Principle investigators: Sigurd Lenzen and Uwe Lendeckel
Co-supervisor: F. Scholz

Diabetes mellitus is the most important metabolic disease worldwide with a dramatically increasing prevalence within the last years. Pancreatic beta cells, which are responsible for insulin synthesis and secretion, are particularly sensitive to oxidative stress due to low expression of the hydrogen peroxide (H₂O₂) inactivating enzymes glutathione, peroxidase and catalase [1]. As a consequence, hydrogen peroxide accumulates intracellularly and forms in an iron (II) ion catalysed reaction highly active hydroxyl radicals. While reactive oxygen species (ROS), which destroy beta cells through action of proinflammatory cytokines in type 1 diabetes (T1D), are generated primarily intramitochondrially [2], ROS generation through lipotoxicity (i.e. palmitate) in type 2 diabetes (T2D) is primarily in the peroxisomes [3; 4]. Additionally high amounts of H₂O₂ are produced by the disulfide isomerase (PDI)/ER oxidoreductin 1 (ERO-1) system during disulfide bond formation in active ER of beta cells to secure high rates of insulin biosynthesis. The described processes of H₂O₂formation in the different organelles were analysed with the tagged variants of the specific sensor fluorescent protein HyPer [5]. Organelles are surrounded by biological membranes which differ in their permeability towards H₂O₂ and sensitivity against oxidation by ROS. The permeability will be determined by newly designed HyPer fusion proteins containing sequence tags for the expression in the membranes of organelles in insulin-producing tissue culture cells and primary murine beta cells. The new HyPer variants together with established organelle specific isoforms will be used in collaboration with project A3 (Mayerle/Lerch) to identify sources of H₂O₂ generation in pancreatic acinar cells during pancreatitis and with project C2 (Lendeckel) for the analyses of ROS sources and mechanisms of toxicity in pancreatic beta cells. For the efficient transduction of tissue culture cells and especially primary non-dividing cells with the HyPer-cDNA the latest lentiviral vector system is available. ROS and nitric oxide (NO) which is formed in pancreatic beta cells during T1D or T2D manifestation can significantly alter the properties of membranes. In cooperation with project B2 (Scholz) and project B1 (Helm) the effects of ROS and NO on fluidity of plasma membranes and membranes of intracellular organelles and the oxidation of cardiolipin will be analysed. Newly developed electrodes will allow in cooperation with project B4 (Kahlert/Lalk) the measurement of hydroxyl radicals and characterisation of new antioxidants for their scavenging properties. Furthermore within project B4 the effects of elevated levels of fatty acids in combination with antioxidants will be investigated.

Literature

1. Lenzen S: Oxidative stress: the vulnerable beta-cell. Biochem Soc Trans 2008;36:343-347

2. Gurgul-Convey E, Mehmeti I, Lortz S, Lenzen S: Cytokine toxicity in insulin-producing cells is mediated by nitro-oxidative stress-induced hydroxyl radical formation in mitochondria. Journal of molecular medicine 2011;89:785-798

3. Elsner M, Gehrmann W, Lenzen S: Peroxisome-generated hydrogen peroxide as important mediator of lipotoxicity in insulin-producing cells. Diabetes 2011;60:200-208

4. Gehrmann W, Elsner M, Lenzen S: Role of metabolically generated reactive oxygen species for lipotoxicity in pancreatic beta-cells. Diabetes, obesity & metabolism 2010;12 Suppl 2:149-158

5. Gehrmann W, Elsner M: A specific fluorescence probe for hydrogen peroxide detection in peroxisomes. Free radical research 2011;45:501-506