Georg Feger

Osteopontin is upregulated during in vivo demyelination and remyelination and enhances myelin formation in vitro.

We have used in vitro oligodendrocyte differentiation and the in vivo remyelination model, the cuprizone model, to identify genes regulating oligodendrocyte function and remyelination. One of the genes we identified, osteopontin (opn), is a secreted glycoprotein with cytokine-like, chemotactic, and anti-apoptotic properties that contains an Arg-Gly-Asp (RGD) cell adhesion motif-mediating interactions with several integrins. Both microglia and astrocytes in demyelinating brain regions of cuprizone-fed mice expressed OPN protein. Recombinant OPN protein produced in a baculovirus expression system induced proliferation of both the rat CG-4 and the mouse Oli-neu oligodendrocyte precursor (OLP)-like cell lines in a dose-dependent manner. In addition, recombinant OPN treatment stimulated both myelin basic protein (MBP) synthesis and myelin sheath formation in mixed cortical cultures from embryonic mouse brain, an in vitro primary culture model of myelination. Interestingly, myelinating mixed cultures prepared from OPN(-/-) mice contained significantly less MBP compared to wild-type cultures after 17 days in culture. We propose that in the central nervous system, OPN may act as a novel regulator of myelination and remyelination.

Detecting the impact of sequencing errors on SAGE data

UNLABELLED SAGE data are obtained by sequencing short DNA tags. Due to the mistakes in DNA sequencing, SAGE data contain errors. We propose a new approach to identify tags whose abundance is biased by sequencing errors. This approach is based on a concept of neighbourhood: abundant tags can contaminate tags whose sequence is very close. The application of our approach reveals that moderately abundant tags can be generated by sequencing errors uniquely. It also allows for detecting correct rare tags. AVAILABILITY Software is available only to non-profit entities and for non-commercial purposes upon request.

MIG--differential gene expression in mouse brain endothelial cells.

Different diseases of the CNS are associated with blood–brain barrier (BBB) damage and mononuclear cell infiltration. In order to study genes that may play a role in endothelial cell regulation in inflammatory CNS diseases, we performed differential gene expression (DGE) analysis using a mouse brain endothelial cell line. We found that interferon-γ (IFNγ)-induced monokine (MIG), a chemokine that plays a role in T lymphocyte and monocyte chemoattraction, is highly expressed in the presence of inflammatory cytokines.We show that MIG, produced by brain endothelial cells in vitro, is biologically active in attracting T lymphocytes and that it is possible to interfere with this mechanism of action using anti-MIG antibodies. We suggest that blocking MIG may be beneficial in CNS inflammation. We detected constitutive expression of the MIG receptor, CXCR3, on the surface of the endothelial cells and therefore hypothesize that it plays a role in maintaining the cytokine gradient at the region of CNS inflammation.

Tumor Necrosis Factor (TNF)-Soluble High-Affinity Receptor Complex as a TNF Antagonist

A novel high-affinity inhibitor of tumor necrosis factor (TNF) is described, which is created by the fusion of the extracellular domains of TNF-binding protein 1 (TBP-1) to both the α and β chains of an inactive version of the heterodimeric protein hormone, human chorionic gonadotropin. The resulting molecule, termed TNF-soluble high-affinity receptor complex (SHARC), self-assembles into a heterodimeric protein containing two functional TBP-1 moieties. The TNF-SHARC is a potent inhibitor of TNF-α bioactivity in vitro and has a prolonged pharmacokinetic profile compared with monomeric TBP-1 in vivo. Consistent with the long half-life, the duration of action in an lipopolysaccharide-mediated proinflammatory mouse model is prolonged similarly. In a collagen-induced arthritis mouse model, this molecule demonstrates improved efficacy over monomeric TBP-1. Based on these results, we demonstrated that inactivated heterodimeric protein hormones are flexible and efficient scaffolds for the creation of soluble high-affinity receptor complexes.

Improving SAGE di-tag processing

BackgroundSAGE is a genome-wide method for obtaining gene expression profiles. It generates tags of 10 nucleotides in length, which are assumed to determine the corresponding gene transcript. In practice however, this is not always sufficient for uniquely identifying a gene.ResultsWe propose an improved processing of SAGE sequences that allows us to obtain one extra base for reasonably abundant tags. This method includes a statistical test for controlling the relevance of extra base predictions.ConclusionsThe improved SAGE sequence processing we present reduces the uncertainty in SAGE tag to gene mapping and can be applied to any SAGE library.