bluuebunny/crossref_metadata_2025_split huggingface dataset:
Generation by the phosphoramidon‐sensitive peptidases, endopeptidase‐24.11 and thermolysin, of endothelin‐1 and C‐terminal fragment from big endothelin‐1
Authors: Murphy Leonard J. , Corder Roger , Mallet Anthony I. , Turner Anthony J. 
Published: July 2012

Phosphoramidon, a potent inhibitor of endopeptidase‐24.11 (E‐24.11) and thermolysin, has been shown to reduce the hypertensive effect of exogenous big endothelin‐1 (big ET‐1) in rats. To examine whether E‐24.11 or thermolysin convert big ET‐1 to endothelin‐1 (ET‐1) and C‐terminal fragment (CTF), the effects on porcine and human big ET‐1 of each of the purified enzymes were compared in vitro.
For E‐24.11, the relative rates of hydrolysis were ET‐1 > CTF >> big ET‐1. The relative half‐lives for hydrolysis of 3 nmol of each peptide by 200 ng enzyme were: big ET‐1 > 24 h; ET‐1, 37 min; CTF, 57 min. For comparison, the half‐life for hydrolysis of substance P under similar conditions was 2.1 min.
For thermolysin the relative rates of hydrolysis were found to be big ET‐1 > CTF > ET‐1. The relative half‐lives for hydrolysis of 3 nmol peptide by 50 ng enzyme were: big ET‐1, 25 min; ET‐1, 56 min; CTF, 47 min.
Because the low rate of conversion of big ET‐1 to ET‐1 by E‐24.11 did not yield sufficient ET‐1 for h.p.l.c. quantification a RIA specific for ET‐1 (16–21) was used to study further the hydrolysis of big ET‐1 by E‐24.11. Incubation of big ET‐1 (0.2–2 nmol) with E‐24.11 (4–400 ng) generated ET‐1 levels of between 1.7 and 33 pmol measured by RIA. Incubation of big ET‐1 (2 nmol) with E‐24.11 (40 ng) for 8 h showed that steady state levels of ET‐1 were achieved after 4 h indicating that the rate of ET‐1 degradation was then equal to the formation of new ET‐1. Characterization of the immunoreactivity by h.p.l.c. and RIA confirmed that authentic ET‐1 had been produced, but the yield was insufficient for verification by mass spectrometry.
Both ET‐1‐like and CTF‐like peaks were detected at 214 nm when the products of big ET‐1 hydrolysis by thermolysin were resolved by h.p.l.c. RIA and mass spectrometry confirmed the production of ET‐1 with amounts in the range 120–160 pmol.
The hydrolysis profile of ET‐1 by E‐24.11 and thermolysin shows that both enzymes have some common cleavage sites consistent with their similar specificities hydrolysing on the amino side of a hydrophobic residue.
Thermolysin, for which 3D structural information is available, may represent a better model for endothelin converting enzyme (ECE) action than E‐24.11 and could be useful for the design of ECE inhibitors. Since E‐24.11 can both synthesize and hydrolyse ET‐1, the presence of E‐24.11 in membrane fractions or in partially purified ECE preparations may produce misleading estimates of ECE activity.
Local cardiac effects of substance P: roles of acetylcholine and noradrenaline
Authors: Chiao Hsi , Caldwell Robert W. 
Published: July 2012

The local cardiac actions of substance P were examined in isolated perfused hearts and atria of the guinea‐pig.
In both hearts and right atria, substance P caused negative inotropic and chronotropic effects.
Atropine (10−6 m) or depletion of acetylcholine, by electrical stimulation and hemicholinium‐3 perfusion, significantly attenuated the negative inotropic and chronotropic effects of substance P. α‐ and β‐adrenoceptor blockade by nadolol and phentolamine (10−6 m each) did not prevent the negative inotropic and chronotropic effects of substance P. This indicates that cholinergic neurones, but not adrenergic neurones, partially mediate the effects of substance P.
There was no significant difference in the effects of substance P observed between groups with acetylcholine depletion and with cholinoceptor blockade. This suggests that substance P elicits its effects mainly through release of acetylcholine.
These results indicate that substance P has negative inotropic and chronotropic effects in guinea‐pig hearts and right atria mediated partly by release of acetylcholine. Substance P also appears to have direct effects on cardiac tissue.
Modulation by nitric oxide of prostaglandin biosynthesis in the rat
Authors: Sautebin Lidia , Ialenti Armando , Ianaro Angela , Di Rosa Massimo 
Published: July 2012

