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Find video protocols related to scientific articles indexed in Pubmed.
Cardiac involvement in sarcoidosis: evolving concepts in diagnosis and treatment.
Semin Respir Crit Care Med
PUBLISHED: 07-09-2014
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Clinically evident sarcoidosis involving the heart has been noted in at least 2 to 7% of patients with sarcoidosis, but occult involvement is much higher (> 20%). Cardiac sarcoidosis is often not recognized antemortem, as sudden death may be the presenting feature. Cardiac involvement may occur at any point during the course of sarcoidosis and may occur in the absence of pulmonary or systemic involvement. Sarcoidosis can involve any part of the heart, with protean manifestations. Prognosis of cardiac sarcoidosis is related to extent and site(s) of involvement. Most deaths due to cardiac sarcoidosis are due to arrhythmias or conduction defects, but granulomatous infiltration of the myocardium may be lethal. The definitive diagnosis of isolated cardiac sarcoidosis is difficult. The yield of endomyocardial biopsies is low; treatment of cardiac sarcoidosis is often warranted even in the absence of histologic proof. Radionuclide scans are integral to the diagnosis. Currently, 18F-fluorodeoxyglucose positron emission tomography/computed tomography and gadolinium-enhanced magnetic resonance imaging scans are the key imaging modalities to diagnose cardiac sarcoidosis. The prognosis of cardiac sarcoidosis is variable, but mortality rates of untreated cardiac sarcoidosis are high. Although randomized therapeutic trials have not been done, corticosteroids (alone or combined with additional immunosuppressive medications) remain the mainstay of treatment. Because of the potential for sudden cardiac death, implantable cardioverter-defibrillators should be placed in any patient with cardiac sarcoidosis and serious ventricular arrhythmias or heart block, and should be considered for cardiomyopathy. Cardiac transplantation is a viable option for patients with end-stage cardiac sarcoidosis refractory to medical therapy.
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Sarcoidosis-associated pulmonary hypertension and lung transplantation for sarcoidosis.
Semin Respir Crit Care Med
PUBLISHED: 07-09-2014
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Pulmonary hypertension (PH) is a significant complication of sarcoidosis, occurring in approximately 6 to?>?20% of cases, and markedly increases mortality among these patients. The clinician should exercise a high index of suspicion for sarcoidosis-associated PH (SAPH) given the nonspecific symptomatology and the limitations of echocardiography in this patient population. The pathophysiology of PH in sarcoidosis is complex and multifactorial. Importantly, there are inherent differences in the pathogenesis of SAPH compared with idiopathic pulmonary arterial hypertension, making the optimal management of SAPH controversial. In this article, we review the epidemiology, diagnosis, prognosis, and treatment considerations for SAPH. Lung transplantation (LT) is a viable therapeutic option for sarcoid patients with severe pulmonary fibrocystic sarcoidosis or SAPH refractory to medical therapy. We discuss the role for LT in patients with sarcoidosis, review the global experience with LT in this population, and discuss indications and contraindications to LT.
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The Prokaryote Ligand-Gated Ion Channel ELIC Captured in a Pore Blocker-Bound Conformation by the Alzheimer's Disease Drug Memantine.
Structure
PUBLISHED: 05-13-2014
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Pentameric ligand-gated ion channels (pLGIC) catalyze the selective transfer of ions across the cell membrane in response to a specific neurotransmitter. A variety of chemically diverse molecules, including the Alzheimer's drug memantine, block ion conduction at vertebrate pLGICs by plugging the channel pore. We show that memantine has similar potency in ELIC, a prokaryotic pLGIC, when it contains an F16'S pore mutation. X-ray crystal structures, using both memantine and its derivative, Br-memantine, reveal that the ligand is localized at the extracellular entryway of the channel pore, and the pore is in a more closed conformation than wild-type ELIC in both the presence and absence of memantine. However, using voltage clamp fluorometry we observe fluorescence changes in opposite directions during channel activation and pore block, revealing an additional conformational transition not apparent from the crystal structures. These results have important implications for drugs such as memantine, which block channel pores.
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Phosphorylation mediated structural and functional changes in pentameric ligand-gated ion channels: implications for drug discovery.
Int. J. Biochem. Cell Biol.
PUBLISHED: 04-02-2014
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Pentameric ligand-gated ion channels (pLGICs) mediate numerous physiological processes, including fast neurotransmission in the brain. They are targeted by a large number of clinically-important drugs and disruptions to their function are associated with many neurological disorders. The phosphorylation of pLGICs can result in a wide range of functional consequences. Indeed, many neurological disorders result from pLGIC phosphorylation. For example, chronic pain is caused by the protein kinase A-mediated phosphorylation of ?3 glycine receptors and nicotine addiction is mediated by the phosphorylation of ?4- or ?7-containing nicotinic receptors. A recent study demonstrated that phosphorylation can induce a global conformational change in a pLGIC that propagates to the neurotransmitter-binding site. Here we present evidence that phosphorylation-induced global conformational changes may be a universal phenomenon in pLGICs. This raises the possibility of designing drugs to specifically treat disease-modified pLGICs. This review summarizes some of the opportunities available in this area.
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Activation of ER? modulates fear generalization through an effect on memory retrieval.
Horm Behav
PUBLISHED: 03-24-2014
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Women are 60% more likely to suffer from an anxiety disorder than men. One hypothesis for this difference may be that females exhibit increased rates of fear generalization. Females generalize fear to a neutral context faster than males, a process driven, in part, by estrogens. In the current study, ovariectomized adult female Long-Evans rats were given acute injections of estradiol benzoate (15?g/0.1mL sesame oil) or sesame oil during a passive avoidance procedure to determine if estrogens increase fear generalization through an effect on fear memory acquisition/consolidation or through fear memory retrieval. Animals injected 1h prior to training generalized to the neutral context 24h later but not 7days after training. Generalization was also seen when injections occurred 24h before testing, but not when tested at immediate (1h) or intermediate (6h) time points. In Experiment 3, animals were injected with estrogen receptor (ER) agonists, PPT or DPN, to determine which ER subtype(s) increased fear generalization. Only the ER? agonist, DPN, increased fear generalization when testing occurred 24h after injection. Our results indicate that estradiol increases fear generalization through an effect on fear memory retrieval mechanisms by activation of ER?.
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Changes in right heart haemodynamics and echocardiographic function in an advanced phenotype of pulmonary hypertension and right heart dysfunction associated with pulmonary fibrosis.
Thorax
PUBLISHED: 01-17-2014
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Pulmonary hypertension (PH)-targeted therapy in the setting of pulmonary fibrosis (PF) is controversial; the main clinical concern is worsening of systemic hypoxaemia. We sought to determine the effects of gentle initiation and chronic administration of parenteral treprostinil on right heart function in patients with PF associated with an advanced PH phenotype.
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GABAA receptor ? and ? subunits shape synaptic currents via different mechanisms.
J. Biol. Chem.
PUBLISHED: 01-14-2014
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Synaptic GABAA receptors (GABAARs) mediate most of the inhibitory neurotransmission in the brain. The majority of these receptors are comprised of ?1, ?2, and ?2 subunits. The amygdala, a structure involved in processing emotional stimuli, expresses ?2 and ?1 subunits at high levels. The effect of these subunits on GABAAR-mediated synaptic transmission is not known. Understanding the influence of these subunits on GABAAR-mediated synaptic currents may help in identifying the roles and locations of amygdala synapses that contain these subunits. Here, we describe the biophysical and synaptic properties of pure populations of ?1?2?2, ?2?2?2, ?1?2?1 and ?2?2?1 GABAARs. Their synaptic properties were examined in engineered synapses, whereas their kinetic properties were studied using rapid agonist application, and single channel recordings. All macropatch currents activated rapidly (<1 ms) and deactivated as a function of the ?-subunit, with ?2-containing GABAARs consistently deactivating ?10-fold more slowly. Single channel analysis revealed that the slower current decay of ?2-containing GABAARs was due to longer burst durations at low GABA concentrations, corresponding to a ?4-fold higher affinity for GABA. Synaptic currents revealed a different pattern of activation and deactivation to that of macropatch data. The inclusion of ?2 and ?1 subunits slowed both the activation and deactivation rates, suggesting that receptors containing these subunits cluster more diffusely at synapses. Switching the intracellular domains of the ?2 and ?1 subunits substantiated this inference. Because this region determines post-synaptic localization, we hypothesize that GABAARs containing ?1 and ?2 use different mechanisms for synaptic clustering.
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Sinorhizobium meliloti flavin secretion and bacteria-host interaction: role of the bifunctional RibBA protein.
Mol. Plant Microbe Interact.
PUBLISHED: 01-11-2014
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Sinorhizobium meliloti, the nitrogen-fixing bacterial symbiont of Medicago spp. and other legumes, secretes a considerable amount of riboflavin. This precursor of the cofactors flavin mononucleotide and flavin adenine dinucleotide is a bioactive molecule that has a beneficial effect on plant growth. The ribBA gene of S. meliloti codes for a putative bifunctional enzyme with dihydroxybutanone phosphate synthase and guanosine triphosphate (GTP) cyclohydrolase II activities, catalyzing the initial steps of the riboflavin biosynthesis pathway. We show here that an in-frame deletion of ribBA does not cause riboflavin auxotrophy or affect the ability of S. meliloti to establish an effective symbiosis with the host plant but does affect the ability of the bacteria to secrete flavins, colonize host-plant roots, and compete for nodulation. A strain missing the RibBA protein retains considerable GTP cyclohydrolase II activity. Based on these results, we hypothesize that S. meliloti has two partly interchangeable modules for biosynthesis of riboflavin, one fulfilling the internal need for flavins in bacterial metabolism and the other producing riboflavin for secretion. Our data also indicate that bacteria-derived flavins play a role in communication between rhizobia and the legume host and that the RibBA protein is important in this communication process even though it is not essential for riboflavin biosynthesis and symbiosis.
