Enzyme Endopeptidase Research Articles

Pro-inflammatory Effect of Quercetin by Dual Blockade of Angiotensin Converting-enzyme and Neutral Endopeptidase In Vivo

The effect of the flavonoid quercetin on substance P- and bradykinin-induced plasma extravasation in rat tissues (duodenum, heart, pancreas, trachea and urinary bladder) was studied, and its modulation by endogenous peptidases. Plasma protein extravasation was assayed by extravasated Evans blue dye. Intravenous injection of substance P (1, 3 and 10 nmol/kg) increased the plasma extravasation in a dose-dependent manner in heart, pancreas, trachea and urinary bladder. Bradykinin (3 and 10 nmol/kg, i.v.) increased plasma extravasation in a dose-dependent manner in duodenum, pancreas, trachea and urinary bladder. Pre-treatment with a selected dose of quercetin potentiated the substance P-induced plasma extravasation in heart, pancreas and urinary bladder, and also the bradykinin-induced plasma extravasation in duodenum, heart, trachea and urinary bladder. The selective pharmacological inhibition of neutral endopeptidase and angiotensin-converting enzyme potentiated the substance P- and bradykinin-induced plasma extravasation, respectively; furthermore, treatment with receptor antagonists showed that the mediators involved in the potentiation of plasma extravasation by quercetin are substance P and bradykinin. Analysis of plasma angiotensin-converting enzyme activity demonstrated that quercetin inhibited this enzyme. These results suggest that quercetin potentiates plasma extravasation induced by substance P and bradykinin, and that this may result from inhibition of the degradative enzymes of these peptides.

Inhibition of both neutral endopeptidase and endothelin converting enzyme lowers pulmonary pressures in congestive heart failure

Inhibition of both neutral endopeptidase and endothelin converting enzyme lowers pulmonary pressures in congestive heart failure

First demonstration in humans of systemic neutral endopeptidase and endothelin converting enzyme inhibition using a new orally active dual metalloprotease inhibitor SLV 306

First demonstration in humans of systemic neutral endopeptidase and endothelin converting enzyme inhibition using a new orally active dual metalloprotease inhibitor SLV 306

Direct inhibition of neutral endopeptidase in vasopeptidase inhibitor-mediated amelioration of cardiac remodeling in rats with chronic heart failure.

Vasopeptidase inhibitors possess dual inhibitory actions on neutral endopeptidase (NEP) and angiotensin-converting enzyme (ACE) and have beneficial effects on cardiac remodeling. However, the contribution of NEP inhibition to their effects is not yet fully understood. To address the role of cardiac NEP inhibition in the anti-remodeling effects of a vasopeptidase inhibitor, we examined the effects of omapatrilat on the development of cardiac remodeling in rats with left coronary artery ligation (CAL) and those on collagen synthesis in cultured fibroblast cells. In vivo treatment with omapatrilat (30 mg/kg/day for 5 weeks) inhibited cardiac NEP activity in rats with CAL, which was associated with a suppression of both cardiac hypertrophy and collagen deposition. In cultured cardiac fibroblasts, omapatrilat (10(-7) to approximately 10(-5) M) inhibited NEP activity and augmented the ANP-induced decrease in [3H]-proline incorporation. ONO-BB, an active metabolite of the NEP selective inhibitor ONO-9902, also augmented the ANP-induced response, whereas captopril, an ACE inhibitor, did not. The angiotensin I-induced increase in [3H]-proline incorporation was prevented by omapatrilat and captopril, but not by ONO-BB. The results suggest that vasopeptidase inhibitor suppressed cardiac remodeling in the setting of chronic heart failure, possibly acting through the direct inhibition of cardiac NEP. Vasopeptidase inhibitors may have therapeutic advantages over the classical ACE and NEP inhibitors alone with respect to the regression of cardiac fibrosis.

Tubular urinary enzymes in acute post-infectious glomerulonephritis.

