RESUMO
OBJECTIVES AND DESIGN: The effects of the endogenous antioxidant alpha-lipoic acid on guinea pig colon smooth muscle contraction (Gpcc) induced by hydrogen peroxide were examined. Having previously shown that the histone deacetylase (HDAC) benzamide inhibitor MGCD0103 inhibits guinea-pig smooth muscle contraction, as do various sulfur-containing antioxidants, we asked whether hybrid compounds possessing both alpha-lipoic acid-derived antioxidant properties and HDAC inhibitory activity could inhibit Gpcc. MATERIALS AND METHODS: Guinea pig colon (Gpc) was incubated at 37 degrees C with Krebs buffer; the four stimulants-hydrogen peroxide, carbachol, histamine, and sodium fluoride-were added independently. The response to each stimulant alone was compared with that in the presence of each of the test compounds: MGCD0103, alpha-lipoic acid, and two of their hybrids, UCL M084 and UCL M109. RESULTS: NaF (10 mM), carbachol (0.05 microM), histamine (0.1 microM), and hydrogen peroxide (1 microM) produced Gpcc of about 50-60% above basal level. With the exception of MGCD0103 against hydrogen peroxide, all four test compounds at 1 microM-MGCD0103, alpha-lipoic acid, UCL M084, and UCL M109-produced a significant inhibition of 35-60% of Gpcc induced by hydrogen peroxide, NaF, and carbachol, although none reduced histamine or ovalbumin-induced Gpcc. Benzalkonium chloride (Bcl), a G-protein inhibitor, reduced the hydrogen peroxide-induced Gpcc by 35%. CONCLUSIONS: Contraction by stimulants used to induce Gpcc is known to involve G-proteins. All four test compounds-MGCD0103, alpha-lipoic acid and two of their hybrids, UCL M084 and UCL M109-reduced Gpcc induced by NaF and carbachol, suggesting that G-protein pathway involvement is relevant to the action of the test compounds, as is also indicated by the Bcl-induced inhibition of hydrogen peroxide-induced contractions. Additionally, alpha-lipoic acid and the two hybrids showed >30% inhibition of hydrogen peroxide-induced contractions, consistent with the antioxidant properties of the 1,2-dithiolane ring.
Assuntos
Antioxidantes/farmacologia , Colo/efeitos dos fármacos , Peróxido de Hidrogênio/farmacologia , Músculo Liso/efeitos dos fármacos , Oxidantes/farmacologia , Animais , Benzamidas/farmacologia , Carbacol/farmacologia , Proteínas de Ligação ao GTP/fisiologia , Cobaias , Histamina/farmacologia , Técnicas In Vitro , Agonistas Muscarínicos/farmacologia , Contração Muscular/efeitos dos fármacos , Pirimidinas/farmacologia , Receptores Histamínicos H1/efeitos dos fármacos , Receptores Histamínicos H1/metabolismo , Fluoreto de Sódio/antagonistas & inibidores , Fluoreto de Sódio/farmacologia , Ácido Tióctico/farmacologiaRESUMO
OBJECTIVES AND DESIGN: The aim of this study was to investigate the ability of (a) antioxidants, some related to alpha-lipoic acid (LA), (b) histone deacetylase (HDAC) inhibitors, and (c) hybrid compounds possessing both alpha-lipoic acid-derived antioxidant properties and HDAC inhibitory activity to inhibit guinea pig smooth muscle contraction. MATERIALS AND METHODS: Guinea pig isolated tracheal rings (GPTR) were prepared and their isometric tension measured using a transducer. Histamine, carbachol and 5-hydroxytryptamine (5-HT) served as agonists. Tests with antigen (ovalbumin) used GPTR from sensitised guinea pigs or rings from non-sensitised animals that had been incubated for at least 2 h with diluted serum from sensitised animals. RESULTS: All antioxidants tested showed a relaxant effect on resting tension and on tension induced by histamine