bluuebunny/crossref_metadata_2025 huggingface dataset:
Stereoselective uptake of β‐lactam antibiotics by the intestinal peptide transporter
Authors: Wenzel Uwe , Thwaites David T. , Daniel Hannelore
Published: July 2012

The stereoselective transport of β‐lactam antibiotics has been investigated in the human intestinal epithelial cell line, Caco‐2, by use of D‐ and L‐enantiomers of cephalexin and loracarbef as substrates.

The L‐isomers of cephalexin, loracarbef and dipeptides displayed a higher affinity for the oligopeptide/H+‐symporter in Caco‐2 cells than the D‐isomers. This was demonstrated by inhibition of the influx of the β‐lactam, [3H]‐cefadroxil.

By measurement of the substrate‐induced intracellular acidification in Caco‐2 cells loaded with the pH‐sensitive fluorescent dye BCECF (2′,7′‐bis(2‐carboxyethyl)‐5‐(6)‐carboxy‐fluorescein), it was demonstrated for the first time that L‐isomers of β‐lactams not only bind to the peptide transporter with high affinity but are indeed transported.

Efficient proton‐coupled transport of L‐β‐lactam antibiotics was also shown to occur in Xenopus laevis oocytes expressing the cloned peptide transporter PepTl from rabbit small intestine.

Both cell systems therefore express a stereoselective transport pathway for β‐lactam antibiotics with very similar characteristics and may prove useful for screening rapidly the oral availability of peptide‐derived drugs.
Cytokine‐mediated induction of cyclo‐oxygenase‐2 by activation of tyrosine kinase in bovine endothelial cells stimulated by bacterial lipopolysaccharide
Authors: Akarasereenont P. , Bakhle Y.S. , Thiemermann C. , Vane J.R.
Published: July 2012

The induction of cyclo‐oxygenase‐2 (COX‐2) afforded by bacterial lipopolysaccharide (LPS, endotoxin) in bovine aortic endothelial cells (BAEC) is mediated by tyrosine kinase. LPS also causes the generation of several cytokines including interleukin‐1β (IL‐1β), tumour necrosis factor‐α (TNF‐α), epidermal growth factor (EGF) and platelet‐derived growth factor (PDGF). This study investigates whether endogenous IL‐1β, TNF‐α, EGF or PDGF contribute to the induction of COX‐2 elicited by LPS in BAEC and if their action is due to activation of tyrosine kinase. Furthermore, we have studied the induction of COX‐2 by exogenous cytokines.

Accumulation of 6‐oxo‐prostaglandin (PG) F1α in cultures of BAEC was measured by radioimmunoassay at 24 h after addition of either LPS (1 μg ml−1) alone or LPS together with a polyclonal antibody to one of the various cytokines. In experiments designed to measure ‘COX activity’, 6‐oxo‐PGF1α generated by BAEC activated with recombinant human IL‐1β, TNF‐α, EGF or PDGF for 12 h was measured after incubation of washed cells with exogenous arachidonic acid (30 μM for 15 min). Western blot analysis determined the expression of COX‐2 protein in BAEC.

The accumulation of 6‐oxo‐PGF1α caused by LPS in BAEC was attenuated by co‐incubation with one of the polyclonal antibodies, anti‐IL‐1β, anti‐TNF‐α, anti‐EGF, anti‐PDGF or with the IL‐1 receptor antagonist, in a dose‐dependent manner. Exogenous IL‐1β, TNF‐α or EGF also caused an increase in COX activity, while PDGF was ineffective. The increase in COX activity elicited by IL‐1β (10 ng ml−1), TNF‐α (100 ng ml−1) or EGF (1000 ng ml−1) in BAEC was attenuated by erbstatin (0.005 to 5 μg ml−1), as was the expression of COX‐2 protein measured by Western blot analysis.

PDGF (10 ng ml−1) significantly augmented the rise in COX activity and COX‐2 protein caused by shorter incubation of BAEC with LPS (1 μg ml−1 for 3 h). Combination of PDGF (10 ng ml−1) with a low concentration of IL‐1β (1 ng ml−1) for 12 h, also increased ‘COX activity’, but combination of PDGF and TNF‐α (10 ng ml−1) did not show any increased activity.

These results suggest that (i) the induction of COX activity and COX‐2 protein elicited by LPS in BAEC is mediated by TNF‐α with lesser contributions from PDGF, EGF or IL‐1β; (ii) exogenous IL‐1β, TNF‐α or EGF alone induce COX‐2 activity and protein in BAEC; (iii) PDGF synergizes with IL‐1β, but not TNF‐α, to cause expression of COX‐2; and (iv) the induction of COX‐2 protein and activity caused by these cytokines involves the activation of tyrosine kinase.
Effect of enalaprilat on bradykinin and des‐Arg9‐bradykinin release following reperfusion of the ischaemic rat heart
Authors: Lamontagne Daniel , Nadeau Réginald , Adam Albert
Published: July 2012

The release of bradykinin (BK) and its metabolite, des‐Arg9‐bradykinin (des‐Arg9‐BK), was studied following reperfusion of a globally ischaemic rat heart.

BK‐like immunoreactivity increased from 13±3 (preischaemic value) to 48±12 fmol min−1 g−1 (P<0.05, n = 14) 30 s after reperfusion. No difference in BK release was found between control hearts and hearts pretreated with the angiotensin converting enzyme (ACE or kininase II) inhibitor, enalaprilat (50 ng ml−1).

No significant change in des‐Arg9‐BK‐like immunoreactivity during reperfusion was observed in control hearts. In contrast, des‐Arg9 ‐ BK ‐ like immunoreactivity rose from 44 ± 15 to 177 ± 61 fmol min−1 g−1 (P<0.05, n = 7) 30 s after reperfusion in enalaprilat‐treated hearts.

