RESUMO
Differences in botulinum neurotoxin manufacturing, formulation, and potency evaluation can impact dose and biological activity, which ultimately affect duration of action. The potency of different labeled vials of incobotulinumtoxinA (Xeomin®; 50 U, 100 U, or 200 U vials; incobotA) versus onabotulinumtoxinA (BOTOX®; 100 U vial; onabotA) were compared on a unit-to-unit basis to assess biological activity using in vitro (light-chain activity high-performance liquid chromatography (LCA-HPLC) and cell-based potency assay (CBPA)) and in vivo (rat compound muscle action potential (cMAP) and mouse digit abduction score (DAS)) assays. Using LCA-HPLC, incobotA units displayed approximately 54% of the protease activity of label-stated equivalent onabotA units. Lower potency, reflected by higher EC50, ID50, and ED50 values (pooled mean ± SEM), was displayed by incobotA compared to onabotA in the CBPA (EC50: incobotA 7.6 ± 0.7 U/mL; onabotA 5.9 ± 0.5 U/mL), cMAP (ID50: incobotA 0.078 ± 0.005 U/rat; onabotA 0.053 ± 0.004 U/rat), and DAS (ED50: incobotA 14.2 ± 0.5 U/kg; onabotA 8.7 ± 0.3 U/kg) assays. Lastly, in the DAS assay, onabotA had a longer duration of action compared to incobotA when dosed at label-stated equivalent units. In summary, onabotA consistently displayed greater biological activity than incobotA in two in vitro and two in vivo assays. Differences in the assay results do not support dose interchangeability between the two products.
Assuntos
Toxinas Botulínicas Tipo A/farmacologia , Músculo Esquelético/efeitos dos fármacos , Fármacos Neuromusculares/farmacologia , Neurônios/efeitos dos fármacos , Potenciais de Ação , Animais , Bioensaio , Toxinas Botulínicas Tipo A/toxicidade , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Rotulagem de Medicamentos , Feminino , Humanos , Camundongos , Músculo Esquelético/fisiopatologia , Fármacos Neuromusculares/toxicidade , Paralisia/induzido quimicamente , Paralisia/fisiopatologia , Ratos Sprague-DawleyRESUMO
Botulin toxin (BTX) is widely used for treating skeletal muscle spasticity. Experimental reports on BTX treatment were mainly focused on the neuromuscular junction, while relatively little is known about toxin effects on the muscle cell itself. We investigated possible impact of BTX type A on skeletal muscle cell transcriptome by microarray analysis in muscle-derived cell cultures (fibroblasts, myoblasts and myotubes) from controls and spastic patients, and results were then validated at transcript and protein level. BTX-A treatment of control cells induced major changes in the myogenic component of the transcriptome, whereas the same treatment had a negligible effect in the fibrogenic component. BTX-A treatment of cell cultures from spastic patients induced an increased number of genes differentially expressed both in the fibrogenic and myogenic components. Specifically, BTX-A had a major effect on cell cycle-related genes in myoblasts, on muscle contraction-related genes in myotubes, and on extracellular matrix-related genes in fibroblasts from spastic patients. Our findings show that in vitro BTX-A treatment differentially affects transcript expression in muscle cells from spastic patients compared to those from controls suggesting a direct effect of BTX-A on muscle-specific functional pathways.
Assuntos
Toxinas Botulínicas Tipo A/toxicidade , Fibras Musculares Esqueléticas/efeitos dos fármacos , Espasticidade Muscular/genética , Fármacos Neuromusculares/toxicidade , Músculo Quadríceps/efeitos dos fármacos , Transcriptoma/efeitos dos fármacos , Adolescente , Adulto , Biópsia , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Humanos , Pessoa de Meia-Idade , Fibras Musculares Esqueléticas/metabolismo , Espasticidade Muscular/metabolismo , Espasticidade Muscular/patologia , Mioblastos Esqueléticos/efeitos dos fármacos , Mioblastos Esqueléticos/metabolismo , Músculo Quadríceps/metabolismo , Músculo Quadríceps/patologia , Adulto JovemRESUMO
Transient muscle paralysis engendered by a single injection of botulinum toxin A (BTxA) rapidly induces profound focal bone resorption within the medullary cavity of adjacent bones. While initially conceived as a model of mechanical disuse, osteoclastic resorption in this model is disproportionately severe compared with the modest gait defect that is created. Preliminary studies of bone marrow following muscle paralysis suggested acute upregulation of inflammatory cytokines, including TNF-α and IL-1. We therefore hypothesized that BTxA-induced muscle paralysis would rapidly alter the inflammatory microenvironment and the osteoclastic potential of bone marrow. We tested this hypothesis by defining the time course of inflammatory cell infiltration, osteoinflammatory cytokine expression, and alteration in osteoclastogenic potential in the tibia bone marrow following transient muscle paralysis of the calf muscles. Our findings identified inflammatory cell infiltration within 24 h of muscle paralysis. By 72 h, osteoclast fusion and pro-osteoclastic inflammatory gene expression were upregulated in tibia bone marrow. These alterations coincided with bone marrow becoming permissive to the formation of osteoclasts of greater size and greater nuclei numbers. Taken together, our data are consistent with the thesis that transient calf muscle paralysis induces acute inflammation within the marrow of the adjacent tibia and that these alterations are temporally consistent with a role in mediating muscle paralysis-induced bone resorption.