or carbachol, with EC(50)(s) of 0.2-5.0 mM and a rank order of potency: LA derivatives > glutathione (GSH) > ascorbic acid (AA). However, low concentrations (<50 microM) of GSH, AA and LA potentiated histamine-induced contractions. The benzamide HDAC inhibitor MGCD0103 inhibited mast cell activation and GPTR contraction produced by antigen and certain agonists, although a 2-6 h pre-incubation was required for those effects to be apparent. Two LA-benzamide HDAC hybrid compounds, UCL M084 and UCL M109 inhibited GPTR contraction after 30 min pre-incubation; however, even after long pre-incubation (up to 6 h) those hybrid compounds showed less potent inhibition of agonist-induced contraction than did MGCD0103. CONCLUSIONS: The results showed that GSH more potently inhibited contraction induced by histamine than that induced by 5-HT or carbachol, whereas LA, and especially UCL M084 and UCL M109, more potently blocked contraction induced by carbachol and 5-HT than that induced by histamine. For GSH, and possibly also for LA-type compounds, the inhibition of agonist-induced tracheal smooth muscle contractions may be due to NO formation. This study did not detect a synergistic relaxant effect in two compounds incorporating the structural union of a benzamide HDAC inhibitor terminus with a LA-derived antioxidant moiety.
Assuntos
Antioxidantes/farmacologia , Músculo Liso/efeitos dos fármacos , Músculos Respiratórios/efeitos dos fármacos , Ácido Tióctico/análogos & derivados , Ácido Tióctico/farmacologia , Traqueia/efeitos dos fármacos , Animais , Ácido Ascórbico/farmacologia , Benzamidas/farmacologia , Carbacol/farmacologia , Sinergismo Farmacológico , Glutationa/farmacologia , Cobaias , Histamina/farmacologia , Antagonistas dos Receptores Histamínicos/farmacologia , Técnicas In Vitro , Contração Isométrica , Agonistas Muscarínicos/farmacologia , Contração Muscular/efeitos dos fármacos , Ovalbumina/farmacologia , Pirimidinas/farmacologia , Coelhos , Serotonina/farmacologiaAssuntos
Colo , Inibidores Enzimáticos/farmacologia , Liberação de Histamina/efeitos dos fármacos , Inibidores de Histona Desacetilases , Mastócitos , Contração Muscular , Animais , Colo/anatomia & histologia , Colo/efeitos dos fármacos , Colo/fisiologia , Feminino , Cobaias , Masculino , Mastócitos/efeitos dos fármacos , Mastócitos/metabolismo , Contração Muscular/efeitos dos fármacos , Contração Muscular/fisiologia , Ratos , Ratos Sprague-DawleyAssuntos
Antígenos/imunologia , Inibidores Enzimáticos/farmacologia , Inibidores de Histona Desacetilases , Contração Muscular , Traqueia , Animais , Antígenos/farmacologia , Inibidores Enzimáticos/química , Humanos , Camundongos , Estrutura Molecular , Contração Muscular/efeitos dos fármacos , Contração Muscular/fisiologia , Traqueia/anatomia & histologia , Traqueia/efeitos dos fármacos , Traqueia/fisiologiaAssuntos
Complemento C5a/metabolismo , Depsipeptídeos/farmacologia , Proteínas de Ligação ao GTP/metabolismo , Mastócitos/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Peptídeos/farmacologia , Animais , Depsipeptídeos/química , Cobaias , Humanos , Mastócitos/metabolismo , Estrutura Molecular , Contração Muscular/efeitos dos fármacos , Contração Muscular/fisiologia , Músculo Liso/fisiologia , Peptídeos/química , RatosAssuntos