In conclusion, BK is released upon reperfusion of the globally ischaemic rat heart. ACE inhibitors, through the inhibition of kininase II, increase the formation of the active metabolite, des‐Arg9‐BK.
Regulation of kinin‐induced contraction and DNA synthesis by inflammatory cytokines in the smooth muscle of the rabbit aorta
Authors: Levesque Luc , Larrivée Jean‐François , Bachvarov Dimcho R. , Rioux Francis , Drapeau Guy , Marceau Francois
Published: July 2012

In rabbit aortic rings, the contractile response to kinins is mediated by the B1 receptors for kinins; the response is upregulated from an initial null level in a time‐ and protein synthesis‐dependent manner. Incubation (3 h) with human recombinant interleukin‐lβ (IL‐lβ) selectively amplified the contractile response to the B1 receptor agonist Sar‐[D‐Phe8]des‐Arg9‐BK, while it did not affect the contractile effect of other agents (angiotensin II, endothelin‐1, phenylephrine).

Oncostatin M (OSM), but not macrophage migration inhibitory factor (MIF), increased the contractile response to the B1 receptor agonist, des‐Arg9‐bradykinin (des‐Arg9‐BK).

Cultured smooth muscle cells derived from the rabbit aorta exhibit a significant des‐Arg9‐BK‐induced increase in [3H]‐thymidine incorporation if pretreated with a cyclo‐oxygenase inhibitor (diclofenac) and concomitantly treated with the cytokines IL‐1 or OSM. Angiotensin II, endothelin‐1 or phenylephrine, alone or in the presence of IL‐lβ, exerted little effect on DNA synthesis in these cells.

The pharmacological characterization of the mitogenic response to kinins using a set of agonist and antagonist analogues is consistent with mediation by B1 receptors. Des‐Arg9‐BK‐induced DNA synthesis is suppressed by prostaglandin E2 by a prostacyclin mimetic (iloprost), by the Ser/Thr protein kinase inhibitor, H‐7, and by a tyrosine kinase inhibitor (i.e. an erbstatin analogue).

B1 receptor‐mediated responses and their capacity to be regulated by cytokines, are retained in rabbit aortic smooth muscle cells. Such responses could be relevant to tissue repair mechanisms and hypertrophic medial responses to injury in arteries.
Structural requirements at the melatonin receptor
Authors: Sugden David , Chong Nelson W.S. , Lewis David F.V.
Published: July 2012

High affinity, specific binding sites for the pineal hormone, melatonin (5‐methoxy N‐acetyltryptamine) can be detected in chick brain membranes by use of the radiolabeled agonist, 2‐[125I]‐iodomelatonin (2‐[125I]‐aMT).

The affinity of a number of analogues of melatonin at the 2‐[125I]‐aMT binding site was determined and compared with an analysis of their electronic structure and significant quantitative relationships obtained.

The best correlations indicated that binding affinity was correlated with ΔE, the difference between the frontier orbital energies, and QNH, the electron density in the highest occupied molecular orbital of the side‐chain nitrogen atom.

These findings suggest that ligand binding may involve hydrogen bonding between the 5‐methoxy and amide moieties of melatonin and complementary amino acid residues, and charge transfer interactions between the indole ring of melatonin and an aromatic amino acid in the receptor binding site.

A molecular model of a putative binding site is proposed based on the predicted amino acid sequence of the cloned Xenopus laevis melanophore melatonin receptor and the quantitative structure‐affinity relationships observed in the present study.
Sustained negative inotropism mediated by α‐adrenoceptors in adult mouse myocardial developmental conversion from positive response in the neonate
Authors: Tanaka Hikaru , Manita Shigeru , Matsuda Tomoyuki , Adachi Mika , Shigenobu Koki
Published: July 2012

Inotropic responses to α‐adrenoceptor stimulation and the effects of antagonists were examined in isolated ventricular preparations from neonatal and adult mice.

Phenylephrine, in the presence of propranolol, produced positive inotropic responses in neonates up to 1 week after birth, while it produced negative inotropic responses in mice older than 3 weeks.

Both positive and negative responses to phenylephrine in neonates and adults, respectively, were antagonized by prazosin, WB4101 (2‐([2,6‐dimethoxyphenoxyethyl]aminomethyl)‐1,4‐benzodioxane) and 5‐methylurapidil, but not by atropine, yohimbine or chlorethylclonidine.

Noradrenaline (NA) produced positive inotropic responses both in the neonate and adult; the responses were observed in a lower concentration‐range in the neonate than in the adult. WB4101 produced a significant leftward shift of the concentration‐response curve for noradrenaline in adult preparations while only a slight rightward shift was observed in the neonate.

Our results demonstrate the presence of α‐adrenoceptor‐mediated inotropic responses in the mouse ventricular myocardia. The response to phenylephrine changes from a positive to a negative effect during postnatal development. The responses are mediated by α1‐adrenoceptors, and modulate the overall inotropic response to NA in the adult.
Pharmacological analysis of the local and reflex responses to bradykinin on rat urinary bladder motility in vivo
Authors: Lecci A. , Giuliani S. , Meini S. , Maggi C.A.
Published: July 2012

The topical application of bradykinin (BK) (0.05–5000 pmol/rat) onto the serosal surface of the urinary bladder in urethane‐anaesthetized rats, evoked low amplitude tonic contractions (not exceeding 25 mmHg) or high amplitude (about 50 mmHg), phasic reflex contractions (chemoceptive micturition reflex) which were abolished by bilateral ablation of the pelvic ganglia. In ganglionectomized rats, BK induced only a local, tonic‐type contraction.

Systemic capsaicin pretreatment (164 μmol kg−1, 4 days before) reduced the incidence of chemoceptive reflex induced by BK (500 pmol/rat) but had no effect on the magnitude of the tonic‐type contraction elicited by BK in ganglionectomized rats. Indomethacin (11 μmol kg−1, 20 min before) reduced the incidence but not the amplitude of the reflex contractions induced by topical application of BK (500 pmol/rat). In ganglionectomized rats, indomethacin (11 μmol kg−1, 20 min before) decreased the amplitude of the tonic contraction evoked by BK. Indomethacin did not affect the chemoceptive reflex induced by topical application of capsaicin (15 nmol/rat) onto the bladder.

