Genomische Transkriptionsanalyse des intrinsischen Alterns der menschlichen Haut.

  • Breitenbach, Michael, (Projektleitung)



    Aging and photoaging of human skin is a currently intensively investigated topic of obvious practical and clinical importance. Up to now, genome-wide transcript analysis has been performed with dermal fibroblasts and with keratinocytes in culture, comparing early passage cell cultures with cells after reaching replicative senescence. We are proposing to study by genome-wide transcript analysis on DNA microarrays skin samples from young and old patients and to also compare areas of skin that are not exposed to sunlight. The relation between replicative senescence in vitro and the actual aging process in vivo are generally (and also in the case of skin cells) not clear. Our investigation will clarify this problem at least in part because the transcript levels measured will be compared with the published cell culture results. Another aspect that we are studying is the role of oxidative stress in skin aging. Oxidative stress is believed to play a major role in skin aging and has also been found to occur in the mother cell-specific aging process of yeast cells. We want to investigate in more detail whether this mechanism is indeed a common feature of aging in these widely divergent eukaryotic cells. On the other hand, we will see which mechanisms (mirrored by differential transcriptional gene expression between young and old skin, and between intrinsically aged and photoaged skin) are specific to skin.

    Data generation
    Skin samples will be obtained from different anatomical regions of volunteers after obtaining informed consent.
    To investigate the expression profile of very small tissue samples or even single cells a method to amplify minute amounts of RNA has been developed by Eberwine and coworkers (1992) .With this approach it is possible to start with total RNA,synthesize cDNA using an oligodT-primer which includes the promoter sequence for the T7 RNA polymerase.After second strand synthesis
    The analysis of the images will be processed by commercial available software as well as by public available algorithms implemented in self-designed database systems. Clustering studies will be applied to the results
    Data analysis
    It is essential for the success of the project to use microarrays corresponding to the largest commercially available number of different human genes. Only if we are as close as possible to the total number of genes in the human genome can we hope to detect new and interesting genes that have to do with the very general and also with some specialized aspects of skin aging. The dataset produced in these projects consists of three comparative analyses: old vs young, photoaged vs. young, and old vs, photoaged. The dataset contains overexpressed as well as underexpressed genes. Modern bioinformatics methods (pattern recognition) are needed to compare these relatively complex datasets

    (1) To determine a set of genes that are specifically under- or over-expressed in aging skin. To analyze this set of genes, in particular with respect to the so-called oxygen hypothesis of aging.
    (2) To compare this dataset with the datasets obtained from dermal fibroblast and keratinocyte cultures and from old yeast mother cells. This comparison will lead us to recognize the genes involved in „public mechanisms of aging“ (for instance the genes for defence against oxidative stress which occurs in all eukaryotic cells) and those involved in skin-specific aging genes.
    (3) Those genes in the „public“ group which are consistently found in aging skin, in replicatively aged dermal fibroblasts and in aged mother yeast cells will be genetically analysed in yeast. A large number of yeast genetics methods are at our disposal including the analysis of interactions partners for the products of those genes which are specifically regulated during senescence. Phenotypic analysis of mutants will be centred around measurements of oxidative stress resistance or sensitivity.
    KurztitelTranskription des in vivo Altern der Haut
    Tatsächlicher Beginn/ -es Ende1/07/0430/06/06

    Systematik der Wissenschaftszweige 2002

    • 1407 Genetik
    • 3914 Gerontologie
    • 1411 Molekularbiologie