Reshaping surveillance for infectious diseases: less chasing of pathogens and more monitoring of drivers.

Animal health surveillance, despite its name, tends to focus on looking for disease. Often this involves searching for cases of infection with known pathogens (â€Ëœpathogen chasing'). Such an approach is both resource intensive and limited by the requirement for prior knowledge of disease likelihood. In this paper, the authors propose the gradual reshaping of surveillance towards the systems level, focusing on the processes (â€Ëœdrivers') that promote disease or health, rather than on the presence or absence of specific pathogens. Examples of relevant drivers include land-use change, increasing global interconnectedness, and finance and capital flows. Importantly, the authors suggest that surveillance should focus on detecting changes in patterns or quantities associated with such drivers. This would generate systems-level, risk-based surveillance information to identify areas where additional attention may be needed, and, over time, inform the implementation of prevention efforts. The collection, integration and analysis of data on drivers is likely to require investment in improving data infrastructures. A period of overlap would allow the two systems (traditional surveillance and driver monitoring) to be compared and calibrated. This would also lead to a better understanding of the drivers and their linkages, and thereby generate new knowledge that can improve surveillance and inform mitigation efforts. Since surveillance of drivers may give signals when changes are occurring, which could act as alerts and enable targeted mitigation, this might even enable disease to be prevented before it happens by directly intervening in the drivers themselves. Such surveillance focused on the drivers could be expected to bring additional benefits, since the same drivers promote multiple diseases. Further, focusing on drivers rather than pathogens should enable control of currently unknown diseases, making this approach particularly timely, given the increasing risk of emergence of new diseases.

La surveillance de la santé animale a tendance, malgré son nom, à se focaliser sur la recherche des maladies. Elle implique souvent de chercher les cas d'infection par des agents pathogènes connus (" chasse aux agents pathogènes "). Ce type d'approche exige non seulement beaucoup de ressources, mais elle est aussi limitée par la nécessité d'avoir une connaissance préalable de la probabilité de survenue de la maladie en question. Dans cet article, les auteurs proposent une refonte progressive de la surveillance pour la déplacer au niveau systémique, en se concentrant sur les processus (" facteurs ") influençant la maladie ou la santé plutôt que sur la présence ou non d'agents pathogènes spécifiques. Parmi les facteurs pertinents, on peut citer les changements dans l'utilisation des sols, l'interconnexion accrue au niveau mondial et les flux financiers et de capitaux. Les auteurs soulignent cet élément important : la surveillance devrait se focaliser sur la détection de changements au niveau des schémas ou des quantités associés à ces facteurs. Cela permettrait d'obtenir des informations de surveillance au niveau systémique et basées sur les risques, afin d'identifier les domaines auxquels il pourrait être nécessaire de porter une attention particulière - ce qui informerait, à terme, la mise en oeuvre des efforts de prévention. Il est probable qu'une amélioration des infrastructures de données soit nécessaire pour assurer la collecte, l'intégration et l'analyse des données sur les facteurs. Une période de chevauchement permettrait de comparer et de calibrer les deux systèmes (surveillance traditionnelle et surveillance des facteurs). Les facteurs et les liens entre eux seraient également mieux compris, ce qui générerait de nouvelles connaissances pouvant améliorer la surveillance et informer les efforts d'atténuation. Grâce à la surveillance des facteurs, des signaux pourraient être identifiés lorsque des changements se produisent, ce qui constituerait une alerte pour que des efforts d'atténuation ciblés soient mis en place afin d'intervenir directement sur les facteurs eux-mêmes et donc prévenir une maladie avant même qu'elle ne survienne. On peut s'attendre à ce que ce type de surveillance centrée sur les facteurs apporte des bénéfices supplémentaires, puisque les mêmes facteurs peuvent favoriser de multiples maladies. De surcroît, cette orientation axée sur les facteurs plutôt que sur les agents pathogènes devrait permettre de contrôler des maladies aujourd'hui inconnues, ce qui rend cette approche d'autant plus opportune, compte tenu du risque croissant d'émergence de nouvelles maladies.