Assuntos
Reabsorção Óssea/fisiopatologia , Inflamação/etiologia , Músculo Esquelético/efeitos dos fármacos , Osteoclastos/patologia , Paralisia/fisiopatologia , Animais , Medula Óssea/patologia , Reabsorção Óssea/etiologia , Toxinas Botulínicas Tipo A/toxicidade , Feminino , Inflamação/fisiopatologia , Camundongos , Camundongos Endogâmicos C57BL , Fármacos Neuromusculares/toxicidade , Paralisia/induzido quimicamente , Paralisia/imunologia , Linfócitos T/imunologiaRESUMO
INTRODUCTION: Botulinum neurotoxin A (BoNT/A) cleaves SNAP-25 and inhibits acetylcholine (ACh) release at the neuromuscular junctions (NMJ) to cause neuroparalysis. Previous reports indicate a dyssynchrony between the inhibitory effect of BoNT/A on ACh release and SNAP-25 cleavage. METHODS: We tested the in vitro (acute; 90 min) and in vivo (chronic; 12 h) effects of BoNT/A on stimulus-evoked ACh release (SEAR), twitch tension, and SNAP-25 cleavage in isolated extensor digitorum longus (EDL) nerve-muscle preparations (NMP). RESULTS: In vitro or in vivo BoNT/A poisoning inhibited SEAR and twitch tension. Conversely, SNAP-25 cleavage and inhibition of spontaneous release frequency were observed only in NMP poisoned with BoNT/A in vivo. Moreover, chronic treatment of BoNT/A inhibited ionomycin stimulated Ca(2+) signals in Neuro 2a cells. CONCLUSIONS: These results demonstrate that the inhibition of SEAR precedes SNAP-25 cleavage and suggest involvement of a more complex mechanism for the inhibitory effect of BoNT/A at the NMJ.
Assuntos
Toxinas Botulínicas Tipo A/toxicidade , Potencial Evocado Motor/efeitos dos fármacos , Fármacos Neuromusculares/toxicidade , Junção Neuromuscular/efeitos dos fármacos , 4-Aminopiridina/análogos & derivados , 4-Aminopiridina/farmacologia , Acetilcolina/metabolismo , Acetilcolinesterase/metabolismo , Amifampridina , Animais , Ionóforos de Cálcio/farmacologia , Linhagem Celular Tumoral , Estimulação Elétrica , Eletromiografia , Técnicas In Vitro , Ionomicina/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Neuroblastoma/patologia , Junção Neuromuscular/metabolismo , Bloqueadores dos Canais de Potássio/farmacologia , Reflexo/efeitos dos fármacos , Fatores de TempoRESUMO
The use of botulinum toxin A for the treatment of wrinkles is increasing. Botulinum toxin A inhibits exocytosis of acetylcholine from 3 to 12 months, depending on the target tissue. Low-dose botulinum toxin A is used to smooth hyperkinetic facial lines. This is especially successful in the upper facial parts, since the target muscles (procerus, corrugator supracilii, frontalis, orbicularis oculi) all directly overlie the osseous structures of the face. This is not the case for the lower facial parts, and more side effects are encountered when treating, for example, wrinkles around the mouth. Contraindications to the use of botulinum toxin A are diseases affecting neuromuscular signal transduction, allergic reactions to components of the solution, therapy with aminoglycosides or acetylsalicylic acid prior to treatment, infections in the planned treatment area, and pregnancy and lactation. Alternative and complementary treatments include erbium-YAG or CO2 laser, as well as augmentation and surgical plastic procedures.
