Project Details
Projekt
Functional intravital imaging platform
Applicant
Dr. Ahmed Ghallab
Subject Area
Gastroenterology
Term
since 2026
Project identifier
Deutsche Forschungsgemeinschaft (DFG) - Project number 546585894
Spatio-temporal analysis of the sequence of events in disease pathogenesis is fundamental for understanding the driving mechanisms and identifying therapeutic interventions. Recently, we have established a functional two-photon-based intravital imaging platform that allowed us to identify pathophysiologically relevant mechanisms responsible for stage transitions of liver disease at a spatio-temporally resolved cellular and subcellular level. Intravital imaging with a spatial resolution of ~200 nm and a temporal resolution in the millisecond rate is possible. In preparation for this project the following intravital imaging toolboxes were established in close cooperation with all other projects of the dangerhep consortium: (i) a toolbox for intravital detection of the mode of cell death. This includes mitochondrial potential markers (e.g. tetramethyl rhodamine ethyl ester, and rhodamine123); and membrane integrity markers (e.g., propidium iodide and Sytox green). This allowed to distinguish lethal versus sublethal necroptosis as well as apoptosis versus necrosis; this toolbox will particularly support projects P01-03 and P05 in the dangerhep consortium. (ii) A toolbox for intravital imaging of DAMP release and the local microenvironment. This includes reporter mice (e.g. the HMGB1-mCherry reporter to detect the release of HMGB1 as a DAMP which will supports P01-03 and P05; and the GFP-LC3 reporter to visualize secretory autophagosomes which is important to achieve the goals of P04); fluorophore-coupled bile acids/bile acid analogues (e.g. 3β-NBD-TCA and cholyl-lysyl-fluorescein) which will be used by P07 to investigate how modification of sphingolipids may alter the apical membrane of hepatocytes and the integrity of the blood-bile-barrier; and nucleic acid dyes (e.g. Hoechst 33258) that will be use for intravital detection of nucleic acids as DAMPs in the various mouse models of the dangerhep consortium. (iii) Intravital imaging of immune cell infiltration in relation to the time and mode of cell death; this includes cell-specific reporter mice and/or antibodies, as well as adoptive transfer of labelled immune cells from donor to acceptor mice. This toolbox will particularly serve P05, P08, and P09. (iv) Intravital imaging of the liver-adipose tissue axis; the tools established in the liver were transferred to the adipose tissue to support the projects focusing on interaction of these two compartments (P05 and P06). This platform will be further developed and adapted according to the needs of the individual dangerhep consortium projects. All generated videos will be quantified using an automated image analysis pipeline. As a central technical platform project, C01 will intensively cooperate with all other projects in the dangerhep consortium, and the established toolbox will be made available to different research fields and future research consortia.
DFG Programme
Research Units