La vigilancia zoosanitaria, pese a lo que su nombre indica, tiende a centrarse en la búsqueda de enfermedades, lo que a menudo pasa por tratar de localizar casos de infección por un patógeno conocido ("persecución de patógenos"). Semejante método no solo exige cuantiosos recursos, sino que además presenta la limitación de que obliga a conocer de antemano la probabilidad de aparición de una enfermedad. Los autores proponen una remodelación gradual de la vigilancia tendente a dotarla de carácter sistémico y a centrarla no tanto en la presencia o ausencia de determinados patógenos, sino en los procesos ("inductores" o "factores de inducción", drivers) que favorecen la enfermedad o la salud. Son ejemplo de tales procesos la evolución de los usos del suelo, el creciente nivel de interconexión mundial o los flujos financieros y de capitales. Un aspecto importante que apuntan los autores es que la vigilancia debería tener por objetivo la detección de cambios en las características o cantidades de esos factores de inducción. Ello generaría información de vigilancia basada en el riesgo de carácter sistémico, que serviría para determinar aquellas zonas a las que convendría prestar más atención y, con el tiempo, fundamentar la realización de actividades de prevención. Es probable que la obtención, integración y análisis de datos sobre los factores de inducción exijan inversiones para mejorar las infraestructuras de datos. Si hubiera una fase de solapamiento, sería posible comparar y valorar los resultados de ambos sistemas (vigilancia tradicional y seguimiento de los factores de inducción). Ello serviría para entender mejor los inductores y su vinculación recíproca, lo que generaría nuevos conocimientos con los que perfeccionar la vigilancia y en los que cimentar las actividades de mitigación. Dado que la vigilancia de los inductores puede generar una señal cuando se estén produciendo cambios, señal que a su vez activaría una alerta y propiciaría medidas selectivas de mitigación, podría ser que ello sirviera incluso para prevenir una enfermedad antes de que surgiera, actuando directamente sobre los propios factores de inducción. Cabría pensar que semejante tipo de vigilancia, centrarse en los inductores, puede tener otros efectos beneficiosos, en la medida en que un mismo inductor alimenta la aparición de varias enfermedades. Además, el hecho de centrarse en los factores de inducción, y no tanto en los patógenos, debería servir para controlar enfermedades actualmente desconocidas, por lo que este planteamiento, ante el creciente riesgo de aparición de nuevas enfermedades, resulta especialmente oportuno.

Surveillance and risk assessment for early detection of emerging infectious diseases in livestock.

Those who work in the area of surveillance and prevention of emerging infectious diseases (EIDs) face a challenge in accurately predicting where infection will occur and who (or what) it will affect. Establishing surveillance and control programmes for EIDs requires substantial and long-term commitment of resources that are limited in nature. This contrasts with the unquantifiable number of possible zoonotic and non-zoonotic infectious diseases that may emerge, even when the focus is restricted to diseases involving livestock. Such diseases may emerge from many combinations of, and changes in, host species, production systems, environments/habitats and pathogen types. Given these multiple elements, risk prioritisation frameworks should be used more widely to support decision-making and resource allocation for surveillance. In this paper, the authors use recent examples of EID events in livestock to review surveillance approaches for the early detection of EIDs, and highlight the need for surveillance programmes to be informed and prioritised by regularly updated risk assessment frameworks. They conclude by discussing some unmet needs in risk assessment practices for EIDs, and the need for improved coordination in global infectious disease surveillance.

