RESUMEN
Novokinin (Arg-Pro-Leu-Lys-Pro-Trp) is a potent hypotensive peptide previously designed based on the structure of ovokinin(2-7) (Arg-Ala-Asp-His-Pro-Phe), a vasorelaxing and hypotensive peptide derived from ovalbumin. Novokinin exhibited an affinity for the angiotensin AT(2) receptor (Ki=7.35 microM). Novokinin significantly lowered systolic blood pressure at a dose of 0.03 and 0.1 mg/kg after intravenous and oral administration, respectively, in spontaneously hypertensive rats (SHRs), and the hypotensive activity was blocked by PD123319, an antagonist of the AT(2) receptor. Novokinin lowered blood pressure in C57BL/6J mice after oral administration at a dose of 50 mg/kg. However, in AT(2) receptor-deficient mice, novokinin did not reduce blood pressure. These results demonstrate that the hypotensive activity of novokinin is mediated by the AT(2) receptor. The hypotensive activity of novokinin in SHRs was completely blocked by indomethacin and CAY10441, an inhibitor of cyclooxygenase and an antagonist of the prostaglandin IP receptor, respectively. These suggest that the hypotensive activity is mediated by prostacyclin and the IP receptor downstream of the AT(2) receptor.
Asunto(s)
Antihipertensivos/farmacología , Oligopéptidos/farmacología , Receptor de Angiotensina Tipo 2/metabolismo , Receptores de Prostaglandina/metabolismo , Administración Oral , Bloqueadores del Receptor Tipo 2 de Angiotensina II , Animales , Antihipertensivos/administración & dosificación , Antihipertensivos/metabolismo , Presión Sanguínea/efectos de los fármacos , Proteínas del Huevo/química , Proteínas del Huevo/metabolismo , Imidazoles/administración & dosificación , Imidazoles/farmacología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Oligopéptidos/administración & dosificación , Oligopéptidos/metabolismo , Fragmentos de Péptidos/química , Fragmentos de Péptidos/metabolismo , Piridinas/administración & dosificación , Piridinas/farmacología , Ratas , Ratas Endogámicas SHR , Ratas Endogámicas WKY , Receptor de Angiotensina Tipo 2/genéticaRESUMEN
Novokinin (Arg-Pro-Leu-Lys-Pro-Trp), which has been designed based on the structure of ovokinin (2-7), significantly reduces the systolic blood pressure at a dose of 100 microg/kg after oral administration in spontaneously hypertensive rats (SHRs). In this study, we generated a transgenic soybean which accumulates novokinin. A vector encoding a modified beta-conglycinin alpha' subunit (4novokinin-alpha') in which four novokinin sequences have been incorporated by site-directed mutagenesis was introduced into somatic embryos by whisker-mediated gene transformation to produce a transgenic soybean. The 4novokinin-alpha' occupied 0.5% of total soluble protein and 5% of the beta-conglycinin alpha' subunit in the transgenic soybean seeds. Protein extracted from the transgenic soybean reduced systolic blood pressure after single oral administration in SHRs at a dose of 0.15 g/kg. Defatted flour from the transgenic soybean also reduced the systolic blood pressure at a dose of 0.25 g/kg. Thus, the 4novokinin-alpha' produced in soybean exhibited an anti-hypertensive activity in SHRs after oral administration.