Anti-Inflamatórios não Esteroides/farmacologia , Fármacos Gastrointestinais/farmacologia , Liberação de Histamina/efeitos dos fármacos , Mastócitos/efeitos dos fármacos , Mesalamina/farmacologia , Fenil-Hidrazinas/farmacologia , Sulfassalazina/farmacologia , Animais , Anti-Inflamatórios não Esteroides/uso terapêutico , Linhagem Celular , Fármacos Gastrointestinais/uso terapêutico , Humanos , Peróxido de Hidrogênio/metabolismo , Doenças Inflamatórias Intestinais/tratamento farmacológico , Mastócitos/metabolismo , Mesalamina/uso terapêutico , Oxidantes/metabolismo , Fenil-Hidrazinas/uso terapêutico , Ratos , Sulfassalazina/uso terapêuticoAssuntos
Inibidores Enzimáticos/farmacologia , Genisteína/farmacologia , Contração Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Animais , Canais de Cálcio Tipo L/efeitos dos fármacos , Canais de Cálcio Tipo L/fisiologia , Genisteína/agonistas , Cobaias , Técnicas In Vitro , Masculino , Contração Muscular/fisiologia , Músculo Liso/fisiologia , Proteína Quinase C/efeitos dos fármacos , Proteína Quinase C/metabolismo , Fluoreto de Sódio/farmacologia , Traqueia/efeitos dos fármacos , Traqueia/fisiologiaAssuntos
Contração Muscular/fisiologia , Proteínas Tirosina Quinases/metabolismo , Receptores Ativados por Proteinase/fisiologia , Animais , Carbacol/farmacologia , Agonistas Colinérgicos/farmacologia , Íleo/efeitos dos fármacos , Íleo/fisiologia , Técnicas In Vitro , Masculino , Músculo Liso/fisiologia , Proteínas Tirosina Quinases/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Receptores Ativados por Proteinase/efeitos dos fármacos , Inibidores de Serina Proteinase/farmacologia , Fluoreto de Sódio/farmacologia , Tosilfenilalanil Clorometil Cetona/farmacologiaAssuntos
Permeabilidade da Membrana Celular , Ciclosporina/farmacologia , Mastócitos/efeitos dos fármacos , Mastócitos/metabolismo , Animais , Permeabilidade da Membrana Celular/efeitos dos fármacos , Células Cultivadas , Humanos , Mastócitos/citologia , Ratos , Ratos Sprague-Dawley , beta-N-Acetil-Hexosaminidases/metabolismoAssuntos
Genisteína/farmacologia , Liberação de Histamina/efeitos dos fármacos , Íleo/efeitos dos fármacos , Mastócitos/efeitos dos fármacos , Mastócitos/metabolismo , Músculo Liso/efeitos dos fármacos , Peritônio/efeitos dos fármacos , Animais , Liberação de Histamina/imunologia , Íleo/fisiologia , Masculino , Mastócitos/imunologia , Contração Muscular/efeitos dos fármacos , Músculo Liso/fisiologia , Peritônio/citologia , Peritônio/imunologia , Ratos , Ratos Sprague-DawleyAssuntos
Íleo/fisiologia , Músculo Liso/fisiologia , Receptores Ativados por Proteinase/efeitos dos fármacos , Animais , Calcimicina/farmacologia , Quimotripsina/antagonistas & inibidores , Quimotripsina/farmacologia , Cromolina Sódica/farmacologia , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Guanidinas/farmacologia , Cobaias , Íleo/efeitos dos fármacos , Técnicas In Vitro , Ionóforos , Masculino , Contração Muscular/efeitos dos fármacos , Contração Muscular/fisiologia , Músculo Liso/efeitos dos fármacos , Óxido Nítrico Sintase/antagonistas & inibidores , Óxido Nítrico Sintase Tipo III , Inibidores de Serina Proteinase/farmacologia , Tosilfenilalanil Clorometil Cetona/farmacologiaAssuntos
Liberação de Histamina/efeitos dos fármacos , Íleo/efeitos dos fármacos , Mastócitos/metabolismo , Músculo Liso/efeitos dos fármacos , Peptídeos/farmacologia , Animais , Compostos de Benzalcônio/farmacologia , Bradicinina/antagonistas & inibidores , Bradicinina/farmacologia , Cátions , Calidina/antagonistas & inibidores , Calidina/farmacologia , Masculino , Mastócitos/efeitos dos fármacos , Contração Muscular/efeitos dos fármacos , Peptídeos/antagonistas & inibidores , Cavidade Peritoneal/citologia , Ratos , Ratos Wistar , Substância P/antagonistas & inibidores , Substância P/farmacologiaRESUMO
