Lymphoid cells from many normal mice of a variety of inbred strains were found to have reactivity, in a 51Cr release cytotoxicity assay, against several syngeneic and allogeneic tumors. Very high reactivity was seen with effector cells from athymic nude mice, which was consistent with other evidence that the reactivity was not T-cell dependent. Target cells susceptible to lysis included tumors induced by oncogenic type-C viruses but also tumors induced by other means and expressing endogenous type-C viruses. The levels of natural reactivity were influenced by age, with highest cytotoxicity produced by cells from 5- to 8-week-old mice. Lymph-node cells, spleen cells, peritoneal exudate cells and peripheral blood lymphocytes all had cytotoxic reactivity. The specificity of the reactions was analyzed in detail by ana inhibition assay. Evidence was obtained for natural reactivty against several different antigens, each apparently associated with expression of murine endogenous type-C viruses.

No abstract

Studies were performed to characterize the effector cells responsible for natural cytotoxicity of mouse lymphoid cells against a variety of syngeneic and allogeneic tumor lines. Since spleen cells from normal nude mice were found to be highly cytotoxic, they were used for most of these experiments. Only a small proportion of the reactivity was affected by treatment with anti-theta serum plus complement. Macrophages dis not appear to be responsible for the reactivity, since treatment with carbonyl iron/magnet or carrageenan did not affect the levels of cytotoxicity. The effector cells were non-adherent, since passage over nylon columns resulted in a considerable increase in activity. The active cells did not have receptors for immunoglobulin or complement, since removal of cells with these receptors by columns or monolayers containing sheep erythrocyte-antibody (EA) complexes or EA-complement complexes did not remove activity. Antibody-dependent cell-mediated cytotoxicity appeared to be ruled out as the mechanism for natural cytotoxicity, since aggregated gamma globulin and a potent anti-immunoglobulin reagent did not inhibit reactivity, and since no role for humoral factors could be demonstrated. The natural effector cell was found to be quite labile at 37 degrees C, losing much of its activity after 4 h. Since no surface markers could be detected on the effector cells, and the mechanism for cytotoxicity appeared distince from others previously described, it is proposed that the natural cytotoxicity against mouse tumor cells is mediated by a unique subpopulation of lymphoid cells, which are tentatively designated N-cells.

In addition to their cytotoxic activities, natural killer (NK) cells can have immunoregulatory functions. We describe a distinct “helper” differentiation pathway of human CD56+CD3− NK cells into CD56+/CD83+/CCR7+/CD25+ cells that display high migratory responsiveness to lymph node (LN)–associated chemokines, high ability to produce interferon-γ upon exposure to dendritic cell (DC)- or T helper (Th) cell–related signals, and pronounced abilities to promote interleukin (IL)-12p70 production in DCs and the development of Th1 responses. This helper pathway of NK cell differentiation, which is not associated with any enhancement of cytolytic activity, is induced by IL-18, but not other NK cell–activating factors. It is blocked by prostaglandin (PG)E2, a factor that induces a similar CD83+/CCR7+/CD25+ LN-homing phenotype in maturing DCs. The current data demonstrate independent regulation of the “helper” versus “effector” pathways of NK cell differentiation and novel mechanisms of immunoregulation by IL-18 and PGE2.

To study the cellular structures involved in NK and lymphokine-activated killer (LAK) cell function, we have produced a panel of mAbs that modulate the cytolytic function of a population of cells with LAK activity that derive from large granular lymphocyte (LGL)/NK cells (adherent LAK [A-LAK] cells). In this report, we describe an mAb (3.2.3; IgG1k) that recognizes a triggering structure that is expressed on rat LGL/NK cells and A-LAK cells. This epitope is also expressed on polymorphonuclear leukocytes (PMN). The expression of the epitope identified by mAb 3.2.3 increased progressively on A-LAK cells after culture in the presence of rIL-2. mAb 3.2.3 enhanced the cytolytic activity of NK and A-LAK cells against FcR+ target cells, but not FcR- target cells. However, this effect was not induced by F(ab')2 fragments of 3.2.3. This antibody also induced the release of N-alpha-benzyloxycarbonyl-L-lysine thiobenzy esteresterase by A-LAK cells. These data suggest that the epitope identified by mAb 3.2.3 is on a triggering structure expressed on rat NK cells and A-LAK cells. The expression of the epitope recognized by mAb 3.2.3 on LGL/NK cells and PMN suggests that this structure may be analogous to that identified by the anti-CD16 (-FcR) mAbs. However, the molecule immunoprecipitated by mAb 3.2.3 was a 60-kD dimer composed of two 30-kD chains. These data suggest that mAb 3.2.3 recognizes a unique triggering structure. As mAb 3.2.3 is the first antibody recognizing a determinant with functional significance, selectively expressed on both rat NK cells and A-LAK cells, it will be a useful tool for the study of NK cell ontogeny and function, and the development of cells with LAK activity from the NK cell compartment.

