CreZOO--the European virtual repository of Cre and other targeted conditional driver strains.

The CreZOO (http://www.crezoo.org/) is the European virtual repository of Cre and other targeted conditional driver strains. These mice serve as tools for researchers to selectively 'switch off' gene expression in mouse models to examine gene function and disease pathology. CreZOO aims to capture and disseminate extant and new information on these Cre driver strains, such as genetic background and availability information, and details pertaining promoter, allele, inducibility and expression patterns, which are also presented. All transgenic strains carry detailed information according to MGI's official nomenclature, whereas their availability [e.g. live mice, cryopreserved embryos, sperm and embryonic stem (ES) cells] is clearly indicated with links to European and International databases and repositories (EMMA, MGI/IMSR, MMRRC, etc) and laboratories where the particular mouse strain is available together with the respective IDs. Each promoter/gene includes IDs and direct links to MGI, Entrez Gene, Ensembl, OMIM and RGD databases depending on their species origin, whereas allele information is presented with MGI IDs and active hyperlinks to redirect the user to the respective page in a new tab. The tissue/cell (special) and developmental (temporal) specificity expression patterns are clearly presented, whereas handling and genotyping details (in the form of documents or hyperlinks) together with all relevant publications are clearly presented with PMID(s) and direct PubMed links. CreZOO's design offers a user-friendly query interface and provides instant access to the list of conditional driver strains, promoters and inducibility details. Database access is free of charge and there are no registration requirements for data querying. CreZOO is being developed in the context of the CREATE consortium (http://www.creline.org/), a core of major European and international mouse database holders and research groups involved in conditional mutagenesis. Database URL: http://www.crezoo.org/; alternative URL: http://www.e-mouse.org/

Mouse Resource Browser--a database of mouse databases.

The laboratory mouse has become the organism of choice for discovering gene function and unravelling pathogenetic mechanisms of human diseases through the application of various functional genomic approaches. The resulting deluge of data has led to the deployment of numerous online resources and the concomitant need for formalized experimental descriptions, data standardization, database interoperability and integration, a need that has yet to be met. We present here the Mouse Resource Browser (MRB), a database of mouse databases that indexes 217 publicly available mouse resources under 22 categories and uses a standardised database description framework (the CASIMIR DDF) to provide information on their controlled vocabularies (ontologies and minimum information standards), and technical information on programmatic access and data availability. Focusing on interoperability and integration, MRB offers automatic generation of downloadable and re-distributable SOAP application-programming interfaces for resources that provide direct database access. MRB aims to provide useful information to both bench scientists, who can easily navigate and find all mouse related resources in one place, and bioinformaticians, who will be provided with interoperable resources containing data which can be mined and integrated. Database URL: http://bioit.fleming.gr/mrb.

Finding and sharing: new approaches to registries of databases and services for the biomedical sciences.

The recent explosion of biological data and the concomitant proliferation of distributed databases make it challenging for biologists and bioinformaticians to discover the best data resources for their needs, and the most efficient way to access and use them. Despite a rapid acceleration in uptake of syntactic and semantic standards for interoperability, it is still difficult for users to find which databases support the standards and interfaces that they need. To solve these problems, several groups are developing registries of databases that capture key metadata describing the biological scope, utility, accessibility, ease-of-use and existence of web services allowing interoperability between resources. Here, we describe some of these initiatives including a novel formalism, the Database Description Framework, for describing database operations and functionality and encouraging good database practise. We expect such approaches will result in improved discovery, uptake and utilization of data resources. Database URL: http://www.casimir.org.uk/casimir_ddf.

Activation of phosphatidylinositol 3-kinase/protein kinase B by corticotropin-releasing factor in human monocytes.

