bluuebunny/crossref_metadata_2025 huggingface dataset:
Can Biliary Carcinoembryonic Antigen Identify Colorectal Cancer Patients with Occult Hepatic Metastases?
Authors: Destri Giovanni Li , Lanteri Raffaele , Santangelo Marco , Torrisi Benedetto , Cataldo Antonio Di , Puleo Stefano
Published: July 2006

Background

Twenty‐five percent of radically treated colorectal cancer patients already have occult hepatic metastases (OHM) that will later be observed during postoperative follow‐up. Instrumental examinations, i.e., intraoperative ultrasound or Doppler perfusion index, have not improved diagnosis. As carcinoembyonic antigen (CEA) levels are useful to reveal hepatic metastases from colorectal cancer, determination of CEA in the bile rather than the blood may allow preclinical diagnosis of OHM thanks to the reduced volume of bile.

Methods

One hundred radically treated colorectal cancer patients were enrolled in the study. Bile was withdrawn from the gallbladder intraoperatively and biliary CEA levels determined using an immuno‐enzymatic method (normal value 0–5 ng/ml). Eighty‐nine fully evaluable patients were followed up for three years postoperatively to monitor hepatic metastases. Preoperative blood CEA, lymph node metastases and biliary CEA were compared in order to assess which procedure was more efficient in identifying patients who would develop hepatic metastases.

Results

Eleven of the 89 evaluable patients developed hepatic metastases: 9/11 presented elevated biliary CEA levels (mean: 12.73; range: 5.1–26.2); 8/11 had high preoperative blood CEA values; and 9/11 were at anatomopathological stage N+. In the 78 patients who did not develop hepatic metastases, biliary CEA was within normal limits in 73/78, preoperative blood CEA was normal in 60/78, and 58/78 patients were at anatomopathological stage N−. Hence, the sensitivity of biliary CEA was 81.8%, specificity was 93.6%, and diagnostic accuracy was 92.1%.

Conclusions

Determination of biliary CEA seems to be more efficient in identifying patients presenting OHM who require frequent clinical examinations or adjuvant cancer treatment.
Effect of Sirolimus in the Prevention of Adhesions Around Intraabdominal Prosthetic Graft
Authors: Kanko Muhip , Ozbudak Ersan , Ozerdem Ali , Aksoy Ahmet , Kilic Metehan , Berki K. Turan
Published: July 2006

Background

Adhesions due to the reactions caused by the grafts used in primary vascular operations can lead to various problems when a secondary operation is necessary. These problems include bleeding, injuries to neighboring organs, and complications occurring due to a prolonged operation. We investigated the effects of sirolimus, which has antiproliferative features, on vascular adhesions.

Methods

The abdominal aorta of rats was explored and abrasions applied. Following the fixation of a polytetrafluoroethylene (PTFE) graft on the abdominal aorta, sirolimus (rapamycin) is applied (in powdered form) onto the grafts of the study group. Eight weeks later a laparotomy was repeated and any adhesions were evaluated.

Results

In the study group the adhesions were determined to be fewer in number and milder in severity. Severe cases of adhesion were determined in the control group.

Conclusions

Therefore, sirolimus applied around the prosthesis in vascular operations was determined to be effective at preventing possible adhesions.
Effects of leaf quality and microhabitat on the survival of a leaf‐rolling weevil (Attelabidae)
Authors: Kobayashi Chisato , Kato Makoto
Published: July 2006

Female Cycnotrachelus roelofsi (Coleoptera: Attelabidae) construct two types of leaf roll, i.e., cut‐off cradles (CCs) and suspended cradles (SCs), in which to lay eggs; these cradles are generally constructed using older and younger leaves, respectively. We conducted two experiments to determine whether the quality of cradle leaves affects egg and larval survival. In the first experiment, we severed all SCs from a tree and placed them on the ground with unmanipulated CCs in the early breeding season. In the second experiment, we resuspended all CCs in a tree with unmanipulated SCs in the late breeding season. We also compared leaf mass per area (LMA), polyphenol content, and nitrogen content between the two cradle types to determine whether there were any differences in leaf quality. Larval mortality, probably caused by cradle herbivory, was significantly greater in severed SCs than in intact CCs in the early season, suggesting that leaf quality had a profound effect on larval mortality in the terrestrial microhabitat. In contrast, larval mortality did not differ between resuspended CCs and intact SCs in the late season, suggesting that leaf quality had little effect on larval mortality in the arboreal microhabitat. LMA was higher in CCs than in SCs, but there were no differences in the nitrogen and polyphenol contents. These results suggest that cradles constructed using mature, tough leaves were more effective against terrestrial cradle herbivores than those constructed using new, soft leaves.
Fingertip Reconstruction with the Laterally Based Thenar Flap: Indications and Long-Term Functional Results
Authors: Rinker Brian
Published: July 2006

