Experimental application of a multistep kinetic model for natural cytotoxicity: determination of rate constants for lytic programming and killer-cell-independent lysis.

A multistep kinetic model for natural cytotoxicity reactions in vitro is described that includes consideration of the effect of nonlytic target-binding lymphocytes on the experimentally determined kinetic parameters. The expression for the maximal velocity, Vmax, obtained using this model is essentially identical to that obtained using simpler models, whereas the expression for the apparent Michaelis constant, KappM, is considerably more complex. As a first step in the application of this model, experiments were designed to determine the relative contribution of some of the component terms to the value of KappM. Methods were developed that allow for experimental determination of the rate constants for both lytic programming and killer-cell-independent lysis (KCIL) steps in the cytolytic process. The results obtained support lytic programming as the rate-determining step in natural cytotoxicity reactions and demonstrate that terms related to nonlytic target-binding lymphocytes contribute significantly to experimentally determined values for KappM. In addition, the methods developed for the determination of the rates of lytic programming and KCIL should prove useful for various studies of the mechanism of cytotoxicity and the effects of drugs and disease on such phenomena.