Studies on the mechanism of T cell-mediated lysis at the single effector cell level. I. Kinetic analysis of lethal hits and target cell lysis in multicellular conjugates

Zagury, D.; Bernard, J.; Jeannesson, P.; Thiernesse, N.; Cerottini, J.C.

Journal of Immunology 123(4): 1604-1609


ISSN/ISBN: 0022-1767
PMID: 314465
Accession: 068526088

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A procedure was previously developed to allow kinetic analysis of target cell lysis in individual conjugates formed between murine alloimmune peritoneal exudate lymphocytes (PEL) and relevant tumor cells in suspension. Under the conditions used in the present work, 60% of the conjugates consisted of 1 lymphocyte bound to 1 target cell, whereas the remaining were multicellular, i.e., contained 1 lymphocyte bound to either 2 (24%), 3 (12%), or 4 (4%) target cells. [EL4 tumor cells in ascites form were used.] Both ultrastructural studies and micromanipulation experiments with individual multicellular conjugates indicated differences in the degree of target cell binding to the same PEL. In more than 80% of the multicellular conjugates, lysis of all conjugated target cells occurred within a 3 h incubation period at C. Kinetic analysis of target cell lysis showed that the time required for target cell death was quite variable, both within the same multicellular conjugate and between different conjugates, ranging from a few minutes to 3 h after conjugation. By comparing the elapsed time necessary for lysis of 50% of the target cells in conjugates containing 1 lymphocyte bound to 3 target cells, it could be estimated that, on the average, lysis of the 1st, 2nd and 3rd target cells occurred after 30, 60 and 90 min, respectively. The time required for lethal hit of each conjugated target cell was measured by adding EDTA to the incubation medium of individual multicellular conjugates at various time intervals. Evidence could be obtained that a CTL induced irreversible lesions in only 1 of the conjugated target cells at a time, i.e., lethal hits were given sequentially rather than simultaneously.