Intrathecal administration of the tachykinin NK1 receptor antagonists, RP 67,580 (10 nmol/rat) or SR 140,333 (10 nmol/rat), abolished the chemoceptive reflex induced by BK without modifying the magnitude of the tonic contraction. SR 140,333 (10 nmol/rat) also abolished the occurrence of the chemoceptive reflex induced by capsaicin.

Intravenous administration of the B2 receptor antagonist, Hoe 140 (35 nmol kg−1, 10 min before) abolished the reflex and local effects induced by BK on bladder motility but failed to modify the chemoceptive reflex induced by topical application of capsaicin (15 nmol/rat).

Intrathecal administration of Hoe 140 (10 nmol/rat) reduced the incidence of the chemoceptive reflex induced by BK but had no effect on the amplitude of the local motor response. Likewise, Hoe 140 (10 nmol/rat, i.t.) reduced the incidence of reflex bladder contractions induced by topical application of capsaicin (15 nmol/rat) without affecting the magnitude of the tonic‐type contraction.

These findings indicate that BK stimulates motility through B2 receptors in the rat urinary bladder. BK activates the reflex response by stimulating capsaicin‐sensitive afferent nerves with a contribution from prostanoids. At the spinal cord level, tachykinin NK1 and BK B2 receptors could also be involved in the chemoceptive reflex induced by BK or capsaicin.
Involvement of hydroxyl radicals in neurogenic airway plasma exudation and bronchoconstriction in guinea‐pigs in vivo
Authors: Lei Yu‐Hong , Barnes Peter J. , Rogers Duncan F.
Published: July 2012

Cigarette smoke induces plasma exudation in the airways of rodents by activation of capsaicin‐sensitive ‘sensory‐efferent’ nerves. The response is mediated predominantly by substance P (SP) and the magnitude of exudation is regulated by neutral endopeptidase (NEP). The component(s) of the smoke responsible for the activation of the nerves may be reactive oxygen radicals. We investigated the effect of the hydroxyl radical scavenger dimethylthiourea (DMTU), a regulator of superoxide anion, superoxide dismutase (SOD), and a regulator of hydrogen peroxide, catalase, on plasma exudation (measured using Evans blue dye) induced by cigarette smoke in guinea‐pig main bronchi in vivo. The effect of DMTU on plasma exudation and non‐cholinergic bronchoconstriction (measured as pulmonary insufflation pressure, PIP) induced by electrical stimulation of the vagus nerves was also assessed. Interaction between hydroxyl radicals and NEP was assessed with the NEP inhibitor phosphoramidon.

In each of the experiments, cigarette smoke increased plasma exudation by ∼200% above air‐exposed controls. Acute administration of DMTU (1.5 g kg−1, i.v. for 20 min) significantly reduced cigarette smoke‐induced plasma exudation by 69%. In contrast, neither SOD (240,000 u kg−1, i.v.) nor catalase (400,000 u kg−1, i.v.) significantly affected the exudative response.

Chronic pretreatment with DMTU (1.25 g kg−1 over 4 days) significantly reduced bronchial plasma exudation induced by cigarette smoke by 72%. Phosphoramidon (1.5 mg kg−1, i.v.) completely reversed the inhibition by DMTU of cigarette smoke‐induced plasma exudation.

Vagal stimulation increased plasma exudation by ∼200% and PIP by ∼250%. Acute treatment with DMTU had no significant inhibitory effect on these responses, whereas chronic pretreatment inhibited them by ∼80%. Phosphoramidon reversed the inhibition by chronic DMTU.

SP (1 nmol kg−1) increased plasma exudation by ∼250%, a response which was not inhibited by either acute or chronic DMTU.

We conclude that hydroxyl radicals, rather than superoxide anion or hydrogen peroxide, are involved in the induction of neurogenic plasma exudation and bronchoconstriction induced by cigarette smoke or by electrical stimulation of the vagus nerves. These radicals also affect the activity of NEP. Acute DMTU may affect directly the neural actions of hydroxyl radicals contained in the cigarette smoke. Chronic pretreatment with DMTU may inhibit the neurogenic airway responses by effects on tachykinin biosynthesis and/or axonal transport.
Effect of soluble P55 tumour‐necrosis factor binding fusion protein on the local Shwartzman and Arthus reactions
Authors: Norman Keith E. , Williams Timothy J. , Feldmann Marc , Rossi Adriano G.
Published: July 2012

In this study, the effects of a protein synthesis inhibitor, cycloheximide, and a soluble tumour necrosis factor (TNF) binding/IgG fusion protein, p55‐sf2, on the priming and challenge stages of the local Shwartzman reaction (LSR) were assessed and compared with their effects on the acute inflammatory response induced by recombinant human tumour necrosis factor‐α (rhTNF), lipopolysaccharide (LPS) and a reversed passive Arthus (RPA) reaction in rabbit skin.

The LSR was induced in skin by giving an intradermal (i.d.) priming injection of LPS followed by two i.v. challenge injections 20 h and 22 h later. Accumulation of 51Cr‐labelled red blood cells and [125I]‐albumin were measured at 24 h as markers of haemorrhage and oedema formation, respectively.

The RPA reaction was induced in the rabbit by giving i.d. injections of Arthus anti‐serum (anti‐bovine‐δ‐globulin, BGG) followed 5 min later by an i.v. injection of the antigen (BGG). Oedema formation and the accumulation of 111In‐labelled neutrophils produced in the RPA reaction and in response to i.d. injection of rhTNF and LPS were measured over the 4 h period after inducing the responses.