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The impact of human hyperekplexia mutations on glycine receptor structure and function.
Mol Brain
PUBLISHED: 01-07-2014
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Hyperekplexia is a rare neurological disorder characterized by neonatal hypertonia, exaggerated startle responses to unexpected stimuli and a variable incidence of apnoea, intellectual disability and delays in speech acquisition. The majority of motor defects are successfully treated by clonazepam. Hyperekplexia is caused by hereditary mutations that disrupt the functioning of inhibitory glycinergic synapses in neuromotor pathways of the spinal cord and brainstem. The human glycine receptor ?1 and ? subunits, which predominate at these synapses, are the major targets of mutations. International genetic screening programs, that together have analysed several hundred probands, have recently generated a clear picture of genotype-phenotype correlations and the prevalence of different categories of hyperekplexia mutations. Focusing largely on this new information, this review seeks to summarise the effects of mutations on glycine receptor structure and function and how these functional alterations lead to hyperekplexia.
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Bacterial community composition of chronic periodontitis and novel oral sampling sites for detecting disease indicators.
Microbiome
PUBLISHED: 01-01-2014
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Periodontitis is an infectious and inflammatory disease of polymicrobial etiology that can lead to the destruction of bones and tissues that support the teeth. The management of chronic periodontitis (CP) relies heavily on elimination or at least control of known pathogenic consortia associated with the disease. Until now, microbial plaque obtained from the subgingival (SubG) sites has been the primary focus for bacterial community analysis using deep sequencing. In addition to the use of SubG plaque, here, we investigated whether plaque obtained from supragingival (SupG) and tongue dorsum sites can serve as alternatives for monitoring CP-associated bacterial biomarkers.
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Practical and Cost-Effective Manufacturing Route for the Synthesis of a ?-Lactamase Inhibitor.
Org. Lett.
PUBLISHED: 12-13-2013
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Compound 1, a potent and irreversible inhibitor of ?-lactamases, is in clinical trials with ?-lactam antibiotics for the treatment of serious and antibiotic-resistant bacterial infections. A short, scalable, and cost-effective route for the production of this densely functionalized polycyclic molecule is described.
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New insights into root causes of pediatric accidental unsupervised ingestions of over-the-counter medications.
Clin Toxicol (Phila)
PUBLISHED: 11-28-2013
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Abstract Context. Changes to regulations, packaging, and labeling and ongoing educational efforts are intended to support appropriate use of medicines. Yet annually poison centers receive > 500 000 reports of accidental or unsupervised exposure to medicines for children under 6 years of age. Objective. To identify root (i.e., fundamental and preventable) causes of accidental unsupervised ingestions (AUIs), we designed a questionnaire and conducted a follow-up survey of caregivers who contacted McNeil Consumer Healthcare (McNeil) following an AUI by a child under 12 years of age. Methods. Reports received between 1 October 2008 and 22 January 2009 were screened retrospectively for specific Medical Dictionary of Regulatory Activities (MedDRA) Preferred Terms relating to AUIs. Using the questionnaire, we collected information about the child, caregiver, medicines involved in AUI, management of AUI, and storage location of medicines. Results. Two hundred twenty reports met inclusion criteria and attempts to contact these caregivers were made throughout a 2-week period in March 2009; caregivers completed the questionnaire for 45 reports. All AUIs occurred in children under 7 years and 56% were boys. In 56% of AUI cases, the child involved was the intended recipient of the medicine; in 71%, a pediatric medicine was involved. Most AUIs occurred in the childs home; most caregivers reported not observing the AUI. Sixty percent of caregivers reported that the medicine involved in AUI was not in the normal storage location when AUI occurred. Among children involved in AUIs, 84% did not experience any symptoms. Seven children experienced mild, self-limiting symptoms which resolved. AUIs often occurred < 24 h after last therapeutic use when the medicine was removed from its normal storage location. Conclusions. These new insights may help guide-targeted interventions and educational efforts to focus caregivers attention to reengaging childproofing mechanisms and returning medicines to a secure location, high and out of sight, immediately after use.
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Serum VEGF-D concentration as a biomarker of lymphangioleiomyomatosis severity and treatment response: a prospective analysis of the Multicenter International Lymphangioleiomyomatosis Efficacy of Sirolimus (MILES) trial.
Lancet Respir Med
PUBLISHED: 10-26-2013
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VEGF-D is a lymphangiogenic growth factor that has a key role in tumour metastasis. Serum VEGF-D concentrations are increased in most patients with lymphangioleiomyomatosis, a rare neoplasm associated with mTOR-activating tuberous sclerosis gene mutations, lymphadenopathy, metastatic spread, and pulmonary cyst formation. We used data from the Multicenter International Lymphangioleiomyomatosis Efficacy of Sirolimus (MILES) trial to assess the usefulness of serum VEGF-D concentration as a marker of severity and therapeutic response to sirolimus in patients with lymphangioleiomyomatosis.
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Sex differences in the generalization of fear as a function of retention intervals.
Learn. Mem.
PUBLISHED: 10-18-2013
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In previous studies using male rodents, context change disrupted a fear response at a short, but not a long, retention interval. Here, we examined the effects of context changes on fear responses as a function of time in male and female rats. Males displayed context discrimination at all intervals, whereas females exhibited generalization by 5 d. Ovariectomized females with no hormone replacement displayed context discrimination at 5 d, whereas those receiving 17?-estradiol generalized their fear response to a neutral context. These results demonstrate that fear generalization for contextual cues occurs faster in female rats and is mediated, in part, by estrogens.
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New hyperekplexia mutations provide insight into glycine receptor assembly, trafficking, and activation mechanisms.
J. Biol. Chem.
PUBLISHED: 10-09-2013
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Hyperekplexia is a syndrome of readily provoked startle responses, alongside episodic and generalized hypertonia, that presents within the first month of life. Inhibitory glycine receptors are pentameric ligand-gated ion channels with a definitive and clinically well stratified linkage to hyperekplexia. Most hyperekplexia cases are caused by mutations in the ?1 subunit of the human glycine receptor (hGlyR) gene (GLRA1). Here we analyzed 68 new unrelated hyperekplexia probands for GLRA1 mutations and identified 19 mutations, of which 9 were novel. Electrophysiological analysis demonstrated that the dominant mutations p.Q226E, p.V280M, and p.R414H induced spontaneous channel activity, indicating that this is a recurring mechanism in hGlyR pathophysiology. p.Q226E, at the top of TM1, most likely induced tonic activation via an enhanced electrostatic attraction to p.R271 at the top of TM2, suggesting a structural mechanism for channel activation. Receptors incorporating p.P230S (which is heterozygous with p.R65W) desensitized much faster than wild type receptors and represent a new TM1 site capable of modulating desensitization. The recessive mutations p.R72C, p.R218W, p.L291P, p.D388A, and p.E375X precluded cell surface expression unless co-expressed with ?1 wild type subunits. The recessive p.E375X mutation resulted in subunit truncation upstream of the TM4 domain. Surprisingly, on the basis of three independent assays, we were able to infer that p.E375X truncated subunits are incorporated into functional hGlyRs together with unmutated ?1 or ?1 plus ? subunits. These aberrant receptors exhibit significantly reduced glycine sensitivity. To our knowledge, this is the first suggestion that subunits lacking TM4 domains might be incorporated into functional pentameric ligand-gated ion channel receptors.
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Analysis of hyperekplexia mutations identifies transmembrane domain rearrangements that mediate glycine receptor activation.
J. Biol. Chem.
PUBLISHED: 10-04-2013
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Pentameric ligand-gated ion channels (pLGICs) mediate numerous physiological processes and are therapeutic targets for a wide range of clinical indications. Elucidating the structural differences between their closed and open states may help in designing improved drugs that bias receptors toward the desired conformational state. We recently showed that two new hyperekplexia mutations, Q226E and V280M, induced spontaneous activity in ?1 glycine receptors. Gln-226, located near the top of transmembrane (TM) 1, is closely apposed to Arg-271 at the top of TM2 in the neighboring subunit. Using mutant cycle analysis, we inferred that Q226E induces activation via an enhanced electrostatic attraction to Arg-271. This would tilt the top of TM2 toward TM1 and hence away from the pore axis to open the channel. We also concluded that the increased side chain volume of V280M, in the TM2-TM3 loop, exerts a steric repulsion against Ile-225 at the top of TM1 in the neighboring subunit. We infer that this steric repulsion would tilt the top of TM3 radially outwards against the stationary TM1 and thus provide space for TM2 to relax away from the pore axis to create an open channel. Because the transmembrane domain movements inferred from this functional analysis are consistent with the structural differences evident in the x-ray atomic structures of closed and open state bacterial pLGICs, we propyose that the model of pLGIC activation as outlined here may be broadly applicable across the eukaryotic pLGIC receptor family.
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CXCR3 ligands are associated with the continuum of diffuse alveolar damage to chronic lung allograft dysfunction.
Am. J. Respir. Crit. Care Med.
PUBLISHED: 09-26-2013
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After lung transplantation, insults to the allograft generally result in one of four histopathologic patterns of injury: (1) acute rejection, (2) lymphocytic bronchiolitis, (3) organizing pneumonia, and (4) diffuse alveolar damage (DAD). We hypothesized that DAD, the most severe form of acute lung injury, would lead to the highest risk of chronic lung allograft dysfunction (CLAD) and that a type I immune response would mediate this process.
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Pulmonary hypertension complicating interstitial lung disease and COPD.
Semin Respir Crit Care Med
PUBLISHED: 09-13-2013
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Pulmonary hypertension (PH) may complicate parenchymal lung disease, specifically interstitial lung diseases and chronic obstructive pulmonary disease, and uniformly increases the mortality risk. The epidemiology and degree of PH is variable and unique to the underlying lung disease. The clinician should exercise a high index of suspicion for PH complicating parenchymal lung disease especially given the nonspecific symptomatology and the limitations of echocardiography in this patient population. In general, PH-specific therapies in this setting have been poorly studied, with concern for increased shunting and/or ventilation/perfusion (V/Q) mismatch and resultant hypoxemia. A better understanding of the mechanisms underlying PH related to parenchymal lung disease may lead to novel pharmacological targets to prevent or treat this serious complication.
