RESUMO
Preeclampsia, a pregnancy-specific syndrome characterized by hypertension, proteinuria and edema, is a major cause of fetal and maternal morbidity and mortality especially in developing countries. Bj-PRO-10c, a proline-rich peptide isolated from Bothrops jararaca venom, has been attributed with potent anti-hypertensive effects. Recently, we have shown that Bj-PRO-10c-induced anti-hypertensive actions involved NO production in spontaneous hypertensive rats. Using in vitro studies we now show that Bj-PRO-10c was able to increase NO production in human umbilical vein endothelial cells from hypertensive pregnant women (HUVEC-PE) to levels observed in HUVEC of normotensive women. Moreover, in the presence of the peptide, eNOS expression as well as argininosuccinate synthase activity, the key rate-limiting enzyme of the citrulline-NO cycle, were enhanced. In addition, excessive superoxide production due to NO deficiency, one of the major deleterious effects of the disease, was inhibited by Bj-PRO-10c. Bj-PRO-10c induced intracellular calcium fluxes in both, HUVEC-PE and HUVEC, which, however, led to activation of eNOS expression only in HUVEC-PE. Since Bj-PRO-10c promoted biological effects in HUVEC from patients suffering from the disorder and not in normotensive pregnant women, we hypothesize that Bj-PRO-10c induces its anti-hypertensive effect in mothers with preeclampsia. Such properties may initiate the development of novel therapeutics for treating preeclampsia.
Assuntos
Anti-Hipertensivos/farmacologia , Bothrops/metabolismo , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Oligopeptídeos/farmacologia , Sequência de Aminoácidos , Animais , Arginina/metabolismo , Argininossuccinato Sintase/metabolismo , Western Blotting , Cálcio/metabolismo , Células Cultivadas , Citrulina/metabolismo , Relação Dose-Resposta a Droga , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/fisiopatologia , Feminino , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Pré-Eclâmpsia/patologia , Pré-Eclâmpsia/fisiopatologia , Gravidez , Complicações Cardiovasculares na Gravidez/patologia , Complicações Cardiovasculares na Gravidez/fisiopatologia , Superóxidos/metabolismo , Venenos de Víboras/farmacologiaRESUMO
Baroreflex sensitivity is disturbed in many people with cardiovascular diseases such as hypertension. Brain deficiency of nitric oxide (NO), which is synthesized by NO synthase (NOS) in the citrulline-NO cycle (with argininosuccinate synthase (ASS) activity being the rate-limiting step), contributes to impaired baroreflex. We recently showed that a decapeptide isolated from Bothrops jararaca snake venom, denoted Bj-PRO-10c, exerts powerful and sustained antihypertensive activity. Bj-PRO-10c promoted vasodilatation dependent on the positive modulation of ASS activity and NO production in the endothelium, and also acted on the central nervous system, inducing the release of GABA and glutamate, two important neurotransmitters in the regulation of autonomic systems. We evaluated baroreflex function using the regression line obtained by the best-fit points of measured heart rate (HR) and mean arterial pressure (MAP) data from spontaneously hypertensive rats (SHRs) treated with Bj-PRO-10c. We also investigated molecular mechanisms involved in this effect, both in vitro and in vivo. Bj-PRO-10c mediated an increase in baroreflex sensitivity and a decrease in MAP and HR. The effects exerted by the peptide include an increase in the gene expression of endothelial NOS and ASS. Bj-PRO-10c-induced NO production depended on intracellular calcium fluxes and the activation of a G(i/o)-protein-coupled metabotropic receptor. Bj-PRO-10c induced NO production and the gene expression of ASS and endothelial NOS in the brains of SHRs, thereby improving baroreflex sensitivity. Bj-PRO-10c may reveal novel approaches for treating diseases with impaired baroreflex function.
