In vitro Stimulation of Natural Killer (NK) Cells by Soluble Factor(s) Generated in Antigen-Stimulated Rhesus Monkey Peripheral Blood Lymphocyte Cultures

Peripheral blood lymphocytes from rhesus monkeys (Macaca mulatta) sensitized to keyhole limpet hemocyanin (KLH), when stimulated in vitro with KLH, developed natural killer (NK) cell activity that was assayed with Rous Sarcoma virus-transformed marmoset fibroblasts as targets in a 4-hr 51Cr-release assay. The supernatant fluids from 24- to 25-hr KLH-activated cultures were capable of stimulating NK development in nonsensitive lymphocyte cultures. The effector cells were neither macrophages nor B cells (plastic and nylon-wool nonadherent) and did not form E-rosettes with neuraminidase-treated sheep red blood cells. Cultures depleted of EA-rosetting cells, i.e., Fc-receptor-bearing lymphocytes, were incapable of generating NK activity when stimulated in vitro. Kinetic studies showed that peak DNA synthesis, as measured by 3H-T incorporation, preceded maximum cytotoxicity. Elimination of dividing cells by 5-bromo-2'deoxyuridine (BrdU) and light treatment during the interval from day 1 to day 4 inhibited the development of cytotoxicity on day 7. Cell replication was required for the induction of NK cells with KLH as well as with antigen-activated culture supernatant fluids. When cultures were left unstimulated for 4 days, NK activity could not be developed subsequently either by adding antigen, mitogen (PHA), or supernatant fluids from activated cultures.