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Pulmonary hypertension complicating connective tissue disease.
Semin Respir Crit Care Med
PUBLISHED: 09-13-2013
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Pulmonary hypertension (PH) may complicate connective tissue disease (CTD), particularly systemic sclerosis (SSc, scleroderma), and markedly increases mortality. More than 70% of cases of PH complicating CTD occur in SSc, which is the major focus of this article. Pulmonary complications (i.e., interstitial lung disease [ILD] and PH) are the leading causes of scleroderma-related deaths. "Isolated" PH (i.e., without ILD) complicates SSc in 7.5 to 20% of cases; secondary PH may also occur in patients with SSc-associated ILD. Several clinical markers and specific autoantibody profiles have been associated with PH in SSc. The role of PH-specific therapy is controversial, as prognosis and responsiveness to therapy are worse in SSc-associated PH compared with idiopathic pulmonary arterial hypertension. We discuss medical therapies for CTD-associated PH and the role of lung transplantation for patients failing medical therapy.
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Comparison of the intrinsic vasorelaxant and inotropic effects of the antiarrhythmic agents vernakalant and flecainide in human isolated vascular and cardiac tissues.
J. Cardiovasc. Pharmacol.
PUBLISHED: 09-04-2013
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This study explored the intrinsic vasorelaxant and inotropic effects of the mixed potassium and sodium channel blocker atrial antiarrhythmic vernakalant and the class IC antiarrhythmic agent flecainide in human isolated subcutaneous resistance artery and in ventricular trabecular muscle preparations. At test concentrations encompassing free plasma concentrations associated with clinical efficacy for conversion of atrial fibrillation, vernakalant (1-10 ?M) displayed no significant direct effects on human resistance artery tone or ventricular contractility. In contrast, tested at equimolar concentrations, flecainide significantly reduced peak isometric contractile force (10 ?M) and maximal rates of force development and decline (3 and 10 ?M) in the human ventricular muscle preparation while displaying no significant effect on human resistance artery tone. The lack of effects of vernakalant on human resistance artery tone and ventricular muscle contractile function suggests that direct vasorelaxant and inotropic effects do not underlie the rare hypotensive events observed clinically with vernakalant, raising the possibility that secondary (eg, reflex) effects may mediate these events. The demonstration of negative inotropic effects with flecainide in the human ventricular muscle preparations in the absence of an effect on resistance artery tone suggests that the hemodynamic effects of flecainide observed clinically result primarily from direct negative inotropic effects.
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Phosphorylation of ?3 Glycine Receptors Induces a Conformational Change in the Glycine-Binding Site.
ACS Chem Neurosci
PUBLISHED: 07-18-2013
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Inflammatory pain sensitization is initiated by prostaglandin-induced phosphorylation of ?3 glycine receptors (GlyRs) that are specifically located in inhibitory synapses on spinal pain sensory neurons. Phosphorylation reduces the magnitude of glycinergic synaptic currents, thereby disinhibiting nociceptive neurons. Although ?1 and ?3 subunits are both expressed on spinal nociceptive neurons, ?3 is a more promising therapeutic target as its sparse expression elsewhere implies a reduced risk of side-effects. Here we compared glycine-mediated conformational changes in ?1 and ?3 GlyRs to identify structural differences that might be exploited in designing ?3-specific analgesics. Using voltage-clamp fluorometry, we show that glycine-mediated conformational changes in the extracellular M2-M3 domain were significantly different between the two GlyR isoforms. Using a chimeric approach, we found that structural variations in the intracellular M3-M4 domain were responsible for this difference. This prompted us to test the hypothesis that phosphorylation of S346 in ?3 GlyR might also induce extracellular conformation changes. We show using both voltage-clamp fluorometry and pharmacology that Ser346 phosphorylation elicits structural changes in the ?3 glycine-binding site. These results provide the first direct evidence for phosphorylation-mediated extracellular conformational changes in pentameric ligand-gated ion channels, and thus suggest new loci for investigating how phosphorylation modulates structure and function in this receptor family. More importantly, by demonstrating that phosphorylation alters ?3 GlyR glycine-binding site structure, they raise the possibility of developing analgesics that selectively target inflammation-modulated GlyRs.
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A rare occurrence of pulmonary alveolar proteinosis after lung transplantation.
Semin Respir Crit Care Med
PUBLISHED: 07-02-2013
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We present a case of pulmonary alveolar proteinosis (PAP) initially diagnosed 28 months after left single-lung transplantation for idiopathic pulmonary fibrosis. The diagnosis was based upon the presence of periodic acid-Schiff (PAS)-positive and surfactant immunostain-positive acellular lipoproteinaceous material within alveoli seen on transbronchial biopsy as well as in bronchoalveolar lavage fluid. The patient eventually also displayed a characteristic "crazy paving" pattern on radiographic imaging. Granulocyte macrophage-colony stimulating factor antibodies were negative, consistent with secondary PAP. PAP is a rare interstitial lung disease with only a few reported cases occurring after lung transplantation. The etiology is thought to be related to a defect in macrophage function caused by immunosuppression. Reduced immunosuppression has been associated with stabilization, but not reversal, of the condition in the case reported here. PAP is an exceptionally rare cause of dyspnea and radiographic infiltrates after lung transplantation and may be related to toxicity of immune-suppressive medications.
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DNA viral infections complicating lung transplantation.
Semin Respir Crit Care Med
PUBLISHED: 07-02-2013
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DNA viruses with potential to cause complications after lung transplantation include the human Herpesviridae family consisting of cytomegalovirus (CMV), herpes simplex virus 1 and 2 (HSV-1, -2), varicella-zoster virus (VZV), human herpesvirus 6, 7, and 8 (HHV-6, -7, -8), and Epstein-Barr virus (EBV); the Polyomaviridae family consisting of BK virus and JC virus; and the Adenoviridae family consisting of more than 50 adenovirus subtypes. This is a diverse group of viruses with equally diverse immediate and long-term impacts on allograft function and clinical outcomes following lung transplantation. This article discusses the individual pathogens, their epidemiology and clinical manifestations, as well as treatment and preventive strategies in this era of antiviral treatment regimens.
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Bronchiolitis obliterans syndrome: the Achilles heel of lung transplantation.
Semin Respir Crit Care Med
PUBLISHED: 07-02-2013
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Lung transplantation is a therapeutic option for patients with end-stage pulmonary disorders. Unfortunately, chronic lung allograft dysfunction (CLAD), most commonly manifest as bronchiolitis obliterans syndrome (BOS), continues to be highly prevalent and is the major limitation to long-term survival. The pathogenesis of BOS is complex and involves alloimmune and nonalloimmune pathways. Clinically, BOS manifests as airway obstruction and dyspnea that are classically progressive and ultimately fatal; however, the course is highly variable, and distinguishable phenotypes may exist. There are few controlled studies assessing treatment efficacy, but only a minority of patients respond to current treatment modalities. Ultimately, preventive strategies may prove more effective at prolonging survival after lung transplantation, but their remains considerable debate and little data regarding the best strategies to prevent BOS. A better understanding of the risk factors and their relationship to the pathological mechanisms of chronic lung allograft rejection should lead to better pharmacological targets to prevent or treat this syndrome.
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Sesterterpene glycinyl-lactams: a new class of glycine receptor modulator from Australian marine sponges of the genus Psammocinia.
Org. Biomol. Chem.
PUBLISHED: 06-11-2013
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Bioassay guided fractionation of three southern Australian marine sponges of the genus Psammocinia, selected for their ability to modulate glycine-gated chloride channel receptors (GlyRs), yielded the rare marine sesterterpenes (-)-ircinianin (1) and (-)-ircinianin sulfate (2), along with the new biosynthetically related metabolites (-)-ircinianin lactam A (3), (-)-ircinianin lactam A sulfate (4), (-)-oxoircinianin (5), (-)-oxoircinianin lactam A (6) and (-)-ircinianin lactone A (7). Acetylation of 1 returned (-)-ircinianin acetate (8). Whole cell patch-clamp electrophysiology on 1-8 established 3 as an exceptionally potent and selective ?3 GlyR potentiator, and 6 as a selective ?1 GlyR potentiator. The discovery and characterization of sesterterpenes 1-8, and in particular the glycinyl-lactams 3 and 6, provide valuable new insights into GlyR pharmacology. These insights have the potential to inform and inspire the development of new molecular tools to probe GlyR distribution and function, and therapeutics to treat a wide array of GlyR mediated diseases and disorders.
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Preoperative cardiac variables of diastolic dysfunction and clinical outcomes in lung transplant recipients.
J Transplant
PUBLISHED: 04-24-2013
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Background. Orthotopic lung transplantation is now widely performed in patients with advanced lung disease. Patients with moderate or severe ventricular systolic dysfunction are typically excluded from lung transplantation; however, there is a paucity of data regarding the prognostic significance of abnormal left ventricular diastolic function and elevated pretransplant pulmonary pressures. Methods. We reviewed the characteristics of 111 patients who underwent bilateral and unilateral lung transplants from 200 to 2009 in order to evaluate the prognostic significance of preoperative markers of diastolic function, including invasively measured pulmonary capillary wedge pressure (PCWP) and echocardiographic variables of diastolic dysfunction including mitral A > E and A > E. Results. Out of 111 patients, 62 were male (56%) and average age was 54.0 ± 10.5 years. Traditional echocardiographic Doppler variables of abnormal diastolic function, including A > E and A > E, did not predict adverse events (P = 0.49). Mildly elevated pretransplant PCWP (16-20?mmHg) and moderately/severely elevated PCWP (>20?mmHg) were not associated with adverse clinical events after transplant (P = 0.30). Additionally, all clinical endpoints did not show any statistical significance between the two groups. Conclusions. Pre-lung transplant invasive and echocardiographic findings of elevated pulmonary pressures and abnormal left ventricular diastolic function are not predictive of adverse posttransplant clinical events.
