Chemotaxis of T lymphocytes on extracellular matrix proteins Analysis of the in vitro method to quantitate chemotaxis of human T cells

The present report compares a variety of T cell purification protocols and chemotaxis procedures in assessing chemokine-induced T cell migration using a microchemotaxis assay. Rapidly purified T cells are capable of directly responding to the β chemokines macrophage inflammatory protein-1α (MIP-1α), MIP-1β, and RANTES in the absence of αCD3 stimulation as previously described (Taub, D.D. and Oppenheim, J.J. (1993) Cytokine 5, 175). However, T cell purification schemes involving prolonged 37° C incubations generally produce non-motile T lymphocytes that require stimulation with αCD3 antibody for 6–12 h in culture to recover chemotactic mobility. This loss of chemotactic potential appears to be due to prolonged 37° C incubations as rapidly purified T cells lose migratory activity upon incubation at 37° C. Radiolabeled binding analysis revealed that β chemokine binding sites are downregulated as short as 2 h after incubation at 37° C. T cells require the presence of extracellular matrix molecules to facilitate T cell migration. While many of these proteins permit chemotactic activity, human plasma and foreskin fibronectin were found to be the most effective matrix molecule for T cell migration. Kinetic analysis of T cell activation revealed that 6–12 h of anti-CD3 stimulation was optimal to restore the ability of purified T cells to migrate in response to the chemokines MIP-1α, MIP-1β, RANTES, and IL-8. However, rapidly dividing T cells ( ≥ 48 h post αCD3 mAb stimulation) fail to migrate in response to any chemotactic stimulus. Together, these results suggest that the measurement of T cell migration, using microchemotaxis chambers, is a multifactorial process with strict environmental and activation requirements.