RESUMEN
Plathymenia reticulata Benth has an anti-inflammatory effect and is capable of neutralizing the neuromuscular blockade induced by Bothrops jararacussu or Crotalus durissus terrificus venoms, probably by precipitating venom proteins (an effect caused by plant tannins). The present study aimed to evaluate the mutagenic activity of P. reticulata by using the Salmonella mutagenicity assay (Ames test) and the micronucleus test in CHO-K1 cells. P. reticulata extract concentrations of 2.84, 5.68, 11.37, and 19.90 mg/plate were assayed by the Ames test using TA97a, TA98, TA100 and TA102 bacterial strains, with (+S9) and without (-S9) metabolic activation. Concentrations of 5, 1.6 and 0.5 μg/mL of P. reticulata extract were used for the micronucleus test. P. reticulata extract was mutagenic to TA98 (-S9) and showed signs of mutagenic activity in TA97a and TA102 (both -S9) strains. Micronucleus test CBPI values showed that the endogenous metabolic system increased the number of viable cells when compared to the non-activated samples and the micronucleus frequency increased when the cells were treated in the absence of S9. We concluded that P. reticulata extract may present direct mutagenic properties.(AU)
Asunto(s)
Humanos , Pruebas de Mutagenicidad/métodos , Extractos Vegetales/uso terapéutico , Antiinflamatorios/farmacologíaRESUMEN
Plathymenia reticulata Benth has an anti-inflammatory effect and is capable of neutralizing the neuromuscular blockade induced by Bothrops jararacussu or Crotalus durissus terrificus venoms, probably by precipitating venom proteins (an effect caused by plant tannins). The present study aimed to evaluate the mutagenic activity of P. reticulata by using the Salmonella mutagenicity assay (Ames test) and the micronucleus test in CHO-K1 cells. P. reticulata extract concentrations of 2.84, 5.68, 11.37, and 19.90 mg/plate were assayed by the Ames test using TA97a, TA98, TA100 and TA102 bacterial strains, with (+S9) and without (-S9) metabolic activation. Concentrations of 5, 1.6 and 0.5 ìg/mL of P. reticulata extract were used for the micronucleus test. P. reticulata extract was mutagenic to TA98 (-S9) and showed signs of mutagenic activity in TA97a and TA102 (both -S9) strains. Micronucleus test CBPI values showed that the endogenous metabolic system increased the number of viable cells when compared to the non-activated samples and the micronucleus frequency increased when the cells were treated in the absence of S9. We concluded that P. reticulata extract may present direct mutagenic properties.
Asunto(s)
Antiinflamatorios , Bothrops , Venenos de Crotálidos , Crotalus cascavella , Solución Hidroalcohólica , Bloqueantes Neuromusculares , Plantas Medicinales , Pruebas de Mutagenicidad/métodosRESUMEN
We have previously isolated a Lys49 phospholipase A(2) homolog (BaTX) from Bothrops alternatus snake venom using a combination of molecular exclusion chromatography and reverse phase HPLC and shown its ability to cause neuromuscular blockade. In this work, we describe a one-step procedure for the purification of this toxin and provide further details of its neuromuscular activity. The toxin was purified by reverse phase HPLC and its purity and molecular mass were confirmed by SDS-PAGE, MALDI-TOF mass spectrometry, amino acid analysis and N-terminal sequencing. BaTX (0.007-1.4 microM) produced time-dependent, irreversible neuromuscular blockade in isolated mouse phrenic nerve-diaphragm and chick biventer cervicis preparations (time to 50% blockade with 0.35 microM toxin: 58+/-4 and 24+/-1 min, respectively; n=3-8; mean+/-S.E.) without significantly affecting the response to direct muscle stimulation. In chick preparations, contractures to exogenous acetylcholine (55 and 110 microM) or KCl (13.4 mM) were unaltered after complete blockade by all toxin concentrations. These results, which strongly suggested a presynaptic mechanism of action for this toxin, were reinforced by (1) the inability of BaTX to interfere with the carbachol-induced depolarization of the resting membrane, (2) a significant decrease in the frequency and amplitude of miniature end-plate potentials, and (3) a significant reduction (59+/-4%, n=12) in the quantal content of the end-plate potentials after a 60 min incubation with the toxin (1.4 microM). In addition, a decrease in the organ bath temperature from 37 degrees C to 24 degrees C and/or the replacement of calcium with strontium prevented the neuromuscular blockade, indicating a temperature-dependent effect possibly mediated by enzymatic activity.
