A 2,3-diphenylpyrido[1,2-a] pyrimidin-4-one derivative inhibits specific angiogenic factors induced by TNF-\u3b1

Low-grade chronic inflammation is a key process of angiogenesis in tumour progression. We investigated whether a synthetic analogue of apigenin, the 2-(3,4-dimethoxyphenyl)-3-phenyl-4H-pyrido[1,2-a] pyrimidin-4-one (called DB103), interfered with the mechanisms involved in the angiogenic process induced by the inflammatory cytokine tumour necrosis factor (TNF\u3b1). In endothelial cells, DB103 but not apigenin reduced the TNF\u3b1-induced oxidative stress. DB103 inhibited the activation of ERK1/2 but not JNK, p38 and Akt kinases, while apigenin was not so selective because it inhibited essentially all examined kinases. Similarly, apigenin inhibited the TNF\u3b1-induced transcription factors CREB, STAT3, STAT5 and NF-kB, while DB103 acted only on NFinhibited the induced-release of angiogenic factors such as monocyte chemotactic protein-1, interleukin-6 (IL-6) and angiopoietin-2 but not IL-8, while apigenin reduced the IL-6 and IL-8 release. DB103 revealed a better ability than apigenin to modulate proangiogenic responses induced by an inflammatory microenvironment

Enteric dysfunctions in experimental Parkinson's disease: alterations of excitatory cholinergic neurotransmission regulating colonic motility in rats

Parkinson's disease (PD) is frequently associated with gastrointestinal symptoms, mostly represented by constipation and defecatory dysfunctions. This study examined the impact of central dopaminergic denervation, induced by injection of 6-hydroxydopamine (6-OHDA) into the medial forebrain bundle, on distal colonic excitatory cholinergic neuromotor activity in rats. Animals were euthanized 4 and 8 weeks after 6-OHDA injection. In vivo colonic transit was evaluated by radiological assay. Electrically and carbachol-induced cholinergic contractions were recorded in vitro from longitudinal and circular muscle colonic preparations, while acetylcholine levels were assayed in their incubation media. Choline acetyltransferase (ChAT), HuC/D (pan-neuronal marker), muscarinic M2 and M3 receptors. As compared with control rats, at week 4 6-OHDA-treated animals displayed the following changes: decreased in vivo colonic transit rate; impaired electrically evoked neurogenic cholinergic contractions; enhanced carbachol-induced contractions; decreased basal and electrically stimulated acetylcholine release from colonic tissues; decreased ChAT immunopositivity in the neuromuscular layer; unchanged density of HuC/D immunoreactive myenteric neurons; increased expression of colonic muscarinic M2 and M3 receptors. The majority of such alterations were detected also at week 8 post-6-OHDA injection. These findings indicate that central nigrostriatal dopaminergic denervation is associated with an impaired excitatory neurotransmission characterized by a loss of myenteric neuronal ChAT positivity and decrease in acetylcholine release, resulting in a dysregulated smooth muscle motor activity, which likely contributes to the concomitant decrease in colonic transit rate

Alteration of colonic excitatory tachykininergic motility and enteric inflammation following dopaminergic nigrostriatal neurodegeneration

Background: Parkinson's disease (PD) is frequently associated with gastrointestinal (GI) symptoms, including constipation and defecatory dysfunctions. The mechanisms underlying such disorders are still largely unknown, although the occurrence of a bowel inflammatory condition has been hypothesized. This study examined the impact of central dopaminergic degeneration, induced by intranigral injection of 6-hydroxydopamine (6-OHDA), on distal colonic excitatory tachykininergic motility in rats. Methods: Animals were euthanized 4 and 8 weeks after 6-OHDA injection. Tachykininergic contractions, elicited by electrical stimulation or exogenous substance P (SP), were recorded in vitro from longitudinal muscle colonic preparations. SP, tachykininergic NK1 receptor, and glial fibrillary acidic protein (GFAP) expression, as well as the density of eosinophils and mast cells in the colonic wall, were examined by immunohistochemical analysis. Malondialdehyde (MDA, colorimetric assay), TNF, and IL-1 beta (ELISA assay) levels were also examined. The polarization of peritoneal macrophages was evaluated by real-time PCR. Results: In colonic preparations, electrically and SP-evoked tachykininergic contractions were increased in 6-OHDA rats. Immunohistochemistry displayed an increase in SP and GFAP levels in the myenteric plexus, as well as NK1 receptor expression in the colonic muscle layer of 6-OHDA rats. MDA, TNF, and IL-1 beta levels were increased also in colonic tissues from 6-OHDA rats. In 6-OHDA rats, the number of eosinophils and mast cells was increased as compared with control animals, and peritoneal macrophages polarized towards a pro-inflammatory phenotype. Conclusions: The results indicate that the induction of central nigrostriatal dopaminergic degeneration is followed by bowel inflammation associated with increased oxidative stress, increase in pro-inflammatory cytokine levels, activation of enteric glia and inflammatory cells, and enhancement of colonic excitatory tachykininergic motility