Modulation of prostaglandin biosynthesis in vivo by either exogenous or endogenous nitric oxide (NO) has been studied in the rat using arachidonic acid (AA)‐induced paw oedema and measuring both the foot volume and the amount of 6‐keto‐prostaglandin F1α (6‐keto‐PGF1α), the stable metabolite of prostacyclin (PGI2), in the oedematous fluid recovered from inflamed paws.
Paw injections of 150 or 300 nmol of AA were virtually inactive whereas 600 nmol produced a moderate oedema which was greatly reduced by the NO synthase inhibitor l‐NG‐nitro arginine methyl ester (l‐NAME, 100 nmol/paw) and the NO scavenger haemoglobin (Hb, 30 μmol/paw), but unaffected by the inhibitor of the soluble guanylate cyclase, methylene blue (Mb, 3 μmol/paw) and L‐arginine (15 μmol/paw).
The NO‐donors (10 μmol/paw) 3‐morpholino‐sydnonimine‐hydrochloride (SIN‐1), S‐nitroso‐N‐acetyl‐d, l‐penicillamine (SNAP) and sodium nitroprusside (SNP) significantly potentiated the paw oedema induced by AA (300 nmol/paw).
SIN‐1 (2.5, 5 and 10 μmol/paw) produced a significant dose‐dependent increase of the oedema induced by AA which was correlated with increased amounts of 6‐keto‐PGF1α in the fluid recovered from inflamed paws.
Both oedema and prostaglandin biosynthesis induced by the combination AA + SIN‐1 were greatly suppressed by either Hb (30 μmol/paw) or indomethacin (3 μmol/paw or 5 mg kg−1 s.c.) but unaffected by Mb (3 μmol/paw).
In LPS‐treated rats (6 mg kg−1, i.p.) doses of AA inactive in normal animals produced a remarkable oedema which was reduced by l‐NAME or Hb, unaffected by Mb and increased by l‐arginine.
These results demonstrate that NO increases prostaglandin biosynthesis in vivo through a guanosine 3′:5′‐cyclic monophosphate (cyclic GMP)‐independent mechanism and suggest that the interaction between NO synthase and cyclo‐oxygenase (COX) pathways may represent an important mechanism for the modulation of the inflammatory response.
Effects of nitric oxide synthase inhibition combined with nitric oxide inhalation in a porcine model of endotoxin shock
Authors: Klemm Peter , Thiemermann Christoph , Winklmaier Gisela , Martorana Piero A. , Henning Rainer 
Published: July 2012

The present investigation compares the effects of intravenous infusion of the NO synthase inhibitor NG‐monomethyl‐l‐arginine (l‐NMMA) with that of an inhalation with NO gas in a porcine model of endotoxin (lipopolysaccharide, LPS) shock. In addition, the effects of the combination of these two treatments were also investigated.
Male pigs were anaesthetized and instrumented for the measurement of haemodynamic parameters. Blood samples were withdrawn at different time intervals for determination of blood gases, pH, and plasma levels of nitrite/nitrate and tumour necrosis factor.
Endotoxin infusion (15 μg kg−1 h−1 for 3 h) caused a progressive fall in mean arterial blood pressure (MABP) and cardiac output (CO) and a biphasic increase in mean pulmonary arterial pressure (MPAP) and pulmonary vascular resistence (PVR). A continuous infusion of l‐NMMA (0.1 mg kg−1 min−1) significantly attenuated the fall in MABP, but did not affect MPAP, CO and PVR. NO‐inhalation (50 p.p.m.) did not affect MABP, but significantly blunted the biphasic increase in MPAP and PVR and significantly delayed the fall in CO. The combination of l‐NMMA infusion (0.1 mg kg−1 min−1) with NO‐inhalation (50 p.p.m.) completely prevented the fall in MABP, significantly improved CO, and attenuated the biphasic increase in MPAP and PVR.
Endotoxin also caused a decline in Pao2 and a rise of Paco2. Infusion of l‐NMMA neither affected the fall in Pao2 nor the increase in Paco2. In contrast, inhalation with NO gas alone as well as the combined administration of l‐NMMA infusion and NO‐inhalation completely prevented the fall in Pao2 and significantly protected against the increase in Paco2.
Infusion of endotoxin for 180 min resulted in a mortality of 58%, which was not affected by l‐NMMA (63%). In contrast, treatment of LPS‐animals with either NO‐inhalation alone or NO‐inhalation plus l‐NMMA completely prevented mortality.
This investigation demonstrates that treatment with NO‐inhalation, in order to prevent the dramatic increase in MPAP, PVR and the alterations in peripheral blood gases combined with systemic l‐NMMA to improve systemic MABP and thus organ perfusion, may be a new therapeutic regimen in the treatment of septic shock.
Protein kinase C and tyrosine kinase pathways regulate lipopolysaccharide‐induced nitric oxide synthase activity in RAW 264.7 murine macrophages
Authors: Paul Andrew , Pendreigh Robert H , Plevin Robin 
Published: July 2012

