Creating local bibliographic databases: new tools for evidence-based health care*

The Internet has created new opportunities for librarians to present literature search results to clinicians. In order to take full advantage of these opportunities, libraries need to create locally maintained bibliographic databases. A simple method of creating a local bibliographic database and publishing it on the Web is described. The method uses off-the-shelf software and requires minimal programming. A hedge search strategy for outcome studies of clinical process interventions is created, and Ovid is used to search MEDLINE. The search results are saved and imported into EndNote libraries. The citations are modified, exported to a Microsoft Access database, and published on the Web. Clinicians can use a Web browser to search the database. The bibliographic database contains 13,803 MEDLINE citations of outcome studies. Most searches take between four and ten seconds and retrieve between ten and 100 citations. The entire cost of the software is under $900. Locally maintained bibliographic databases can be created easily and inexpensively. They significantly extend the evidence-based health care services that libraries can offer to clinicians.

Discrimination of the closely related A and D genomes of the hexaploid oat Avena sativa L.

A satellite DNA sequence, As120a, specific to the A-genome chromosomes in the hexaploid oat, Avena sativa L., was isolated by subcloning a fragment with internal tandem repeats from a plasmid, pAs120, that had been obtained from an Avena strigosa (As genome) genomic library. Southern and in situ hybridization showed that sequences with homology to sequences within pAs120 were dispersed throughout the genome of diploid (A and C genomes), tetraploid (AC genomes), and hexaploid (ACD genomes) Avena species. In contrast, sequences homologous to As120a were found in two A-genome species (A. strigosa and Avena longiglumis) and in the hexaploid A. sativa whereas this sequence was little amplified in the tetraploid Avena murphyi and was absent in the remaining A- and C-genome diploid species. In situ hybridization of pAs120a to hexaploid oat species revealed the distribution of elements of the As120a repeated family over both arms of 14 of 42 chromosomes of this species. By using double in situ hybridization with pAs120a and a C genome-specific probe, three sets of 14 chromosomes were revealed corresponding to the A, C, and D genomes of the hexaploid species. Simultaneous in situ hybridizations with pAs120a and ribosomal probes were used to assign the SAT chromosomes of hexaploid species to their correct genomes. This work reports a sequence able to distinguish between the closely related A and D genomes of hexaploid oats. This sequence offers new opportunities to analyze the relationships of Avena species and to explore the possible evolution of various polyploid oat species.

Subtraction hybridization identifies a transformation progression-associated gene PEG-3 with sequence homology to a growth arrest and DNA damage-inducible gene

Cancer is a progressive multigenic disorder characterized by defined changes in the transformed phenotype that culminates in metastatic disease. Determining the molecular basis of progression should lead to new opportunities for improved diagnostic and therapeutic modalities. Through the use of subtraction hybridization, a gene associated with transformation progression in virus- and oncogene-transformed rat embryo cells, progression elevated gene-3 (PEG-3), has been cloned. PEG-3 shares significant nucleotide and amino acid sequence homology with the hamster growth arrest and DNA damage-inducible gene gadd34 and a homologous murine gene, MyD116, that is induced during induction of terminal differentiation by interleukin-6 in murine myeloid leukemia cells. PEG-3 expression is elevated in rodent cells displaying a progressed-transformed phenotype and in rodent cells transformed by various oncogenes, including Ha-ras, v-src, mutant type 5 adenovirus (Ad5), and human papilloma virus type 18. The PEG-3 gene is transcriptionally activated in rodent cells, as is gadd34 and MyD116, after treatment with DNA damaging agents, including methyl methanesulfonate and γ-irradiation. In contrast, only PEG-3 is transcriptionally active in rodent cells displaying a progressed phenotype. Although transfection of PEG-3 into normal and Ad5-transformed cells only marginally suppresses colony formation, stable overexpression of PEG-3 in Ad5-transformed rat embryo cells elicits the progression phenotype. These results indicate that PEG-3 is a new member of the gadd and MyD gene family with similar yet distinct properties and this gene may directly contribute to the transformation progression phenotype. Moreover, these studies support the hypothesis that constitutive expression of a DNA damage response may mediate cancer progression.

