GiSAO.db: a database for ageing research

<p>Abstract</p> <p>Background</p> <p>Age-related gene expression patterns of <it>Homo sapiens </it>as well as of model organisms such as <it>Mus musculus</it>, <it>Saccharomyces cerevisiae</it>, <it>Caenorhabditis elegans </it>and <it>Drosophila melanogaster </it>are a basis for understanding the genetic mechanisms of ageing. For an effective analysis and interpretation of expression profiles it is necessary to store and manage huge amounts of data in an organized way, so that these data can be accessed and processed easily.</p> <p>Description</p> <p>GiSAO.db (Genes involved in senescence, apoptosis and oxidative stress database) is a web-based database system for storing and retrieving ageing-related experimental data. Expression data of genes and miRNAs, annotation data like gene identifiers and GO terms, orthologs data and data of follow-up experiments are stored in the database. A user-friendly web application provides access to the stored data. KEGG pathways were incorporated and links to external databases augment the information in GiSAO.db. Search functions facilitate retrieval of data which can also be exported for further processing.</p> <p>Conclusions</p> <p>We have developed a centralized database that is very well suited for the management of data for ageing research. The database can be accessed at <url>https://gisao.genome.tugraz.at</url> and all the stored data can be viewed with a guest account.</p

Synthesis of (±)-trans-2,5-Diisopropylborolane

The cyclic hydroboration of 2,7-dimethyl-2,6-octadiene (6) was studied. It was found that the stereochemical outcome of the reaction was dependent upon the solvent, temperature, time and the nature of the borane reagent. Pure racemic trans-2,5-diisopropylborolane (14) was isolated following selective complexation of the cis-2,5-diisopropylborolane (15) with 1-(2-hydroxyethyl)-pyrrolidine

Controversial Issue: Is it safe to employ mesenchymal stem cells in cell-based therapies?

The prospective clinical use of multipotent mesenchymal stromal stem cells (MSC) holds enormous promise for the treatment of a large number of degenerative and age-related diseases. However, the challenges and risks for cell-based therapies are multifaceted. The risks for patients receiving stem cells, which have been expanded in vitro in the presence of xenogenic compounds, can hardly be anticipated and methods for the culture and manipulation of “safe” MSC ex vivo are being investigated. During in vitro expansion, stem cells experience a long replicative history and are thus subject to damage from intracellular and extracellular influences. While murine MSC are prone to cellular transformation in culture, human MSC do not transform. One reason for this striking difference is that during long-term culture, human MSC finally become replicatively senescent. In consequence, this greatly restricts their proliferation and differentiation efficiency. It however also limits the yield of sufficient numbers of cells needed for therapy. Another issue is to eliminate contamination of expanding cells with serumbound pathogenic agents in order to reduce the risks for infection. A recent technical advancement, which applies human serum platelet lysates as an alternative source for growth factors and essential supplements, allows the unimpaired proliferation of MSC in the absence of animal sera. Here, we present an update regarding cellular senescence of MSC and recent insights concerning potential risks associated with their clinical use

Leptin receptor/CD295 is upregulated on primary human mesenchymal stem cells of advancing biological age and distinctly marks the subpopulation of dying cells

During the lifetime of an adult organism, stem cells face extrinsic and intrinsic aging. Mesenchymal stem cells (MSC) can be expanded in culture, and the proliferation potential of individual cell isolates before growing senescent appear to be dependent on fitness and age of the donor, respectively. To date no molecular markers are available, which specifically reflect the degree of cellular aging in a population of MSC. Employing a genomic approach, we noticed that the gene encoding leptin receptor (also termed OBR) is differentially regulated in MSC derived from aged donors as well as in MSC that had been stressed due to cultivation under hyperoxic conditions. We further observed that the leptin receptor transcript levels in primary MSC isolates are inversely correlated with the prospective number of generations that are ahead of these cells in culture, i.e., the number of population doublings that will occur in long term culture prior to cessation of growth due to replicative senescence. The MSC subpopulation, which exhibited distinctly elevated levels of leptin receptor or CD295 at the cell surface, is indistinguishable from dying cells. Considered together with the observation that primary MSC derived from healthy individuals showed proliferation capacities that declined at differentially increasing rates, we concluded that attenuation of MSC proliferation potential during aging greatly relies on the strictly increasing withdrawal of cells due to cell death

Changes of the Functional Capacity of Mesenchymal Stem Cells due to Aging or Age-Associated Disease – Implications for Clinical Applications and Donor Recruitment

In contrast to stem cells of embryonic origin, autologous tissue-specific stem cells are easier to introduce into the clinical practice. In this context however, molecular and cellular changes, which alter tissue-specific stem cell properties with age, are of particular interest, since elderly patients represent the main target group for cellbased therapies. The clinical use of mesenchymal stem cells is an emerging field, especially because this stem cell type appears to be amenable for the treatment of a large number of diseases, such as non-healing bone defects and fractures, inflammatory relieve during arthritis, and the repair of suspensory ligament tears. More than that, mesenchymal stem cells provoke effective immune suppression in the context of graft-versus-host disease. Here, we present a comprehensive overview of the recent findings with special attention to age-related changes of mesenchymal stem cells properties and the consequential impact involved on tissue regeneration and repair, together with the current perception concerning their therapeutic application potential as well as challenges associated with their clinical use

High-Yield Recombinant Expression of the Extremophile Enzyme, Bee Hyaluronidase In Pichia Pastoris

Hyaluronidase from honey bee was recombinantly expressed as a secreted glycoprotein in Pichia pastoris. The active enzyme was produced in milligram quantities per liter of primary culture. When changing the codons of the original transcript to triplet sequences preferred by P. pastoris, no further increase of protein product could be achieved. After expression of a fusion protein by linking hyaluronidase and human serum albumin together with the recognition sequence for the protease, factorXa, fragmented protein products were obtained in the culture supernatant. Only after replacement of the hinge region with a serine-glycine-rich linker, stable full-length fusion protein could be generated. The protein products were purified by cation exchange chromatography at pH 5.0 and pure enzyme fractions were further characterized in detail. The biochemical properties of the product matched those of crude hyaluronidase within bee venom: the native and the recombinant enzyme exhibited activity over a pH range from 3 to 8 (maximum: 3.8), at temperatures as low as 4°C and up to 90°C (maximum 62°C), and at ionic strength as high as 2 M salt. Recombinant bee hyaluronidase efficiently degrades 6-S-chondroitin sulfate (chondroitin sulfate C) as well as 4-S-chondroitin sulfate (chondroitin sulfate A), the latter to a lesser extent. Only very little hydrolase activity towards chondroitin sulfate B (dermatan sulfate) was detectable