Migration of CXCR3+ T regulatory cells towards the chemokine IP-10

Introduction Chemotaxis is the process where cells migrate under influence of specific chemoattractant mediators (chemokines) toward an extracellular gradient. The study of in vitro chemotaxis with traditional methods has its limitations, however a new promising method to study cell migration on single cell level is with microfluidics. I designed a microfluidic model to study migratory behavior of a CD4+ T regulatory cell subset (Tregs). Tregs are essential for tempering immune reactions. The traffic of T regulatory cells to local inflammatory sites is now known to be important for suppression and tolerance, therefore migration of Tregs from blood to the inflamed tissue is essential for the suppression of alloimmunity. Cell migration is promoted by chemokines; a well-established T cell chemoattractant is interferon-inducible protein 10 (IP-10). One of its ligands, CXCR3, is expressed on activated human CD4+ T cells and is important in T effector cell trafficking. The presence of CXCR3+ T cells and IP-10 within allograft biopsies is associated with rejection. However, the Briscoe lab recently found that CXCR3 is also expressed on Tregs. With help of microfluidics I proof that CXCR3+ Tregs migrate to the chemokine IP-10. Materials and Methods For my migration studies I isolated human CD4+CD25+CD127dim/- cells from PBMC and stained them with an anti-CXCR3 antibody. With FACS I sorted the Tregs into CXCR3+ and CXCR3- populations. I introduced these populations in our microfluidic device, which consist out of a microfluidic network array of up to 450 parallel micro channels connected to one main channel. The device was primed with a chemokine solution of IP-10 (100 nM) and fibronectin (250 nM). The cells were monitored for 3 hours using time-lapse microscopy. Results I designed a microfluidic device that enabled us to study cell migratory behavior on a single cell level. Using my device we found that CXCR3+ Tregs migrate towards IP-10 and that the directional persistence of CXCR3+ is significantly greater (P