Imaging, myeloid precursor immortalization, and genome editing for defining mechanisms of leukocyte recruitment .

Recruitment of leukocytes from the blood to sites of inflammation poses a promising target for new diagnostic and therapeutic approaches. We aimed to develop a novel method to non-invasively analyze molecular mechanisms of leukocyte migration in pre-clinical models of inflammation . We used the ER-HoxB8 system to transiently immortalize murine myeloid precursors from and CD18- as well as MRP14-deficient mice. A cell line was generated by CRISPR/Cas9-mediated gene editing. We analyzed the migration of and leukocytes by optical and nuclear imaging in mice with irritant contact dermatitis, cutaneous granuloma, experimental arthritis and myocardial infarction. Transient immortalization, gene editing and imaging can be combined to analyze migratory mechanisms of murine leukocytes, even for gene deletions resulting in lethal phenotypes in mice. We reliably confirmed known migratory defects of leukocytes deficient for the adhesion molecules CD18 or VLA4α. Also, using our new method we identified a new role of the most abundant calcium-binding proteins in phagocytes and major alarmins in many inflammatory diseases, MRP8 and MRP14, for transmigration . We provide a combinatorial approach to rapidly characterize molecular mechanisms of leukocyte recruitment with the potential to aid in identification of diagnostic and therapeutic targets in inflammatory pathologies.