RESUMEN
Two plasma kallikrein-kinin system inhibitors in the salivary glands of the kissing bug Triatoma infestans, designated triafestin-1 and triafestin-2, have been identified and characterized. Reconstitution experiments showed that triafestin-1 and triafestin-2 inhibit the activation of the kallikrein-kinin system by inhibiting the reciprocal activation of factor XII and prekallikrein, and subsequent release of bradykinin. Binding analyses showed that triafestin-1 and triafestin-2 specifically interact with factor XII and high molecular weight kininogen in a Zn2+-dependent manner, suggesting that they specifically recognize Zn2+-induced conformational changes in factor XII and high molecular weight kininogen. Triafestin-1 and triafestin-2 also inhibit factor XII and high molecular weight kininogen binding to negatively charged surfaces. Furthermore, they interact with both the N-terminus of factor XII and domain D5 of high molecular weight kininogen, which are the binding domains for biological activating surfaces. These results suggest that triafestin-1 and triafestin-2 inhibit activation of the kallikrein-kinin system by interfering with the association of factor XII and high molecular weight kininogen with biological activating surfaces, resulting in the inhibition of bradykinin release in an animal host during insect blood-feeding.
Asunto(s)
Proteínas de Insectos/genética , Sistema Calicreína-Quinina/efectos de los fármacos , Glándulas Salivales/metabolismo , Proteínas y Péptidos Salivales/genética , Triatoma/genética , Secuencia de Aminoácidos , Animales , Coagulación Sanguínea/efectos de los fármacos , Clonación Molecular , ADN Complementario/química , ADN Complementario/genética , Relación Dosis-Respuesta a Droga , Electroforesis en Gel de Poliacrilamida , Factor XII/antagonistas & inhibidores , Factor XII/química , Factor XII/metabolismo , Proteínas de Insectos/metabolismo , Proteínas de Insectos/farmacología , Cinética , Cininas/antagonistas & inhibidores , Cininas/sangre , Datos de Secuencia Molecular , Peso Molecular , Filogenia , Calicreína Plasmática/antagonistas & inhibidores , Precalicreína/antagonistas & inhibidores , Precalicreína/química , Precalicreína/metabolismo , Unión Proteica/efectos de los fármacos , Proteínas Recombinantes/química , Proteínas Recombinantes/farmacología , Proteínas y Péptidos Salivales/metabolismo , Proteínas y Péptidos Salivales/farmacología , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Triatoma/metabolismo , Tiempo de Coagulación de la Sangre Total , Zinc/farmacologíaRESUMEN
OBJECTIVE: To study the effect of compound Chinese drug Bailong on the transcription of Cyclin Dependent Kinase Inhibitor (CKI) p16INK4a, p21 and Rb, c-myc genes, and the relationship between gene expression and cAMP-PKA pathway. METHODS: Using the traditional molecular biology methods (cell synchronization, molecular hybridization--Western blotting, Northern blotting, etc.) examine the gene expression. RESULTS: Bailong promoted the expression (both mRNA and protein) of p16INK4a obviously in G1 phase cells. When prekallikrein (PKA) inhibitor was added in the cells which were treated by Bailong, the mRNA and protein level of p16INK4a decreased. It was shown that the inhibited proliferation of BGC82-3 cell by Bailong may come from the enhanced p16INK4a gene expression in G1 phase. Being same as p16INK4a, tumor suppressor genes Rb, p21 and oncogene c-myc expression were all affected by Bailong. When PKA inhibitor was added, the results were reversed. CONCLUSION: Bailong can affect many anticancer genes (including p16INK4a, p21 and Rb genes) and oncogenes (including c-myc) transcription by regulating cAMP-PKA pathway.
Asunto(s)
Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Medicamentos Herbarios Chinos/farmacología , Genes Supresores de Tumor , Precalicreína/antagonistas & inhibidores , Transducción de Señal , Neoplasias Gástricas/genética , Genes de Retinoblastoma/genética , Genes myc , Genes p16 , Humanos , Interfase , Proteína Oncogénica p21(ras)/biosíntesis , Proteína Oncogénica p21(ras)/genética , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patología , Células Tumorales CultivadasRESUMEN
The 70% ethanol extract (KS-ext) from Kochiae Fructus (dried fruits of Kochia scoparia L.) was screened for its activity on nociceptive and inflammatory responses in experimental animals. Although KS-ext at an oral administration of 500 mg/kg had an antinociceptive effect on writhing responses induced by acetic acid, it was ineffective on nociceptive response in the hot plate test. Oleanolic acid oligoglycoside, momordin Ic isolated from Kochiae Fructus significantly decreased the frequency of licking behavior within a unit of time at the late phase without affecting that of the early phase in the formalin test. Also, KS-ext inhibited the rise of vascular permeability induced by acetic acid, the increase of paw edema induced by carrageenin, histamine, serotonin or bradykinin and ear swelling induced by arachidonic acid. Momordin Ic also exhibited an inhibitory effect on carrageenin-induced edema. These results indicated that Kochiae Fructus has a peripheral antinociceptive effect mediated by antiinflammatory action, and that its active component can be partially attributed to momordin Ic.