Les personnes travaillant dans le domaine de la surveillance et de la prévention des maladies infectieuses émergentes (MIE) sont confrontées à la difficulté de prédire avec exactitude le lieu d'émergence d'une maladie, ainsi que l'espèce, le système ou le site affectés. La mise en place de programmes de surveillance et de lutte contre les MIE exige une mobilisation conséquente et durable de ressources nécessairement limitées. Par contraste, le nombre des maladies infectieuses zoonotiques et non zoonotiques pouvant se déclarer est impossible à quantifier, même si l'on s'en tient aux seules maladies affectant les animaux d'élevage. Ces maladies surviennent à la faveur des nombreuses et diverses configurations, associations ou modifications qui peuvent se produire parmi les espèces hôtes, les systèmes de production, les environnements ou habitats et les types d'agents pathogènes. Compte tenu de la multiplicité de ces éléments, il devrait être fait plus largement appel à des cadres de priorisation du risque afin de soutenir les processus de prise de décision et d'allocation des ressources en matière de surveillance. Les auteurs s'appuient sur des exemples récents d'événements liés à des MIE pour faire le point sur les méthodes de surveillance appliquées pour la détection précoce de ces maladies et soulignent l'importance de documenter et de prioriser les programmes de surveillance en procédant à des mises à jour régulières des cadres utilisés pour l'évaluation du risque. Ils concluent en évoquant certains aspects importants que les pratiques actuelles d'évaluation du risque ne permettent pas de couvrir lorsqu'il s'agit de MIE, ainsi que l'importance d'améliorer la coordination de la surveillance des maladies infectieuses au niveau mondial.

Cuantos trabajan en el ámbito de la vigilancia y la prevención de enfermedades infecciosas emergentes (EIE) tienen dificultades para predecir con precisión dónde va a surgir y a quién (o qué) afectará una infección. La instauración de programas de vigilancia y control de EIE exige una inversión sustancial y duradera de recursos que por definición son escasos, sobre todo teniendo en cuenta el número incalculable de enfermedades infecciosas zoonóticas y no zoonóticas que pueden aparecer, aun considerando solo aquellas que afectan al ganado. Este tipo de enfermedades pueden surgir como resultado de muchas combinaciones distintas de especie hospedadora, sistema productivo, medio/hábitat y tipo de patógeno o por efecto de cambios que se den en cualquiera de estos elementos. En vista de la multiplicidad de factores que concurren, convendría emplear de modo más generalizado un sistema de jerarquización de los riesgos en el cual fundamentar las decisiones de vigilancia y la distribución de los recursos destinados a ella. Los autores, valiéndose de ejemplos recientes de episodios infecciosos emergentes que afectaron al ganado, pasan revista a distintos métodos de vigilancia para la detección temprana de EIE y recalcan que los programas de vigilancia deben reposar en procedimientos de determinación del riesgo periódicamente actualizados y en las prioridades fijadas a partir de estos procedimientos. Por último, los autores se detienen en algunas necesidades desatendidas en la praxis de la determinación del riesgo de EIE y en la necesidad de una mejor coordinación de la vigilancia mundial de las enfermedades infecciosas.

Regulation of bone mineral density in the grey squirrel, Sciurus carolinensis: Bioavailability of calcium oxalate, and implications for bark stripping.

The damage caused when grey squirrels strip the outer bark off trees and ingest the underlying phloem can result in reduced timber quality or tree death. This is extremely costly to the UK forestry industry and can alter woodland composition, hampering conservation efforts. The calcium hypothesis proposes that grey squirrels ingest phloem to ameliorate a seasonal calcium deficiency. Calcium in the phloem predominantly takes the form of calcium oxalate (CaOx), however not all mammals can utilise CaOx as a source of calcium. Here, we present the results of a small-scale study to determine the extent to which grey squirrels can utilise CaOx. One of three custom-made diets containing calcium in varying forms and quantities (CaOx diet, Low-calcium carbonate (CaCO3 ) diet and Control diet) were fed to three treatment groups of six squirrels for 8 weeks. Bone densitometric properties were measured at the end of this time using peripheral quantitative computed tomography and micro-computed tomography. Pyridinoline-a serum marker of bone resorption-was measured regularly throughout the study. Bone mineral density and cortical mineralisation were lower in squirrels fed the CaOx diet compared to the Control group but similar to that of those on the Low-calcium diet, suggesting that calcium from calcium oxalate was not effectively utilised to maintain bone mineralisation. Whilst no differences were observed in serum pyridinoline levels between individuals on different diets, females had on average higher levels than males throughout the study. Future work should seek to determine if this apparent lack of ability to utilise CaOx is common to a large sample of grey squirrels and if so, whether it is consistent across all areas and seasons.

Quantitative interferon-gamma responses predict future disease progression in badgers naturally infected with Mycobacterium bovis.