Tubular function of 17 pediatric patients with a mild form of acute post-infectious glomerulonephritis was prospectively evaluated by assessment of the urinary activity of proximal and distal tubule enzymes. Neutral-like endopeptidase (NEP-like) and angiotensin-converting enzyme (ACE) were the proximal tubule enzymes assessed, while prolyl-endopeptidase (PE) and serine-endopeptidase H1 and H2 were the distal tubule enzymes analyzed. Urine was collected at diagnosis (T0) and after 2 (T2) and 6 (T6) months of follow-up. NEP-like enzyme activity (nmol/mg creatinine; median+/-quartile range) was increased at diagnosis, and this remained stable during the first 6 months (T0 18.30+/-83.26, T2 17.32+/-49.56, T6 23.38+/-107.18). Urinary activity of the other enzymes was as follows: ACE (mU/ml per mg creatinine) T0 0.08+/-0.16, T2 0.06+/-0.10, T6 0.18+/-0.29; PE (nmol/mg creatinine) T0 6.70+/-84.87, T2 9.55+/-69.00, T6 13.67+/-28.70; serine-endopeptidase H1 (nmol/mg creatinine) T0 7.86+/-26.95, T2 17.17+/-59.37, T6 18.19+/- 79.14; and serine-thiol-endopeptidase H2 (nmol/mg creatinine) T0 3.06+/-21.97, T2 12.06+/-32.42, T6 16.22+/- 44.06. Thirty other healthy children matched for age and gender were considered as a control group. This group was assessed once and the results were: NEP-like activity 6.05+/-10.54, ACE 0.11+/-0.22, PE 7.10+/-13.36, H1 5.00+/-17.30, and H2 6.00+/-20.16. In conclusion, we observed that NEP-like and H1 enzymes exhibited significant increased urinary activity 6 months after the diagnosis. This increase occurred in spite of the disappearance of clinical symptoms, which occurred 2 months after the diagnosis. We believe that the increase in urinary enzymatic activity could be a manifestation of a silent tubular dysfunction following an episode of acute post-infectious glomerulonephritis.

Atheroma-inhibiting and vasoprotective properties of CO-targeting neutral endopeptidase and angiotensin converting enzyme

Atheroma-inhibiting and vasoprotective properties of CO-targeting neutral endopeptidase and angiotensin converting enzyme

Effects of age and gender on the pharmacodynamics of omapatrilat in healthy volunteers.

Omapatrilat is the most clinically advanced member of a new class of cardiovascular drugs, vasopeptidase inhibitors. Omapatrilat is a single molecule that simultaneously inhibits neutral endopeptidase and angiotensin-converting enzyme, thus preserving vasodilator peptides and inhibiting production of the vasoconstrictor angiotensin II. In healthy male volunteers, omapatrilat decreased blood pressure while being generally well tolerated, with no serious adverse events. This study was undertaken to determine the effect of age and gender on the pharmacodynamics of omapatrilat. Healthy male or female volunteers between the ages of 18 and 80 were given a single oral dose of omapatrilat 40 mg. Neither age nor gender affected the vasopeptidase inhibition by omapatrilat. There were no differences between subject groups in the effect of omapatrilat on supine systolic, diastolic, or mean arterial blood pressure. Based on this study of healthy subjects, it can be concluded that it is not necessary to adjust the initial dose of omapatrilat for the treatment of hypertension based solely on age or gender.

Retinoic acid treatment enhances the acetylcholine contents in the human teratocarcinoma cell line NTera-2

Human NTera-2/clone D1 teratocarcinoma cells are induced by retinoic acid (RA) to differentiate into postmitotic cells with morphological and biochemical characteristics of embryonic human neurones. Currently only limited information concerning peptide-contents and neurotransmitter pools of these cells is available. Zeller and Strauss [Int. J. Dev. Neurosci. 1995;13(5):437] described an increase in choline acetyltransferase (ChAT) activity in RA-treated, but not in untreated NTera-2 cells, suggesting the induction of a cholinergic phenotype during treatment with RA. In the present study we investigated the effect of RA-differentiation on the amount of the neurotransmitters acetylcholine (ACh), and dopamine in NTera-2 in order to specify the transmitter phenotype induced by RA-differentiation. We found that a 4-week treatment of NTera-2 cells with 10 μM RA markedly increased the ACh-content of these cells, while dopamine levels were unchanged. Depolarisation with potassium (60 mM) enhanced ACh-outflow in the differentiated cells in a Ca ++ dependent way. Also neuropeptides like substance P and NPY were detectable in the undifferentiated NTera-2 cells, while vasointestinal peptide (VIP) could not be found in either precursor or RA-differentiated cells. Differentiation was accompanied by a marked reduction of neutral endopeptidase enzyme activity and aminopeptidase activity. From these observations it was concluded that RA induces a cholinergic neurochemical differentiation of this human teratocarcinoma cell line, and that these cells might provide a model system to investigate cholinergic properties of human origin.