We have studied the effect of nitric oxide (NO) and hydrogen peroxide (H(2)O(2)), two reactive oxygen species (ROS) on histamine release (HR) from RBL-2H3 cells, a rat mucosal-type mast cell line. Marked HR was elicited by antigen (DNP-HSA), calcium ionophore A23187, sodium fluoride or phospholipase C, but not with compound 48/80 or 1,2-dioctanoyl-sn-glycerol. The NO-synthase substrate L-arginine and its inactive enantiomer (D-arginine), each on its own, induced a small but significant increase in HR above the basal level. However, the NO-donors (sodium nitroprusside or NaNO(3)) or the NO-synthase inducer lipopolysaccharide did not induce HR. Moreover, methylene blue (MB), which inhibits guanylate cyclase and N(omega)-nitro-L-arginine (L-NA), an inhibitor of NO synthase, were also without effect on either the basal HR or the L-arginine-induced HR. HR induced by A23187, DNP-HSA, sodium fluoride or phospholipase C was markedly reduced by MB, but mildly by L-NA (both at 1-100 microM). H(2)O(2) (0.01-1.0 mM) on its own did not induce HR, but it had a potent inhibitory effect on DNP-HSA- or A23187-induced HR, which was not reversed by L-NA (1-100 microM). Taken together, it seems that neither the stimulatory nor the inhibitory effects of the NO-related compounds on HR can be attributed to NO, but rather to other mechanisms. The inhibition of HR by H(2)O(2) also does not involve NO and suggests a negative feedback regulatory role for the peroxide in the allergic inflammation.
Assuntos
Histamina/metabolismo , Peróxido de Hidrogênio/farmacologia , Óxido Nítrico/farmacologia , Animais , Calcimicina/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Interações Medicamentosas , Inibidores Enzimáticos/farmacologia , Azul de Metileno/farmacologia , Nitroarginina/farmacologia , Ratos , Fluoreto de Sódio/farmacologiaRESUMO
Using the ratiometric Ca2+ indicator, indo-1, the antigen-induced increase in intracellular Ca2+ concentration ([Ca2+]i) was measured in individual RBL-2H3 cells which had been passively sensitized with monoclonal antibody to the dintrophenyl (DNP) haptenic group. Antigenic stimulation using DNP-human serum albumin conjugate (DNP-HSA) induced concentration-dependent asynchronous Ca2+ oscillations, or irregular spikes. To achieve a quantitative comparison of the effects of different concentrations of antigen on changes in Ca2+[i, the area under the curve (AUC) of Ca2+ oscillations in each cell was calculated. The dose-response curve of the calculated AUC is consistent with the bell-shaped dose-response curve for antigen-induced mediator release, depolarization and 86Rb(+)-efflux. Ca2+ oscillations induced by antigenic stimulation were abolished by removal of external Ca2+ and the subsequent reintroduction of external Ca2+ caused their resumption. To investigate the role of Ca2+ oscillations in the secretory response, changes in [Ca2+]i induced by concanavalin A (Con-A), A23187, thapsigargin and NECA were also monitored. Con-A mimicked the response induced by antigen, whilst A23187 and thapsigargin induced a large transient non-oscillatory response. NECA, an adenosine receptor agonist, induced only a small transient rise in Ca2+[i without oscillatory behaviour. Since all these stimuli accept NECA-induced degranulation in these cells, it is suggested that, although Ca2+ oscillations are not essential for the initiation of secretion, they probably underlie the in-vivo physiological response of mast cells and basophils to an antigenic challenge. They also seem to enhance the efficacy of the Ca2+ signal.