Corticotropin-releasing factor (CRF) exerts proinflammatory effects in peripheral tissues, whereas the intracellular pathways mediating these effects have not been completely characterized yet. We have previously shown that CRF induces nuclear factor-kappaB DNA-binding activity in mouse and human leukocytes. Here we demonstrate that in the human monocytic THP-1 cells, CRF activates the phosphatidylinositol 3-kinase (PI3K)/Akt and ERK1/2 pathways. These effects of CRF are mediated by corticotropin-releasing factor receptor 2 (CRF2), as suggested by their abolishment after treatment with the specific CRF2 antagonist, astressin 2B. The CRF-mediated PI3K/Akt activation induces cell survival as suggested by the stimulation of the antiapoptotic factor Bcl-2. ERK1/2 activation results in up-regulation of IL-8 expression, an effect inhibited by the CRF-induced activation of PI3K/Akt. These studies demonstrate novel effects of CRF in human monocytes mediated by the activation of PI3K/Akt. Moreover, they reveal pathway-specific effects of the CRF/CRF2 system in chemokine activation and cell survival that may be of importance for the development of novel therapeutics for inflammatory diseases.

Models for financial sustainability of biological databases and resources.

Following the technological advances that have enabled genome-wide analysis in most model organisms over the last decade, there has been unprecedented growth in genomic and post-genomic science with concomitant generation of an exponentially increasing volume of data and material resources. As a result, numerous repositories have been created to store and archive data, organisms and material, which are of substantial value to the whole community. Sustained access, facilitating re-use of these resources, is essential, not only for validation, but for re-analysis, testing of new hypotheses and developing new technologies/platforms. A common challenge for most data resources and biological repositories today is finding financial support for maintenance and development to best serve the scientific community. In this study we examine the problems that currently confront the data and resource infrastructure underlying the biomedical sciences. We discuss the financial sustainability issues and potential business models that could be adopted by biological resources and consider long term preservation issues within the context of mouse functional genomics efforts in Europe.

Terbutaline inhibits corticotropin-releasing hormone (CRH) expression in human trophoblast cells.

OBJECTIVE: To evaluate the effect of beta-adrenergic agonists on the regulation of the expression of the placental corticotropin-releasing hormone (CRH) gene. STUDY DESIGN: Term placentae were collected at the time of elective cesarean section, and trophoblast cells were harvested, isolated, and cultured. The isolated trophoblasts were plated, cultured and subsequently treated with cortisol, terbutaline, RU486, or vehicle control. CRH expression, mRNA abundance of CRH, and the housekeeping gene beta-actin were evaluated by Northern blot analysis. RESULTS: Exposure of the trophoblasts to terbutaline (10(-8) M) inhibited the expression of the CRH gene as depicted by Northern blot analysis. Co-addition of terbutaline (10(-8) M) and RU486 (10(-6) M) did not block the stimulatory effects of RU486 on placental trophoblast cells. CONCLUSION: The beta-adrenergic agonist terbutaline inhibits the expression of CRH in human trophoblasts. This finding may provide insight into the mechanism of action of terbutaline as a tocolytic agent.

Role for prostaglandins in the regulation of type 1 11beta-hydroxysteroid dehydrogenase in human granulosa-lutein cells.

11beta-hydroxysteroid dehydrogenase (11betaHSD) enzymes regulate glucocorticoid availability in target tissues. 11betaHSD1 is the predominant isoenzyme expressed and active in human granulosa-lutein (hGL) cells. This study investigated the effects of pharmacological inhibitors of prostaglandin (PG) synthesis on 11betaHSD1 activities and expression in hGL cells. The consequences for 11betaHSD1 of increasing exposure of hGL cells to PGs, either by treatment with exogenous PGs or by challenging cells with IL-1beta, were also assessed. Suppression of basal PG synthesis using four different inhibitors of PG H synthase enzymes [indomethacin, niflumic acid, meclofenamic acid (MA) and N-(2-cyclohexyloxy-4-nitorophenyl) methane sulfonamide (NS-398)] each resulted in significant decreases in both cortisol oxidation and cortisone reduction. Both activities of 11betaHSD1 were suppressed by up to 64+/-6% (P<0.05). Over 4 and 24 h, neither MA nor NS-398 affected the expression of 11betaHSD1 protein, suggesting enzyme regulation by PGs at the posttranslational level. When cells were cotreated for 4 h with PGHS inhibitors plus 30 nm PGD2, PGF2alpha, or PGE2, each PG overcame the suppression of cortisol oxidation by indomethacin or MA. Treatment of hGL cells with IL-1beta increased the concentrations of both PGE2 and PGF2alpha, accompanied by a 70+/-25% increase in net cortisol oxidation. All three responses to IL-1beta were abolished when cells were cotreated with MA. These findings suggest a role for PGs in the posttranslational regulation of 11betaHSD1 activities in hGL cells.