Synthesis, Characterization and Biodegradability of a Water-Soluble Poly(ethylene oxide) Derivative Polymer Bearing Carboxylic Acid Side Chain Function

A water-soluble poly(ethylene oxide) derivative polymer bearing carboxylic acid side chain function was synthesized and characterized by elemental analysis, nuclear magnetic resonance spectroscopy and Fourier transform infrared spectroscopy. Gel permeation chromatography (GPC) results indicated a narrow molecular weight distribution but could not give the absolute molecular weight. However, the electrospray mass spectrometry data indicated a molecular mass of 5500 for the polymer. Using GPC molecular weight distributions, we have been able to study the biodegradability of the polymer by papain and proteinase A in a buffer at pH 7.81 and at a temperature of 37°C. Both endopeptidase enzymes were active in the degradation of the polymer but at a very low rate. However, proteinase A showed continuous activity up to 48 hours, whereas the papain effect seemed to culminate after 24 hours.

Improvement by phosphoramidon of damaged endothelial function in porcine coronary artery

Background. The bradykinin (BK)-induced endothelium-dependent relaxation is impaired in the presence of elevated potassium concentration enhancing the vasospastic tendency of large coronary arteries. Inhibition of the angiotensin-converting enzyme responsible for bradykinin degradation was found to enhance the endothelium-dependent relaxation by BK. The aim of the present study was to investigate the effect of phosphoramidon, known to inhibit a BK-metabolizing neutral endopeptidase enzyme, on relaxation of porcine-isolated coronary artery in depolarizing solution. Methods. Endothelium intact porcine coronary artery rings were studied in organ chambers. The rings were isometrically contracted with potassium chloride (30 mmol/L) and the response to BK (1 to 1,000 nmol/L)-induced relaxation was investigated in the presence of nitric oxide synthase inhibitor N ω-nitro- L-arginine (300 μmol/L) alone and in combination with the cyclooxygenase inhibitor indomethacin (10 μmol/L), and that of the inhibitor of calcium-dependent potassium channels tetraethylammonium (7 mmol/L). Under these conditions, phosphoramidon (10 μmol/L), an inhibitor of a neutral endopeptidase enzyme (EC.3.4.24.11.), which is responsible for the degradation of BK, was used to enhance the endothelium-dependent relaxation. Results. Phosphoramidon potentiated the maximum vasorelaxant effect of BK in N ω-nitro- L-arginine (control 26.6% ± 10.86% versus phosphoramidon 49.05% ± 4.52%; n = 6, p < 0.05) or in N ω-nitro- L-arginine + indomethacin-pretreated rings (control 20.7% ± 9.92% versus phosphoramidon 42.0% ± 12.26%; n = 5, p < 0.05) and this increased vasodilation was not modified by tetraethylammonium. Conclusions. In the present study phosphoramidon potentiated the effect of BK in the absence of nitric oxide and prostaglandins in porcine-isolated coronary artery. This effect did not depend on tetraethylammonium-sensitive potassium channels. Phosphoramidon may be a useful pharmacologic tool for preserving the vasorelaxing capacity of coronary arteries after cardioplegia.

Combined neutral endopeptidase and angiotensin-converting enzyme inhibition attenuates left ventricular remodeling without hypotension in a rat coronary artery ligation model of heart failure

Combined neutral endopeptidase and angiotensin-converting enzyme inhibition attenuates left ventricular remodeling without hypotension in a rat coronary artery ligation model of heart failure

Role of neutral endopeptidase and angiotensin converting enzyme in asthma

Role of neutral endopeptidase and angiotensin converting enzyme in asthma

Neutral endopeptidase and angiotensin-converting enzyme inhibitors increase nitric oxide production in isolated canine coronary microvessels by a kinin-dependent mechanism.

