom proteins antigenically related to Bothrops asper myotoxins.

1. The presence of proteins antigenically related to Bothrops asper myotoxins in various snake venoms, mainly from South America, was investigated by using polyclonal and monoclonal antibodies. 2. Myotoxin-like components were detected in ten Bothrops venoms from South America, and in the venoms of Crotalus atrox (North America), Trimeresurus flavoviridis (Japan), and Micrurus alleni (Costa Rica). 3. Cross-reactive components detected in several Bothrops venoms show a common subunit of 15-16 kDa by sodium dodecyl sulphate-polyacrylamide gel electrophoresis, although significant charge variations are evident by immunoelectrophoresis. 4. It is concluded that proteins antigenically related to B. asper myotoxins are relatively common in the genus Bothrops and, in the light of findings discussed, are likely to possess myotoxic activity.

Animals↗
PubMed1990

[The serum of newborn Clelia clelia (Serpentes: Colubridae) neutralizes the hemorrhagic action of Brothrops asper venom (Serpentes: Viperidae)].

The ability of serum from nine newborn specimens of Clelia clelia (Colubridae) to neutralize hemorrhagic action of Bothrops asper venom was tested. All serum samples neutralized completely the hemorrhagic effect of the venom in mice. This finding shows that the neutralizing ability of C. clelia serum towards the hemorrhagic activity of B. asper venom is innate.

Animals↗
PubMed1989

Histopathological and biochemical alterations induced by intramuscular injection of Bothrops asper (terciopelo) venom in mice.

The local and systemic pathological changes induced by an i.m. injection of 100 micrograms of Bothrops asper venom in mice were studied histologically and by following the changes in serum levels of enzymes, proteins, ATP and lactate, as well as alterations in hematocrit and clotting time. B. asper venom induced a rapid and marked increase in serum levels of creatine kinase, aspartate aminotransferase and lactate dehydrogenase, but not alanine aminotransferase or alkaline phosphatase. A local myonecrosis and hemorrhage was observed, with the lungs collapsing by 24 hr and the kidneys showing glomerular congestion and vacuolar degeneration of tubular cells. Only minor histopathological changes were observed in cardiac muscle and liver. Both ATP and lactate blood levels decreased after venom injection, whereas there were no changes in serum protein concentration. Blood incoagulability was observed 1 and 3 hr after envenomation. Antivenom neutralized venom-induced increases in serum enzyme levels following preincubation with venom, indicating that antivenom contains antibodies against tissue-damaging toxins. However, when antivenom was administered i.v. at different time intervals after venom injection, neutralization was only partial, with the exception of defibrinating activity, which was totally neutralized even after a delay of 1 hr in administering antivenom.

Adenosine Triphosphate↗
PubMed1989

A new muscle damaging toxin, myotoxin II, from the venom of the snake Bothrops asper (terciopelo).

A new muscle damaging toxin, myotoxin II, was purified from the venom of Bothrops asper by ion-exchange chromatography on CM-Sephadex C-25. The toxin is a dimeric, basic protein with a monomer mol.wt of 13,341, according to the amino acid composition, and 16,000 on the basis of SDS-polyacrylamide gel electrophoretic mobility. It has a high number of aspartate and lysine residues, as well as of hydrophobic amino acids. Upon i.m. injection into mice, the toxin induces myonecrosis and increase in serum creatine kinase levels. In addition, myotoxin II induces edema in the mouse foot pad. Immunochemical tests, mol.wt, and amino acid composition indicate a high degree of homology between myotoxin II and a previously characterized myotoxin from this venom, myotoxin I. However, in contrast to myotoxin I, myotoxin II lacks phospholipase A2 and anticoagulant activities in vitro.

Amino Acids↗
PubMed1989

Myonecrosis induced in mice by a basic myotoxin isolated from the venom of the snake Bothrops nummifer (jumping viper) from Costa Rica.

The mode of action of a basic myotoxin isolated from Bothrops nummifer venom was studied. This myotoxin is a basic polypeptide of 13,000 mol.wt, with a high content of lysine and aspartate, as well as of hydrophobic amino acids. It lacked phospholipase A2 activity when tested on several substrates at different pH values. Upon i.m. injection into mice, the toxin induced early morphological alterations typified by 'delta lesions' in the periphery of muscle fibers, an indication that the plasma membrane was the first cellular structure to be affected. Afterwards, necrotic cells had a clumped appearance, which then changed to a more hyaline histological pattern. Removal of necrotic material by phagocytes was followed by skeletal muscle regeneration, with the presence of myoblasts, myotubes and fully regenerated myofibers. The toxin induced a rapid and drastic drop in muscle creatine and creatine kinase contents of injected muscle, as well as an increase in serum levels of the enzymes lactic dehydrogenase and creatine kinase. Moreover, total muscle calcium increased significantly after toxin administration. Myotoxin induced a dose-dependent release of peroxidase entrapped in liposomes made from muscle phospholipids. The lack of phospholipase A2 activity in this toxin, together with the observation that it behaved as an amphiphilic protein in charge-shift electrophoresis, suggests that it might penetrate and disorganize muscle plasma membrane by means of a hydrophobic interaction.

Animals↗