Assuntos
Barorreflexo/efeitos dos fármacos , Pressão Sanguínea/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Venenos de Crotalídeos/farmacologia , Frequência Cardíaca/efeitos dos fármacos , Óxido Nítrico/biossíntese , Análise de Variância , Animais , Argininossuccinato Sintase/genética , Argininossuccinato Sintase/metabolismo , Barorreflexo/fisiologia , Pressão Sanguínea/fisiologia , Bothrops , Encéfalo/citologia , Células Cultivadas , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Expressão Gênica , Frequência Cardíaca/fisiologia , Masculino , Óxido Nítrico Sintase Tipo III/genética , Óxido Nítrico Sintase Tipo III/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Endogâmicos SHR , Ratos Wistar , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Angiotensin-converting enzyme (ACE) inhibitors were developed based on proline-rich oligopeptides found in the venom of Bothrops jararaca (Bj) previously known as bradykinin-potentiating peptides (BPPs). However, the exact mechanism of action of BPPs remains unclear. The role of the ACE in the cardiovascular effects of two of naturally proline-rich oligopeptides (Bj-BPP-7a and Bj-BPP-10c) was evaluated in vitro and in vivo. Bj-BPP-7a does not potentiate the cardiovascular response to bradykinin and is a weak inhibitor of ACE C and N sites (K(i) = 40,000 and 70,000 nM, respectively), whereas Bj-BPP-10c is a strong bradykinin potentiator and inhibitor of the ACE C site (K(i) = 0.5 versus 200 nM for N site). Strikingly, both peptides, in doses ranging from 0.47 to 71 nmol/kg, produced long-lasting reduction (>6 h) in the mean arterial pressure of conscious spontaneously hypertensive rats (maximal change, 45 +/- 6 and 53 +/- 6 mm Hg for Bj-BPP-7a and Bj-BPP-10c, respectively). The fall in blood pressure was accompanied by variable degrees of bradycardia. In keeping with the absence of relationship between ACE-inhibitory and antihypertensive activities, no changes in the pressor effect of angiotensin I or in the hypotensive effect of bradykinin were observed at the peak of the cardiovascular effects of both peptides. Our results indicate that the antihypertensive effect of two Bj-BPPs containing the motif Ile-Pro-Pro is unrelated to their ability for inhibiting ACE or potentiating bradykinin (BK), indicating as a major component ACE and BK-independent mechanisms. These results are in line with previous observations suggesting ACE inhibition-independent mechanisms for angiotensin I-converting enzyme inhibitor.
Assuntos
Inibidores da Enzima Conversora de Angiotensina/farmacologia , Sistema Cardiovascular/efeitos dos fármacos , Venenos de Crotalídeos/química , Oligopeptídeos/farmacologia , Peptidil Dipeptidase A/metabolismo , Angiotensina I/farmacologia , Inibidores da Enzima Conversora de Angiotensina/química , Inibidores da Enzima Conversora de Angiotensina/metabolismo , Animais , Anti-Hipertensivos/química , Anti-Hipertensivos/metabolismo , Anti-Hipertensivos/farmacologia , Pressão Sanguínea/efeitos dos fármacos , Bothrops , Bradicinina/farmacologia , Células CHO , Cricetinae , Cricetulus , Relação Dose-Resposta a Droga , Feminino , Cobaias , Frequência Cardíaca/efeitos dos fármacos , Humanos , Íleo/efeitos dos fármacos , Íleo/fisiologia , Técnicas In Vitro , Masculino , Contração Muscular/efeitos dos fármacos , Oligopeptídeos/química , Oligopeptídeos/metabolismo , Peptidil Dipeptidase A/genética , Ratos , Ratos Endogâmicos SHR , Ratos WistarRESUMO
Bradykinin potentiating peptides (BPPs) from Bothrops jararaca venom were first described in the middle of 1960s and were the first natural inhibitors of the angiotensin-converting enzyme (ACE). BPPs present a classical motif and can be recognized by their typical pyroglutamyl (Pyr)/proline rich sequences presenting, invariably, a proline residue at the C-terminus. In the present study, we describe the isolation and biological characterization of a novel BPP isolated from the skin secretion of the Brazilian tree-frog Phyllomedusa hypochondrialis. This new BPP, named Phypo Xa presents the sequence Pyr-Phe-Arg-Pro-Ser-Tyr-Gln-Ile-Pro-Pro and is able to potentiate bradykinin activities in vivo and in vitro, as well as efficiently and competitively inhibit ACE. This is the first canonical BPP (i.e. Pyr-Aaa(n)-Gln-Ile-Pro-Pro) to be found not only in the frog skin but also in any other natural source other than the snake venoms.
Assuntos
Anuros/metabolismo , Bradicinina/metabolismo , Oligopeptídeos/isolamento & purificação , Sequência de Aminoácidos , Inibidores da Enzima Conversora de Angiotensina/isolamento & purificação , Inibidores da Enzima Conversora de Angiotensina/metabolismo , Inibidores da Enzima Conversora de Angiotensina/farmacologia , Animais , Anuros/genética , Pressão Sanguínea/efeitos dos fármacos , Bradicinina/farmacologia , Sinergismo Farmacológico , Feminino , Cobaias , Íleo/efeitos dos fármacos , Técnicas In Vitro , Masculino , Oligopeptídeos/genética , Oligopeptídeos/metabolismo , Oligopeptídeos/farmacologia , Ratos , Ratos Wistar , Pele/metabolismoRESUMO
Hemorphins are biologically active peptides, derived from hemoglobin, which presents a number of physiological activities. Proteolytic generation of these peptides is not fully understood; however, among their roles, is to provoke reduction on blood pressure. In this work, this particular biological effect was chosen as the monitor for the selection of mammalian vasoactive peptides. By combining high-performance liquid chromatography and mass spectrometry, including 'de novo' sequencing, several hemorphin-like peptides were identified presenting bradykinin potentiating activity. Moreover, taking LVV-hemorphin-7 as model compound, we evaluated its biological effect on blood pressure of anaesthetized rats. By summarizing all the results, it is possible to present the hemorphins as a family of proteolytically generated peptides that are able to potentiate bradykinin activity in vivo.