The thenar flap is a time-tested method of fingertip reconstruction, but functional outcome data are scarce in the literature. The purpose of this study was to analyze the long-term function following fingertip reconstruction with a laterally based thenar flap and to compare these results with other established methods. Nineteen patients underwent a thenar flap between 2001 and 2004. Patients ranged in age from 3 to 48 years. The mean angle of proximal interphalangeal immobilization was 66° (range 30–85°) and was greater for radial digits. Time to division ranged from 11 to 15 days. Seventeen patients underwent follow-up evaluation of range of motion, two-point discrimination, and sensory threshold (Semmes-Weinstein). A questionnaire measured patient satisfaction in three areas: sensibility, function, and appearance. The mean follow-up was 20 months. Reconstructive goals were met in all cases. The mean metacarpalphalangeal and proximal interphalangeal motion in the reconstructed fingers was not significantly reduced, compared to the unaffected side. The distal interphalangeal motion was 42°, compared to 55° in the contralateral side ( p < 0.01). The mean static two-point discrimination in the flap was 6.8 mm, compared to 3.8 mm in the contralateral side. Fourteen of 17 patients exhibited monofilament thresholds of 33.1 g/mm2 or less. There were no hypertrophic or tender donor scars. This study does not support the contention that thenar flaps are associated with problematic donor scars and flexion contractures, even for adults or ulnar digits. Sensory recovery compared favorably to published results of cross-finger and homodigital flaps. When sound technical principles are followed, excellent outcomes can be expected.
Chloroplast heat shock protein Cpn60 from Chlamydomonas reinhardtii exhibits a novel function as a group II intron‐specific RNA‐binding protein
Authors: Balczun Carsten , Bunse Astrid , Schwarz Christian , Piotrowski Markus , Kück Ulrich
Published: July 2006

Intron‐binding proteins in eukaryotic organelles are mainly encoded by the nuclear genome and are thought to promote the maturation of precursor RNAs. Here, we present a biochemical approach that enable the isolation of a novel nuclear‐encoded protein from Chlamydomonas reinhardtii showing specific binding properties to organelle group II intron RNA. Using FPLC chromatography of chloroplast protein extracts, a 61‐kDa RNA‐binding protein was isolated and then tentatively identified by mass spectrometry as the chloroplast heat shock protein Cpn60. Heterologous Cpn60 protein was used in RNA protein gel mobility shift assays and revealed that the ATPase domains of Cpn60 mediates the specific binding of two group II intron RNAs, derived from the homologous chloroplast psaA gene and the heterologous mitochondrial LSU rRNA gene. The function of Cpn60 as a general organelle splicing factor is discussed.
Movements and survival of sonically tagged farmed Atlantic salmon released in Cobscook Bay, Maine, USA
Authors: Whoriskey Frederick G. , Brooking Paul , Doucette Gino , Tinker Stephen , Carr Jonathan W.
Published: July 2006

We sonically tagged and released farmed Atlantic salmon (Salmo salar) from a cage site in Cobscook Bay, Maine, USA. The fish were released in January (n = 75) and in April and May (n = 198) 2004 to study their movement patterns and survival and to assess the possibility of recapturing them. Inshore and offshore waters in this region are subject to intense tidal currents. Tagged salmon dispersed >1 km from the cage site within a few hours of their release. Mortality was high within Cobscook Bay and the surrounding coastal region (56% of the winter (January) releases; 84% of the spring (March) releases), probably the result of seal predation. Most surviving fish exited the coastal zone and entered the Bay of Fundy along the routes of the dominant tidal currents, passing through Canadian waters. No tagged fish were detected during the wild salmon spawning season in autumn 2004 in any of the 43 monitored salmon rivers draining into the Bay of Fundy, or during 2005 either in the Magaguadavic River, the site of the hatchery in which the fish were reared to the smolt stage, or by a limited coastal receiver array.
Unfree Labor, Apprenticeship and the Rise of the Victorian Hull Fishing Industry: An Example of the Importance of Law and the Local State in British Economic Change
Authors: Steinberg Marc W.
Published: July 2006

Within the last decade there has been considerable renewed attention on the importance of British master and servant law in the eighteenth and nineteenth centuries as a means of labor discipline and control. This article argues for further analyses of how the law was used within local contexts and specific industries and calls for increased focus on the role of the local state in labor relations. It argues that unfree labor played an important role in the development of some industries, and challenges claims of the demise of apprenticeship in later nineteenth-century England. Through an analysis of the Hull fish trawling industry in 1864–1875 it demonstrates that the exploitation of apprentice labor, and the control of fishing apprentices through punitive master–servant prosecutions were vital to the expansion of the trade.
Beckwith–Wiedemann syndrome: multiple molecular mechanisms
Authors: Enklaar Thorsten , Zabel Bernhard U. , Prawitt Dirk
Published: July 2006

Beckwith–Wiedemann syndrome (BWS) is a congenital overgrowth condition with an increased risk of developing embryonic tumours, such as Wilms' tumour. The cardinal features are abdominal wall defects, macroglossia and gigantism. BWS is generally sporadic; only 10–15% of cases are familial. A variety of molecular aberrations have been associated with BWS. The only mutations within a gene are loss-of-function mutations in the CDKN1C gene, which codes for an imprinted cell-cycle regulator. CDKN1C mutations appear to be particularly associated with umbilical abnormalities, but not with increased predisposition to Wilms' tumour. In the remaining BWS subgroups, a disturbance of the tight epigenetic regulation of gene expression (patUPD 11p, microdeletions or epimutations) seems to be the cause of the syndrome. Here we describe the clinical presentation of BWS and its dissociation from phenotypically overlapping overgrowth syndromes. We then review the current concepts of causative molecular genetic and epigenetic mechanisms, and discuss future directions of research.
Role of tachykinins in castor oil diarrhoea in rats
Authors: Croci Tiziano , Landi Marco , Emonds‐Alt Xavier , Le Fur Gérard , Maffrand Jean‐Pierre , Manara Luciano
Published: July 2006

We set out to ascertain the role of tachykinins, neurokinin A and substance P, in castor oil‐induced diarrhoea in rats as disclosed by the inhibitory effect of the non‐peptide NK1‐ and NK2‐receptor antagonists, SR 140333 and SR 48968, respectively.

