We develop technologies for intravital imaging and live cell imaging to achieve new insights into chronic inflammatory processes
Recently, we developed and refined an endogenous fluorescence method for in vivo microscopy to monitor the NAD(P)H-dependent enzymatic activity in cells. This method allows us to distinguish between various classes of enzymes based on the spatially-resolved fluorescence lifetime of the coenzymes NADH and NADPH. Using this technique, we were able to define the concept of “oxidative stress memory” in the context of chronic neuroinflammation and neurodegeneration, both in mouse models and in patients. We found that astrocytes and microglia within the central nervous system (CNS) of diseased mice show a partially irreversible phenotypic shift towards oxidative stress generation, which is also present in monocytes of MS patients. Due to the persistence of plasma cells in the CNS in the late disease phases, we assumed and found that antibodies maintain this phenotypic shift of astrocytes in human brain slices treated with a NMDA-R antibody. This project is a collaboration with Dr. H. Radbruch and Prof. Dr. F. Heppner (Neuropathology, Charité), Prof. Dr. F. Paul (Neuroimmunology, Charité) and Prof. Dr. A.E. Hauser (DRFZ). Since we expect a similar phenotypic shift in renal tissue of mice affected by lupus, we are currently performing longitudinal NAD(P)H fluorescence lifetime imaging of the kidney in collaboration with Prof. Dr. R. Voll (Erlangen, CRC130).
Long-lived plasma cells are central players in forming both physiologic and pathologic immunological memory. Due to a lack of technological tools, these cells could not be investigated over their whole lifetime in their natural environment – the bone marrow of long bones. We therefore developed LIMB: Longitudinal Intravital Microscopy of the femoral Bone marrow. In this way, we were able to monitor plasma cells over 157 days in the deep cavity of murine femurs. This technology has also allowed us to repeatedly monitor monocytes and macrophages, as well as vasculature remodelling in models of tissue regeneration after bone injury.
Independent of the organ of interest, intravital microscopic investigation faces the challenge of capturing the high complexity of cellular subpopulations and extracellular tissue structures in a dynamic manner. This requires simultaneous multiplexed imaging of the living tissue, as designed and demonstrated by us imaging germinal center reactions in murine popliteal lymph nodes.
Intravital multi-photon microscopy
Functional in vivo imaging
Prof. Dr. rer. nat. Raluca A. Niesner
Dr. Asylkhan Rakhymzhan
Volker Andresen, LaVision Biotec – Milteny, Bielefeld/Germany
Dr. Marcus Beutler, APE, Berlin/Germany
Prof. Georg Duda, Julius-Wolf-Institute, Charité /Berlin/Germany
Dr. David Entenberg, Albert Einstein College of Medicine/Intravital Imaging/ New York/US
Prof. Dr. Susanne Hartmann, Freie Universität, Berlin /Veterinary Medicine/Berlin/Germany
Prof. Dr. Anja E. Hauser, DRFZ and Charité/Immune Dynamics/ Berlin/Germany
Prof. Dr. Frank Heppner, Charité /Neuropathology/Berlin/Germany
Dr. Julia Jellusowa, University Freiburg/Faculty for Biology/Freiburg/Germany
Romano Matthys, RISystem, Davos/Switzerland
Prof. Dr. Dirk Mielenz, University Nürenberg-Erlangen / Immunology/ Erlangen/Germany
Prof. Dr. Friedemann Paul, Charité /Neuroimmunology/Berlin/Germany
Dr. Helena Radbruch, Charité /Neuropathology/Berlin/Germany
Prof. Dr. Tim Schulz, DIfE / Potsdam/Germany
Dr. Heinrich Spiecker, LaVision Biotec – Milteny, Bielefeld/Germany
Prof. Dr. Reinhard Voll, University Hospital Freiburg/Rheumatology/Freiburg/Germany
Prof. Dr. Carolina Wählby, Uppsala University/Quantitative Microscopy/Upsalla/Sweden
Prof. Dr. Jürgen Zentek, Freie Universität, Berlin / Veterinary Medicine /Berlin/Germany
DFG: TRR130, C01
DFG: FOR2165-2, TP7
ISAC: ISAC Scholar Collaboration Grant together with Carolina Wählby
- “Synergistic Strategy for Multicolor Two-photon Microscopy: Application to the Analysis of Germinal Center Reactions In Vivo.“ Rakhymzhan A, Leben R, Zimmermann H, Günther R, Mex P, Reismann D, Ulbricht C, Acs A, Brandt AU, Lindquist RL, Winkler TH, Hauser AE, Niesner RA. Sci Rep. 2017 Aug 2;7(1):7101. doi: 10.1038/s41598-017-07165-0.
- “Analyzing Nicotinamide Adenine Dinucleotide Phosphate Oxidase Activation in Aging and Vascular Amyloid Pathology.“ Radbruch H, Mothes R, Bremer D, Seifert S, Köhler R, Pohlan J, Ostendorf L, Günther R, Leben R, Stenzel W, Niesner RA*, Hauser AE.* Front Immunol. 2017 Jul 31;8:844. doi: 10.3389/fimmu.2017.00844. eCollection 2017.
- “Longitudinal intravital imaging of the femoral bone marrow reveals plasticity within marrow vasculature.“ Reismann D, Stefanowski J, Günther R, Rakhymzhan A, Matthys R, Nützi R, Zehentmeier S, Schmidt-Bleek K, Petkau G, Chang HD, Naundorf S, Winter Y, Melchers F, Duda G, Hauser AE*, Niesner RA*. Nat Commun. 2017 Dec 18;8(1):2153. doi: 10.1038/s41467-017-01538-9.
- „Phasor-Based Endogenous NAD(P)H Fluorescence Lifetime Imaging Unravels Specific Enzymatic Activity of Neutrophil Granulocytes Preceding NETosis.“ Leben R, Ostendorf L, van Koppen S, Rakhymzhan A, Hauser AE, Radbruch H, Niesner RA. Int J Mol Sci. 2018 Mar 29;19(4). pii: E1018. doi: 10.3390/ijms19041018.
- „Multiplexed fluorescence microscopy reveals heterogeneity among stromal cells in mouse bone marrow sections“ Holzwarth K., Köhler R., Philipsen L., Tokoyoda K., Ladyhina V., Wählby C., Niesner R.A., Hauser A.E. Cytometry Part A, 2018