Assuntos
Toxinas Botulínicas Tipo A/uso terapêutico , Face , Fármacos Neuromusculares/uso terapêutico , Envelhecimento da Pele/efeitos dos fármacos , Adulto , Animais , Toxinas Botulínicas Tipo A/administração & dosagem , Toxinas Botulínicas Tipo A/efeitos adversos , Toxinas Botulínicas Tipo A/toxicidade , Terapia Combinada , Contraindicações , Estética , Músculos Faciais , Feminino , Haplorrinos , Humanos , Injeções Intramusculares , Lactação , Terapia a Laser , Dose Letal Mediana , Pessoa de Meia-Idade , Fármacos Neuromusculares/administração & dosagem , Fármacos Neuromusculares/efeitos adversos , Fármacos Neuromusculares/toxicidade , Postura , Gravidez , Ritidoplastia/métodos , Cirurgia Plástica , Fatores de Tempo , Resultado do TratamentoRESUMO
Many medications are available today by prescription or in over-the-counter preparations. This article reviews the pharmacokinetics, mechanism of action, toxicity, clinical signs, and management procedures necessary for some oral medications. The medications reviewed include selective serotonin reuptake inhibitors, benzodiazepines, amphetamines or amphetamine like drugs, carprofen, cyclooxygenase-2 inhibitors, pseudoephedrine, calcium channel blockers, and baclofen.
Assuntos
Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/veterinária , Administração Oral , Animais , Anti-Inflamatórios não Esteroides/toxicidade , Benzodiazepinas/toxicidade , Bloqueadores dos Canais de Cálcio/toxicidade , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/etiologia , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/terapia , Fármacos Neuromusculares/toxicidade , Medicamentos sem Prescrição/toxicidade , Moduladores Seletivos de Receptor Estrogênico/toxicidadeRESUMO
Botulinum neurotoxin (BoNT) is known to produce cationic channels in artificial bilayers. This study examined ion channels formed by BoNT in native membranes from cultured PC12 cells under conditions approximating those thought to occur during toxin internalization. Membrane patches were excised from PC12 cells using patch electrodes and exposed to symmetrical solutions containing either 200 mM CsCl, RbCl or KCl. The patch pipettes also contained 1-5 microg/ml BoNT buffered to pH 5.3 while the bath solutions were buffered to pH 7.0. In the presence of toxin, bursts of ion channel openings were observed. These toxin-induced channels were most active with a negative voltage applied to the same side as the toxin (cis). The increased activity at negative voltages was due to an increase in mean open time of e-fold per 120 mV and a decrease in mean closed time between bursts of e-fold per 110 mV. The shorter mean closed time within a burst was independent of membrane voltage. While BoNT-induced ion channels started as a single conductance level of 27 pS (KCl), 34 pS (RbCl) or 46 pS (CsCl) they typically increased in roughly equal steps to five or more times the original channel conductance. These higher conductance BoNT 'channels' opened and closed synchronously and could be distinguished from superposition of multiple independent channels. Despite differences in putative transmembrane sequences between BoNT/A and BoNT/E, both serotypes evidenced the same channel conductance and mean open time.
Assuntos
Toxinas Botulínicas Tipo A/toxicidade , Toxinas Botulínicas/toxicidade , Membrana Celular/efeitos dos fármacos , Canais Iônicos/efeitos dos fármacos , Fármacos Neuromusculares/toxicidade , Sequência de Aminoácidos , Animais , Toxinas Botulínicas/química , Toxinas Botulínicas Tipo A/química , Césio/farmacologia , Cloretos/farmacologia , Dados de Sequência Molecular , Células PC12 , Técnicas de Patch-Clamp , Cloreto de Potássio/farmacologia , Ratos , Rubídio/farmacologiaRESUMO
In hippocampal neurons, neurotransmitter release can be regulated by protein kinase A (PKA) through a direct action on the secretory machinery. To identify the site of PKA modulation, we have taken advantage of the ability of the neurotoxin Botulinum A to cleave the synaptic protein SNAP-25. Cleavage of this protein decreases the Ca2+ responsiveness of the secretory machinery by partially uncoupling Ca2+-sensing from fusion per se. This is expressed as a shift toward higher Ca2+ levels of the Ca2+ to neurotransmitter release relationship and as a perturbation of synaptic delay under conditions where secretion induced by the Ca2+-independent secretagogue ruthenium red is unimpaired. We find that SNAP-25 cleavage also perturbs PKA-dependent modulation of secretion; facilitation of ruthenium red-evoked neurotransmitter release by the adenylyl cyclase activator forskolin is blocked completely after Botulinum toxin A action. Together with our observation that forskolin modifies the Ca2+ to neurotransmitter release relationship, our results suggest that SNAP-25 acts as a functional linker between Ca2+ detection and fusion and that PKA modulates an early step in the secretory machinery related to calcium sensing to facilitate synaptic transmission.