Governing step of metastasis visualized in vitro

Metastasis is the ultimate life-threatening stage of cancer. The lack of accurate model systems thwarted studies of the metastatic cell’s basic biology. To follow continuously the succeeding stages of metastatic colony growth, we heritably labeled cells from the human lung adenocarcinoma cell line ANIP 973 with green fluorescent protein (GFP) by transfection with GFP cDNA. Labeled cells were then injected intravenously into nude mice, where, by 7 days, they formed brilliantly fluorescing metastatic colonies on mouse lung [Chishima, T., Miyagi, Y., Wang, X., Yang, M., Tan, Y., Shimada, H., Moossa, A. R. & Hoffman, R. M. (1997) Clin. Exp. Metastasis 15, 547–552]. The seeded lung tissue was then excised and incubated in the three-dimensional sponge-gel-matrix-supported histoculture that maintained the critical features of progressive in vivo tumor colonization while allowing continuous access for measurement and manipulation. Tumor progression was continuously visualized by GFP fluorescence in the same individual cultures over a 52-day period, during which the tumors spread throughout the lung. Histoculture tumor colonization was selective for lung cancer cells to grow on lung tissue, because no growth occurred on histocultured mouse liver tissue, which was also observed in vivo. The ability to support selective organ colonization in histoculture and visualize tumor progression by GFP fluorescence allows the in vitro study of the governing processes of metastasis [Kuo, T.-H., Kubota, T., Watanbe, M., Furukawa, T., Teramoto, T., Ishibiki, K., Kitajima, M., Moossa, A. R., Penman, S. & Hoffman, R. M. (1995) Proc. Natl. Acad. Sci. USA 92, 12085–12089]. The results presented here provide significant, new opportunities to understand and to develop treatments that prevent and possibly reverse metastasis.

Building a dictionary for genomes: Identification of presumptive regulatory sites by statistical analysis

The availability of complete genome sequences and mRNA expression data for all genes creates new opportunities and challenges for identifying DNA sequence motifs that control gene expression. An algorithm, “MobyDick,” is presented that decomposes a set of DNA sequences into the most probable dictionary of motifs or words. This method is applicable to any set of DNA sequences: for example, all upstream regions in a genome or all genes expressed under certain conditions. Identification of words is based on a probabilistic segmentation model in which the significance of longer words is deduced from the frequency of shorter ones of various lengths, eliminating the need for a separate set of reference data to define probabilities. We have built a dictionary with 1,200 words for the 6,000 upstream regulatory regions in the yeast genome; the 500 most significant words (some with as few as 10 copies in all of the upstream regions) match 114 of 443 experimentally determined sites (a significance level of 18 standard deviations). When analyzing all of the genes up-regulated during sporulation as a group, we find many motifs in addition to the few previously identified by analyzing the subclusters individually to the expression subclusters. Applying MobyDick to the genes derepressed when the general repressor Tup1 is deleted, we find known as well as putative binding sites for its regulatory partners.

Creating a Web-accessible, point-of-care, team-based information system (PoinTIS): the librarian as publisher*

The Internet has created new opportunities for librarians to develop information systems that are readily accessible at the point of care. This paper describes the multiyear process used to justify, fund, design, develop, promote, and evaluate a rehabilitation prototype of a point-of-care, team-based information system (PoinTIS) and train health care providers to use this prototype for their spinal cord injury and traumatic brain injury patient care and education activities. PoinTIS is a successful model for librarians in the twenty-first century to serve as publishers of information created or used by their parent organizations and to respond to the opportunities for information dissemination provided by recent technological advances.

