IL 2 Activated Killer 細胞の Heterogeneity に関する研究

Lymphokine activated killer (LAK) cells are generated by a culture of lymphocytes with interleukin 2 (IL 2) without the need for any additional antigenic stimulation. LAK cells can lyse a wide spectrum of allogeneic and syngeneic tumor cells regardless of whether the tumor target cells are sensitive or resistant to fresh natural killer (NK) cells. In these studies, the phenotypic and functional characteristics of LAK cells by using metabolic inhibitors and various monoclonal antibodies were investigated. The following results were obtained: (1) Human peripheral blood lymphocytes were cultured in the presence of 2 unit/ml of human recombinant IL 2 (TGP-3, Takeda, Osaka), the dose of which is routinely used for the activation of LAK cells in human pilot studies. The killing activity was generated within 4 days and continued for more than 14 days. The maximum consumption of r-IL 2 was observed at the 5 th day of culture. (2) The subfractionation of lymphocytes by Percoll discontinuous gradient centrifugation revealed that the main LAK activity was located in the low density fraction of less than 1.061 g/ml. (3) LAK cells generated by a higher dose of r-IL 2 (2 unit/ml), are resistant to inhibitors for lipid peroxidation such as dimethylsulfoxide, ethyl alcohol, and nordihydroguaiaretic acid, while NK lineage killers such as fresh NK cells and killer cells activated by a lower dose of r-IL 2 (approximately 0.02 unit/ ml) according to the method of Itoh et al. are sensitive to those reagents. (4) Sodium azide, an inhibitor of mitochondrial oxidation blocked the induction of LAK activity even at a concentration of 3.3×10-5 mole/L (0.001%). (5) By a negative selection of lymphocytes using complement and monoclonal antibodies, CD 3 and CD 8 tends to inhibit LAK activity at 2 unit/ml of r-IL 2. (6) By a positive selection using a fluororescent activated cell sorter, LAK cell activity was obtained in all the sorted fractions of low density lymphocytes including CD 16(+), CD 16(-), CD 3 dull(+) and CD 3(-) cells. LAK cells newly derived from CD 16(-) cells were insensitive to lipid peroxidation inhibitors. The CD 3 dull(+) cells changed to CD 3 strong (+) after cultivation with IL-2. It is concluded that LAK cells induced by a reatively higher dose of r-IL 2 have different lytic pathways from killer cells of NK lineage, and LAK activity appears to be derived from phenotypically heterogenous population and confined to any single subpopulation