Bradykinin is a substrate for both neutral endopeptidase 24.11 (NEP) and angiotensin-converting enzyme (ACE). Our previous studies showed that ACE inhibitors can stimulate nitric oxide production in coronary microvessels, which is mediated by local kinins. Whether inhibition of NEP also can affect local vascular NO production has not been established. To determine the role of NEP in the control of NO production, coronary microvessels were isolated from seven mongrel dogs. Two NEP inhibitors, phosphoramidon and thiorphan, and an ACE inhibitor, ramiprilat, were used. Nitrite, the metabolite of NO in aqueous solution, was measured by using the Griess reaction. Phosphoramidon and thiorphan (10(-6) M) increased nitrite production from 80 +/- 6 to 136 +/- 6 and 144 +/- 7 pmol/mg, respectively. Ramiprilat (10(-8) M) increased nitrite production from 78 +/- 6 to 155 +/- 7 pmol/mg wet weight. The effect of these agents on nitrite release was blocked by L-NAME, which inhibits NO synthase, HOE-140, which blocks bradykinin B2-receptor, and dichloroisocoumarin, which blocks kinin-forming enzymes. These results clearly indicate that inhibition of kinin metabolism by using neutral endopeptidase inhibitors increases NO production from coronary microvessels. Thus neutral endopeptidase plays an important role in local kinin-modulated NO production in the coronary microcirculation and NEP inhibitors may be useful clinical tools in treatment of cardiovascular disease.

Neutral endopeptidase modulates VIP-induced vasodilation in hamster cheek pouch vessels in situ

The purpose of this study was to determine whether vasoactive intestinal peptide (VIP) dilates resistance arterioles in the in situ systemic circulation and whether inhibitors of neutral endopeptidase (NEP) and angiotensin I-converting enzyme (ACE), two membrane-bound metalloenzymes that are widely distributed in the microcirculation and cleave and inactive VIP, potentiate this response. Using intravital microscopy, we found that VIP (0.05 and 0.1 nmol) induced significant vasodilation in the hamster cheek pouch (13 +/- 1 and 20 +/- 2% increase from baseline, respectively; mean +/- SE; P < 0.05). These responses were significantly potentiated by topical application of phosphoramidon and thiorphan, two relatively selective NEP inhibitors, but not by captopril, a relatively selective ACE inhibitor. Furthermore, suffusion of a mixture of proteinase inhibitors consisting of leupeptin, Bestatin, and DL-2-mercaptomethyl-3-guanidinoethylthiopropanoic acid to inhibit serine proteinases, including mast cell tryptase, aminopeptidases, and carboxypeptidase N, respectively, had no significant effects on VIP-induced responses. These data indicate that VIP elicits vasodilation in the in situ systemic microcirculation and that NEP modulates this response.

Dual inhibition of neutral endopeptidase and angiotensin converting enzyme delays the progression of experimental heart failure

Dual inhibition of neutral endopeptidase and angiotensin converting enzyme delays the progression of experimental heart failure

Dual inhibition of neutral endopeptidase (NEP) and angiotensin converting enzyme (ACE) suppresses atherogenesis and improves endothelial function in hypercholesterolemic rabbits

Dual inhibition of neutral endopeptidase (NEP) and angiotensin converting enzyme (ACE) suppresses atherogenesis and improves endothelial function in hypercholesterolemic rabbits

Alterations in plasma dipeptidyl peptidase IV enzyme activity in depression and schizophrenia: effects of antidepressants and antipsychotic drugs.

Recently, our laboratory reported that the activity of dipeptidyl-peptidase IV (DPP IV) was significantly lower in the peripheral blood of major depressed patients than in normal controls. The present study examines plasma DPP IV activity in 43 major depressed and 13 schizophrenic subjects versus 21 normal controls and the effects of antidepressants and antipsychotic drugs on plasma DPP IV activity. DPP IV activity was significantly lower in major depressed subjects than in normal controls and schizophrenic subjects. There was a trend towards higher DPP IV activity in schizophrenic patients than in normal controls. There were no significant effects of antidepressants or neuroleptics on plasma DPP IV activity in depressed and schizophrenic patients, respectively. There were no significant relationships between plasma DPP IV activity and plasma cortisol or immune-inflammatory markers, such as serum interleukin-6 (IL-6) or soluble IL-2 receptor. A significant and positive correlation was found between plasma DPP IV and prolyl endopeptidase (PEP) enzyme activity in the study group as a whole and in schizophrenic subjects. The results support the hypothesis that lower and higher plasma DPP IV activities are trait markers of major depression and schizophrenia, respectively. It is concluded that alterations in the enzyme activity of peptidases, such as DPP IV and PEP, play a role in the pathophysiology of major depression and schizophrenia.