A single local injection of cycloheximide (10 μ/site) did not inhibit neutrophil accumulation or oedema formation produced by 100% Arthus anti‐sera. Although LPS injected i.d. induced a marked dose‐dependent neutrophil accumulation, there was little associated plasma leakage. Cycloheximide (10 μ/site) did not significantly inhibit the neutrophil accumulation induced by LPS (0.1 μ/site). In the LSR, priming i.d. injections of LPS caused a dose‐dependent increase in haemorrhage and plasma leakage at skin sites after challenge with LPS (two injections of 100 μ, i.v.). Co‐injection of a single dose of cycloheximide (10 μ/site) with LPS (30 μ/site) caused a marked reduction in the amount of haemorrhage. Local cycloheximide (10 μ/site) administered immediately before LSR challenge did not affect the responses produced in the LSR.

Neutrophil accumulation induced by TNF (0.17 μ/site) was abolished by co‐administration of p55‐sf2 (3 μ/site) whereas neutrophil accumulation induced by i.d. LPS and produced in the RPA reaction was not affected. In the LSR, haemorrhage and oedema formation were inhibited by p55‐sf2 (3 μ/site) when it was administered i.d. with the LPS priming injection, but not when given i.d. immediately before LSR challenge.

These data suggest that the acute neutrophil accumulation produced in the RPA reaction and in response to i.d. LPS may not be dependent on local protein synthesis or TNF production. On the other hand, haemorrhage appears to be dependent on local protein synthesis during the priming phase but not during the challenge stage of the LSR. Importantly, haemorrhage and plasma leakage appear to be dependent on local TNF generation during the priming phase but not during the challenge stage of the LSR. Thus TNF appears to play a key role in the LSR in rabbit skin.
In vitro pharmacological profile of YM‐43611, a novel D2‐like receptor antagonist with high affinity and selectivity for dopamine D3 and D4 receptors
Authors: Hidaka Kazuyuki , Tada Shoko , Matsumoto Mitsuyuki , Ohmori Junya , Tasaki Yoshikazu , Nomura Tamako , Usuda Shinji , Yamaguchi Tokio
Published: July 2012

We investigated some neurochemical properties of a novel benzamide, YM‐43611, [(S)‐S‐(1‐benzyl‐3‐pyrrolidinyl)‐5‐chloro‐4‐cyclopropylcarbonyl‐2‐methoxybenzamide] in comparison with putative D2‐like receptor antagonists using both rat and human cloned dopamine D2‐like receptors in vitro.

Receptor binding studies revealed that YM‐43611 had appropriately potent affinities for both rat and human D2‐like receptors, with moderate selectivity for D3 receptors and high selectivity for D4 receptors over D2 receptors (Ktvalues (nM) for rat receptors: D2, 165; D3, 35.5; D4, 1.85, and for human receptors: D2 42.9; D3, 11.2; D4, 2.10).

YM‐43611 displayed weak or negligible affinity for other neurotransmitter receptors, namely D1, D5, α1, α2, β, 5‐HT1A, 5‐HT2A, 5‐HT3, H1, M1 and M2 receptors.

Dopamine stimulated low‐Km GTPase activity on membranes from Chinese hamster ovary (CHO) cells expressing the human D2‐like receptor subtype. This response to dopamine of low‐Km GTPase activity was inhibited by use of putative D2‐like receptor antagonists. YM‐43611 showed a moderate selectivity for D3 receptors (Ki=45.5 nM) and a high selectivity for D4 receptors (Ki=3.28 nM) over D2 receptors (Ki=70.6 nM).

Dopamine inhibited forskolin‐stimulated adenylate cyclase in intact CHO cells expressing the human D2‐like receptor subtype. YM‐43611 shifted the inhibition curve of dopamine on respective D2‐like receptor subtype‐mediated cyclic AMP formation to the right in a parallel fashion, showing a pA2 value of 7.42 (38.1 nM) for D2 receptors, a pKB> value of 8.06 (8.68 nM) for D3 receptors, and a pA2 value of 8.42 (3.77 nM) for D4 receptors.

YM‐43611 but not the other D2‐like receptor antagonists exhibited good selectivity with respect to dual antagonism for D3 and D4 receptors in both receptor binding and functional assays.

These results indicate that YM‐43611 is a novel D2‐like receptor antagonist with high potency and selectivity for both D3 and D4 receptors. YM‐43611 is therefore expected to be valuable in exploration of the physiological role of D3 and D4 receptors.
Failure of BQ123, a more potent antagonist of sarafotoxin 6b than of endothelin‐1, to distinguish between these agonists in binding experiments
Authors: Maguire Janet J. , Kuc Rhoda E. , Rous Brian A. , Davenport Anthony P.
Published: July 2012

In homogenates of human saphenous vein, [125I]‐ET‐1 and [125I]‐S6b each labelled a single population of high affinity binding sites with KD values of 0.64±0.11 nM and 0.55±0.08 nM respectively. Hill slopes were close to one. However, the density of receptors labelled by [125I]‐ET‐1 was significantly greater than that by [125I]‐S6b (187.6±23.0 compared to 91.7±23.6 fmol mg−1 protein, P<0.02).

BQ123, an ETA‐selective antagonist, inhibited specific [125I]‐ET‐1 and [125I]‐S6b binding with equal affinity. BQ123 competed in a biphasic manner for both [125I]‐ET‐1 (0.1 nM) and [125I]‐S6b (0.1 nM) with ETA KD values of 0.55±0.17 nM and 0.52±0.02 nM and ETB KD values of 14.4±2.60 μm and 11.2±0.31 μm respectively. S6b monophasically inhibited 0.1 nM [125I]‐ET‐1 (KD 1.16±0.9 nM) but competed for 0.25 nM [125I]‐ET‐1 in a bipahasic manner (KD high affinity site 1.99±0.84 nM, KD low affinity site 0.68±0.63 μm, ratio 67%: 33%).

BQ123 antagonized the vasoconstrictor responses of ET‐1 with a pKB value of 6.47 whereas BQ123 exhibited 50 fold higher affinity against S6b‐mediated vasoconstriction with a pKB value of 8.18. Regression slopes were 0.80±0.13 and 1.08±0.11 respectively.