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Asymmetric synthesis of cis-2,5-disubstituted pyrrolidine, the core scaffold of ?3-AR agonists.
Org. Lett.
PUBLISHED: 03-01-2013
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A practical, enantioselective synthesis of cis-2,5-disubstituted pyrrolidine is described. Application of an enzymatic DKR reduction of a keto ester, which is easily accessed through a novel intramolecular N?C benzoyl migration, yields syn-1,2-amino alcohol in >99% ee and >99:1 dr. Subsequent hydrogenation of cyclic imine affords the cis-pyrrolidine in high diastereoselectivity. By integrating biotechnology into organic synthesis and isolating only three intermediates over 11 steps, the core scaffold of ?3-AR agonists is synthesized in 38% overall yield.
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Australian marine sponge alkaloids as a new class of glycine-gated chloride channel receptor modulator.
Bioorg. Med. Chem.
PUBLISHED: 02-25-2013
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Chemical analysis of a specimen of the sponge Ianthella cf. flabelliformis returned two new sesquiterpene glycinyl lactams, ianthellalactams A (1) and B (2), the known sponge sesquiterpene dictyodendrillin (3) and its ethanolysis artifact ethyl dictyodendrillin (4), and five known sponge indole alkaloids, aplysinopsin (5), 8E-3-deimino-3-oxoaplysinopsin (6), 8Z-3-deimino-3-oxoaplysinopsin (7), dihydroaplysinopsin (8) and tubastrindole B (9). The equilibrated mixture 6/7 exhibited glycine-gated chloride channel receptor (GlyR) antagonist activity with a bias towards ?3 over ?1 GlyR, while tubastrindole B (9) exhibited a bias towards ?1 over ?3 GlyR. At low- to sub-micromolar concentrations, 9 was also a selective potentiator of ?1 GlyR, with no effect on ?3 GlyR-a pharmacology that could prove useful in the treatment of movement disorders such as spasticity and hyperekplexia. Our investigations into the GlyR modulatory properties of 1-9 were further supported by the synthesis of a number of structurally related indole alkaloids.
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Fluorescence-based high-throughput functional profiling of ligand-gated ion channels at the level of single cells.
PLoS ONE
PUBLISHED: 02-06-2013
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Ion channels are involved in many physiological processes and are attractive targets for therapeutic intervention. Their functional properties vary according to their subunit composition, which in turn varies in a developmental and tissue-specific manner and as a consequence of pathophysiological events. Understanding this diversity requires functional analysis of ion channel properties in large numbers of individual cells. Functional characterisation of ligand-gated channels involves quantitating agonist and drug dose-response relationships using electrophysiological or fluorescence-based techniques. Electrophysiology is limited by low throughput and high-throughput fluorescence-based functional evaluation generally does not enable the characterization of the functional properties of each individual cell. Here we describe a fluorescence-based assay that characterizes functional channel properties at single cell resolution in high throughput mode. It is based on progressive receptor activation and iterative fluorescence imaging and delivers >100 dose-responses in a single well of a 384-well plate, using ?1-3 homomeric and ?? heteromeric glycine receptor (GlyR) chloride channels as a model system. We applied this assay with transiently transfected HEK293 cells co-expressing halide-sensitive yellow fluorescent protein and different GlyR subunit combinations. Glycine EC50 values of different GlyR isoforms were highly correlated with published electrophysiological data and confirm previously reported pharmacological profiles for the GlyR inhibitors, picrotoxin, strychnine and lindane. We show that inter and intra well variability is low and that clustering of functional phenotypes permits identification of drugs with subunit-specific pharmacological profiles. As this method dramatically improves the efficiency with which ion channel populations can be characterized in the context of cellular heterogeneity, it should facilitate systems-level analysis of ion channel properties in health and disease and the discovery of therapeutics to reverse pathological alterations.
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Ceftazidime-avibactam: a novel cephalosporin/?-lactamase inhibitor combination.
Drugs
PUBLISHED: 02-02-2013
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Avibactam (formerly NXL104, AVE1330A) is a synthetic non-?-lactam, ?-lactamase inhibitor that inhibits the activities of Ambler class A and C ?-lactamases and some Ambler class D enzymes. This review summarizes the existing data published for ceftazidime-avibactam, including relevant chemistry, mechanisms of action and resistance, microbiology, pharmacokinetics, pharmacodynamics, and efficacy and safety data from animal and human trials. Although not a ?-lactam, the chemical structure of avibactam closely resembles portions of the cephem bicyclic ring system, and avibactam has been shown to bond covalently to ?-lactamases. Very little is known about the potential for avibactam to select for resistance. The addition of avibactam greatly (4-1024-fold minimum inhibitory concentration [MIC] reduction) improves the activity of ceftazidime versus most species of Enterobacteriaceae depending on the presence or absence of ?-lactamase enzyme(s). Against Pseudomonas aeruginosa, the addition of avibactam also improves the activity of ceftazidime (~fourfold MIC reduction). Limited data suggest that the addition of avibactam does not improve the activity of ceftazidime versus Acinetobacter species or most anaerobic bacteria (exceptions: Bacteroides fragilis, Clostridium perfringens, Prevotella spp. and Porphyromonas spp.). The pharmacokinetics of avibactam follow a two-compartment model and do not appear to be altered by the co-administration of ceftazidime. The maximum plasma drug concentration (C(max)) and area under the plasma concentration-time curve (AUC) of avibactam increase linearly with doses ranging from 50 mg to 2,000 mg. The mean volume of distribution and half-life of 22 L (~0.3 L/kg) and ~2 hours, respectively, are similar to ceftazidime. Like ceftazidime, avibactam is primarily renally excreted, and clearance correlates with creatinine clearance. Pharmacodynamic data suggest that ceftazidime-avibactam is rapidly bactericidal versus ?-lactamase-producing Gram-negative bacilli that are not inhibited by ceftazidime alone.Clinical trials to date have reported that ceftazidime-avibactam is as effective as standard carbapenem therapy in complicated intra-abdominal infection and complicated urinary tract infection, including infection caused by cephalosporin-resistant Gram-negative isolates. The safety and tolerability of ceftazidime-avibactam has been reported in three phase I pharmacokinetic studies and two phase II clinical studies. Ceftazidime-avibactam appears to be well tolerated in healthy subjects and hospitalized patients, with few serious drug-related treatment-emergent adverse events reported to date.In conclusion, avibactam serves to broaden the spectrum of ceftazidime versus ß-lactamase-producing Gram-negative bacilli. The exact roles for ceftazidime-avibactam will be defined by efficacy and safety data from further clinical trials. Potential future roles for ceftazidime-avibactam include the treatment of suspected or documented infections caused by resistant Gram-negative-bacilli producing extended-spectrum ß-lactamase (ESBL), Klebsiella pneumoniae carbapenemases (KPCs) and/or AmpC ß-lactamases. In addition, ceftazidime-avibactam may be used in combination (with metronidazole) for suspected polymicrobial infections. Finally, the increased activity of ceftazidime-avibactam versus P. aeruginosa may be of clinical benefit in patients with suspected or documented P. aeruginosa infections.
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The new antiarrhythmic drug vernakalant: ex vivo study of human atrial tissue from sinus rhythm and chronic atrial fibrillation.
Cardiovasc. Res.
PUBLISHED: 01-22-2013
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Vernakalant is a newly developed antiarrhythmic drug against atrial fibrillation (AF). However, its electrophysiological actions on human myocardium are unknown.
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Interaction between Pseudomonas and CXC chemokines increases risk of bronchiolitis obliterans syndrome and death in lung transplantation.
Am. J. Respir. Crit. Care Med.
PUBLISHED: 01-17-2013
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Rationale: Pseudomonas aeruginosa is the most commonly isolated gram-negative bacterium after lung transplantation and has been shown to up-regulate glutamic acid-leucine-arginine-positive (ELR(+)) CXC chemokines associated with bronchiolitis obliterans syndrome (BOS), but the effect of pseudomonas on BOS and death has not been well defined. Objectives: To determine if the influence of pseudomonas isolation and ELR(+) CXC chemokines on the subsequent development of BOS and the occurrence of death is time dependent. Methods: A three-state model was developed to assess the likelihood of transitioning from lung transplant (state 1) to BOS (state 2), from transplant (state 1) to death (state 3), and from BOS (state 2) to death (state 3). This Cox semi-Markovian approach determines state survival rates and cause-specific hazards for movement from one state to another. Measurements and Main Results: The likelihood of transition from transplant to BOS was increased by acute rejection, CXCL5, and the interaction between pseudomonas and CXCL1. The pseudomonas effect in this transition was due to infection rather than colonization. Movement from transplant to death was facilitated by pseudomonas infection and single lung transplant. Transition from BOS to death was affected by the length of time in state 1 and by the interactions between any pseudomonas isolation and CXCL5 and aspergillus, either independently or in combination. Conclusions: Our model demonstrates that common post-transplantation events drive movement from one post-transplantation state to another and influence outcomes differently depending upon when after transplantation they occur. Pseudomonas and the ELR(+) CXC chemokines may interact to negatively influence lung transplant outcomes.
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Evolution of antimicrobial resistance among Enterobacteriaceae (focus on extended spectrum ?-lactamases and carbapenemases).
Expert Opin Pharmacother
PUBLISHED: 01-15-2013
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Bacteria within the family Enterobacteriaceae are important pathogens in nosocomial and community settings. Over the past two decades, antimicrobial resistance among Enterobacteriaceae dramatically escalated worldwide. The authors review the mechanisms of antimicrobial resistance among Enterobacteriaceae, epidemiology and global spread of resistance elements and discuss therapeutic options.