Viral infection potentiates the increase in airway blood flow produced by substance P.

We examined the effect of respiratory tract infection with Sendai virus on the responsiveness of airway blood flow to substance P (SP) in rats. Pathogen-free rats were inoculated with either Sendai virus suspension or sterile viral growth medium into each nostril. Five days later, we measured airway and esophageal blood flows before and immediately after injection of SP or histamine into the left ventricle of rats in both groups using a modification of the reference-sample microsphere technique. Viral infection potentiated the increase in airway blood flow evoked by SP but not by histamine. We also examined the effect of neutral endopeptidase (NEP) and angiotensin-converting enzyme (ACE) on the SP-induced increase in airway blood flow. Both phosphoramidon (NEP inhibitor) and captopril (ACE inhibitor) potentiated the increase in airway blood flow produced by SP in pathogen-free rats. In the presence of both peptidase inhibitors, a submaximal dose of SP increased blood flow to a similar level in infected and pathogen-free rats. Thus decreased activity of both ACE and NEP may be involved in the exaggerated increase in airway blood flow evoked by SP in virus-infected rats.

Differential ability of tachykinin NK-1 and NK-2 agonists to produce scratching and grooming behaviours in mice

Potent and selective NK-1 and NK-2 agonists as well as compounds with lower selectivity and affinity for NK-1 binding sites were compared in their ability to produce scratching and grooming behaviours when injected intracerebroventricularly in mice. Septide, an agonist with a low affinity for NK-1 binding sites, [Sar 9, Met(O 2) 11]SP and to a lesser extent [Pro 9]SP, two potent and selective NK-1 agonists were the most effective drugs in stimulating these behaviours. Only high doses of [Apa 9,10]SP and [Lys 5, Tyr 7, Pro 8]NKA(4–10), two agonists with low affinity for NK-1 binding sites, produced scratching and grooming responses. Similarly, only high doses of [Lys 5, MeLeu 9, NLe 10]NKA(4–10), a potent NK-2 agonist, produced grooming behaviour. When coinjected with the endopeptidase enzyme inhibitor phosphoramidon, the effects of [Apa 9,10]SP, [Lys 5, Tyr 7, Pro 8]NKA(4–10) and [Pro 9]SP were markedly enhanced. Analyses of the potency of the different agents to displace 3H-SP binding in mouse subcortical structures revealed that the affinities of the agonists for NK-1 receptors are similar to those previously reported in rat brain. The efficacy of the agonists at producing behavioural responses was not equivalent to their potency to bind to central NK-1 receptors. These findings therefore suggest that a stimulation of NK-1 but also non classical NK-1 receptors are involved in the induction of scratching and grooming behaviours.

Lower serum prolyl endopeptidase enzyme activity in major depression: Further evidence that peptidases play a role in the pathophysiology of depression

Prolyl endopeptidase (PEP) is a serine proteinase, which may cleave peptides that are involved in the pathophysiology of major depression, such as arginine vasopressin, β-endorphin, luteinizing hormone-releasing hormone, thyrotropin-releasing hormone, and maybe corticotropin-releasing hormone. PEP may be involved in activation of cell-mediated immunity, autoimmune and inflammatory responses, which repeatedly occur in severe depression. The present study investigates serum PEP activity in 33 normal controls, 16 minor, 14 simple major, and 18 melancholic depressed subjects. Pre-dexamethasone and post-dexamethasone (DST) intact adrenocorticotropic hormone (ACTH) and cortisol values were determined in 33 depressed subjects. Serum PEP activity was siginificantly lower in depressed subjects compared to normal controls and in melancholic depressed subjects compared to minor and simple major depressed subjects. Up to 61.1% of the melancholic patients had serum PEP activities below the mean PEP values of normal controls minus two SDs. In the depressed study group, significant negative correlations between serum PEP activity and severity of illness, post-DST cortisol, and ACTH values were observed. There was a trend toward higher serum PEP activity with increasing age. It is hypothesized that lower serum PEP activity, and lower serum activity of other peptidases, may play a role in the neuroendocrine and immune pathophysiology of major depression.