SR 48968 (0.02 to 20 μg kg−1, s.c. or p.o.), and the opioid receptor agonist loperamide (1–10 mg kg−1, p.o.), dose‐dependently prevented castor oil effects: % inhibition vs castor oil, diarrhoea 0 to 100, increase in faecal mass 7 to 90 and water content 16 to 90. SR 140333 (0.02 to 20 μg kg−1, s.c.) and the platelet activating factor antagonist SR 27417 (5 to 500 μg kg−1, p.o.) did not prevent the increase in faecal water content, but reduced faecal mass (35 to 66%) and diarrhoea (0 to 57%).

The R‐enantiomers of tachykinin NK1 and NK2 receptor antagonists, SR 140603 and SR 48605 (both at 2 or 20 μg kg−1, s.c.) had no effect other than reducing faecal mass at the highest dose tested.

SR 48968 (20 μg kg−1, p.o.) but not loperamide (10 mg kg−1, p.o.) given 24 h before castor oil, still slightly but significantly reduced by 30% the increase of faecal mass output; both treatments significantly reduced (30 to 70%) the effect of castor oil on faecal water content, although the incidence of diarrhoea was only slightly less than in controls.

In castor oil‐treated rats, naloxone (2 mg kg−1, s.c.) completely blocked the antidiarrhoeal action of loperamide (10 mg kg−1, p.o.) but not of SR 48968 (20 μg kg−1, p.o.); a similar result was obtained on faecal mass and water content.

Castor oil strongly increased the occurrence of manometrically recorded propulsive giant contractions (500 to 1000% over control values) of transverse and distal colon, this effect being significantly prevented (80 to 100%) by SR 48968 and loperamide and partially by SR 140333 (35% distal colon, 70% transverse colon).

In castor oil free rats, loperamide but not SR 48968 or SR 140333 significantly reduced by 50% the gastrointestinal transit of a charcoal test meal, as well as 24 h faecal mass output. Consistently, loperamide, unlike the tachykinin receptor antagonists, had a dramatic effect on manometric recordings of intestinal motility, reducing all kinds of colonic contractions.

Our findings suggest that castor oil diarrhoea in rats entails activation of NK1 and NK2 receptors by endogenous tachykinins, whose antagonists may have a potential as antidiarrhoeal agents free from the constipating action of opioids.
Role of 5‐ht7 receptors in the long‐lasting hypotensive response induced by 5‐hydroxytryptamine in the rat
Authors: Terrón José A.
Published: July 2006

The receptor mediating the long‐lasting hypotensive effect of intravenous (i.v.) 5‐hydroxytryptamine (5‐HT) in the rat was originally classified as 5‐HT1‐like. Since some pharmacological properties of this receptor are closely similar to those for the cloned 5‐ht7 receptor, the present study investigated the effects of several 5‐HT receptor agonists and antagonists showing high affinity for the cloned 5‐ht7 receptor in pithed rats with artificially raised blood pressure.

I.v. bolus administration of 5‐HT, 5‐carboxamidotryptamine (5‐CT), 5‐methoxytryptamine, lisuride and sumatriptan to bilaterally vagotomized pithed rats pretreated with ketanserin (0.18 μmol kg−1, i.v.), the diastolic blood pressure of which had been raised by a continuous i.v. infusion of methoxamine (60–80 nmol kg−1 min−1), produced dose‐dependent hypotensive responses; only 5‐HT and 5‐CT displayed similar maximum effects. In addition to mimicking the hypotensive action of 5‐HT with a lower maximum effect, lisuride strongly antagonized the 5‐CT‐induced hypotensive responses thus suggesting a partial agonist effect. The rank order of hypotensive agonist potency was 5‐CT >> 5‐HT 5‐methoxytryptamine lisuride >> sumatriptan.

In experiments with antagonists, i.v. treatment with metergoline (2.48 μmol kg−1), mesulergine (2.76 μmol kg−1), methysergide (2.13 μmol kg−1), lisuride (0.22 μmol kg−1), methiothepin (0.68 μmol kg−1), mianserin (10.6 μmol kg−1), or the atypical antipsychotic drugs, clozapine (11.0 μmol kg−1) or risperidone (78.0 nmol kg−1), produced significant rightward displacements of the dose‐response curve for 5‐CT in methoxamine‐infused pithed animals pretreated with ketanserin (0.18 μmol kg−1, i.v.); lisuride, methiothepin and risperidone behaved as non‐competitive antagonists as they elicited a significant reduction of the maximum effect to 5‐CT. In contrast, blockade of 5‐HT1, 5‐HT3 and 5‐HT4 receptors with i.v. propranolol (3.38 μmol kg−1), MDL‐72222 (1.59 μmol kg−1) and GR125487 (1.91 μmol kg−1), respectively, did not alter 5‐CT‐induced hypotensive responses; ketanserin (0.18 μmol kg−1, i.v.) failed to modify the dose‐response curve for 5‐CT in saline‐pretreated animals. Lastly, inhibition of the prostaglandin‐forming cyclo‐oxygenase and nitric oxide synthase with indomethacin (14 μmol kg−1, i.v.) and NG‐nitro‐L‐arginine methyl ester (L‐NAME, 120 μmol kg−1, i.v.), respectively, had no significant effects on 5‐CT‐induced hypotensive effects.