In desensitization experiments, S6b (300 nM) did not contract preparations which were no longer responsive to ET‐1 whereas a small contraction to ET‐1 (300 nM) was obtained in preparations rendered unresponsive to S6b.

Medial sections of non‐diseased human aorta, which express only ETA receptors, were used to compare dissociation rates of the two agonists. The time course for the dissociation of [125I]‐ET‐1 and [125I]‐S6b was similar with 20–30% of each ligand dissociating at 4 h.

These data suggest that whilst BQ123, in common with other endothelin antagonists, is a much more potent blocker of S6b contractile responses than of ET‐1 contractile responses, this is not reflected by the equal affinity of BQ123 determined in competition binding experiments against both [125I]‐ET‐1 and [125I]‐S6b. This discrepancy in antagonist potency is probably not due to a marked difference in the rate of dissociation of [125I]‐ET‐1 and [125I]‐S6b from endothelin receptors. One possible explanation is that ET‐1 is activating an additional population of receptors which may have lower affinity for BQ123. This is suggested by the discrepancy in receptor density identified by [125I]‐ET‐1 and [125I]‐S6b.
Mechanical and electrophysiological effects of 8‐oxoberberine (JKL1073A) on atrial tissue
Authors: Chi Jo‐Feng , Chu Shu‐Hsun , Lee Chih‐Shone , Chou Nai‐Kaun , Su Ming‐Jai
Published: July 2012

The effects of 8‐oxoberberine (JKL1073A) on contractions and electrophysiological characteristics of atrial tissues were examined.

In driven left atria of the rat JKL1073A (10–100 μm) increased twitch tension dose‐dependently. In spontaneously beating right atria, JKL1073A increased twitch tension but decreased beating rate slightly.

The positive inotropic and the negative chronotropic effect of 30 μm JKL1073A was not affected by prazosin (1 μm), propranolol (1 μm) and 3‐isobutyl‐1‐methyl‐xanthine (10 μm) but significantly suppressed by 4‐aminopyridine (2 mM 4‐AP).

Current‐clamp study revealed that JKL1073A prolonged rat atrial action potential duration (APD). This prolongation of APD by JKL1073A was decreased by pretreating the cells with 2 mM 4‐AP. Voltage‐clamp study showed that JKL1073A inhibited the integral of the transient outward current (Ito) dose‐dependently with a KD value of 3.66 ± 0.93 μm in rat atrial myocytes. The equilibrium dissociation constant (Kd) for JKL1073A bindings to open state Ito was 0.50 ± 0.08 μm. The suppression of Ito by 3 μm JKL1073A was accompanied by shortening of its inactivation time constant from 52.5 ± 0.9 ms to 16.8 ± 0.7 ms. V0.5 for the steady‐state inactivation curve of Ito was shifted from −25.7 ± 3.3 mV to −34.8 ± 3.2 mV.

In human atrial cells, similar inhibition of Ito and prolongation of APD by JKL1073A was found. The KD value of JKL1073A for inhibition of the integral of Ito in human atrial cells is 4.03 ± 0.02 μm. The Kd for bindings to open state Ito is 0.5 μm.

Currents through K1 channels of rat and human atrial myocytes were not inhibited by JKL1073A at concentrations up to 10 μm.

These results indicate that JKL1073A exerts a positive inotropic effect by inhibition of Ito. JKL1073A inhibit Ito by binding to open state channels or shifting of the steady‐state inactivation curve of Ito.
Inhibition of nitric oxide synthesis by NG‐nitro‐L‐arginine methyl ester (L‐NAME): requirement for bioactivation to the free acid, NG‐nitro‐L‐arginine
Authors: Pfeiffer Silvia , Leopold Eva , Schmidt Kurt , Brunner Friedrich , Mayer Bernd
Published: July 2012

The L‐arginine derivatives NG‐nitro‐L‐arginine (L‐NOARG) and NG‐nitro‐L‐arginine methyl ester (L‐NAME) have been widely used to inhibit constitutive NO synthase (NOS) in different biological systems. This work was carried out to investigate whether L‐NAME is a direct inhibitor of NOS or requires preceding hydrolytic bioactivation to L‐NOARG for inhibition of the enzyme.

A bolus of L‐NAME and L‐NOARG (0.25 μmol) increased coronary perfusion pressure of rat isolated hearts to the same extent (21 ± 0.8 mmHg; n = 5), but the effect developed more rapidly following addition of L‐NOARG than L‐NAME (mean half‐time: 0.7 vs. 4.2 min). The time‐dependent onset of the inhibitory effect of L‐NAME was paralleled by the appearance of L‐NOARG in the coronary effluent.

Freshly dissolved L‐NAME was a 50 fold less potent inhibitor of purified brain NOS (mean IC50 = 70 μm) than L‐NOARG (IC50 = 1.4 μm), but the apparent inhibitory potency of L‐NAME approached that of L‐NOARG upon prolonged incubation at neutral or alkaline pH. H.p.l.c. analyses revealed that NOS inhibition by L‐NAME closely correlated with hydrolysis of the drug to L‐NOARG.

Freshly dissolved L‐NAME contained 2% of L‐NOARG and was hydrolyzed with a half‐life of 365 ± 11.2 min in buffer (pH 7.4), 207 ± 1.7 min in human plasma, and 29 ± 2.2 min in whole blood (n = 3 in each case). When L‐NAME was preincubated in plasma or buffer, inhibition of NOS was proportional to formation of L‐NOARG, but in blood the inhibition was much less than expected from the rates of L‐NAME hydrolysis. This was explained by accumulation of L‐NOARG in blood cells.