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Molecular determinants of ivermectin sensitivity at the glycine receptor chloride channel.
J. Biol. Chem.
PUBLISHED: 10-27-2011
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Ivermectin is an anthelmintic drug that works by activating glutamate-gated chloride channel receptors (GluClRs) in nematode parasites. GluClRs belong to the Cys-loop receptor family that also includes glycine receptor (GlyR) chloride channels. GluClRs and A288G mutant GlyRs are both activated by low nanomolar ivermectin concentrations. The crystal structure of the Caenorhabditis elegans ? GluClR complexed with ivermectin has recently been published. Here, we probed ivermectin sensitivity determinants on the ?1 GlyR using site-directed mutagenesis and electrophysiology. Based on a mutagenesis screen of transmembrane residues, we identified Ala288 and Pro230 as crucial sensitivity determinants. A comparison of the actions of selamectin and ivermectin suggested the benzofuran C05-OH was required for high efficacy. When taken together with docking simulations, these results supported a GlyR ivermectin binding orientation similar to that seen in the GluClR crystal structure. However, whereas the crystal structure shows that ivermectin interacts with the ? GluClR via H-bonds with Leu218, Ser260, and Thr285 (? GluClR numbering), our data indicate that H-bonds with residues homologous to Ser260 and Thr285 are not important for high ivermectin sensitivity or direct agonist efficacy in A288G ?1 GlyRs or three other GluClRs. Our data also suggest that van der Waals interactions between the ivermectin disaccharide and GlyR M2-M3 loop residues are unimportant for high ivermectin sensitivity. Thus, although our results corroborate the ivermectin binding orientation as revealed by the crystal structure, they demonstrate that some of the binding interactions revealed by this structure do not pertain to other highly ivermectin-sensitive Cys-loop receptors.
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Activation and desensitization induce distinct conformational changes at the extracellular-transmembrane domain interface of the glycine receptor.
J. Biol. Chem.
PUBLISHED: 09-14-2011
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Most ligand-gated channels exhibit desensitization, which is the progressive fading of ionic current in the prolonged presence of agonist. This process involves conformational changes that close the channel despite continued agonist binding. Despite the physiological and pathological importance of desensitization, little is known about the conformational changes that underlie this process in any Cys-loop ion channel receptor. Here we employed voltage clamp fluorometry to identify conformational changes that occur with a similar time course as the current desensitization rate in both slow- and fast-desensitizing ?1 glycine receptor chloride channels. Voltage clamp fluorometry provides a direct indication of conformational changes that occur in the immediate vicinity of residues labeled with environmentally sensitive fluorophores. We compared the rates of current desensitization and fluorescence changes at nine labeled extracellular sites in both wild type slow-desensitizing and mutated (A248L) fast-desensitizing glycine receptors. As labels attached to three sites at the interface between the ligand binding domain and transmembrane domain reported fluorescence responses that changed in parallel with the current desensitization rate, we concluded that they experienced local conformational changes associated with desensitization. These labeled sites included A52C in loop 2, Q219C in the pre-M1 domain, and M227C in the M1 domain. Activation and desensitization were accompanied by physically distinct conformational changes at each labeled site. Because activation is mediated by a specific reorganization of molecular interactions at the extracellular-transmembrane domain interface, we propose that desensitization is mediated by a distinct set of conformational changes that prevents this reorganization from occurring, thereby favoring channel closure.
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A comparison of glycine- and ivermectin-mediated conformational changes in the glycine receptor ligand-binding domain.
Int. J. Biochem. Cell Biol.
PUBLISHED: 09-12-2011
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Glycine receptor chloride channels are Cys-loop receptor proteins that isomerize between a low affinity closed state and a high affinity ion-conducting state. There is currently much interest in understanding the mechanisms that link affinity changes with conductance changes. This essentially involves an agonist binding in the glycine receptor ligand-binding site initiating local conformational changes that propagate in a wave towards the channel gate. However, it has proved difficult to convincingly distinguish those agonist-induced domain movements that are critical for triggering activation from those that are simply local deformations to accommodate ligands in the site. We employed voltage-clamp fluorometry to compare conformational changes in the ligand-binding site in response to activation by glycine, which binds locally, and ivermectin, which binds in the transmembrane domain. We reasoned that ivermectin-mediated activation should initiate a conformational wave that propagates from the pore-lining domain towards the ligand-binding domain, eliciting conformational changes in those extracellular domains that are allosterically linked to the gate. We found that ivermectin indeed elicited conformational changes in ligand-binding domain loops C, D and F. This implies that conformational changes in these domains are important for activation. This result also provides a mechanism to explain how ivermectin potentiates glycine-induced channel activation.
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Adenovirus.
Semin Respir Crit Care Med
PUBLISHED: 08-19-2011
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Adenoviruses (AdV) are DNA viruses that typically cause mild infections involving the upper or lower respiratory tract, gastrointestinal (GI) tract, or conjunctiva. Rare manifestations of AdV infections include hemorrhagic cystitis, hepatitis, hemorrhagic colitis, pancreatitis, nephritis, or encephalitis. Adenovirus infections are more common in young children, owing to lack of humoral immunity. Epidemics of AdV infections may occur in healthy children or adults in closed or crowded settings (particularly military recruits). The disease is more severe, and dissemination is more likely in patients with impaired immunity (eg, organ transplant recipients, human immunodeficiency virus infection, congenital immunodeficiency syndromes). Fatality rates for untreated severe AdV pneumonia or disseminated disease may exceed 50%. More than 50 serotypes of AdV have been identified. Different serotypes display different tissue trophisms and correlate with clinical manifestations of infection. The predominant serotypes differ among countries or regions and change over time. Transmission of novel strains between countries or across continents and replacement of dominant serotypes by new strains may occur. Treatment of AdV infections is controversial because prospective, randomized therapeutic trials have not been done. Cidofovir is considered the drug of choice for severe AdV infections, but not all patients require treatment. Vaccines have been shown to be highly efficacious in reducing the risk of respiratory AdV infection but are currently not available.
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Respiratory viral infections in hematopoietic stem cell and solid organ transplant recipients.
Semin Respir Crit Care Med
PUBLISHED: 08-19-2011
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Respiratory viral infections (RVIs) are common causes of mild illness in immunocompetent children and adults with rare occurrences of significant morbidity or mortality. Complications are more common in the very young, very old, and those with underlying lung diseases. However, RVIs are increasingly recognized as a cause of morbidity and mortality in recipients of hematopoietic stem cell transplants (HSCT) and solid organ transplants (SOTs). Diagnostic techniques for respiratory syncytial virus (RSV), parainfluenza, influenza, and adenovirus have been clinically available for decades, and these infections are known to cause serious disease in transplant recipients. Modern molecular technology has now made it possible to detect other RVIs including human metapneumovirus, coronavirus, and bocavirus, and the role of these viruses in causing serious disease in transplant recipients is still being worked out. This article reviews the current information regarding epidemiology, pathogenesis, clinical presentation, diagnosis, and treatment of these infections, as well as the aspects of clinical significance of RVIs unique to HSCT or SOT.
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Influenza: epidemiology, clinical features, therapy, and prevention.
Semin Respir Crit Care Med
PUBLISHED: 08-19-2011
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Influenza A and B are important causes of respiratory illness in all age groups. Influenza causes seasonal outbreaks globally and, less commonly, pandemics. In the United States, seasonal influenza epidemics account for >200,000 hospitalizations and >30,000 deaths annually. More than 90% of deaths occur in the elderly population. Interestingly, in the novel 2009 H1N1 influenza pandemic, attack rates were highest among children and young adults. Fewer than 10% of cases occurred in adults >60 years old, likely because preexisting antibodies against other H1N1 viruses afforded protection. Despite concerns about a high lethality rate with the novel 2009 H1N1 strain, most illnesses caused by the 2009 H1N1 viruses were mild (overall case fatality rate <0.5%). Clinical features of influenza infection overlap with other respiratory pathogens (particularly viruses). The diagnosis is often delayed due to low suspicion and the limited use of specific diagnostic tests. Rapid diagnostic tests are widely available and allow detection of influenza antigen in respiratory secretions within 1 hour; however, sensitivity ranges from 50 to 90%. Neuraminidase inhibitors (NAIs) (eg, oseltamivir and zanamivir) are effective for treating influenza A or B and for prophylaxis in selected adults and children. Resistance to NAIs is rare, but influenza strains resistant to oseltamivir have been detected. Vaccines are the cornerstone of influenza control. Currently, trivalent inactivated vaccine (TIV) and live attenuated influenza vaccine (LAIV) are available. These agents reduce mortality and morbidity in high-risk patients (i.e., the elderly or patients with comorbidities), and expanding the use of vaccines to healthy children and adults reduces the incidence of influenza, pneumonia, and hospitalizations due to respiratory illnesses in the community.
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Comparison of electrophysiological and antiarrhythmic effects of vernakalant, ranolazine, and sotalol in canine pulmonary vein sleeve preparations.
Heart Rhythm
PUBLISHED: 08-10-2011
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Vernakalant (VER) is a relatively atrial-selective antiarrhythmic drug capable of blocking potassium and sodium currents in a frequency- and voltage-dependent manner. Ranolazine (RAN) is a sodium-channel blocker shown to exert antiarrhythmic effects in pulmonary vein (PV) sleeves. dl-Sotalol (SOT) is a ?-blocker commonly used in the rhythm-control treatment of atrial fibrillation. This study evaluated the electrophysiological and antiarrhythmic effects of VER, RAN, and SOT in canine PV sleeve preparations in a blinded fashion.
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? Subunit M2-M3 loop conformational changes are uncoupled from ?1 ? glycine receptor channel gating: implications for human hereditary hyperekplexia.