Taken together, the present pharmacological data suggest that the long‐lasting vasodepressor action of 5‐HT in the rat involves activation of receptors closely similar to the cloned 5‐ht7 subtype. Since no evidence for an indirect mechanism could be obtained, these receptors may be primarily located in the vascular smooth muscle of the systemic resistance vessels. These findings represent further evidence favouring the functional role of the 5‐ht7 receptor.
Inhibition by mibefradil, a novel calcium channel antagonist, of Ca2+‐ and volume‐activated Cl− channels in macrovascular endothelial cells
Authors: Nilius Bernd , Prenen Jean , Kamouchi Masahiro , Viana Felix , Voets Thomas , Droogmans Guy
Published: July 2006

We have studied the effects of mibefradil, a novel calcium antagonist, on the resting potential and ion channel activity of macrovascular endothelial cells (calf pulmonary artery endothelial cells, CPAE). The patch clamp technique was used to measure ionic currents and the Fura‐II microfluorescence technique to monitor changes in the intracellular Ca2+ concentration, [Ca2+]i.

Mibefradil (10 μM) hyperpolarized the membrane potential of CPAE cells from its mean control value of −26.6±0.6 mV (n=7) to −59.8±1.7 mV (n=6). A depolarizing effect was observed at higher concentrations (−13.7±0.6 mV, n=4, 30 μM mibefradil).

Mibefradil inhibited Ca2+‐activated Cl− currents, ICl,Ca, activated by loading CPAE cells via the patch pipette with 500 nM free Ca2+ (Ki=4.7±0.18 μM, n=8).

Mibefradil also inhibited volume‐sensitive Cl− currents, ICl,vol, activated by challenging CPAE cells with a 27% hypotonic solution (Ki=5.4±0.22 μM, n=6).

The inwardly rectifying K+ channel, IRK, was not affected by mibefradil at concentrations up to 30 μM.

Ca2+ entry activated by store depletion, as assessed by the rate of [Ca2+]i‐increase upon reapplication of 10 mM extracellular Ca2+ to store‐depleted cells, was inhibited by 17.6±6.5% (n=8) in the presence of 10 μM mibefradil.

Mibefradil inhibited proliferation of CPAE cells. Half‐maximal inhibition was found at 1.7±0.12 μM (n=3), which is similar to the concentration for half‐maximal block of Cl− channels.

These actions of mibefradil on Cl− channels and the concomitant changes in resting potential might, in addition to its effect on T‐type Ca2+ channels, be an important target for modulation of cardiovascular function under normal and pathological conditions.
Cytokines, nerve growth factor and inflammatory hyperalgesia: the contribution of tumour necrosis factor α
Authors: Woolf C. J. , Allchorne A. , Safieh‐Garabedian B. , Poole S.
Published: July 2006

Peripheral inflammation is characterized by heightened pain sensitivity. This hyperalgesia is the consequence of the release of inflammatory mediators, cytokines and growth factors. A key participant is the induction of the neurotrophin nerve growth factor (NGF) by interleukin‐1β (IL‐1β).

Tumour necrosis factor α (TNFα) has been shown both to produce hyperalgesia and to upregulate IL‐1β. We have now examined whether the induction of TNFα in inflammatory lesions contributes to inflammatory sensory hypersensitivity by inducing IL‐1β and NGF.

The intraplantar injection of complete Freund's adjuvant (CFA) in adult rats produced a localized inflammation of the hindpaw with a rapid (3 h) reduction in withdrawal time in the hot plate test and in the mechanical threshold for eliciting the flexion withdrawal reflex.

The CFA‐induced inflammation resulted in significant elevation in the levels of TNFα, IL‐1β and NGF in the inflamed paw. In the case of TNFα, an elevation was detected at 3 h, rose substantially at 6 h, peaked at 24 h and remained elevated at 5 days, with similar but smaller changes in the contralateral non‐inflamed hindpaw. No increase in serum TNFα was detected at 24 h post CFA injection.

Intraplantar recombinant murine TNFα injections produce a short‐lived (3–6 h) dose‐dependent (50–500 ng) increase in thermal and mechanical sensitivity which was significantly attenuated by prior administration of anti‐NGF antiserum.

Intraplantar TNFα (100–500 ng) also elevated at 6 but not 48 h the levels of IL‐1β and NGF in the hindpaw.

A single injection of anti‐TNFα antiserum, 1 h before the CFA, at a dose sufficient to reduce the effects of a 100 ng intraplantar injection of TNFα, significantly delayed the onset of the resultant inflammatory hyperalgesia and reduced IL‐1β but not NGF levels measured at 24 h.

