FP7 Project 202222 - Predict IV

    Participant 3: IMU

    Innsbruck Medical University, Dept of Physiology and Medical Physics, Division Physiology

    Participant leader: Prof. Dr. Walter Pfaller and Dr. Paul Jennings

    Relevant experience and laboratory resources:

    The Renal Physiology-Group has ample experience in characterization of metabolic properties of renal cell cultures and is pioneer in the development of perfusion and co-cultures of human renal and pulmonary cell systems. It has further ample experience in assessment of key epithelial function like electrophysiological properties, trans- and para-cellular transport of solutes and its regulation by external signals (cytokines). In addition expertise has been developed in the field of mitogenic signals controlling growth and differentiation as well as function of renal epithelial cells. The laboratory has experience in utilizing morphological methods at all levels, i.e. electron and cryo-immuno-electronmicroscopy, conventional and confocal immunofluorescence, image analysis and stereology, real time video imaging to assess intracellular changes in Ca++, mitochondrial membane potentials (JC-1) etc. The Physiology laboratory is further equipped for performing transport studies, electro-physiological (resistance), biochemical (enzyme, metabolite assays) and molecular biology (e.g. western blotting) assays. Expertise has been acquired recently in the assessment of gene expression from human renal cell cultures using DNA array and PCR techniques.

    Principal staff involved:

    Prof. W. Pfaller (M) and Dr. Jennings (M) have extensive experience in perfusion culturing of mono- and co-culture systems as well as in renal in vivo and in vitro toxicology. Furthermore experience exists with regard to morphological and immuno- cytochemical characterization of a wide variety of renal cell culture systems (Ms E. Nemati, F). Expertise also is available with respect to the electrophysiological characterization of renal epithelial barriers exposed to a variety of nephrotoxins (Dr. J. Lechner, F).

    Prof. Gstraunthaler (M) and Dr. Lechner (F) are experienced in the assessment of molecular intracellular signalling pathways regulating metabolism cell growth and differentiation. Dr. Jennings (M) and Mag. Aydin (F) are experienced in genomic analyses of cells and quantitative PCR.

    Prof. Pfaller is a member of the ECVAM Taskforce in Chronic toxicty, a member of ERGATT and has participated in several European Commission programmes (BRIDGE, FP-4, FP-6). He has coordinated an in vitro pharmacotoxicity project during the 4th Framework. In addition he currently is acting Chairman of the Austrian "Centre for Alternatives in Animal Experimentation – zet" (NGO).

    Role of the participant in the project and main tasks:

    1) Generation of well characterized stable mono- and co-cultures of proximal tubular renal epithelial cell lines (HK-2) and human primary dermal microvascular endothelial cells and fibroblasts (preferably renal interstitial fibroblasts) under classical static cell culture conditions utilising filter inserts; 2) Establishment of the above mono- and co-cultures under perfusion conditions, which will allow for continuous and repeated administration of test compounds; 3) Standardization of cell culture techniques to be used and decision on Standard Operation procedures (SOP´s) for the renal cell culture methodologies applied. 4) Identification of changes in renal cellular function following exposure of cell culture systems to test compounds selected. 5) Assessment of changes in intracellular signalling pathways triggered by the nephrotoxicants and identification of their potency in gene activation. 6) Development of non target organ based cell culture models. 7) Development of a modular co-culture system for the investigation of bioactivation of test compounds by liver cells expressing CYPs and the effect on down stream target organ related cell systems.

    A recently developed cell culture apparatus (EpiFlow developed in a FP-4 project), which allows maintenance of cell cultures under conditions of continuous medium replenishment and removal over prolonged periods of time even when using media with different composition at the two sides of an epithelial layer (blood /urine), will be refined and adopted for the specific needs of this project in a way that information on dysfunction of those renal tissue components involved in and interacting during chronic renal disease states will be obtained.

    This group will undertake the leadership of WP2, will be primarily involved in WP2.2, WP2.4 and WP2.5 and will collaborate in WP3 and WP4. Prof. Pfaller will act as a member of steering committee of the project.

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