Julie Y. Djeu

Inhibition of rat mixed lymphocyte cultures by suppressor macrophages

Abstract Normal rat spleens contain suppressor cells which can inhibit proliferative and cytotoxic responses of lymphocytes to alloantigens in vitro. The suppressor cells are adherent, phagocytic, resistant to treatment with ATS and C, radioresistant, resistant to treatment with mitomycin C, apparently absent from the thymus, and found in very high concentrations in peritoneal exudates. These characteristics indicate that the suppressor cell is a macrophages and not a T cell. When suppressor cells were removed from spleen cell suspensions, strong in vitro proliferative and cytotoxic responses to alloantigens could consistently be observed.

The effect of immunopharmacological agents on mouse natural cell-mediated cytotoxicity and on its augmentation by poly I:C

Treatment of young mice with lethal X-irradiation (900 R), cyclophosphamide, or hydrocortisone significantly depressed their spontaneous NK activity. The same treatments, however, did not inhibit the augmentation of NK function by poly I:C, suggesting the existence of a treatment resistant pre-NK cell. Both the spontaneous activity and its augmentation were readily inhibited by macrophage-toxic agents such as silica and carrageenan in vivo. Since the NK cells themselves were not macrophages, as shown by the inability of silica or carrageenan to block in vitro cytolysis of target cells, we postulated that macrophages were required to maintain NK activity in vivo and that they were essential accessory cells in the augmentation. The induction of interferon by poly I:C was also inhibited by silica and carrageenan. The augmentation of NK activity induced by poly I:C was consistently accompanied by the rise in serum IF levels of the treated mice, and its inhibition by the macrophage-toxic agents was followed by decreased production of interferon. These observations support our hypothesis that macrophages, in response to poly I:C, produced interferon which in turn activated NK cells to become cytolytic.

INTERFERON AND NK CELLS

Abstract: Interferon (IFN) has been shown to play a major role in the positive regulation of the activity of natural killer (NK) cells. All human IFN species tested to date of each of the three types, including homogeneous material and recombinant species, have had the ability to augment human NK activity. The mechanism for augmentation of this effector cell activity has been shown to be complex, with the potential for increasing the numbers of cells recognizing NK-susceptible targets, activating the lytic activity of target-binding cells, accelerating the rate of cytolysis of targets, and increasing the numbers of target cells killed by each effector cell. In addition, IFN has been shown to influence the growth and differentiation of NK cells from their progenitors: on the one hand, increasing the responsiveness of NK cells to interleukin-2 and thereby increasing cloning efficiency, and on the other hand, inhibiting generation of NK cells via a T cell-dependent mechanism. Some preliminary evidence has been obtained for the involvement of the 2′-5′A pathway in the mechanism of the augmentation of NK activity by IFN, since triphosphorylated 2′-5′A, after exposure of the cells to calcium chloride, caused an increase in NK activity. Nk cells appear to not only respond to IFN but also to produce IFN, in response to a wide range of stimuli. Overnight incubation of a highly purified population of human NK cells with viruses (herpes simplex or influenza), mycoplasma-contaminated tumor cell lines, BCG or C. parvum produced IFN-2, whereas under the same conditions, various mitogens (PHA, Con A, Staph enterotoxin) induced mainly IFN-α, and some IFN-β. Thus NK cells appear to be a major component of both the IFN-producing system and of the host effector mechanisms modulated by IFN. These observations have important implications in regard to clinical therapeutic trials with IFN or IFN inducers.

Participation of three lymphoid cell types in the in vitro activation of cell-mediated immunity to a syngeneic gross virus-induced lymphoma in rats

Abstract W/Fu rats inoculated with the syngeneic Gross-virus induced lymphoma, (C58NT)D, had transient lymphocyte-mediated specific cytotoxicity against the tumor cells at 7–15 days after tumor injection. Spleen cells 40 days after immunization (spleen 40) were unreactive by a direct 4-hr 51 Cr release assay, but activity appeared after in vitro culture of these spleen eclls by themselves for 18–24 hr. The nature of the cells involved in the activation of cytotoxicity and the characteristics of the effector cells themselves were studied. Significant differences were seen in the cell types involved in the activation phase and the effector phase. Activation appeared to require the cooperation of three cell types. Induction of activity was lost by treatment of cells with ATS plus complement, by passage over an EAC-column, or by treatment with carbonyl iron. Thus, T cells, CRL and macrophages were necessary for full activation of cytotoxicity in spleen 40. In contrast, after activation, only CRL seemed to be required for cytotoxicity, and treatment wih ATS or carbonyl iron had little effect. The effector cell detected after in vitro activation was quite distinct from that seen in the direct cytotoxicity assay with spleen cells at 10 days after tumor cell inoculation. The early, direct cytotoxic reactivity was dependent on T cells, being eliminated by treatment with ATS and complement but not by EAC columns or carbonyl iron. It appears therefore that the in vitro activation is a separate mechanism for cytotoxicity against tumor cells, rather than a simple recovery of T cells from in vivo inhibition.

MODELS FOR MECHANISMS OF NATURAL CYTOTOXICITY

Publisher Summary The concept of natural cell-mediated cytotoxicity in humans derived from observations that peripheral blood mononuclear cells of normal healthy donors, and cancer patients as well, mediated direct cytotoxic reactions against a variety of neoplastic and normal cell lines. The presence of these natural cytotoxic cells has complicated the study of specific, disease-related direct cytotoxic reactions. On the other hand, these natural killer (NK) cells are themselves of considerable interest and they have been extensively studied in both murine and human experimental systems. Until recently, NK activity was thought to be mediated by lymphocytes devoid of detectable cell surface markers, which, therefore, appeared distinct from cytotoxic T-lymphocytes and from antibody-dependent cell-mediated cytotoxicity (ADCC). More recently, studies of human and mouse NK cells (have revealed that NK cells do bear an Fc receptor (FcR) and are in fact not separable from K lymphocytes; thus, they may well represent the very same cells. Both types of activities can be adsorbed via FcR to immobilized antigen–antibody complexes, and this is reversed by the protein A of Staphylococcus aureus. This chapter discusses three models for mechanisms of natural cytotoxicity: (1) natural antibodies ARM FcR2 on FcRl-FcR2-positive cells; (2) NK due to a specific membrane receptor on FcR-positive cells; and (3) IgG bound to FcR turns on or augments mechanisms of NK reactivity.