The elevation of TNFα in inflammation, by virtue of its capacity to induce IL‐1β and NGF, may contribute to the initiation of inflammatory hyperalgesia.
Cardiovascular effects of a low‐dose combination of ramipril and felodipine in spontaneously hypertensive rats
Authors: Mervaala Eero M. A. , Teräväinen Terttu‐Liisa , Malmberg Lena , Laakso Juha , Vapaatalo Heikki , Karppanen Heikki
Published: July 2006

Cardiovascular effects of submaximal antihypertensive doses of the angiotensin converting enzyme inhibitor, ramipril (0.25 mg kg−1 day−1 in the food), and the calcium channel blocker, felodipine (0.4 mg kg−1 day−1 subcutaneously by osmotic minipump), both alone and in combination, were examined in spontaneously hypertensive rats (SHR) in a four‐week study.

Both ramipril and felodipine as monotherapy decreased systolic blood pressure. The antihypertensive effect of the drug combination was more than that of ramipril treatment alone, but not significantly better than that of felodipine monotherapy. Ramipril or felodipine treatments did not significantly affect the heart rate, either alone or in combination.

The beneficial effect of ramipril monotherapy on left ventricular hypertrophy was more prominent than that of felodipine. The cardioprotective effect of felodipine was improved when combined to ramipril. The systolic blood pressure at the end of the experimental period correlated only weakly with left ventricular hypertrophy.

Responses of mesenteric arterial rings in vitro were examined at the end of the four‐week study. Ramipril and felodipine monotherapies as well as their combination markedly improved the endothelium‐dependent vascular relaxation responses to acetylcholine. The combination of ramipril and felodipine slightly enhanced the endothelium‐independent vascular relaxation responses to sodium nitroprusside. Ramipril treatment alone slightly diminished the vascular contractile responses to noradrenaline. Neither ramipril nor felodipine alone or in combination affected the vascular contractile responses to potassium chloride.

Ramipril treatment, both alone and in combination with felodipine, caused a three fold increase in plasma renin activity. Serum aldosterone, fasting blood glucose level, serum insulin and the 24 hour urinary excretions of sodium, potassium, magnesium, calcium, phosphorus or protein were not significantly affected by the drug treatments.

Our findings suggest that a better overall control of hypertension and end‐organ damages, without an increase in adverse effects, can be achieved by the combination of submaximal antihypertensive doses of felodipine and ramipril than by monotherapy with either drug alone.
Inhibitory responses to exogenous adenosine in murine proximal and distal colon
Authors: Zizzo Maria Grazia , Mulè Flavia , Serio Rosa
Published: July 2006

The aims of the present study were firstly, to characterize pharmacologically the subtypes of P1purinoreceptors involved in the inhibitory effects induced by exogenous adenosine in longitudinal smooth muscle of mouse colon, and secondly, to examine differences in the function and distribution of these receptors between proximal and distal colon.

Adenosine (100 μM–3 mM) caused a concentration‐dependent reduction of the amplitude of spontaneous contractions in the proximal colon, and muscular relaxation in the distal colon. In the proximal colon, adenosine effects were antagonized by a selective A1receptor antagonist, 1,3‐dipropyl‐8‐cyclopentylxanthine (DPCPX, 10 nM), but were not modified by 3,7‐dimethyl‐1‐propargylxanthine (DMPX, 10 μM) or by 9‐chloro‐2‐(2‐furanyl)‐5‐((phenylacetyl)amino)‐ [1,2,4]triazolo[1,5‐c]quinazoline (MRS 1220, 0.1 μM), selective A2and A3receptor antagonists, respectively. In the distal colon, adenosine effects were antagonized by DPCPX, DMPX, and by a selective A2Breceptor antagonist, 8‐[4‐[((4‐cyanophenyl)carbamoylmethyl)oxy]phenyl]‐1,3‐di(n‐propyl) xanthine (MRS 1754, 10 μM), but not by 8‐(3‐chlorostyryl)‐caffeine (CSC, 10 μM), a selective A2Areceptor antagonist, or by MRS 1220.

Tetrodotoxin (TTX 1 μM), the nitric oxide (NO) synthase inhibitor,N‐nitro‐L‐arginine methyl ester (L‐NAME, 100 μM), or 1H‐[1,2,4]oxadiazolo[4,3‐a]quinoxalin‐1‐one (10 μM), an inhibitor of soluble guanylyl cyclase, reduced adenosine effects only in distal colon. In addition,L‐NAME induced a further reduction of adenosine relaxation in the presence of DPCPX, but not in the presence of MRS 1754.

From these results we conclude that, in the murine proximal colon, adenosine induces inhibitory effectsviaTTX‐insensitive activation of A1receptor. In the distal colon, adenosine activates both A1and A2Breceptors, the latter located on enteric inhibitory neurons releasing NO.