In RAW 264.7 macrophages, lipopolysaccharide (LPS) and γ‐interferon (IFNγ) alone or in combination stimulated the induction of nitric oxide synthase (iNOS) activity and increased the expression of the 130 kDa isoform of NOS.
LPS‐induced NOS activity was reduced by incubation with CD14 neutralising antibodies and abolished in macrophages deprived of serum.
LPS stimulated a small increase in protein kinase C (PKC) activity in RAW 264.7 macrophages which was dependent on the presence of serum. However, IFNγ did not potentiate LPS‐stimulated PKC activity.
The protein kinase C inhibitor, Ro‐318220, abolished both LPS‐ and IFNγ‐stimulated protein kinase C activity and the induction of NOS activity.
LPS‐ and IFNγ‐induced NOS activity was reduced by the tyrosine kinase inhibitor genestein. Genestein also reduced LPS‐stimulated protein kinase C activity but did not affect the response to the protein kinase C activator, tetradecanoylphorbol acetate (TPA).
Nicotinamide, an inhibitor of poly‐ADP ribosylation, abolished LPS‐ and IFNγ‐induced NOS activity.
Brefeldin A, an inhibitor of a factor which stimulates nucleotide exchange activity on the 21 kDa ADP‐ribosylation factor, ARF, reduced LPS‐ and IFNγ‐induced NOS activity by approximately 80%.
These results suggest the involvement of protein kinase C, tyrosine kinase and ploy‐ADP ribosylation pathways in the regulation of the induction of nitric oxide synthase in RAW 264.7 macrophages by LPS and IFNγ.
Comparisons of the effects of nicorandil, pinacidil, nicardipine and nitroglycerin on coronary vessels in the conscious dog: role of the endothelium
Authors: Ghaleh Bijan , Dubois‐Randé Jean‐Luc , Hittinger Luc , Giudicelli Jean‐François , Berdeaux Alain 
Published: July 2012

The vasodilator properties of nicorandil on large and small coronary arteries were compared to those of nicardipine, pinacidil, nitroglycerin and acetylcholine in six conscious dogs.
Intravenous bolus injections of acetylcholine (0.1 μg kg−1), nitroglycerin (0.3–3 μg kg−1), pinacidil (10–100 μg kg−1), nicardipine (3–30 μg kg−1) and nicorandil (10–100 μg kg−1) dose‐dependently increased circumflex coronary artery diameter and decreased coronary vascular resistance, indicating vasodilator effects on both conduit and resistance coronary arteries.
Three days after removal of the endothelium of the circumflex coronary artery (balloon angioplasty), pinacidil‐ and nicardipine‐induced dilatation of large coronary arteries was greatly reduced (both − 76%, P<0.01) whereas that produced by nitroglycerin and nicorandil was decreased only slightly and to a similar extent for both drugs (− 19%, P<0.01 and −28%, P<0.05, respectively).
Thus in conscious dogs, nicardipine‐ and pinacidil‐induced dilatation of large coronary arteries is endothelium‐dependent. In contrast, the vasodilator effects of nitroglycerin and nicorandil on conduit vessels are endothelium‐independent.
Finally, our results demonstrate that nicorandil dilates the large coronary arteries through its nitrate‐like action and that the ATP‐potassium channel opening properties of the drug are not involved in this effect in the conscious dog.
Unexpected prosecretory action component of loperamide at μ–opioid receptors in the guinea‐pig colonic mucosa in vitro
Authors: Kromer Wolfgang 
Published: July 2012