The diagnosis and control of Mycobacterium bovis infection (bovine tuberculosis: TB) continues to present huge challenges to the British cattle industry. A clearer understanding of the magnitude and duration of immune response to M. bovis infection in the European badger (Meles meles) - a wildlife maintenance host - may assist with the future development of diagnostic tests, and vaccination and disease management strategies. Here, we analyse 5280 diagnostic test results from 550 live wild badgers from a naturally-infected population to investigate whether one diagnostic test (a gamma interferon release [IFNγ] assay, n = 550 tests) could be used to predict future positive results on two other tests for the same disease (a serological test [n = 2342 tests] and mycobacterial culture [n = 2388 tests]) and hence act as an indicator of likely bacterial excretion or disease progression. Badgers with the highest IFNγ optical density (OD) values were most likely to subsequently test positive on both serological and culture tests, and this effect was detectable for up to 24 months after the IFNγ test. Furthermore, the higher the original IFNγ OD value, the greater the chance that a badger would subsequently test positive using serology. Relationships between IFNγ titres and mycobacterial culture results from different types of clinical sample suggest that the route of infection may affect the magnitude of immune response in badgers. These findings identify further value in the IFNγ test as a useful research tool, as it may help us to target studies at animals and groups that are most likely to succumb to more progressive disease.

Active animal health surveillance in European Union Member States: gaps and opportunities.

Animal health surveillance enables the detection and control of animal diseases including zoonoses. Under the EU-FP7 project RISKSUR, a survey was conducted in 11 EU Member States and Switzerland to describe active surveillance components in 2011 managed by the public or private sector and identify gaps and opportunities. Information was collected about hazard, target population, geographical focus, legal obligation, management, surveillance design, risk-based sampling, and multi-hazard surveillance. Two countries were excluded due to incompleteness of data. Most of the 664 components targeted cattle (26·7%), pigs (17·5%) or poultry (16·0%). The most common surveillance objectives were demonstrating freedom from disease (43·8%) and case detection (26·8%). Over half of components applied risk-based sampling (57·1%), but mainly focused on a single population stratum (targeted risk-based) rather than differentiating between risk levels of different strata (stratified risk-based). About a third of components were multi-hazard (37·3%). Both risk-based sampling and multi-hazard surveillance were used more frequently in privately funded components. The study identified several gaps (e.g. lack of systematic documentation, inconsistent application of terminology) and opportunities (e.g. stratified risk-based sampling). The greater flexibility provided by the new EU Animal Health Law means that systematic evaluation of surveillance alternatives will be required to optimize cost-effectiveness.

Diagnosis of tuberculosis in groups of badgers: an exploration of the impact of trapping efficiency, infection prevalence and the use of multiple tests.

Accurate detection of infection with Mycobacterium bovis in live badgers would enable targeted tuberculosis control. Practical challenges in sampling wild badger populations mean that diagnosis of infection at the group (rather than the individual) level is attractive. We modelled data spanning 7 years containing over 2000 sampling events from a population of wild badgers in southwest England to quantify the ability to correctly identify the infection status of badgers at the group level. We explored the effects of variations in: (1) trapping efficiency; (2) prevalence of M. bovis; (3) using three diagnostic tests singly and in combination with one another; and (4) the number of badgers required to test positive in order to classify groups as infected. No single test was able to reliably identify infected badger groups if 80% sensitive, at least 94% specific, and able to be performed rapidly in the field.

Patterns of direct and indirect contact between cattle and badgers naturally infected with tuberculosis.

Tuberculosis (TB) due to infection with Mycobacterium bovis is transmitted between cattle and badgers (Meles meles) in the UK and Ireland but it is unclear where or when transmission occurs. We investigated direct and indirect interactions between badgers and cattle using automated proximity loggers on animals and at badger latrines located on pasture, in an area of south-west England with a high-density badger population. Direct contacts (interactions within 1.4 m) between badgers and cattle at pasture were very rare (four out of >500000 recorded animal-to-animal contacts) despite ample opportunity for interactions to occur. Indirect interactions (visits to badger latrines by badgers and cattle) were two orders of magnitude more frequent than direct contacts: 400 visits by badgers and 1700 visits by cattle were recorded. This suggests that indirect contacts might be more important than direct contacts in terms of disease transmission at pasture. The TB infection status of individual badgers (ascribed with 93% accuracy using three diagnostic tests) did not affect the frequency or duration of their visits to latrines located on pasture grazed by cattle. Nevertheless, there was wide variation in contact behaviour between individuals, which highlights the importance of understanding heterogeneity in contact patterns when developing strategies to control disease spread in wildlife and livestock.