Early detection of ovarian cancer might improve clinical outcome. Some studies have shown the role of cytokines as a new group of tumor markers for ovarian cancer. We hypothesized that a panel comprised of multiple cytokines, which individually may not show strong correlation with the disease, might provide higher diagnostic power. To evaluate the diagnostic utility of cytokine panel, we used a novel multianalyte LabMAP profiling technology that allows simultaneous measurement of multiple markers. Concentrations of 24 cytokines (cytokines/chemokines, growth, and angiogenic factors) in combination with cancer antigen-125 (CA-125), were measured in sera of 44 patients with earlystage ovarian cancer, 45 healthy women, and 37 patients with benign pelvic tumors. Six markers, i.e., interleukin (IL)-6, IL-8, epidermal growth factor (EGF), vascular endothelial growth factor (VEGF), monocyte chemoattractant protein-1 (MCP-1), and CA-125, showed significant differences in serum concentrations between ovarian cancer and control groups. Out of this group, IL-6, IL-8, VEGF, EGF, and CA-125, were used in a classification tree analysis that resulted in 84% sensitivity at 95% specificity. The receiver operator characteristic curve created using the combination of markers produced sensitivities between 90% and 100% in the area of 80% to 90% specificity, whereas the receiver operator characteristic curve for CA-125 alone resulted in sensitivities of 70% to 80%. The classification tree analysis for discrimination of benign condition from ovarian cancer used CA-125, granulocyte colony-stimulating factor (G-CSF), IL-6, EGF, and VEGF resulting in 86.5% sensitivity and 93.0% specificity. The presented data show that simultaneous testing of a panel of serum cytokines and CA-125 using LabMAP technology may present a promising approach for ovarian cancer detection.

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The ability of BALB/c nude and C57BL/6 mice to eliminate tumor cells from the blood stream was severely impaired after a single inoculation of 0.2 ml of anti-asialo BMI (asGMI) serum, diluted 1:40 to 1:320. The number of i.v.-inoculated YAC-I cells surviving in the lungs of BALB/c nude mice pretreated with anti-asGMI serum was 28 times higher than in the control nude mice. In this respect, nude mice treated with anti-asGMI behaved similarly to beige mice. The increase in the initial survival of tumor cells in the mice that was induced by pre-treatment with anti-asGMI resulted in a substantial increase in the number of artificial lung metastases that developed. In C57BL/6 +/+ mice treated with anti-asGMI and in C57BL/6 beige mice, i.v. inoculation of B16 melanoma cells induced 10 times more metastatic foci in the lungs than in the control C57BL/6 +/+ mice. In contrast, in nude mice which possess higher levels of NK reactivity, metastatic growth was suppressed 7-fold in comparison with intact C57BL/6 +/+ mice. In beige mice and in C57BL/6 +/+ mice treated with anti-asGMI, multiple metastatic foci developed in the liver, whereas in control C57BL/6 +/+ and nude mice, no extrapulmonary metastases were found. These data indicate that B16 melanoma cells are able to grow in the liver, but their growth is ordinarily prevented by NK cells. The antimetastatic defense of C57BL/6 mice treated by anti-asGMI could be restored by transplantation of 40 X 10(6) normal spleen cells. This antimetastatic effect of transplanted spleen cells was mediated by asGMI-bearing cells, since after in vitro pre-treatment of normal spleen cells with anti-asGMI and complement, they lost their ability to inhibit the development of artificial metastases in the lungs of C57BL/6 mice. Suppression of NK reactivity by multiple injections of anti-asGMI (every 4 to 5 days), in C57BL/6 mice inoculated intrafootpad (i.f.p.) with B16 melanoma or 3LL tumor cells, did not influence the growth of local tumors, but dramatically accelerated the development of spontaneous pulmonary metastases. These data demonstrate that NK cells may play an important role in resistance to the dissemination of tumor cells, and therefore contribute to the control of metastasis formation in mice.