PLoS ONE
PUBLISHED: 06-30-2011
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Hereditary hyperekplexia, or startle disease, is a neuromotor disorder caused mainly by mutations that either prevent the surface expression of, or modify the function of, the human heteromeric ?1 ? glycine receptor (GlyR) chloride channel. There is as yet no explanation as to why hyperekplexia mutations that modify channel function are almost exclusively located in the ?1 to the exclusion of ? subunit. The majority of these mutations are identified in the M2-M3 loop of the ?1 subunit. Here we demonstrate that ?1 ? GlyR channel function is less sensitive to hyperekplexia-mimicking mutations introduced into the M2-M3 loop of the ? than into the ?1 subunit. This suggests that the M2-M3 loop of the ? subunit dominates the ? subunit in gating the ?1 ? GlyR channel. A further attempt to determine the possible mechanism underlying this phenomenon by using the voltage-clamp fluorometry technique revealed that agonist-induced conformational changes in the ? subunit M2-M3 loop were uncoupled from ?1 ? GlyR channel gating. This is in contrast to the ? subunit, where the M2-M3 loop conformational changes were shown to be directly coupled to ?1 ? GlyR channel gating. Finally, based on analysis of ?1 ? chimeric receptors, we demonstrate that the structural components responsible for this are distributed throughout the ? subunit, implying that the ? subunit has evolved without the functional constraint of a normal gating pathway within it. Our study provides a possible explanation of why hereditary hyperekplexia-causing mutations that modify ?1 ? GlyR channel function are almost exclusively located in the ?1 to the exclusion of the ? subunit.
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Immunosuppressive and cytotoxic therapy: pharmacology, toxicities, and monitoring.
Semin Respir Crit Care Med
PUBLISHED: 06-14-2011
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Treatment strategies for anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) are evolving. Cyclophosphamide (CYC) plus corticosteroids (CSs) is the mainstay of therapy for generalized, multisystemic AAV. Historically, the combination of CYC plus CS was used for a minimum of 12 months, but concern about late toxicities associated with CYC has led to novel treatment approaches. Currently, short-course (3 to 6 months) induction treatment with CYC plus CS, followed by maintenance therapy with less toxic agents (eg, methotrexate, azathioprine, mycophenolate mofetil) is recommended. Further, methotrexate combined with CS may be adequate for limited, non-life-threatening AAV. Recent studies suggest that rituximab may be useful for induction therapy or for CYC-refractory AAV. This article reviews the key agents used to treat AAV, with a focus on pharmacology, toxicities, and monitoring.
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Wegener granulomatosis (granulomatosis with polyangiitis): evolving concepts in treatment.
Semin Respir Crit Care Med
PUBLISHED: 06-14-2011
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Wegener granulomatosis (WG), the most common of the pulmonary granulomatous vasculitides, typically involves the upper respiratory tract, lower respiratory tract (bronchi and lung), and kidney, with varying degrees of disseminated vasculitis. THE TERM GRANULOMATOSIS WITH POLYANGIITIS (WEGENER) WAS RECENTLY PROPOSED TO REPLACE THE OLDER TERM, WG. THE TERM GRANULOMATOSIS WITH POLYANGIITIS CAN BE ABBREVIATED TO GPA, WITH THE IDEA THAT THE EPONYM WEGENER WOULD BE OMITTED OVER TIME. Cardinal histologic features include a necrotizing vasculitis involving small vessels, extensive "geographic" necrosis, and granulomatous inflammation. Clinical manifestations of WG are protean; virtually any organ can be involved. The spectrum and severity of the disease are heterogeneous, ranging from indolent disease involving only one site to fulminant, multiorgan vasculitis. The pathogenesis of WG has not been elucidated, but both cellular and humoral components are involved. Circulating antibodies against cytoplasmic components of neutrophils [anti-neutrophil cytoplasmic antibodies (c-ANCAs)] likely play a role in the pathogenesis, and often correlate with activity of the disease. Treatment strategies are evolving. Cyclophosphamide (CYC) plus corticosteroids (CSs) is the mainstay of therapy for generalized, multisystemic WG. Historically, the combination of CYC plus CS was used for a minimum of 12 months, but concern about late toxicities associated with CYC has led to novel treatment approaches. Currently, short-course (3 to 6 months) induction treatment with CYC plus CS, followed by maintenance therapy with less toxic agents (e.g., methotrexate, azathioprine) is recommended. Further, methotrexate combined with CS may be adequate for limited, non-life-threatening WG. Recent studies suggest that rituximab may be useful for induction therapy or CYC-refractory WG. The role of other immunomodulatory agents (including trimethoprim-sulfamethoxazole) is also explored.
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Efficacy and safety of sirolimus in lymphangioleiomyomatosis.
N. Engl. J. Med.
PUBLISHED: 03-16-2011
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Lymphangioleiomyomatosis (LAM) is a progressive, cystic lung disease in women; it is associated with inappropriate activation of mammalian target of rapamycin (mTOR) signaling, which regulates cellular growth and lymphangiogenesis. Sirolimus (also called rapamycin) inhibits mTOR and has shown promise in phase 1-2 trials involving patients with LAM.
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Lung transplantation and coronary artery disease.
Ann. Thorac. Surg.
PUBLISHED: 02-09-2011
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Coronary artery disease (CAD) remains a relative contraindication to lung transplantation. We have offered lung transplantation and coronary revascularization to selected patients with discrete CAD and preserved left ventricular function. The purpose of this report is the following: (1) to examine the short-term and medium-term outcome of patients after coronary revascularization and lung transplantation; and (2) to compare the short-term and medium-term outcome of this cohort to a matched group of lung transplant recipients without CAD.
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A cation-? interaction at a phenylalanine residue in the glycine receptor binding site is conserved for different agonists.
Mol. Pharmacol.
PUBLISHED: 01-25-2011
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Cation-? interactions have been demonstrated to play a major role in agonist-binding in Cys-loop receptors. However, neither the aromatic amino acid contributing to this interaction nor its location is conserved among Cys-loop receptors. Likewise, it is not clear how many different agonists of a given receptor form a cation-? interaction or, if they do, whether it is with the same aromatic amino acid as the major physiological agonist. We demonstrated previously that Phe159 in the glycine receptor (GlyR) ?1 subunit forms a strong cation-? interaction with the principal agonist, glycine. In the current study, we investigated whether the lower efficacy agonists of the human GlyR ?-alanine and taurine also form cation-? interactions with Phe159. By incorporating a series of unnatural amino acids, we found cation-? interactions between Phe159 and the amino groups of ?-alanine and taurine. The strengths of these interactions were significantly weaker than for glycine. Modeling studies suggest that ?-alanine and taurine are orientated subtly differently in the binding pocket, with their amino groups further from Phe159 than that of glycine. These data therefore show that similar agonists can have similar but not identical orientations and interactions in the binding pocket and provide a possible explanation for the lower potencies of ?-alanine and taurine.
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Immune response CC chemokines CCL2 and CCL5 are associated with pulmonary sarcoidosis.
Fibrogenesis Tissue Repair
PUBLISHED: 01-17-2011
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Pulmonary sarcoidosis involves an intense leukocyte infiltration of the lung with the formation of non-necrotizing granulomas. CC chemokines (chemokine (C-C motif) ligand 2 (CCL2)-CCL5) are chemoattractants of mononuclear cells and act through seven transmembrane G-coupled receptors. Previous studies have demonstrated conflicting results with regard to the associations of these chemokines with sarcoidosis. In an effort to clarify previous discrepancies, we performed the largest observational study to date of CC chemokines in bronchoalveolar lavage fluid (BALF) from patients with pulmonary sarcoidosis.
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Thermal effects of glenoid reaming during shoulder arthroplasty in vivo.
J Bone Joint Surg Am
PUBLISHED: 01-07-2011
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Glenoid component loosening is a common cause of failure of total shoulder arthroplasty. It has been proposed that the heat generated during glenoid preparation may reach temperatures capable of producing osteonecrosis at the bone-implant interface. We hypothesized that temperatures sufficient to induce thermal necrosis can be produced with routine drilling and reaming during glenoid preparation for shoulder arthroplasty in vivo. Furthermore, we hypothesized that irrigation of the glenoid during reaming can reduce this temperature increase.
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Non-tuberculous mycobacterium infection after lung transplantation is associated with increased mortality.
J. Heart Lung Transplant.
PUBLISHED: 01-03-2011
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Pulmonary non-tuberculous mycobacterial (NTM) infection is relatively common after lung transplantation, but the effect on mortality remains undetermined. Herein we describe our experience with pulmonary NTM infection after lung transplantation and hypothesized that non-tuberculous mycobacterial infection after lung transplantation would be associated with increased mortality.
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Ligand- and subunit-specific conformational changes in the ligand-binding domain and the TM2-TM3 linker of {alpha}1 {beta}2 {gamma}2 GABAA receptors.
J. Biol. Chem.
PUBLISHED: 10-11-2010
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Cys-loop receptor ligand binding sites are located at subunit interfaces where they are lined by loops A-C from one subunit and loops D-F from the adjacent subunit. Agonist binding induces large conformational changes in loops C and F. However, it is controversial as to whether these conformational changes are essential for gating. Here we used voltage clamp fluorometry to investigate the roles of loops C and F in gating the ?1 ?2 ?2 GABA(A) receptor. Voltage clamp fluorometry involves labeling introduced cysteines with environmentally sensitive fluorophores and inferring structural rearrangements from ligand-induced fluorescence changes. Previous attempts to define the roles of loops C and F using this technique have focused on homomeric Cys-loop receptors. However, the problem with studying homomeric receptors is that it is difficult to eliminate the possibility of bound ligands interacting directly with attached fluorophores at the same site. Here we show that ligands binding to the ?2-?1 interface GABA binding site produce conformational changes at the adjacent subunit interface. This is most likely due to agonist-induced loop C closure directly altering loop F conformation at the adjacent ?1-?2 subunit interface. However, as antagonists and agonists produce identical ?1 subunit loop F conformational changes, these conformational changes appear unimportant for gating. Finally, we demonstrate that TM2-TM3 loops from adjacent ?2 subunits in ?1 ?2 receptors can dimerize via K24C disulfides in the closed state. This result implies unexpected conformational mobility in this crucial part of the gating machinery. Together, this information provides new insights into the activation mechanisms of Cys-loop receptors.
