Migration of innate immune cells is an essential mechanism for the resolution of tissue injuries and inflammation. This process is orchestrated by chemokines including IL-8 and SDF-1α that attract neutrophils and monocytes, respectively, to the site of damage. Dysregulation in this process can contribute to the development of conditions, such as asthma, arthritis and psoriasis.
We developed chemotaxis assays using blood-derived human neutrophils or monocytes to test the effects of drug candidates. Neutrophils were isolated from blood donors using Ficoll and dextran-based separation methods, while monocytes were separate with a CD14 magnetic bead-based approach. Isolated cells were seeded in the upper chamber of a 96-well transwell system containing 5.0 μm pore size, and cell migration to the lower chamber was determined by a luminescent-based ATP assay.
Neutrophils stimulated with IL-8 showed consistent and dose-dependent migration reaching a peak at 10 nM of IL-8 after 1 hour. This migratory effect was inhibited by the addition of the CXCR1/2 antagonist Sch527123 in a dose-dependent fashion. Monocytes showed a strong migratory response upon stimulation with SDF-1α after 4 hours. The addition of the CXCR4 antagonist AMD3100, resulted in a dose-dependent inhibition of SDF-1α-induced migration. IC50 values were consistent across donors demonstrating strong reproducibility for both assays.
Our data demonstrate a robust and donor independent chemokine-induced migration that is inhibited by selective compounds and provide a rapid method to evaluate the in vitro potency therapeutic candidates for the treatment of acute and chronic inflammatory diseases.
Lisa Scandiuzzi– Sr. Scientist, Charles River Laboratories
Monique Hazenoot– Scientist, Charles River Laboratories
Yasmin Habani– Scientist, Charles River Laboratories
Sanne Demandt– Group Leader, Charles River Laboratories
Jeroen DeGroot– Senior Director, Charles River Laboratories
Jamil Aarbiou– Group Leader, Charles River Laboratories