These results suggest that L‐NAME represents a prodrug lacking NOS inhibitory activity unless it is hydrolyzed to L‐NOARG. Bioactivation of L‐NAME proceeds at moderate rates in physiological buffers, but is markedly accelerated in tissues such as blood or vascular endothelium.
A pharmacological and histochemical study of hamster urethra and the role of urothelium
Authors: Pinna Christian , Ventura Sabatino , Puglisi Lina , Burnstock Geoffrey
Published: July 2012

Electrical field stimulation (EFS) of circular strips of hamster proximal urethra caused frequency‐dependent relaxations at raised tone. Phentolamine (10−6 m), propranolol (10−6 m) and atropine (10−6 m) were present throughout the experiment. Neurogenic relaxation was attenuated by L‐NG‐nitroarginine methyl ester (L‐NAME) (10−4 m), was restored by L‐arginine (3 × 10−3 m) but not by D‐arginine (3 × 10−3 m) and completely blocked by tetrodotoxin (10−6 m). Neurogenic relaxation was also reduced by suramin (10−4 m) and totally blocked by suramin together with L‐NAME. Strips of hamster urethra devoid of urothelium showed little, if any, relaxant response to EFS.

An immunohistochemical study showed nitric oxide synthase‐immunoreactive nerves in the smooth muscle layers and in the lamina propria, just beneath the urothelium, but no nitric oxide synthase (NOS) staining in the urothelial layer.

Noradrenaline elicited a significantly greater contraction in strips without urothelium than in control strips. L‐NAME (10−4 m) did not affect noradrenaline‐induced contraction in both control and urothelium‐free strips. The contractile response to acetylcholine was not dependent on the presence or absence of urothelium. Nevertheless the response induced by exogenous acetylcholine (10−3 m) was increased by L‐NAME (10−4 m), both in intact and in urothelium‐free strips.

Prostaglandin E2 (10−8‐5 × 10−6 m) and 2‐methyl‐thio‐ATP (10−9‐10−5 m) relaxed proximal urethra. Suramin (10−4 m) significantly inhibited the relaxation induced by 2‐methyl‐thio‐ATP. The amplitude of these responses was not significantly different between intact and urothelium‐free strips and was not blocked by L‐NAME (10−4 m).

These results suggest that nitric oxide (NO) is the principal transmitter involved in the non‐adrenergic, non‐cholinergic (NANC) relaxation of hamster proximal urethra possibly together with another inhibitory transmitter released from nerves. NO can be released from nerves located in the circular smooth muscle layer and in the lamina propria rather than in the urothelium. The reduced neurogenic relaxation in urothelium‐free preparations suggests that a NO‐dependent inhibitory factor is released from the urothelium. In addition, ATP and prostaglandin E2 may be involved, together with NO, in the urethra during micturition.
Inhibition by memantine of the development of persistent oral dyskinesias induced by long‐term haloperidol treatment of rats
Authors: Andreassen Ole A. , Aamo Trond O. , Jørgensen Hugo A.
Published: July 2012

Tardive dyskinesia (TD) is a serious side‐effect of long‐term treatment with neuroleptics. To investigate if neuroleptic‐induced excessive stimulation of striatal glutamate receptors may underlie TD development, the effect of the NMDA antagonist, memantine (1‐amino‐3,5‐dimethyladamantane), was studied in a rat model of TD.

In an acute experiment, six groups of rats were treated daily for 1 week with either vehicle or memantine 20 or 40 mg kg−1 day−1, and on the seventh day they received one injection of either haloperidol 1.0 mg kg−1 i.p. or saline i.p. In a subsequent long‐term experiment lasting 20 weeks, the same treatment was continued, except that haloperidol was injected i.m. as decanoate (38 mg kg−1 every 4 weeks) and control rats received sesame oil. The behaviour was videotaped and scored at intervals during both experiments, and for 16 weeks after cessation of the long‐term treatment.

In the acute experiment, haloperidol decreased motor activity and memantine increased moving and tended to attenuate the immobility induced by haloperidol. Memantine also enhanced the haloperidolinduced increase in the putative TD‐analogue vacuous chewing movements (VCM).

In the long‐term experiment, the most marked effect of haloperidol was a gradual increase in VCM and the increase persisted significantly for 12 weeks after cessation of treatment. Memantine dose‐dependently increased VCM and moving during long‐term treatment. However, only one week after stopping treatment, both these effects of memantine disappeared. In contrast to rats previously treated with haloperidol alone, rats co‐treated with memantine (both doses) and haloperidol had VCM at the level of controls two weeks after stopping treatment. The blood levels of drugs were within the therapeutic range achieved in human subjects.

These results suggest that long‐lasting changes induced by haloperidol are prevented by memantine, which supports the theory that excessive NMDA receptor stimulation may be a mechanism underlying the development of persistent VCM in rats and maybe also TD in human subjects.
Four‐dimensional temperature distributions in red blood cells withdrawn from storage and exposed to ambient temperature: a magnetic resonance thermometry study
Authors: Reiter Ursula , Reiter Gert , Wagner Thomas , Kozma Noemi , Roland Jörg , Schöllnast Helmut , Ebner Franz , Lanzer Gerhard
Published: July 2012

BACKGROUND: Recommended by current guidelines, red blood cell (RBC) temperature should not exceed 10°C during transport. Since warming is a generically three‐dimensional process that is not homogeneous, it is necessary to clarify the term “temperature during warming.” The purpose of this study was therefore to investigate laws and relations between surface, mean, and core temperature and the corresponding times when they exceed 10°C during warm‐up.

STUDY DESIGN AND METHODS: Time‐resolved three‐dimensional temperature distributions of 53 resuspended RBC units (mean volume, 253 ± 17 mL) were measured noninvasively by magnetic resonance thermometry. Warm‐up temperature maps were visualized and analyzed by dedicated software.

RESULTS: Mean times when surface, mean, and core temperature exceeded 10°C were 16 ± 4, 24 ± 5, and 36 ± 7 minutes, respectively. Times strongly correlated with each other (r = 0.78‐0.95) and their variances mainly depended on RBC storage temperature and RBC pouch width (R2 = 0.81‐0.89). Measured mean temperature time courses were well described by a lumped capacitive model of heat transfer with a sample width‐dependent time constant τRBC = 56.3 ± 3.5 minutes (mean R2 = 0.996).