Evaluation of animal and public health surveillance systems: a systematic review.

Disease surveillance programmes ought to be evaluated regularly to ensure they provide valuable information in an efficient manner. Evaluation of human and animal health surveillance programmes around the world is currently not standardized and therefore inconsistent. The aim of this systematic review was to review surveillance system attributes and the methods used for their assessment, together with the strengths and weaknesses of existing frameworks for evaluating surveillance in animal health, public health and allied disciplines. Information from 99 articles describing the evaluation of 101 surveillance systems was examined. A wide range of approaches for assessing 23 different system attributes was identified although most evaluations addressed only one or two attributes and comprehensive evaluations were uncommon. Surveillance objectives were often not stated in the articles reviewed and so the reasons for choosing certain attributes for assessment were not always apparent. This has the potential to introduce misleading results in surveillance evaluation. Due to the wide range of system attributes that may be assessed, methods should be explored which collapse these down into a small number of grouped characteristics by focusing on the relationships between attributes and their links to the objectives of the surveillance system and the evaluation. A generic and comprehensive evaluation framework could then be developed consisting of a limited number of common attributes together with several sets of secondary attributes which could be selected depending on the disease or range of diseases under surveillance and the purpose of the surveillance. Economic evaluation should be an integral part of the surveillance evaluation process. This would provide a significant benefit to decision-makers who often need to make choices based on limited or diminishing resources.

Safety, tolerability and pharmacokinetics of intravaginal pentamycin.

BACKGROUND/AIMS: Intravaginal pentamycin is a polyene macrolide with a broad spectrum of antimicrobial activity and is effective in various forms of infectious vaginitis. We evaluated the safety, tolerability and pharmacokinetics of escalating doses of this product. METHODS: Nineteen healthy volunteers were randomized to receive double blind one of five doses of intravaginal pentamycin (3, 10, 30, 60 or 100 mg) or the corresponding dose of pentamycin vehicle daily for 6 days. Patients with symptomatic vaginitis received a single dose of 60 (n = 6) or 100 mg (n = 6) of intravaginal pentamycin. Safety and tolerability parameters were monitored throughout the study. Plasma concentrations of pentamycin were measured daily in the healthy volunteers and on the day of drug application in the patients. RESULTS: The most frequently reported adverse events were mild or moderate vaginal discharge and mild symptoms of vaginal irritation (mainly pruritus or burning sensation), which also occurred in women who applied the vehicle. No patient with symptomatic vaginitis reported treatment-related adverse events. The plasma levels of pentamycin were below the quantification limit in all samples. CONCLUSION: Intravaginal pentamycin does not cause adverse reactions compared with vehicle and is not absorbed through the intact or the inflamed vagina.

Synaptic transmission between dissociated adult mammalian neurons and attached synaptic boutons.

In most studies of synaptic currents in mammalian central neurons, preparations have been used in which synaptic currents are recorded at some distance from the synapse itself. This procedure introduces problems in interpretation of the kinetics and voltage-dependent properties of the synaptic current. These problems have now been overcome by the development of a preparation in which presynaptic vesicle-containing boutons have been coisolated with the soma of individual neurons, thus providing the opportunity to study synaptic currents under conditions of both adequate voltage control and internal ionic perfusion. Spontaneous synaptic currents mediated by gamma-aminobutyric acid and excitatory amino acids were recorded from neurons isolated from a mammalian medial solitary tract nucleus. Calcium- and depolarization-dependent spontaneous currents of several to hundreds of picoamperes occurred with rapid rise times of 0.8 to 3 milliseconds and decays at least ten times as long.

Exchange of conduction pathways between two related K+ channels.