British Journal of Pharmacology(2006)148, 956–963. doi:10.1038/sj.bjp.0706808
Protective effects of relaxin in ischemia/reperfusion‐induced intestinal injury due to splanchnic artery occlusion
Authors: Masini Emanuela , Cuzzocrea Salvatore , Mazzon Emanuela , Muià Carmelo , Vannacci Alfredo , Fabrizi Francesca , Bani Daniele
Published: July 2006

Splanchnic artery occlusion (SAO) followed by reperfusion causes endothelial injury and inflammation which contribute to the pathophysiology of shock. We investigated the effects of relaxin (RLX), known to afford protection against the deleterious effects of cardiac ischemia/reperfusion, given to rats subjected to splanchnic artery occlusion and reperfusion (SAO/R)‐induced splanchnic injury.

RLX (30 ng kg−1, 15 min. before reperfusion) significantly reduced the drop of blood pressure and high mortality rate caused by SAO/R. RLX also reduced histopathological changes, leukocyte infiltration (myeloperoxidase) and expression of endothelial cell adhesion molecules in the ileum. RLX counteracted free radical‐mediated tissue injury, as judged by significant decrease in the tissue levels of peroxidation and nitration products (malondialdehyde, nitrotyrosine), DNA damage markers (8‐hydroxy‐2′‐deoxyguanosine, poly‐ADP‐ribosylated DNA) and consumption of tissue antioxidant enzymes (superoxide dismutase). As a result, RLX led to a reduction of ileal cell apoptosis (caspase 3, terminal deoxynucleotidyltransferase‐mediated UTP end labeling). The effects of RLX appear specific, as inactivated RLX substituted for the bioactive hormone had no effects.

In conclusion, these results show that RLX exerts a clear‐cut protective effect in SAO/R‐induced splanchnic injury, likely due to endothelial protection, decreased leukocyte recruitment and hindrance of free radical‐mediated tissue injury leading to cell death, lethal complications and high mortality rate. Thus, RLX could be used therapeutically in intestinal ischemia.

British Journal of Pharmacology (2006) 148, 1124–1132. doi:10.1038/sj.bjp.0706811
Quantifying the association and dissociation rates of unlabelled antagonists at the muscarinic M3receptor
Authors: Dowling Mark R , Charlton Steven J
Published: July 2006

Slow receptor dissociation kinetics has been implicated in the long clinical duration of action of the muscarinic receptor antagonist tiotropium. However, despite the potential benefits of new drugs with slow dissociation kinetics, the rate parameters of new compounds are seldom measured due to technical difficulties and financial implications associated with radiolabeling multiple ligands. Here we describe the development and optimisation of a medium throughput assay which is capable of measuring the kinetic parameters of novel, unlabelled compounds.

Radioligand binding studies were performed with membranes derived from CHO cells recombinantly expressing the human M3muscarinic receptor.

Initial characterisation of the radioligand [3H]‐NMS yielded on and off rates of 4.1±0.2 × 108 M−1 min−1and 0.015±0.0005 min−1, respectively.

The specific binding of [3H]‐NMS was measured over time in the presence and absence of several concentrations of unlabelled competitor compounds. These data were analysed using a competition kinetic model to provide on and off rates for the unlabelled competitor. Comparison of the kinetically derived Kd (koff/kon) withKivalues generated at equilibrium showed an excellent correlation (r2=0.99), providing good validation of the method.

The on and off rates were also used in theoretical computer simulations to successfully predict the effect of incubation time on apparent IC50values.

This study demonstrates that a medium‐throughput competition kinetic binding assay can be used to determine accurate on and off rates of unlabelled compounds, providing the opportunity to optimise for kinetic parameters early in the drug discovery process.

British Journal of Pharmacology(2006)148, 927–937. doi:10.1038/sj.bjp.0706819
Lipid lowering effects of Momordica charantia (Bitter Melon) in HIV‐1‐protease inhibitor‐treated human hepatoma cells, HepG2
Authors: Nerurkar Pratibha V , Lee Yun Kyung , Linden Ellen H , Lim Steven , Pearson Laurel , Frank Jennifer , Nerurkar Vivek R
Published: July 2006

Hyperlipidemic effects of HIV‐1‐protease inhibitors (PI) are associated with increased hepatic production of triglyceride (TG)‐rich lipoproteins, rather than lipoprotein clearance. PI are known to increase apolipoprotein B (apoB) secretion, apoC‐III mRNA expression and decrease apoA‐1 secretion. Nutritional therapy remains an important strategy to manage PI‐associated hyperlipidemia.

This study investigated the in vitro efficacy of Asian vegetable, Momordica charantia or bitter melon (BM) to ameliorate PI‐associated apoB and lipid abnormalities in HepG2 cells.

Our study demonstrates that bitter melon juice (BMJ) significantly reduced apoB secretion and apoC‐III mRNA expression and normalized apoA‐I expression in PI‐treated HepG2 cells. BMJ also significantly reduced cellular TG and microsomal TG transfer protein, suggesting that lipid bioavailability and lipidation of apoB assembly may play a role in decreased apoB secretion.

Identifying molecular targets of BM may offer alternative dietary strategies to decrease PI‐associated hyperlipidemia and improve quality of life among HIV‐1‐infected patients.