Exploring the genetics of irritable bowel syndrome: A GWA study in the general population and replication in multinational case-control cohorts

OBJECTIVE: IBS shows genetic predisposition, but adequately powered gene-hunting efforts have been scarce so far. We sought to identify true IBS genetic risk factors by means of genome-wide association (GWA) and independent replication studies. DESIGN: We conducted a GWA study (GWAS) of IBS in a general population sample of 11\u2005326 Swedish twins. IBS cases (N=534) and asymptomatic controls (N=4932) were identified based on questionnaire data. Suggestive association signals were followed-up in 3511 individuals from six case-control cohorts. We sought genotype-gene expression correlations through single nucleotide polymorphism (SNP)-expression quantitative trait loci interactions testing, and performed in silico prediction of gene function. We compared candidate gene expression by real-time qPCR in rectal mucosal biopsies of patients with IBS and controls. RESULTS: One locus at 7p22.1, which includes the genes KDELR2 (KDEL endoplasmic reticulum protein retention receptor 2) and GRID2IP (glutamate receptor, ionotropic, delta 2 (Grid2) interacting protein), showed consistent IBS risk effects in the index GWAS and all replication cohorts and reached p=9.31 710(-6) in a meta-analysis of all datasets. Several SNPs in this region are associated with cis effects on KDELR2 expression, and a trend for increased mucosal KDLER2 mRNA expression was observed in IBS cases compared with controls. CONCLUSIONS: Our results demonstrate that general population-based studies combined with analyses of patient cohorts provide good opportunities for gene discovery in IBS. The 7p22.1 and other risk signals detected in this study constitute a good starting platform for hypothesis testing in future functional investigations. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions

L-histidine decarboxylase decreases its own transcription through downregulation of ERK activity

A poorly defined negative feedback loop decreases transcription of theL-histidine decarboxylase (HDC) gene. To help understand this regulation, we have studied the effect of HDC protein expression on HDC gene transcription in transfected AGS-B cells. Expression of the rat HDC protein inhibited HDC promoter activity in a dose-dependent fashion. The region of the HDC promoter mediating this inhibitory effect corresponded to a previously defined gastrin and extracellular signal-related kinase (ERK)-1 response element. Overexpression of the HDC protein reduced nuclear factor binding in this region. Experiments employing specific histamine receptor agonists indicated that the inhibitory effect was not dependent on histamine production, and studies with the HDC inhibitor \u3b1-fluoromethylhistidine revealed that inhibition was unrelated to enzyme activity. Instead, an enzymatically inactive region at the amino terminal of the HDC enzyme (residues 1\u2013271) was shown to mediate inhibition. Fluorescent chimeras containing this domain were not targeted to the nucleus, arguing against specific inhibition of the HDC transcription machinery. Instead, we found that overexpression of HDC protein decreased ERK protein levels and ERK activity and that the inhibitory effect of HDC protein could be overcome by overexpression of ERK1. These data suggest a novel feedback-inhibitory role for amino terminal sequences of the HDC protei

H3 receptor-mediated inhibition of intestinal acetylcholine release: pharmacological characterization of signal transduction pathways