The structure of the ion conduction pathway or pore of voltage-gated ion channels is unknown, although the linker between the membrane spanning segments S5 and S6 has been suggested to form part of the pore in potassium channels. To test whether this region controls potassium channel conduction, a 21-amino acid segment of the S5-S6 linker was transplanted from the voltage-activated potassium channel NGK2 to another potassium channel DRK1, which has very different pore properties. In the resulting chimeric channel, the single channel conductance and blockade by external and internal tetraethylammonium (TEA) ion were characteristic of the donor NGK2 channel. Thus, this 21-amino acid segment controls the essential biophysical properties of the pore and may form the conduction pathway of these potassium channels.

Bupivacaine concentrations in the lumbar cerebrospinal fluid of patients during spinal anaesthesia.

BACKGROUND: Data on bupivacaine concentrations in the cerebral spinal fluid (CSF) during spinal anaesthesia are scarce. The purpose of this study was to determine the concentration of bupivacaine in the lumbar CSF of patients with an adequate level of spinal anaesthesia after injection of plain bupivacaine 0.5%. METHODS: Sixty patients with an adequate level of spinal block after standardized administration of plain bupivacaine 20 mg in men and of 17.5 mg in women were studied. To measure the CSF bupivacaine concentration, we performed a second lumbar spinal puncture and obtained a CSF sample at a randomized time point 5-45 min after the bupivacaine injection. In addition, we calculated the half-life of bupivacaine in the CSF and tested the hypothesis that the level of spinal block is related to the lumbar CSF bupivacaine concentration. RESULTS: Men and women had CSF bupivacaine concentrations ranging from 95.4 to 773.0 microg ml(-1) (median 242.4 microg ml(-1)) and from 25.9 to 781.0 microg ml(-1) (median 187.6 microg ml(-1)), respectively. The large variability of bupivacaine concentrations obtained at similar times after subarachnoid administration made calculation of a meaningful half-life of bupivacaine in CSF impossible. There was no association between CSF bupivacaine concentration and spinal block level, and CSF bupivacaine concentrations for the same spinal block level differed between patients by six-fold. CONCLUSIONS: There is a large variability of CSF bupivacaine concentrations in patients with an adequate level of spinal anaesthesia.

Pathology of Mycobacterium bovis infection in wild meerkats (Suricata suricatta).

Pathological lesions associated with Mycobacterium bovis infection (bovine tuberculosis; bTB) in free-living meerkats (Suricata suricatta) in the Kalahari Desert of South Africa are described. The pathology of bTB in meerkats was determined through detailed post-mortem examinations of 57 animals (52 meerkats showing clinical signs of bTB, and five not showing signs of disease). Lymph nodes and tissue lesions thought to be associated with bTB were cultured for mycobacteria. All 52 bTB-infected meerkats showed gross or microscopical granulomatous lesions, but M. bovis was cultured from only 42% (22/52) of these animals. The majority (96%, 50/52) of diseased meerkats had lesions in multiple sites, the pattern of which suggested haematogenous spread of M. bovis infection in this species. The histological characteristics of the tuberculous lesions, together with the gross pathology and the wide range of body systems affected, indicate that infection in meerkats is acquired principally via the respiratory and oral routes, whereas excretion is most likely via the respiratory tract and suppurating skin wounds. Urine and faeces appear to be unlikely sources of infection. The findings of this study provide information on the transmission, pathogenesis and epidemiology of bTB in meerkats that is likely to be relevant to the understanding of M. bovis infection in other social mammal species such as the European badger (Meles meles).

Alteration and restoration of K+ channel function by deletions at the N- and C-termini.

Voltage-dependent ion channels are thought to consist of a highly conserved repeated core of six transmembrane segments, flanked by more variable cytoplasmic domains. Significant functional differences exist among related types of K+ channels. These differences have been attributed to the variable domains, most prominently the N- and C-termini. We have therefore investigated the functional importance of both termini for the delayed rectifier K+ channel from rat brain encoded by the drk1 gene. This channel has an unusually long C-terminus. Deletions in either terminus affected both activation and inactivation, in some cases profoundly. Unexpectedly, more extensive deletions in both termini restored gating. We could therefore define a core region only slightly longer than the six transmembrane segments that is sufficient for the formation of channels with the kinetics of a delayed rectifier.