Asunto(s)
Glycine max/metabolismo , Oligopéptidos/metabolismo , Plantas Modificadas Genéticamente/metabolismo , Semillas/metabolismo , Animales , Antihipertensivos/aislamiento & purificación , Antihipertensivos/metabolismo , Antihipertensivos/farmacología , Presión Sanguínea/efectos de los fármacos , Southern Blotting , Immunoblotting , Masculino , Oligopéptidos/aislamiento & purificación , Oligopéptidos/farmacología , Plantas Modificadas Genéticamente/genética , Regiones Promotoras Genéticas/genética , Ratas , Ratas Endogámicas SHR , Semillas/química , Semillas/genética , Proteínas de Soja/aislamiento & purificación , Proteínas de Soja/metabolismo , Proteínas de Soja/farmacología , Glycine max/genéticaRESUMEN
In this study, we found that novokinin (Arg-Pro-Leu-Lys-Pro-Trp), a potent hypotensive peptide acting through the AT(2) receptor, has vasorelaxing activity in the mesenteric artery isolated from spontaneously hypertensive rats. The vasorelaxing activity was significantly blocked by PD123319, indomethacin, and CAY10441, which are an AT(2) receptor antagonist, a cyclooxygenase inhibitor, and an IP receptor antagonist, respectively. N(G)-nitro-L-arginine methyl ester, an inhibitor of nitric oxide synthase, did not block the vasorelaxing activity. These results suggest that the vasorelaxing activity of novokinin, which contributes to the hypotensive effect, is mainly mediated by prostaglandin I(2) (prostacyclin) and the IP receptor downstream of the AT(2) receptor.
Asunto(s)
Arterias Mesentéricas/fisiología , Oligopéptidos/farmacología , Vasodilatación/efectos de los fármacos , Animales , Antihipertensivos/farmacología , Arterias Mesentéricas/efectos de los fármacos , NG-Nitroarginina Metil Éster/farmacología , Ratas , Ratas Endogámicas SHRRESUMEN
Four potent angiotensin converting enzyme (ACE) inhibitory peptides, IY, RIY, VW and VWIS, were isolated from subtilisin digest of rapeseed protein. Among them RIY and VWIS are new peptides with IC(50) 28 and 30 microM, respectively. All isolated peptides lowered blood pressure of spontaneously hypertensive rats (SHR) following oral administration. The maximum effect in the case of RIY was observed 4h after administration, while maximum effect of other peptides on blood pressure occurred 2h after administration. Furthermore, the antihypertensive effect of RIY was observed even in old rats, in which ACE inhibitors become less effective, suggesting that a different mechanism other than ACE inhibition is also involved in lowering blood pressure by this peptide. Subtilisin digest of rapeseed protein also significantly lowered blood pressure of SHR after oral administration of a single dosage 0.15 g/kg, exerting maximum antihypertensive effect 4h after administration. This digest appears promising as a functional food, which may be useful in the prevention and treatment of hypertension.
Asunto(s)
Antihipertensivos/aislamiento & purificación , Antihipertensivos/farmacología , Brassica rapa/química , Péptidos/aislamiento & purificación , Péptidos/farmacología , Administración Oral , Inhibidores de la Enzima Convertidora de Angiotensina/aislamiento & purificación , Animales , Antihipertensivos/administración & dosificación , Presión Sanguínea/efectos de los fármacos , Bovinos , Relación Dosis-Respuesta a Droga , Concentración 50 Inhibidora , Fragmentos de Péptidos/química , Fragmentos de Péptidos/aislamiento & purificación , Péptidos/administración & dosificación , Ratas , Ratas Endogámicas SHR , Porcinos , Factores de TiempoRESUMEN
Four new inhibitory peptides for angiotensin I-converting enzyme (ACE), that is, MRWRD, MRW, LRIPVA, and IAYKPAG, were isolated from the pepsin-pancreatin digest of spinach Rubisco with the use of HPLC. IC(50) values of individual peptides were 2.1, 0.6, 0.38, and 4.2 microM, respectively. MRW and MRWRD had an antihypertensive effect after oral administration to spontaneously hypertensive rats. Maximal reduction occurred 2 h after oral administration of MRW, whereas MRWRD showed maximal decrease 4 h after oral administration at doses of 20 and 30 mg/kg, respectively. IAYKPAG also exerted antihypertensive activity after oral administration at the dose of 100 mg/kg, giving a maximum decrease 4 h after oral administration. IAYKP, IAY, and KP, the fragment peptides of IAYKPAG, also exerted antihypertensive activity. LRIPVA [corrected] did not show any antihypertensive effect at a dose of 100 mg/kg despite its potent ACE-inhibitory activity.