British Journal of Pharmacology (2006) 148, 1156–1164. doi:10.1038/sj.bjp.0706821
Investigation on the relationship between cannabinoid CB1and opioid receptors in gastrointestinal motility in mice
Authors: Carai Mauro A M , Colombo Giancarlo , Gessa Gian Luigi , Yalamanchili Ratnakumar , Basavarajppa Balapal S , Hungund Basalingappa L
Published: July 2006

This study investigated whether (a) cannabinoid CB1receptor knockout (CB1−/−) mice displayed altered gastrointestinal transit and (b) cannabinoid CB1and opioid receptors functionally interact in the regulation of gastrointestinal transit.

Gastrointestinal transit was assessed by the Whole Gastrointestinal Transit, measuring the excretion time of an intragastrically administered marker (whole intestine), and the Upper Gastrointestinal Transit, measuring the distance covered by the marker in the small intestine.

CB1−/−and homozygous CB1+/+(CB1+/+) mice did not differ in both whole gut and small intestine transit. CB1−/−and CB1+/+mice were equally responsive to the inhibitory effect of morphine (10 mg kg−1) and loperamide (3 mg kg−1) on whole gut transit.

Additionally, in CD1 mice the cannabinoid CB1receptor antagonist, rimonabant (0–0.5 mg kg−1), failed to block the inhibitory effect of morphine (0–1.25 mg kg−1) and loperamide (0–0.5 mg kg−1) on transit in small and whole intestine. Similarly, the opioid receptor antagonists, naloxone (0–1 mg kg−1) and naltrexone (0–10 mg kg−1), failed to block the inhibitory effect of the cannabinoid WIN 55,212‐2 (0–3 mg kg−1) on transit in small and whole intestine.

These results suggest that (a) compensatory mechanisms likely developed in CB1−/−mice to overcome the lack of inhibitory function of endocannabinoid system; (b) cannabinoid and opioid receptor systems did not interact in regulating gastrointestinal transit in mice.

British Journal of Pharmacology(2006)148, 1043–1050. doi:10.1038/sj.bjp.0706824
Neuropeptide B and W: neurotransmitters in an emerging G‐protein‐coupled receptor system
Authors: Singh Gurminder , Davenport Anthony P
Published: July 2006

Deorphanised G‐protein‐coupled receptors represent new and expanding targets for drug development. Neuropeptide B (NPB) and W (NPW) have recently been identified as the cognate endogenous ligands for the orphan receptor GPR7, now designated as NPBW1. NPB and NPW also bound to a second related orphan receptor, GPR8, now designated as NPBW2 that is present in humans but not rats or mice. In humans, high levels of NPW mRNA have been visualised in the substantia nigra, whereas moderate expression levels have been detected in the amygdala and hippocampus. In peripheral tissues, expression of NPW mRNA has been confirmed in the progenital system, comprising the kidney, testis, uterus, ovary and placenta, and also in stomach homogenates. Immunocytochemical, molecular biological and autoradiography techniques have revealed a discrete CNS distribution for NPBW1 in human, mouse and rat. Highest expression of NPBW1 mRNA and protein was identified in the amygdala and hypothalamic nuclei known to regulate feeding behaviour. [125I]‐NPW bound with a single high affinity to rat amygdala, KD=0.44 nM and 150 fmol mg−1 protein. Physiological studies demonstrate that intracerebroventricular infusion of NPBW1 ligands modulates feeding behaviour, regulates the release of corticosterone, prolactin and growth hormone while also manipulating pain pathway. Mouse knockout models of the gene encoding either NPB or NPBW1 have a gender‐specific phenotype, with moderate obesity evident in males but not females. Further investigation is required to elucidate the precise physiological role of NPB and NPW as neurotransmitters.

British Journal of Pharmacology (2006) 148, 1033–1041. doi:10.1038/sj.bjp.0706825
Vitamin A active metabolite, all‐trans retinoic acid, induces spinal cord sensitization. I. Effects after oral administration
Authors: Romero‐Sandoval E A , Molina C , Alique M , Moreno‐Manzano V , Lucio F J , Herrero J F
Published: July 2006

Background and purpose:

Retinoic acid is an active metabolite of vitamin A involved in the modulation of the inflammatory and nociceptive responses. The aim of the present study was to analyze the properties of spinal cord neuronal responses of male Wistar rats treated with all‐trans retinoic acid (ATRA) p.o. in the normal situation and under carrageenan‐induced inflammation. We also studied the expression and distribution of cyclooxygenases (COX) in the spinal cord.

Experimental approach:

Properties of spinal cord neurons were studied by means of the single motor unit technique. The expression of COX enzymes in the spinal cord was assessed by Western blot analysis and immunohistochemistry.

Key results:

Intensity thresholds for mechanical and electrical stimulation (C‐fibers) were significantly lower in animals treated with ATRA than vehicle, either in normal rats or in rats with inflammation. The size of cutaneous receptive fields was also larger in animals treated with ATRA in the normal and inflammatory conditions. The expression of COX‐2 enzyme, but not COX‐1, was significantly higher in animals treated with ATRA. COX‐2 labeling was observed in dorsal horn cells and in ventral horn motoneurons.

Conclusions and implications:

In conclusion, the oral treatment with ATRA in rats induces a sensitization‐like effect on spinal cord neuronal responses similar to that observed in animals with inflammation and might explain the enhancement of allodynia and hyperalgesia observed in previously published behavioral experiments. The mechanism of action involves an over‐expression of COX‐2, but not COX‐1, in dorsal and ventral horn areas of the lumbar spinal cord.