Segmental exchanges define 4-aminopyridine binding and the inner mouth of K+ pores.

4-Aminopyridine (4AP) blocks the intracellular mouth of voltage-gated K+ channels. We identified critical regions for 4AP binding with chimeric channels in which segments of a low affinity clone (Kv2.1, IC50 = 18 mM) were replaced with those of a high affinity clone (Kv3.1, IC50 = 0.1 mM). 4AP sensitivity was not transferred with the S5-S6 linker (pore or P region). Instead, a chimera of the cytoplasmic half of S6 increased block 20-fold, without affecting gating. A double chimera of the cytoplasmic halves of S5 and S6 fully transferred 4AP sensitivity. Because 4AP block was inhibited by tetrapentylammonium, we conclude that determinants of 4AP binding lie in the S6 segment that forms the cytoplasmic vestibule of the pore and that this site may overlap a quaternary ammonium site.

Differences between the deep pores of K+ channels determined by an interacting pair of nonpolar amino acids.

The pore of a chimeric K+ channel, CHM, differed from its parental host channel, Kv2.1, by 9 amino acids. Four were located in a putative deep region and 5 in a nearby outer mouth. Point reversions were without restorative effects, and reversions V369I or L374V in the deep pore produced novel phenotypes. Among double mutations, only V369I and L374V were effective in restoring the Kv2.1 pore phenotype. Adding a change in charge at Q382K in the outer pore fully restored the parental phenotype. Thus, the pore appears to have an inner, deep region where ions such as K+ and TEA+ may be regulated by nonpolar residues and an outer region where ions may be regulated by charged residues.

Pharmacokinetics and pharmacodynamic effects of oral GLP-1 and PYY3-36: a proof-of-concept study in healthy subjects.

This proof-of-concept study was performed in order to establish the pharmacokinetics and pharmacodynamics of increasing oral doses of the satiety peptides glucagon-like peptide-1 (GLP-1) and peptide YY3-36 (PYY3-36). Six healthy male subjects were given oral doses of either a placebo or GLP-1 in a dose-escalating schedule (doses of 0.5, 1.0, 2.0, and 4.0 mg). Next, another group of six healthy male subjects were given oral doses of either a placebo or PYY3-36 in the same pattern of escalating doses (doses of 0.25, 0.5, 1.0, 2.0, and 4.0 mg). In healthy male volunteers, (i) oral administration of either of the peptides induced a rapid and dose-dependent increase in plasma drug concentrations; (ii) oral administration of GLP-1 induced a potent effect on insulin release; and (iii) both peptides suppressed ghrelin secretion. In conclusion, this study showed, for the first time, that satiety peptides such as GLP-1 and PYY3-36 can be orally delivered safely and effectively in humans.

Effects of budesonide on P-glycoprotein expression in intestinal cell lines.

BACKGROUND AND PURPOSE: P-glycoprotein (P-gp) is an important efflux transporter that supports the barrier function of the gut against invading antigens and against administered drugs. Since glucocorticoids, such as budesonide, are frequently used during inflammatory bowel disease we investigated how budesonide influences P-gp expression in different intestinal cell lines. EXPERIMENTAL APPROACH: LS180 and Caco-2 cells were incubated with budesonide and changes in P-gp expression were determined on mRNA, protein and functional level. The mRNA expression levels of glucocorticoid receptor (GR) and pregnane X receptor (PXR) were determined in these cell lines. PXR receptor was transiently transfected into Caco-2 cells. KEY RESULTS: Budesonide showed an induction of P-gp in LS180 cells and a down-regulation in Caco-2 cells. Expression levels of nuclear receptors revealed high expression of PXR only in LS180 cells and exclusive expression of GR in Caco-2 cells. Mifepristone, an anti-glucocorticoid, could not reverse the down-regulation of P-gp by budesonide in Caco-2 cells. In PXR-transfected Caco-2 cells the budesonide-mediated down-regulation of P-gp was abolished. Furthermore the expression of cytochrome P450 3A4 (CYP3A4), another PXR target gene, was induced in PXR-transfected Caco-2 cells after budesonide treatment. CONCLUSIONS AND IMPLICATIONS: Budesonide has the potential to influence MDR1 expression in vitro. In LS180 cells, the induction of MDR1 by budesonide probably is mediated via PXR. The mechanism of the down-regulation in Caco-2 cells still remains unclear, but GR does not seem to be involved. Further studies are required to evaluate how budesonide alters P-gp expression in vivo.