In a voltage clamp setting (Ussing chamber), the antidiarrhoeal drug, loperamide (Lop) slightly augmented prostaglandin E1 (PGE1) plus theophylline‐stimulated net chloride secretion above control values at low concentrations (10−10 and 10−9 m) but inhibited it at higher concentrations (10−6 and 10−5 m). The apparently weak prosecretory action component of Lop was turned into a clear cut antisecretory effect by pretreatment with 2 times 10−7 m naloxonazine plus 10−7 m CTOP‐NH2 , two selective μ opioid receptor antagonists. This indicates a prosecretory effect of loperamide at μ opioid receptors. The antisecretory effect of low Lop concentrations, uncovered by μ opioid receptor blockade, was prevented by additional blockade of κ opioid receptors by 5 times 10−9 m nor‐binaltorphmiine (nor‐BNI).
The nonselective opioid antagonist, naloxone, at 10−6 m did not significantly reduce either PGE1 plus theophylline‐stimulated net chloride secretion in Lop‐free controls or the antisecretory action of Lop. By contrast, the partial agonist ethylketocyclazocine (EKC), which activates κ but blocks μ opioid receptors, concentration‐dependently inhibited PGE1 plus theophylline‐stimulated net chloride secretion without any consistent prosecretory action component. Nor‐BNI at 5 times 10−8 m significantly blocked the antisecretory action of EKC.
It is concluded that, in the guinea‐pig colonic mucosa under the present conditions, μ opioid receptors mediate enhancement and κ opioid receptors inhibition of PGE1‐stimulated net chloride secretion by low Lop concentrations. The two opposite actions are largely masked by superimposition. An opioid receptor‐independent mechanism of action contributes to the antisecretory effect of Lop at high concentrations.
Effects of portal hypertension on responsiveness of rat mesenteric artery and aorta
Authors: Cawley Teresa , Geraghty J. , Osborne H. , Docherty J.R. 
Published: July 2012

We have examined the effects of pre‐hepatic portal hypertension on the responsiveness of rat small mesenteric arteries and aorta. Rats were made portal hypertensive by creating a calibrated portal vein stenosis, or sham‐operated.
In rat mesenteric arteries, there was no significant difference between portal hypertensive and sham‐operated animals in the contractile potency of noradrenaline (NA), but the maximum contractile responses to NA, U46619 and KCl were significantly increased in vessels from portal hypertensive animals. This altered maximum contractile response was not due to alterations in smooth muscle mass.
In rat mesenteric arteries, there were no significant differences between portal hypertensive and sham‐operated animals in endothelium‐dependent relaxations to acetylcholine (ACh). The difference between portal hypertensive and sham‐operated rats in the maximum response to U46619 was maintained following a combination of methylene blue (1 μm) and NG‐monomethyl‐l‐arginine (100 μm), suggesting that any differences in endothelial function do not explain differences in the response to vasoconstrictors.
In rat aorta, there were no significant differences between portal hypertensive and sham‐operated animals in the contractile response to NA or KCl or in the endothelium‐dependent relaxations to ACh.
In pithed rats, there was no difference between portal hypertensive and sham‐operated animals in the pressor potency of NA.
It is concluded that portal hypertension produces an increase in the contractile response to the vasoconstrictors NA, U46619 and KCl in rat mesenteric arteries but not in the aorta. This suggests that the diminished responsiveness to vasoconstrictors reported in portal hypertensive rats in vivo is not due to a diminished responsiveness at the level of the vascular smooth muscle.
The inhibition of CYP enzymes in mouse and human liver by pilocarpine
Authors: Kimonen Tanja , Juvonen Risto O. , Alhava Esko , Pasanen Markku 
Published: July 2012

Pilocarpine is a cholinomimetic natural alkaloid. Its interactions with testosterone hydroxylations, coumarin 7‐hydroxylase (COH), dimethylnitrosamine N‐demethylase (DMNA), pentoxyresorufin O‐dealkylase (PROD) and 7‐ethoxyresorufin O‐deethylase (EROD), which are indicative of the activities of cytochrome P4502A5 (CYP2A5) or 6, 2E1, 2B, 1A, were examined in mouse and human liver microsomes.
In mouse liver microsomes the IC50 values of pilocarpine were 6 μm for COH and testosterone 15a‐hydroxylase (T15αOH) activities, 4 μm for PROD, ≅ 100 μm for DMNA and testosterone 60‐hydroxylase (T6βOH) activities and > 1 mm for EROD activity.
In human liver microsomes, the IC50 value for COH was 6 μm and for DMNA 10 μm; T15αOH and PROD activities were not detectable but T6βOH and testosterone 16β/2β‐hydroxylase activities were moderately inhibited (IC50 70 μm).
These results suggest that pilocarpine has (i) a high affinity towards phenobarbitone‐inducible CYP2A4/5 and CYP2B activities in mouse liver, (ii) a high affinity towards CYP2A6 in human liver microsomes and (iii) a moderate affinity towards CYP3A enzyme(s) in both microsomal preparations.
The low IC50 concentrations in vitro indicate potential metabolic interactions between pilocarpine and several P450 enzymes.
Dysfunction of muscarinic M2 receptors after the early allergic reaction: possible contribution to bronchial hyperresponsiveness in allergic guinea‐pigs
Authors: ten Berge Donald E.J. , Santing Ruud E. , Hamstra Jacob Jan , Roffel Ad F. , Zaagsma Johan 
Published: July 2012