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Effect of calcitonin gene-related peptide receptor antagonism on the systemic blood pressure responses to mechanistically diverse vasomodulators in conscious rats.
J. Cardiovasc. Pharmacol.
PUBLISHED: 08-24-2010
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The effects of calcitonin gene-related peptide (CGRP) receptor antagonism with CGRP 8-37 on blood pressure changes evoked by the intravenous administration of the vasoactive modulators angiotensin II, phenylephrine, adenosine, nitroglycerine, and sodium nitroprusside were assessed in conscious rats. The effects of sumatriptan and dihydroergotamine on the blood pressure responses evoked by these vasomodulators also were assessed. The intravenous test dose of CGRP 8-37 was validated through block of depressor responses to intravenous CGRP in conscious rats, whereas the intravenous test doses of sumatriptan and dihydroergotamine were validated by reductions in carotid blood flow in anesthetized rats. CGRP 8-37 had no significant effects on blood pressure dose-response profiles and individual dose blood pressure responses to any of the vasomodulators tested. In contrast, sumatriptan altered the blood pressure dose-response profiles to angiotensin II and sodium nitroprusside (P < 0.03) and dihydroergotamine altered the blood pressure dose-response profile to sodium nitroprusside (P < 0.02) and tended to alter that of phenylephrine (P = 0.06). Both sumatriptan and dihydroergotamine displayed frequent alterations of individual dose blood pressure responses to all vasomodulators. These findings are consistent with concerns for sumatriptan and dihydroergotamine to alter systemic hemodynamics, whereas CGRP receptor antagonism did not display the same hemodynamic liability.
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Exercise-induced pulmonary hypertension associated with systemic sclerosis: four distinct entities.
Arthritis Rheum.
PUBLISHED: 08-20-2010
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Exercise-induced pulmonary hypertension (PH) may represent an early but clinically relevant phase in the spectrum of pulmonary vascular disease. There are limited data on the prevalence of exercise-induced PH determined by right heart catheterization in scleroderma spectrum disorders. We undertook this study to describe the hemodynamic response to exercise in a homogeneous population of patients with scleroderma spectrum disorders at risk of developing pulmonary vascular disease.
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Design, synthesis and SAR of a series of 1,3,5-trisubstituted benzenes as thrombin inhibitors.
Bioorg. Med. Chem. Lett.
PUBLISHED: 08-14-2010
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A novel 1,3,5-trisubstituted benzamide thrombin inhibitor template was designed via hybridization of a known aminopyridinoneacetamide and a known 1,3,5-trisubstituted phenyl ether. Optimization of this lead afforded a novel potent series of biaryl 1,3,5-trisubstituted benzenes with excellent functional anticoagulant potency.
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P3 optimization of functional potency, in vivo efficacy and oral bioavailability in 3-aminopyrazinone thrombin inhibitors bearing non-charged groups at the P1 position.
Bioorg. Med. Chem. Lett.
PUBLISHED: 08-14-2010
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Although the S3 pocket of the thrombin active site is lined with lipophilic amino acid residues, the accommodation of polarity within the lipophilic P3 moiety of small molecule inhibitors is possible provided that the polar functionality is capable of pointing away from the binding pocket outwards toward solvent while simultaneously allowing the lipophilic portion of the P3 ligand to interact with the S3 amino acid residues. Manipulation of this motif provided the means to effect optimization of functional potency, in vivo antithrombotic efficacy and oral bioavailability in a series of 3-aminopyrazinone thrombin inhibitors which contained non-charged groups at the P1 position.
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Exon-skipping splice variants of excitatory amino acid transporter-2 (EAAT2) form heteromeric complexes with full-length EAAT2.
J. Biol. Chem.
PUBLISHED: 08-05-2010
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The glial transporter excitatory amino acid transporter-2 (EAAT2) is the main mediator of glutamate clearance in brain. The wild-type transporter (EAAT2wt) forms trimeric membrane complexes in which each protomer functions autonomously. Several EAAT2 variants are found in control and Alzheimer-diseased human brains; their expression increases with pathological severity. These variants might alter EAAT2wt-mediated transport by abrogating membrane trafficking, or by changing the configuration or functionality of the assembled transporter complex. HEK293 cells were transfected with EAAT2wt; EAAT2b, a C-terminal variant; or either of two exon-skipping variants: alone or in combination. Surface biotinylation studies showed that only the exon-7 deletion variant was not trafficked to the membrane when transfected alone, and that all variants could reach the membrane when co-transfected with EAAT2wt. Fluorescence resonance energy transfer (FRET) studies showed that co-transfected EAAT2wt and EAAT2 splice variants were expressed in close proximity. Glutamate transporter function was measured using a whole cell patch clamp technique, or by changes in membrane potential indexed by a voltage-sensitive fluorescent dye (FMP assay): the two methods gave comparable results. Cells transfected with EAAT2wt or EAAT2b showed glutamate-dependent membrane potential changes consistent with functional expression. Cells transfected with EAAT2 exon-skipping variants alone gave no response to glutamate. Co-transfection of EAAT2wt (or EAAT2b) and splice variants in various ratios significantly raised glutamate EC(50) and decreased Hill coefficients. We conclude that exon-skipping variants form heteromeric complexes with EAAT2wt or EAAT2b that traffic to the membrane but show reduced glutamate-dependent activity. This could allow glutamate to accumulate extracellularly and promote excitotoxicity.
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Pathophysiological mechanisms of dominant and recessive GLRA1 mutations in hyperekplexia.
J. Neurosci.
PUBLISHED: 07-16-2010
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Hyperekplexia is a rare, but potentially fatal, neuromotor disorder characterized by exaggerated startle reflexes and hypertonia in response to sudden, unexpected auditory or tactile stimuli. This disorder is primarily caused by inherited mutations in the genes encoding the glycine receptor (GlyR) alpha1 subunit (GLRA1) and the presynaptic glycine transporter GlyT2 (SLC6A5). In this study, systematic DNA sequencing of GLRA1 in 88 new unrelated human hyperekplexia patients revealed 19 sequence variants in 30 index cases, of which 21 cases were inherited in recessive or compound heterozygote modes. This indicates that recessive hyperekplexia is far more prevalent than previous estimates. From the 19 GLRA1 sequence variants, we have investigated the functional effects of 11 novel and 2 recurrent mutations. The expression levels and functional properties of these hyperekplexia mutants were analyzed using a high-content imaging system and patch-clamp electrophysiology. When expressed in HEK293 cells, either as homomeric alpha1 or heteromeric alpha1beta GlyRs, subcellular localization defects were the major mechanism underlying recessive mutations. However, mutants without trafficking defects typically showed alterations in the glycine sensitivity suggestive of disrupted receptor function. This study also reports the first hyperekplexia mutation associated with a GlyR leak conductance, suggesting tonic channel opening as a new mechanism in neuronal ligand-gated ion channels.
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Chimera construction using multiple-template-based sequential PCRs.
J. Neurosci. Methods
PUBLISHED: 07-10-2010
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Chimera construction between different proteins is a useful method for investigating protein structure and function relationships. However, this technique, by its traditional application, has been daunting because of its complex procedure and low success rate. Here we describe a protocol for constructing chimeras between proteins that does not require the existence of restriction sites, or the purification of intermediate PCR products, which are essential in the traditional protocols. By introducing the "multiple-template" concept, this protocol only requires the use of two or three simple PCRs followed by general subcloning steps. Most importantly, the success rate is nearly 100%.
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A glycine residue essential for high ivermectin sensitivity in Cys-loop ion channel receptors.
Int. J. Parasitol.
PUBLISHED: 06-30-2010
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Ivermectin exerts its anthelmintic effect by activating nematode Cys-loop glutamate-gated receptors. Here we show that a glycine residue at a specific transmembrane domain location is essential for high ivermectin sensitivity in both glycine- and glutamate-gated Cys-loop receptors. We also show that ivermectin sensitivity can be conferred on an ivermectin-insensitive receptor by introducing a glycine at this position. Furthermore, comparison of amino acid sequences of ivermectin-sensitive and -resistant receptors reveals that the presence of a glycine reliably predicts ivermectin sensitivity. By providing a means of identifying ivermectin-sensitive receptors, this finding should help in characterising ivermectin-resistance mechanisms and identifying new anthelmintic targets.
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Streptococcus pneumoniae: epidemiology and risk factors, evolution of antimicrobial resistance, and impact of vaccines.
Curr Opin Pulm Med
PUBLISHED: 04-09-2010
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Streptococcus pneumoniae (pneumococcus) remains an important cause of pneumonia, meningitis, bacteremias, and acute otitis media worldwide. Antimicrobial resistance among pneumococci has escalated dramatically over the past three decades, and is influenced by patterns of antibiotic use, population density, and spread of a few international clones.
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Chronic allograft rejection: epidemiology, diagnosis, pathogenesis, and treatment.
Semin Respir Crit Care Med
PUBLISHED: 03-30-2010
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Lung transplantation is a therapeutic option for patients with end-stage pulmonary disorders. Unfortunately, chronic lung allograft rejection, in the form of obliterative bronchiolitis and its clinical correlate bronchiolitis obliterans syndrome (BOS), continues to be highly prevalent and is the major limitation to long-term survival. The pathogenesis of BOS is complex and involves alloimmune and nonalloimmune pathways. The airway obstruction involved is classically progressive and unresponsive to treatment; however, the course is highly variable, and distinguishable phenotypes may exist. A better understanding of the risk factors and their relationship to the pathological mechanisms of chronic lung allograft rejection should lead to better pharmacological targets to prevent or treat this syndrome.