The present study investigates the mechanisms through which prejunctional histamine H3 receptors modulate intestinal cholinergic neurotransmission. The experiments were performed on longitudinal muscle-myenteric plexus preparations of guinea pig ileum, preincubated with [3H]choline, superfused with physiological salt solution containing hemicholinium-3, and subjected to electrical field stimulation. The stimulation-induced outflow of radioactivity was taken as an index of endogenous acetylcholine release. The electrically induced [3H]acetylcholine release was inhibited by histamine (EC50)=33.5 nM) or the H3 receptor agonist R-alpha-methylhistamine (EC50=41.6 nM), whereas it was not affected by pyridylethylamine (H1 agonist), impromidine (H2 agonist), pyrilamine (H1 antagonist), cimetidine (H2 antagonist), thioperamide or clobenpropit (H3 antagonists). The inhibitory effects of histamine or R-alpha-methylhistamine were antagonized by thioperamide (pKd= 8.31 and 8.53, respectively) or clobenpropit (pKd=9.44 and 9.32, respectively), but not by pyrilamine or cimetidine. The modulatory action of histamine on the evoked tritium outflow was attenuated by pertussis toxin and abolished by N-ethylmaleimide, two selective blockers of Gi/Go proteins. Tetraethylammonium or 4-aminopyridine, acting as inhibitors of voltage-dependent K+ channels, enhanced the evoked tritium outflow when tested alone, and apparently counteracted the inhibitory effect of histamine. However, the blocking actions of tetraethylammonium and 4-aminopyridine were no longer evident when their enhancing actions were compensated by appropriate reductions of Ca2+ concentration in the superfusion medium. Histamine-induced inhibition of evoked tritium output was enhanced by omega-conotoxin, a selective blocker of N-type Ca2+ channels, or low Ca2+ concentration, whereas it was not modified by nifedipine, an antagonist of L-type Ca2+ channels. In addition, the inhibitory effect of histamine was not significantly affected by forskolin (activator of adenylyl cyclase), 8-bromo-cyclic AMP (a stable analog of cyclic AMP), rolipram (a selective blocker of type IV phosphodiesterase), phorbol myristate acetate (activator of protein kinase C), H-89 (N-(2-[p-bromocinnamylamino]ethyl)-5-isoquinolinesulfonamide, inhibitor of protein kinase A), Ro-31-8220 (2-(1-[3-(amidinothio)propyl]-1H-indol-3-yl)-3-(1-methylindol-3-yl)-maleimide, inhibitor of protein kinase C), KT5823 (N-methyl-(8R*,9S*,11S*)-(-)-9-methoxy-9-methoxycarbonyl-8-methyl-2,3,9,10-tetrahydro-8,11-epoxy-1H,8H,11H-2,7b,11a-triazadibenzo [a,g]cycloocta[c,d,e]-trinden-1-one, inhibitor of protein kinase G), or lavendustin A (inhibitor of tyrosine kinase). The present results indicate that histamine inhibits intestinal cholinergic neurotransmission through presynaptic H3 receptors coupled to Gi/Go proteins. It is suggested that adenylyl cyclase, serine-threonine protein kinase and tyrosine kinase pathways are not implicated in this regulatory action, and that Gi/Go proteins modulate the activity of N-type Ca2+ channels through a direct link, thus causing a reduced availability of extracellular Ca2+ at the level of ileal cholinergic nerve terminals

Central administration of cholecystokinin stimulates gastric pepsinogen secretion from anaesthetized rats.

Intracerebroventricular administration of CCK-8S was associated with a stimulation of gastric pepsinogen secretion from anaesthetized rats; similar effects were induced by CCK-8S given intravenously. The excitatory effect of intracerebroventricular CCK-8S was not modified by central injection of L-364,718 or L-365,260, whereas both these antagonists, given by intravenous route, prevented the pepsigogue action of parenteral CCK-8S. Intracerebroventricular or intravenous CCK-8S also increased basal acid secretion, this latter effect being prevented by parenteral L-365,260 but not L-364,718. It is suggested that centrally applied CCK-8S evokes pepsinogen secretion through the activation of peripheral CCK-A and CCK-B receptors

Efficay and tolerability of meloxicam, a COX-2 preferential nonsteroidal anti-inflammatory drug: a review