CONCLUSION: Times when RBC surface, mean, and core temperature exceed 10°C can be estimated from each other. Moreover RBC mean temperature can be calculated for arbitrary storage and ambient temperatures. Findings might serve as a helpful tool in RBC temperature monitoring.
Fracture resistance of inter‐joined zirconia abutment of dental implant system with injection molding technique
Authors: Yang* Jianjun , Wang* Ke , Liu Guangyuan , Wang Dashan
Published: July 2012

Objectives

Zirconia powder in nanometers can be fabricated into inter‐joined abutment of dental implant system with the injection shaping technique. This study was to detect the resistance of inter‐joined zirconia abutment with different angle loading for clinical applications.

Materials and Methods

The inter‐joined abutments were shaped with the technique of injection of zirconia powder in nanometers. Sixty Osstem GSII 5 × 10 mm implants were used with 30 zirconia abutments and 30 Osstem GSII titanium abutments for fixation using 40 N torque force. The loading applications included 90°, 30°, and 0° formed by the long axis of abutments and pressure head of universal test machine. The fracture resistances of zirconia and titanium abutments were documented and analyzed.

Results

The inter‐joined zirconia abutments were assembled to the Osstem GSII implants successfully. In the 90° loading mode, the fracture resistance of zirconia abutment group and titanium abutment group were 301.5 ± 15.4 N and 736.4 ± 120.1 N, respectively. And those in the 30°groups were 434.7 ± 36.1 N and 1073.1 ± 74 N, correspondingly. Significant difference in the two groups was found using t‐test and Wilcoxon test. No damage on the abutments of the two groups but S‐shaped bending on the implants was found when the 0° loading was 1300–2000N.

Conclusions

Through the assembly of Zirconia abutments and implants, all the components presented sufficient resistance acquired for the clinical application under loadings with different angle.
Methylphenidate improves the behavioral and cognitive deficits of neurogranin knockout mice
Authors: Huang F. L. , Huang K.‐P.
Published: July 2012

Neurogranin (Ng), a brain‐specific calmodulin‐binding protein, is expressed highly in hippocampus, and is important for cognitive function. Deletion of the Ng gene from mice caused attenuation of signal reaction cascade in hippocampus, impairments in learning and memory and high frequency stimulation‐induced long‐term potentiation (LTP). Environmental enrichment alone failed to improve cognitive function. In this study, behavioral testing revealed that Ng knockout (NgKO) mice were both hyperactive and socially withdrawn. Methylphenidate (MPH) was given to mice while they were also kept under an enrichment condition. MPH treatment reduced the hyperactivity of NgKO mice tested in both the open field and forced swim chamber. MPH improved their social abilities such that mice recognized and interacted better with novel subjects. The cognitive memories of MPH‐treated mutants were improved in both water maze and contextual fear conditioning tests. High frequency stimulation‐induced LTP of NgKO mice was also improved by MPH. The present treatment regimen, however, did not fully reverse the deficits of the mutant mice. In contrast, MPH exerted only a minimal effect on the wild type mice. At the cellular level, MPH increased the number of glial fibrillary acidic protein‐positive cells in hippocampus, particularly within the dentate gyrus of NgKO mice. Therefore it will be of interest to determine the nature of MPH‐mediated astrocyte activation and how it may modulate behavior in future studies. Taken together these NgKO mice may be useful for the development of better drug treatment to improve cognitive and behavioral impairments.
Assessment of nutritional status and nutritional risk in hospitalized Iranian children
Authors: Moeeni Vesal , Walls Tony , Day Andrew S
Published: July 2012

Aims:  This study aimed to define the nutritional state of children admitted to a tertiary Iranian hospital and to evaluate nutritional risk score tools in these children.

Methods:  The anthropometry of hospitalized and healthy children from the same community was determined. Three nutritional risk score tools were applied to all inpatients.

Results:  One hundred and nineteen inpatients were recruited along with a comparison group of 100 children. The prevalence of under‐nutrition in the inpatient group was 25.2% and 3% in the community group (p < 0.0001). Obesity/overweight was more prevalent in the community group than the inpatients (22% versus 2.5%: p = 0.04). Severely malnourished children had a longer hospital stay than those with normal nutrition (p < 0.0001). The nutritional risk score tools identified between 83% and 90% of the malnourished patients in the moderate and high‐risk groups. The STRONGkids tool correlated more strongly with anthropometric measurements than the other tools. The length of hospital stay was associated with risk status (p = 0.004).

Conclusion:  Hospitalized Iranian children have higher rates of under‐nutrition than healthy children from the same community. NRS tools were able to identify children at nutritional risk; however, variable utility was observed. Further assessment of NRS tools in the developing setting is required.
The Alberta Pregnancy Outcomes and Nutrition (APrON) cohort study: rationale and methods
Authors: Kaplan Bonnie J. , Giesbrecht Gerald F. , Leung Brenda M.Y. , Field Catherine J. , Dewey Deborah , Bell Rhonda C. , Manca Donna P. , O'Beirne Maeve , Johnston David W. , Pop Victor J. , Singhal Nalini , Gagnon Lisa , Bernier Francois P. , Eliasziw Misha , McCargar Linda J. , Kooistra Libbe , Farmer Anna , Cantell Marja , Goonewardene Laki , Casey Linda M. , Letourneau Nicole , Martin Jonathan W. ,   the APrON Study Team
Published: July 2012

The Alberta Pregnancy Outcomes and Nutrition (APrON) study is an ongoing prospective cohort study that recruits pregnant women early in pregnancy and, as of 2012, is following up their infants to 3 years of age. It has currently enrolled approximately 5000 Canadians (2000 pregnant women, their offspring and many of their partners). The primary aims of the APrON study were to determine the relationships between maternal nutrient intake and status, before, during and after gestation, and (1) maternal mood; (2) birth and obstetric outcomes; and (3) infant neurodevelopment. We have collected comprehensive maternal nutrition, anthropometric, biological and mental health data at multiple points in the pregnancy and the post‐partum period, as well as obstetrical, birth, health and neurodevelopmental outcomes of these pregnancies. The study continues to follow the infants through to 36 months of age. The current report describes the study design and methods, and findings of some pilot work. The APrON study is a significant resource with opportunities for collaboration.
“OUR SHARED VALUES” IN SINGAPORE: A CONFUCIAN PERSPECTIVE
Authors: Tan Charlene
Published: July 2012