Glucagon-like peptide 1 receptor expression in primary porcine proximal tubular cells.

BACKGROUND: GLP-1 is secreted into the circulation after food intake. The main biological effects of GLP-1 include stimulation of glucose dependent insulin secretion and induction of satiety feelings. Recently, it was demonstrated in rats and humans that GLP-1 can stimulate renal excretion of sodium. Based on these data, the existence of a renal GLP-1 receptor (GLP-1R) was postulated. However, the exact localization of the GLP-1R and the mechanism of this GLP-1 action have not yet been investigated. METHODS: Primary porcine proximal tubular cells were isolated from porcine kidneys. Expression of GLP-1R was measured at the mRNA level by quantitative RT-PCR. Protein expression of GLP-1R was verified with immunocytochemistry, immunohistochemistry and Western blot analysis. Functional studies included transport assessments of sodium and glucose using three different GLP-1 concentrations (200 pM, 2 nM and 20 nM), 200 pM exendin-4 (GLP-1 analogue) and an inhibitor of the dipeptidylpeptidase IV (DPPIV) enzyme (P32/98 at 10 microM). Finally, the expression of NHE3, the predominant Na(+)/H(+) exchanger in proximal tubular cells, was also investigated. RESULTS: GLP-1R, NHE3 and DPPIV were expressed at the mRNA level in porcine proximal tubular kidney cells. GLP-1R expression was confirmed at the protein level. Staining of human and pig kidney cortex revealed that GLP-1R was predominantly expressed in proximal tubular cells. Functional assays demonstrated an inhibition of sodium re-absorption with GLP-1 after 3 h of incubation. Exendin-4 and GLP-1 in combination with P32/98 co-administration had no clear influence on glucose and sodium uptake and transport. CONCLUSION: GLP-1R is functionally expressed in porcine proximal tubular kidney cells. Addition of GLP-1 to these cells resulted in a reduced sodium re-absorption. GLP-1 had no effect on glucose re-absorption. We conclude that GLP-1 modulates sodium homeostasis in the kidney most likely through a direct action via its GLP-1R in proximal tubular cells.

Comparison of two antacid preparations on intragastric acidity--a two-centre open randomised cross-over placebo-controlled trial.

BACKGROUND: Rennie and Riopan Gel are 2 of the most well-known and popular over-the-counter antacids; for heartburn symptoms, pain relief is fast with both preparations. A direct comparison with respect to intragastric acidity has not been done yet. The aim of our study was therefore to compare the effects of both preparations on intragastric acidity of fasting volunteers. METHODS: The study was conducted as an open, randomised, placebo-controlled, 2-centre cross-over study. On different days, 24 healthy adult volunteers (11 males and 13 females) received equimolar acid-neutralising amounts of either Riopan Gel (800 mg magaldrate) or 2 tablets of Rennie (680 mg calcium carbonate and 80 mg magnesium carbonate) or no drug (control) with a wash-out period of at least 4 days between applications. The intragastric pH was measured for 3 h by intragastric pH-metry. The primary endpoint was the median time lag before intragastric pH >3.0 was reached for 10 consecutive min after drug administration. RESULTS: For both antacids, the median pH during the first 30 min after drug administration was statistically significantly different from placebo (p < 0.05), but there was a statistically significant increase in pH during the first 5 min for Riopan Gel only. CONCLUSION: Compared to placebo, both antacids (Rennie and Riopan Gel) have short-lasting effects on intragastric acidity. There is no statistically significant difference between the 2 preparations, except in the first 5 min, indicating a faster onset of action for Riopan Gel. We conclude that the antacid formulation (tablet or liquid) has little influence on intragastric acidity.