Meloxicam is an enolcarboxamide with preferential COX-2 inhibitory activity. In vitro studies with human tissues have confirmed the high affinity of meloxicam for COX-2, whereas COX-1 was inhibited only at the highest concentrations (ratio of 50% inhibitory concentration for COX-2 : COX-1 = 0.09 in whole blood assays).Meloxicam has a bioavailability of 89% after oral administration, is strongly bound to plasma proteins, and its half-life is 20-24 hours. It readily penetrates into synovial fluid, reaching 45-57% of plasma concentrations. Meloxicam pharmacokinetics are not significantly altered in elderly patients or in those with mild renal/hepatic impairment. The efficacy and tolerability of meloxicam in the treatment of pain and inflammation associated with rheumatic and musculoskeletal disorders has been evaluated in numerous studies comparing meloxicam 7.5-15 mg/day (up to 22.5 mg/day in ankylosing spondylitis), administered for 2 weeks to 12 months, with placebo or other nonsteroidal anti-inflammatory drugs (NSAIDs). Overall, the efficacy of meloxicam was significantly superior to that of placebo and similar to that of other NSAIDs, whereas the incidence of adverse events (especially gastrointestinal) was lower than with other NSAIDs. Intramuscular meloxicam provided faster pain relief than the oral drug in patients with rheumatoid arthritis. Meloxicam also appears to be effective in the prevention of postoperative pain, as shown in patients undergoing abdominal hysterectomy or inguinal hernia repair. Meloxicam may also have a cardioprotective role: in patients with acute coronary syndrome without ST-segment elevation, a lower incidence of cardiovascular events was observed in those who received meloxicam plus aspirin and heparin versus aspirin and heparin alone, both during coronary care stay and at 90-day follow-up. Meloxicam has demonstrated a favourable tolerability profile in large-scale comparative trials, where its gastrointestinal tolerability was superior to that of nonselective NSAIDs. In particular, meloxicam 7.5 mg/day was associated with a lower incidence of gastrointestinal adverse events compared with diclofenac (13% vs 19%; p < 0.001) or piroxicam (10.3% vs 15.4%; p < 0.001). This was confirmed by a prescription-event monitoring study that found a relatively low incidence of dyspepsia, upper gastrointestinal haemorrhage and peptic ulcer (28.3, 0.4 and 0.3 per 1000 patient-months, respectively) among first-time users of meloxicam. In conclusion, meloxicam is at least as effective as nonselective NSAIDs in the treatment of rheumatic disease or postoperative pain, but has a more favourable gastrointestinal tolerability profile. Further investigations into the potential role of meloxicam as a cardioprotective agent are warranted

Isotopic effects in sub-barrier fusion of Si plus Si systems

Background: Recent measurements of fusion cross sections for the 28 Si + 28 Si system revealed a rather unsystematic behavior; i.e., they drop faster near the barrier than at lower energies. This was tentatively attributed to the large oblate deformation of 28 Si because coupled-channels (CC) calculations largely underestimate the 28 Si + 28 Si cross sections at low energies, unless a weak imaginary potential is applied, probably simulating the deformation. 30 Si has no permanent deformation and its low-energy excitations are of a vibrational nature. Previous measurements of this system reached only 4 mb, which is not sufficient to obtain information on effects that should show up at lower energies. Purpose: The aim of the present experiment was twofold: (i) to clarify the underlying fusion dynamics by measuring the symmetric case 30 Si + 30 Si in an energy range from around the Coulomb barrier to deep sub-barrier energies, and (ii) to compare the results with the behavior of 28 Si + 28 Si involving two deformed nuclei. Methods: 30 Si beams from the XTU tandem accelerator of the Laboratori Nazionali di Legnaro of the Istituto Nazionale di Fisica Nucleare were used, bombarding thin metallic 30 Si targets (50 µ g / cm 2 ) enriched to 99 . 64% in mass 30. An electrostatic beam deflector allowed the detection of fusion evaporation residues (ERs) at very forward angles, and angular distributions of ERs were measured. Results: The excitation function of 30 Si + 30 Si was measured down to the level of a few microbarns. It has a regular shape, at variance with the unusual trend of 28 Si + 28 Si. The extracted logarithmic derivative does not reach the L CS limit at low energies, so that no maximum of the S factor shows up. CC calculations were performed including the low-lying 2 + and 3 - excitations. Conclusions: Using a Woods-Saxon potential the experimental cross sections at low energies are overpredicted, and this is a clear sign of hindrance, while the calculations performed with a M3Y + repulsion potential nicely fit the data at low energies, without the need of an imaginary potential. The comparison with the results for 28 Si + 28 Si strengthens the explanation of the oblate shape of 28 Si being the reason for the irregular behavior of that system.Peer Reviewe