British Journal of Pharmacology (2006) 149, 56–64. doi:10.1038/sj.bjp.0706829
Identification of a nuclear-localized nuclease from wheat cells undergoing programmed cell death that is able to trigger DNA fragmentation and apoptotic morphology on nuclei from human cells
Authors: Domínguez Fernando , Cejudo Francisco J.
Published: July 2006

PCD (programmed cell death) in plants presents important morphological and biochemical differences compared with apoptosis in animal cells. This raises the question of whether PCD arose independently or from a common ancestor in plants and animals. In the present study we describe a cell-free system, using wheat grain nucellar cells undergoing PCD, to analyse nucleus dismantling, the final stage of PCD. We have identified a Ca2+/Mg2+ nuclease and a serine protease localized to the nucleus of dying nucellar cells. Nuclear extracts from nucellar cells undergoing PCD triggered DNA fragmentation and other apoptotic morphology in nuclei from different plant tissues. Inhibition of the serine protease did not affect DNA laddering. Furthermore, we show that the nuclear extracts from plant cells triggered DNA fragmentation and apoptotic morphology in nuclei from human cells. The inhibition of the nucleolytic activity with Zn2+ or EDTA blocked the morphological changes of the nucleus. Moreover, nuclear extracts from apoptotic human cells triggered DNA fragmentation and apoptotic morphology in nuclei from plant cells. These results show that degradation of the nucleus is morphologically and biochemically similar in plant and animal cells. The implication of this finding on the origin of PCD in plants and animals is discussed.
In-frame deletion in the EGF receptor alters kinase inhibition by gefitinib
Authors: Sakai Kazuko , Yokote Hideyuki , Murakami-Murofushi Kimiko , Tamura Tomohide , Saijo Nagahiro , Nishio Kazuto
Published: July 2006

The existence of an in-frame deletion mutant correlates with the sensitivity of lung cancers to EGFR (epidermal growth factor receptor)-targeted tyrosine kinase inhibitors. We reported previously that the in-frame 15-bp deletional mutation (delE746–A750 type deletion) was constitutively active in cells. Kinetic parameters are important for characterizing an enzyme; however, it remains unclear whether the kinetic parameters of deletion mutant EGFR are similar to those of wild-type EGFR. We analysed autophosphorylation in response to ATP and inhibition of gefitinib for deletion mutant EGFR and wild-type EGFR. Kinetic studies, examining autophosphorylation, were carried out using EGFR fractions extracted from 293-pΔ15 and 293-pEGFR cells transfected with deletion mutant EGFR and wild-type EGFR respectively. We demonstrated the difference in activities between unstimulated wild-type (Km for ATP=4.0±0.3 μM) and mutant EGFR (Km for ATP=2.5±0.2 μM). There was no difference in Km values between EGF-stimulated wild-type EGFR (Km for ATP=1.9±0.1 μM) and deletion mutant EGFR (Km for ATP=2.2±0.2 μM). These results suggest that mutant EGFR is active without ligand stimulation. The Ki value for gefitinib of the deletion mutant EGFR was much lower than that of wild-type EGFR. These results suggest that the deletion mutant EGFR has a higher affinity for gefitinib than wild-type EGFR.
Structure–function relationships for Schizophyllum commune trehalose phosphorylase and their implications for the catalytic mechanism of family GT-4 glycosyltransferases
Authors: Goedl Christiane , Griessler Richard , Schwarz Alexandra , Nidetzky Bernd
Published: July 2006

The cDNA encoding trehalose phosphorylase, a family GT-4 glycosyltransferase from the fungus Schizophyllum commune, was isolated and expressed in Escherichia coli to yield functional recombinant protein in its full length of 737 amino acids. Unlike the natural phosphorylase that was previously obtained as a truncated 61 kDa monomer containing one tightly bound Mg2+, the intact enzyme produced in E. coli is a dimer and not associated with metal ions [Eis, Watkins, Prohaska and Nidetzky (2001) Biochem. J. 356, 757–767]. MS analysis of the slow spontaneous conversion of the full-length enzyme into a 61 kDa fragment that is fully active revealed that critical elements of catalysis and specificity of trehalose phosphorylase reside entirely in the C-terminal protein part. Intact and truncated phosphorylases thus show identical inhibition constants for the transition state analogue orthovanadate and α,α-trehalose (Ki≈1 μM). Structure-based sequence comparison with retaining glycosyltransferases of fold family GT-B reveals a putative active centre of trehalose phosphorylase, and results of site-directed mutagenesis confirm the predicted crucial role of Asp379, His403, Arg507 and Lys512 in catalysis and also delineate a function of these residues in determining the large preference of the wild-type enzyme for the phosphorolysis compared with hydrolysis of α,α-trehalose. The pseudo-disaccharide validoxylamine A was identified as a strong inhibitor of trehalose phosphorylase (Ki=1.7±0.2 μM) that displays 350-fold tighter binding to the enzyme–phosphate complex than the non-phosphorolysable substrate analogue α,α-thio-trehalose. Structural and electronic features of the inhibitor that may be responsible for high-affinity binding and their complementarity to an anticipated glucosyl oxocarbenium ion-like transition state are discussed.