Relation between locomotion, chemotaxis and clustering of immune cells

Experiments were designed to discover whether locomotor or chemotactic events are needed for clustering of lymphocytes with accessory cells or, conversely, whether clustering precedes the activation of lymphocyte locomotion. The time-courses of clustering and locomotor activation were compared and the behaviour of moving cells during cluster formation was filmed. Human lymphocytes direct from blood were activated by culture for 24-48 hr with anti-CD3 antibody or in allogeneic mixed leucocyte reactions (AMLR). The proportion of clustered and locomotor lymphocytes was low at the beginning of culture. Clusters appeared during the first few hours, before the increase in numbers of locomotor lymphocytes. Filming gave no evidence that the cells attracted one another chemotactically to form clusters. Rather, cells made chance contact by random locomotion and then remained adherent, though lymphocytes very close (less than or equal to 10 microns) to clusters did show increased pseudopod formation towards the cluster. However, the behaviour of motile lymphocytes responding to monocytes or macrophages given a phagocytic stimulus was different. Human monocytes which ingested opsonized zymosan released a material during but not following phagocytosis which caused an immediate increase in polar shape-change in lymphocytes. Macrophages from Corynebacterium parvum-induced mouse peritoneal exudates, given a phagocytic stimulus (opsonized Candida albicans), acted as sources of chemotactic gradients which attracted nearby lymphocytes to form clusters. This was due to brief release of a material immediately following phagocytosis, but after 15 min or so the macrophages no longer attracted nearby cells. These experiments suggest that, during induction of an immune response to a non-phagocytic stimulus, clusters form slowly by random contact followed by preferential adhesion. However, after phagocytosis, there may be a chemotactic response to the ingesting macrophage. This may help to focus lymphocytes onto macrophages which present microbial antigens.