The requirement for monocytes in spontaneous cytotoxicity by lymphocytes from healthy donors and melanoma patients

Lymphocytes from 12 healthy donors were depleted of monocytes by velocity sedimentation at unit gravity (average monocyte contamination 0.2%) and tested for spontaneous cytotoxicity (SC) against tumor target cells in 24 h microcytotoxicity assays. Sixty percent of all lymphocytes were recovered in this monocyte-de-pleted lymphocyte fraction (LF). In contrast, with the corresponding unfractionated lymphocytes (UL) the LF cells were not spontaneously cytotoxic on T₂₄ bladder carcinoma, NKI-1, NKI-7, NKI-8, NKI-1O melanoma, and MC-1 mammary carcinoma cell line cells and target cells derived from two short-term melanoma cultures (Me 215 and Me 223). SC of the LF could be restored by reconstitution with autologous monocytes, which were obtained >80% pure in the same velocity sedimentation procedure. Addition of 4–8% monocytes was sufficient to restore the SC of the LF to the level achieved with the corresponding UL, whereas maximal induction of SC was observed after addition of 8–16% monocytes. Higher numbers of monocytes had suboptimal effects. SC of the LF could also be restored by monocyte culture supernatants. The spontaneous cytotoxic effector cells were characterized as non-E rosette-forming, membrane slg-negative Fc-receptor-bearing lymphocytes, a proportion of which probably also bear C₃-receptors. Since the LF was not spontaneously cytotoxic, in spite of the presence of Fc-receptor bearing lymphocytes, it is concluded that monocyte help, mediated via soluble factors, is required. Evidence is presented that the lack of SC by the LF might be attributable to the failure of these cells to make appropriate contact with the target cells. LF/target cell contact followed by target cell kill occurred after addition of monocytes. Effector/target cell contacts, artificially established by agglutinins, did not result in target cell kill. Thus the effector/target cell contacts observed in the presence of monocytes suggest recognition of particular membrane determinants involved in SC. An identical requirement for monocytes was observed with LF cells from 5 stage I and 5 stage II melanoma patients tested for SC against both melanoma and non-melanoma target cells. This indicates that either the cytotoxicity against melanoma cells with lymphocytes from melanoma patients is of the same nature as the SC of lymphocytes from healthy donors, or that specific melanoma-associated cellular cytotoxicity also requires monocyte help.