Corticotropin-releasing hormone contributes to the peripheral inflammatory response in experimental autoimmune encephalomyelitis.

Peripheral corticotropin-releasing hormone (CRH) is thought to have proinflammatory effects. We used the model of experimental autoimmune encephalomyelitis (EAE) to study the role of CRH in an immune-mediated disease. We showed that CRH-deficient mice are resistant to EAE, with a decrease in clinical score as well as decreased cellular infiltration in the CNS. Furthermore, Ag-specific responses of primed T cells as well as anti-CD3/anti-CD28 TCR costimulation were decreased in crh(-/-) mice with decreased production of Th1 cytokines and increased production of Th2 cytokines. Wild-type mice treated in vivo with a CRH antagonist showed a decrease in IFN-gamma production by primed T cells in vitro. This effect of CRH is independent of its ability to increase corticosterone production, because adrenalectomized wild-type mice had similar disease course and severity as control mice. We found that IkappaBalpha phosphorylation induced by TCR cross-linking was decreased in crh(-/-) T cells. We conclude that peripheral CRH exerts a proinflammatory effect in EAE with a selective increase in Th1-type responses. These findings have implications for the treatment of Th1-mediated diseases such as multiple sclerosis.

Roles for prostaglandins in the steroidogenic response of human granulosa cells to high-density lipoproteins.

In human granulosa-lutein cells, high-density lipoproteins (HDL) can stimulate progesterone synthesis. The objective of the present study was to establish whether prostaglandins (PGs) participate in the steroidogenic response to HDL. Both HDL and apolipoprotein AI (ApoAI) stimulated concentration-dependent increases in PGE2, cAMP and progesterone accumulation. The minimum concentrations of HDL and ApoAI required to elevate PGE2 production were the same as those required to stimulate cAMP accumulation and progesterone synthesis. Concentrations of PGE2 were elevated within 10 min in cells exposed to HDL and rose progressively over 24 h, whereas cAMP and progesterone were only increased significantly after 24 h of treatment with HDL. Co-treatment with prostaglandin H synthase inhibitors (meclofenamic acid and indomethacin) abolished the cAMP and progesterone responses to both HDL and ApoAI. Hence, the ability of HDL to stimulate progesterone synthesis can be mimicked by ApoAI and appears to involve increased generation of one or more luteotrophic PGs, possibly acting via cAMP.

NF-kappaB participates in the corticotropin-releasing, hormone-induced regulation of the pituitary proopiomelanocortin gene.

Corticotropin-releasing hormone is a main regulator of mammalian stress response by stimulating pituitary proopiomelanocortin (POMC) gene expression, and thus adrenocorticotropic hormone (ACTH) secretion, which then causes glucocorticoid release from the adrenal. In a recent study in the pituitary corticotroph cell line AtT20, oxidative stress stimulated the activity of nuclear transcription factor B (NF-kappaB), whereas corticotropin-releasing hormone (CRH) inhibited both the constitutive and the oxidative stress-induced NF-kappaB DNA-binding activity. To further investigate the role of NF-kappaB on the CRH-induced pituitary POMC gene activation, AtT20 cells were transiently transfected with a POMC-luciferase construct mutated at an NF-kappaB binding site. After treatment with CRH, intracellular POMC-luciferase activity was significantly higher from the stimulation observed with transfection of the parental POMC-luciferase construct. In agreement with a previous report, CRH inhibited the constitutive NF-kappaB DNA-binding activity in AtT20 cells, as shown by electrophoretic mobility-shift assay, as soon as within 15 min of treatment. These effects of CRH were blocked by the CRH-R1 antagonist CP154,256. Our findings provide evidence that the regulation of corticotroph NF-kappaB activity by CRH is related to the activation of the pituitary POMC gene and, thus, may play an important role in stress response.