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Pulmonary arterial hypertension and lung transplantation.
Semin Respir Crit Care Med
PUBLISHED: 03-30-2010
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Heart-lung transplantation (HLT) and lung transplantation (LT) remain important therapies for idiopathic pulmonary arterial hypertension (IPAH), but recent advances in medical therapy can substantially delay or even obviate the need for transplantation, especially in certain PAH populations. By the early 1990s, the advent of epoprostenol, initially introduced as a bridge therapy to transplantation, in fact resulted in a survival advantage for IPAH. These benefits were comparable to those of HLT, and many patients who were thought to be destined for HLT were subsequently removed from active listing. Since 2005, however, the impact of the new lung allocation score (LAS) on IPAH has increased waiting list mortality. In the new millennium, the balance between the role of available medical therapies for PAH, the existing issues of the current LAS regarding the PAH patient, and the inherent morbidity associated with transplantation of PAH, will be critical to optimizing patient outcomes. The following discussion mainly focuses on adult IPAH, with some reference to congenital heart disease (CHD) and secondary PAH.
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An improved ivermectin-activated chloride channel receptor for inhibiting electrical activity in defined neuronal populations.
J. Biol. Chem.
PUBLISHED: 03-22-2010
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The ability to silence the electrical activity of defined neuronal populations in vivo is dramatically advancing our understanding of brain function. This technology may eventually be useful clinically for treating a variety of neuropathological disorders caused by excessive neuronal activity. Several neuronal silencing methods have been developed, with the bacterial light-activated halorhodopsin and the invertebrate allatostatin-activated G protein-coupled receptor proving the most successful to date. However, both techniques may be difficult to implement clinically due to their requirement for surgically implanted stimulus delivery methods and their use of nonhuman receptors. A third silencing method, an invertebrate glutamate-gated chloride channel receptor (GluClR) activated by ivermectin, solves the stimulus delivery problem as ivermectin is a safe, well tolerated drug that reaches the brain following systemic administration. However, the limitations of this method include poor functional expression, possibly due to the requirement to coexpress two different subunits in individual neurons, and the nonhuman origin of GluClR. Here, we describe the development of a modified human alpha1 glycine receptor as an improved ivermectin-gated silencing receptor. The crucial development was the identification of a mutation, A288G, which increased ivermectin sensitivity almost 100-fold, rendering it similar to that of GluClR. Glycine sensitivity was eliminated via the F207A mutation. Its large unitary conductance, homomeric expression, and human origin may render the F207A/A288G alpha1 glycine receptor an improved silencing receptor for neuroscientific and clinical purposes. As all known highly ivermectin-sensitive GluClRs contain an endogenous glycine residue at the corresponding location, this residue appears essential for exquisite ivermectin sensitivity.
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Discovery of triarylethanolamine inhibitors of the Kv1.5 potassium channel.
Bioorg. Med. Chem. Lett.
PUBLISHED: 02-26-2010
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A series of triarylethanolamine inhibitors of the Kv1.5 potassium channel have been prepared and evaluated for their effects in vitro and in vivo. The structure-activity relationship (SAR) studies described herein led to the development of potent, selective and orally active inhibitors of Kv1.5.
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Comparison of the vasoconstrictor effects of the calcitonin gene-related peptide receptor antagonist telcagepant (MK-0974) and zolmitriptan in human isolated coronary arteries.
J. Cardiovasc. Pharmacol.
PUBLISHED: 02-19-2010
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Studies were conducted in human isolated coronary arteries to explore the vascular effects of the calcitonin gene-related peptide (CGRP) receptor antagonist telcagepant and to compare its coronary vasoconstrictive potential to that of zolmitriptan. KCl precontracted coronary vessels were shown to relax to human alphaCGRP, with the CGRP-mediated vasorelaxation completely blocked with 30 microM telcagepant. In coronary vessels at basal tone, zolmitriptan caused a concentration-dependent contraction (pEC50 = 6.9 +/- 0.1; slope 0.94), with the greatest contraction obtained between 1 and 10 microM in most tissues. In contrast, telcagepant at concentrations up to 30 microM evoked no change in contractile tone. These findings suggest the potential for CGRP receptor antagonists to exert antimigraine efficacy in the absence of adverse effects on coronary tone.
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Ircinialactams: subunit-selective glycine receptor modulators from Australian sponges of the family Irciniidae.
Bioorg. Med. Chem.
PUBLISHED: 02-08-2010
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Screening an extract library of >2500 southern Australian and Antarctic marine invertebrates and algae for modulators of glycine receptor (GlyR) chloride channels identified three Irciniidae sponges that yielded new examples of a rare class of glycinyl lactam sesterterpene, ircinialactam A, 8-hydroxyircinialactam A, 8-hydroxyircinialactam B, ircinialactam C, ent-ircinialactam C and ircinialactam D. Structure-activity relationship (SAR) investigations revealed a new pharmacophore with potent and subunit selective modulatory properties against alpha1 and alpha3 GlyR isoforms. Such GlyR modulators have potential application as pharmacological tools, and as leads for the development of GlyR targeting therapeutics to treat chronic inflammatory pain, epilepsy, spasticity and hyperekplexia.
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Glenohumeral chondrolysis: a systematic review of 100 cases from the English language literature.
J Shoulder Elbow Surg
PUBLISHED: 01-19-2010
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Chondrolysis can be a devastating complication of shoulder arthroscopy. We undertook a review of the 100 cases reported in the English language to test the hypothesis that common factors could be identified and that the identification of these factors could suggest strategies for avoiding this complication.
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Pharmacological properties of MK-3207, a potent and orally active calcitonin gene-related peptide receptor antagonist.
J. Pharmacol. Exp. Ther.
PUBLISHED: 01-11-2010
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Calcitonin gene-related peptide (CGRP) has long been hypothesized to play a key role in migraine pathophysiology, and the advent of small-molecule antagonists has clearly demonstrated a clinical link between blocking the CGRP receptor and migraine efficacy. 2-[(8R)-8-(3,5-Difluorophenyl)-10-oxo-6,9-diazaspiro[4.5]dec-9-yl]-N-[(2R)-2-oxo-1,1,2,3-tetrahydrospiro[indene-2,3-pyrrolo[2,3-b]pyridin]-5-yl]acetamide (MK-3207) represents the third CGRP receptor antagonist to display clinical efficacy in migraine trials. Here, we report the pharmacological characterization of MK-3207, a potent and orally bioavailable CGRP receptor antagonist. In vitro, MK-3207 is a potent antagonist of the human and rhesus monkey CGRP receptors (K(i) = 0.024 nM). In common with other CGRP receptor antagonists, MK-3207 displays lower affinity for CGRP receptors from other species, including canine and rodent. As a consequence of species selectivity, the in vivo potency was assessed in a rhesus monkey pharmacodynamic assay measuring capsaicin-induced changes in forearm dermal blood flow via laser Doppler imaging. MK-3207 produced a concentration-dependent inhibition of dermal vasodilation, with plasma concentrations of 0.8 and 7 nM required to block 50 and 90% of the blood flow increase, respectively. The tritiated analog [3H]MK-3207 was used to study the binding characteristics on the human CGRP receptor. [3H]MK-3207 displayed reversible and saturable binding (K(D) = 0.06 nM), and the off-rate was determined to be 0.012 min(-1), with a t(1/2) value of 59 min. In vitro autoradiography studies on rhesus monkey brain slices identified the highest level of binding in the cerebellum, brainstem, and meninges. Finally, as an index of central nervous system penetrability, the in vivo cerebrospinal fluid/plasma ratio was determined to be 2 to 3% in cisterna magna-ported rhesus monkeys.
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Burkholderia cepacia complex: impact on the cystic fibrosis lung lesion.
Semin Respir Crit Care Med
PUBLISHED: 09-16-2009
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Burkholderia cepacia complex (Bcc) is a group of phenotypically similar nonfermenting, aerobic, gram-negative rods that infect 2 to 8% of patients with cystic fibrosis (CF). Bcc comprises several distinct species (formerly termed genomovars). Infection with Bcc has been associated with worse outcomes and survival in CF patients. Additionally, colonization with Bcc has been associated with heightened mortality following lung transplantation. Currently, the role of lung transplantation in Bcc-infected patients remains controversial. Antimicrobial therapy for Bcc is often ineffective because most strains are multidrug resistant. Combination therapy with two or three agents is typically administered, but optimal therapy has not been elucidated. Certain strains (particularly B. cenocepacia) have been associated with heightened transmissibility and virulence. Several genes or gene products have important roles in virulence, persistence of the organism(s), and transmissibility. The incidence of Bcc and specific species fluctuates over time and between centers. Bcc can be transmitted by social contact, and several major epidemic strains have disseminated globally. Strict infection control measures, including cohorting (segregation) of infected patients, dramatically curtail transmission and have become the standard of care, but such measures cause social isolation, stigmatism, and psychological impact among infected patients.
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Risk factors for femoral nonunion after femoral shaft fracture.
J Trauma
PUBLISHED: 08-26-2009
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The purpose of this study was to evaluate risk factors for nonunion after femoral nailing of femoral shaft fractures. A case-control study with two to one matching was conducted. Forty-five patients with 46 femoral nonunions (cases) and 92 patients with healed femoral shaft fractures (controls) were identified from our orthopedic trauma registry. All cases and controls were initially managed with reamed, statically locked femoral nails. The characteristics that were significantly different between the two groups were open fracture, delay to weight bearing, and tobacco use. Fracture classification, gender, direction of nail insertion (antegrade vs. retrograde), and Injury Severity Score were not predictive of nonunion. We conclude that open fracture, tobacco use, and delayed weight bearing are risk factors for femoral nonunion after intramedullary nailing for diaphyseal femur fractures.
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JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

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In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.