Using a guinea‐pig model of allergic asthma, in which the animals display early (0–5 h) and late phase (8–23 h after antigen challenge) bronchoconstrictor reactions, the function of prejunctional inhibitory M2 and postjunctional M3 receptors in isolated tracheal preparations have been investigated. In addition, cardiac M2 receptor function in vitro and bronchial responsiveness to histamine in vivo were evaluated.
Sensitivity to inhaled histamine was increased 3.1 fold and 1.6 fold after the early and late allergic reactions (i.e. at 5 h and 23 h after a single ovalbumin challenge), respectively. At 23 h after the last of four allergen challenges, executed on four consecutive days, bronchial hyperresponsiveness to histamine was diminished to 1.3 fold.
After the early response, there was no change in cardiac muscarinic M2 receptor function, since in left atria pD2 (‐log EC50) and Emax values of pilocarpine and pKB values of AQ‐RA 741, a selective M2 receptor antagonist, were not significantly different from controls (unchallenged sensitized animals), and this also applied to methacholine pD2 values for muscarinic M3 receptors in tracheal smooth muscle.
Prejunctional inhibitory muscarinic M2 autoreceptors in airway smooth muscle were markedly dysfunctional after the early allergic response, since potentiation of electrically evoked twitch contractions of tracheal preparations by low concentrations of the M2‐selective muscarinic receptor antagonists, gallamine, methoctramine, AQ‐RA 741 and AF‐DX 116, which is the result of M2 receptor blockade, was clearly and significantly diminished compared to controls. However, after the late response, both in single and repeatedly challenged animals, twitch potentiation was not significantly different from and similar to controls, indicating restoration of M2 receptor function during the late allergic reaction.
It is concluded that dysfunction of muscarinic M2 autoreceptors in the airways of sensitized and challenged guinea‐pigs is already present after the early allergic reaction, and that it has recovered after the late response. Since histamine‐induced bronchoconstriction involves vagal pathways, the present results suggest that bronchial hyperresponsiveness to histamine is partly due to M2 autoreceptor dysfunction, leading to increased release of acetylcholine.
Role of endothelium in the human uterine arteries during normal menstrual cycle
Authors: Azuma Hiroshi , Obayashi Satoshi , Hamasaki Hidehisa , Koyama Takao , Aso Takeshi 
Published: July 2012

The present experiments were designed to investigate the role of endothelium in the human uterine arteries during the normal menstrual cycle.
Acetylcholine (ACh) produced a concentration‐dependent relaxation response during the higher level of plasma 17β‐oestradiol (E2) (follicular and luteal phases, E2 = 131.9 ± 15.9 pg ml−1, n = 13; group I). However, the agent did not produce a definite relaxation, but produced a slight contraction during the ovulatory and menstruation phases (E2 = 19.8 ± 2.9 pg mg−1, n = 5; group II). During the follicular and luteal phases (E2 = 181.1 ± 9.0 pg ml−1, n = 6), ACh produced a slight contraction, but not relaxation in 6 cases (group III). Relaxation in response to A23187 in group II was not different from that in group I, while it was significantly (P<0.05 and P <0.005) reduced in group III. Sodium nitroprusside (SNP)‐induced relaxation was similar in the three groups.
Correlation between the maximum response to ACh and the plasma E2 was highly significant (γ = 0.8142, P <0.001) in 18 cases of groups I and II, but not in all 24 cases including group III (γ = 0.1183, NS).
Relaxations in response to ACh in group I or A23187 in all groups were abolished after removal of the endothelium. In group I, ACh‐ and A23187–induced relaxations were greatly inhibited by methylene blue or NG‐nitro‐l‐arginine (l‐NOARG) and partially inhibited by indomethacin. None of these treatments except for methylene blue modified the SNP‐induced relaxation, which was significantly inhibited by methylene blue.
The A23187‐induced relaxation was hardly affected by methylene blue or l‐NOARG in group III, but was partially inhibited by these agents in group II. The effect of indomethacin in inhibiting the A23187 induced‐relaxation was most potent (58.9%) in group III and least (16.9%) in group I.
There were no histological changes in 14 cases out of 18 (groups I and II), but very slight intimal thickening was observed in 4 cases in group I. On the other hand, severe intimal thickening was observed in all 6 cases in group III.
These results indicate that, in human uterine artery strips, ACh and A23187 cause endothelium‐dependent relaxations, which are mediated mainly through EDRF/NO in group I, mainly prostacyclin (PGI2) in group III, or both in group II. It is suggested that lack of the production/release of EDRF/NO and/or of interaction between EDRF/NO and PGI2 might play a role in the formation of intimal thickening in human uterine arteries.