In this essay Charlene Tan offers a philosophical analysis of the Singapore state's vision of shared citizenship by examining it from a Confucian perspective. The state's vision, known formally as “Our Shared Values,” consists of communitarian values that reflect the official ideology of multiculturalism. This initiative included a White Paper, entitled Shared Values, which presented pejorative assessments of the ideals of “individual rights” and “individual interests” as antithetical to national interests. Rejecting this characterization, Tan argues that a dominant Confucian perspective recognizes the correlative rights of all human beings that are premised on the inherent right to human dignity, worth, and equality. Furthermore, Confucianism posits that it is in everyone's interest to attain the Confucian ethical ideal of becoming a noble person in society through self‐cultivation. Tan concludes by highlighting two key implications for Singapore from a Confucian perspective on the Shared Values: first, schools in Singapore should place greater emphasis on individual moral development of their students, and second, more avenues should be provided for residents to contribute actively to the development of the vision of shared citizenship.
MAD HATTERS, JACKBOOTED MANAGERS, AND THE MASSIFICATION OF HIGHER EDUCATION
Authors: Sidorkin Alexander M.
Published: July 2012

In this review of three recent books on higher education, Alexander Sidorkin shows how the disinterested discourse that appears to be anticapitalist and anticommercial is actually a way of obtaining income from state subsidies. What links the books under review—Cary Nelson's No University Is an Island: Saving Academic Freedom, Frank Donoghue's The Last Professors: The Corporate University and the Fate of the Humanities, and Jennifer Washburn's University, Inc.: The Corporate Corruption of Higher Education—is their critical evaluation of the corporatization and commercialization of higher education. In his analysis of this common theme, Sidorkin considers discourse as a means of production, and he maintains that the semiotic fields produced by discourse may create inflationary bubbles unless they engage in innovative discursive practices. Higher education is shaped by the trend toward massification, which makes the innovative discourse essential. Sidorkin concludes that the discursive energy of proponents of higher education should be focused on solving the numerous problems that arise from the massification of higher education rather than trying to reverse the trend and return to some golden age of academia.
Inhibition of XMRV and HIV‐1 proteases by pepstatin A and acetyl‐pepstatin
Authors: Matúz Krisztina , Mótyán János , Li Mi , Wlodawer Alexander , Tőzsér József
Published: July 2012

The kinetic properties of two classical inhibitors of aspartic proteases (PRs), pepstatin A and acetyl‐pepstatin, were compared in their interactions with HIV‐1 and xenotropic murine leukemia virus related virus (XMRV) PRs. Both compounds are substantially weaker inhibitors of XMRV PR than of HIV‐1 PR. Previous kinetic and structural studies characterized HIV‐1 PR–acetyl‐pepstatin and XMRV PR–pepstatin A complexes and suggested dramatically different binding modes. Interaction energies were calculated for the possible binding modes and suggested a strong preference for the one‐inhibitor binding mode for HIV‐1 PR–acetyl‐pepstatin and the two‐inhibitor binding mode for XMRV PR–pepstatin A interactions. Comparison of the molecular models suggested that in the case of XMRV PR the relatively unfavorable interactions at S3′ and the favorable interactions at S4 and S4′ sites with the statine residues may shift the ground state binding towards the two‐inhibitor binding mode, whereas the single molecule ground state binding of statines to the HIV‐1 PR appear to be more favorable. The preferred single molecular binding to HIV‐1 PR allows the formation of the transition state complex, represented by substantially better binding constants. Intriguingly, the crystal structure of the complex of acetyl‐pepstatin with XMRV PR has shown a mixed type of binding: the unusual binding mode of two molecules of the inhibitor to the enzyme, in a mode very similar to the previously determined complex with pepstatin A, together with the classical binding mode found for HIV‐1 PR. The structure is thus in good agreement with the very similar interaction energies calculated for the two types of binding.

Database
 The final coordinates of the crystal structure of XMRV protease complexed with acetyl‐pepstatin are available in the Protein Data Bank under the accession number 4EXH

Structured digital abstract

 HIV‐1 PR and HIV‐1 PR bind by biochemical (View interaction)

 XMRV PR cleaves MLV Gag by enzymatic study (View interaction)

XMRV PR and XMRV PR bind by biochemical (View interaction)

XMRV PR and XMRV PR bind by x‐ray crystallography (View interaction)
Brain orexin promotes obesity resistance
Authors: Kotz Catherine , Nixon Joshua , Butterick Tammy , Perez‐Leighton Claudio , Teske Jennifer , Billington Charles
Published: July 2012

Resistance to obesity is becoming an exception rather than the norm, and understanding mechanisms that lead some to remain lean in spite of an obesigenic environment is critical if we are to find new ways to reverse this trend. Levels of energy intake and physical activity both contribute to body weight management, but it is challenging for most to adopt major long‐term changes in either factor. Physical activity outside of formal exercise, also referred to as activity of daily living, and in stricter form, spontaneous physical activity (SPA), may be an attractive modifiable variable for obesity prevention. In this review, we discuss individual variability in SPA and NEAT (nonexercise thermogenesis, or the energy expended by SPA) and its relationship to obesity resistance. The hypothalamic neuropeptide orexin (hypocretin) may play a key role in regulating SPA and NEAT. We discuss how elevated orexin signaling capacity, in the context of a brain network modulating SPA, may play a major role in defining individual variability in SPA and NEAT. Greater activation of this SPA network leads to a lower propensity for fat mass gain and therefore may be an attractive target for obesity prevention and therapy.