2010 Articles

CD8+ T cell concentration determines their efficiency in killing cognate antigen-expressing syngeneic mammalian cells in vitro and in mouse tissues

Budhu, Sadna; Loike, John D.; Pandolfi, Ashley; Han, Soo; Catalano, Geoffrey; Constantinescu, Andrei; Clynes, Raphael A.; Silverstein, Samuel C.

We describe a quantitative model for assessing the cytolytic activity of antigen-specific CD8+ T cells in vitro and in vivo in which the concentration of antigen-specific CD8+ T cells determines the efficiency with which these cells kill cognate antigen-expressing melanoma cells in packed cell pellets, in three-dimensional collagen-fibrin gels in vitro, and in established melanomas in vivo. In combination with a clonogenic assay for melanoma cells, collagen-fibrin gels are 4,500-5,500-fold more sensitive than the packed cell pellet-type assays generally used to measure CD8+ T cell cytolytic activity. An equation previously used to describe neutrophil bactericidal activity in vitro and in vivo also describes antigen-specific CD8+ T cell-mediated cytolysis of cognate antigen-expressing melanoma cells in collagen-fibrin gels in vitro and in transplanted tumors in vivo. We have used this equation to calculate the critical concentration of antigen-specific CD8+ T cells, which is the concentration of these cells required to hold constant the concentration of a growing population of cognate antigen-expressing melanoma cells. It is ~3.5 × 10^5/ml collagen-fibrin gel in vitro and ~3 × 10^6/ml or /g melanoma for previously published studies of ex vivo-activated adoptively transferred tumor antigen-specific CD8+ T cell killing of cognate antigen-expressing melanoma cells in established tumors in vivo. The antigen-specific CD8+ T cell concentration required to kill 100% of 2 × 10^7/ml cognate antigen-expressing melanoma cells in collagen fibrin gels is ≥10^7/ml of gel.