Asunto(s)
Bothrops , Venenos de Crotálidos/enzimología , Bloqueantes Neuromusculares/farmacología , Unión Neuromuscular/efectos de los fármacos , Fosfolipasas A2/farmacología , Terminales Presinápticos/efectos de los fármacos , Animales , Calcio/química , Embrión de Pollo , Agonistas Colinérgicos/farmacología , Cromatografía Líquida de Alta Presión , Venenos de Crotálidos/química , Venenos de Crotálidos/aislamiento & purificación , Venenos de Crotálidos/farmacología , Diafragma/efectos de los fármacos , Diafragma/inervación , Relación Dosis-Respuesta a Droga , Estimulación Eléctrica , Electroforesis en Gel de Poliacrilamida , Masculino , Ratones , Potenciales Postsinápticos Miniatura , Peso Molecular , Bloqueantes Neuromusculares/química , Bloqueantes Neuromusculares/aislamiento & purificación , Fosfolipasas A2/química , Fosfolipasas A2/aislamiento & purificación , Nervio Frénico/efectos de los fármacos , Análisis de Secuencia de Proteína , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Transmisión Sináptica/efectos de los fármacos , Temperatura , Factores de TiempoRESUMEN
Ethanolic extract of leaves of Galactia glauscescens (GGE) at concentration of 100 and 500 microg/ml prevented the neuromuscular paralysis induced by Crotalus durissus terrificus venom on mouse phrenic nerve-diaphragm preparation.
Asunto(s)
Venenos de Crotálidos/toxicidad , Crotalus/fisiología , Fabaceae/química , Extractos Vegetales/farmacología , Animales , Diafragma/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Masculino , Ratones , Bloqueo Neuromuscular , Unión Neuromuscular/efectos de los fármacos , Nervio Frénico/efectos de los fármacos , Extractos Vegetales/química , Hojas de la Planta/químicaRESUMEN
The pharmacological effects of Bothrops neuwiedi pauloensis venom on mouse phrenic nerve-diaphragm (PND) preparations were studied. Venom (20 mug/ml) irreversibly inhibited indirectly evoked twitches in PND preparations (60 ± 10 percent inhibition, mean ± SEM; p<0.05; n=6). At 50 mug/ml, the venom blocked indirectly and directly (curarized preparations) evoked twitches in mouse hemidiaphragms. In the absence of Ca2+, venom (50 mug/ml), produced partial blockade only after an 80 min incubation, which reached 40.3 ± 7.8 percent (p<0.05; n=3) after 120 min. Venom (20 mug/ml) increased (25 ± 2 percent, p< 0.05) the frequency of giant miniature end-plate potentials in 9 of 10 end-plates after 30 min and the number of miniature end-plate potentials which was maximum (562 ± 3 percent, p<0.05) after 120 min. During the same period, the resting membrane potential decreased from - 81 ± 1.4 mV to - 41.3 ± 3.6 mV 24 fibers; p<0.01; n=4) in the end-plate region and from - 77.4 ± 1.4 to -44.6 ± 3.9 mV (24 fibers; p<0.01; n=4) in regions distant from the end-plate. These results indicate that B. n. pauloensis venom acts primarily at presynaptic sites. They also suggest that enzymatic activity may be involved in this pharmacological action.(AU)