Transcriptional and posttranslational plasticity and the generation of inflammatory pain

Inflammatory pain manifests as spontaneous pain and pain hypersensitivity. Spontaneous pain reflects direct activation of specific receptors on nociceptor terminals by inflammatory mediators. Pain hypersensitivity is the consequence of early posttranslational changes, both in the peripheral terminals of the nociceptor and in dorsal horn neurons, as well as later transcription-dependent changes in effector genes, again in primary sensory and dorsal horn neurons. This inflammatory neuroplasticity is the consequence of a combination of activity-dependent changes in the neurons and specific signal molecules initiating particular signal-transduction pathways. These pathways phosphorylate membrane proteins, changing their function, and activate transcription factors, altering gene expression. Two distinct aspects of sensory neuron function are changed as a result of these processes, basal sensitivity, or the capacity of peripheral stimuli to evoke pain, and stimulus-evoked hypersensitivity, the capacity of certain inputs to generate prolonged alterations in the sensitivity of the system. Posttranslational changes largely alter basal sensitivity. Transcriptional changes both potentiate the system and alter neuronal phenotype. Potentiation occurs as a result of the up-regulation in the dorsal root ganglion of centrally acting neuromodulators and simultaneously in the dorsal horn of their receptors. This means that the response to subsequent inputs is augmented, particularly those that induce stimulus-induced hypersensitivity. Alterations in phenotype includes the acquisition by A fibers of neurochemical features typical of C fibers, enabling these fibers to induce stimulus-evoked hypersensitivity, something only C fiber inputs normally can do. Elucidation of the molecular mechanisms responsible provides new opportunities for therapeutic approaches to managing inflammatory pain.

Ecological approaches and the development of “truly integrated” pest management

Recent predictions of growth in human populations and food supply suggest that there will be a need to substantially increase food production in the near future. One possible approach to meeting this demand, at least in part, is the control of pests and diseases, which currently cause a 30–40% loss in available crop production. In recent years, strategies for controlling pests and diseases have tended to focus on short-term, single-technology interventions, particularly chemical pesticides. This model frequently applies even where so-called integrated pest management strategies are used because in reality, these often are dominated by single technologies (e.g., biocontrol, host plant resistance, or biopesticides) that are used as replacements for chemicals. Very little attention is given to the interaction or compatibility of the different technologies used. Unfortunately, evidence suggests that such approaches rarely yield satisfactory results and are unlikely to provide sustainable pest control solutions for the future. Drawing on two case histories, this paper demonstrates that by increasing our basic understanding of how individual pest control technologies act and interact, new opportunities for improving pest control can be revealed. This approach stresses the need to break away from the existing single-technology, pesticide-dominated paradigm and to adopt a more ecological approach built around a fundamental understanding of population biology at the local farm level and the true integration of renewable technologies such as host plant resistance and natural biological control, which are available to even the most resource-poor farmers.

Molecular cloning of violaxanthin de-epoxidase from romaine lettuce and expression in Escherichia coli.

Plants need to avoid or dissipate excess light energy to protect photosystem II (PSII) from photoinhibitory damage. Higher plants have a conserved system that dissipates excess energy as heat in the light-harvesting complexes of PSII that depends on the transthylakoid delta pH and violaxanthin de-epoxidase (VDE) activity. To our knowledge, we report the first cloning of a cDNA encoding VDE and expression of functional enzyme in Escherichia coli. VDE is nuclear encoded and has a transit peptide with characteristic features of other lumen-localized proteins. The cDNA encodes a putative polypeptide of 473 aa with a calculated molecular mass of 54,447 Da. Cleavage of the transit peptide results in a mature putative polypeptide of 348 aa with a calculated molecular mass of 39,929 Da, close to the apparent mass of the purified enzyme (43 kDa). The protein has three interesting domains including (i) a cysteine-rich region, (ii) a lipocalin signature, and (iii) a highly charged region. The E. coli expressed enzyme de-epoxidizes violaxanthin sequentially to antheraxanthin and zeaxanthin, and is inhibited by dithiothreitol, similar to VDE purified from chloroplasts. This confirms that the cDNA encodes an authentic VDE of a higher plant and is unequivocal evidence that the same enzyme catalyzes the two-step mono de-epoxidation reaction. The cloning of VDE opens new opportunities for examining the function and evolution of the xanthophyll cycle, and possibly enhancing light-stress tolerance of plants.