Virus-mediated induction in human lymphocytes of antibody-independent cytotoxicity (VDCC) and enhancement of antibody-dependent cytotoxicity (ADCC) against natural killer-resistant tumor target cells

The effect of Parotis virus on the in vitro cytotoxicity of human lymphocytes against NK-resistant mouse mastocytoma cells was studied. In the ⁵¹Cr-release assay, treatment of lymphocytes with virus induced a rapid cytotoxicity in the absence of anti-P8 15 antibody (virus-dependent cellular Cytotoxicity, VDCC) and strongly enhanced antibody-dependent cytotoxicity (ADCC). At the effector cell level, virus treatment was found to increase the frequency of target-binding cells (TBC) as well as the proportion thereof mediating VDCC and/ or ADCC, indicating recruitment of active effector cells. The recruited cells were heterogeneous but contained a major fraction bearing the T-cell-associated antigen T3. Virus was found to decrease rather than to increase the recycling capacity of the cytotoxic lymphocytes, suggesting that VDCC induction and ADCC enhancement were due to a virus-mediated improvement of effector cell-target cell interactions. VDCC and ADCC enhancement may be of protective importance in early phases of virus infection as well as for the production of nonspecific tissue injuries associated with viral disease.     

Antibody-dependent cell-mediated cytotoxicity to target cells infected with type 1 and type 2 herpes simplex virus.

The phenomenon of antibody-dependent cell-mediated cytoxicity (ADCC) has been extended to include target cells acutely infected with herpes simplex type 1 virus (HSV-1) or herpes simplex type 2 virus (HSV-2) in an in vitro system that employs immune human serum and human blood mononuclear cells. The cytotoxic reaction was detectable after 1 hr of incubation and was complete between 4 and 8 hr. The amount of ADCC noted was directly proportional to the logarithm(10) of the effector: target cell ratio (E:T), and ADCC was noted at E:T as low as 1:1. The mononuclear effector cell was present in the blood of both HSV immune and non-immune individuals. The immune serum factor was demonstrated to be an antibody with specificity for HSV membrane antigen(s) and was reactive with target cells infected with either of the two HSV types. The antibody rendered the mononuclear cell cytotoxic by sensitization of the target cell rather than by direct attachment to or "arming" of the mononuclear cell. The physiochemical properties of the antibody as well as its presence in cord blood demonstrated that it is an immunoglobulin on the IgG class.     

Lymphocyte-mediated cytotoxicity to autologous cells infected with measles virus. II. Specificity of the cytotoxic reaction and characterization of the effector cells involved.

The mechanism of lymphocyte-mediated cytotoxicity to cells infected with measles virus was investigated. Cytotoxicity was measured in a direct assay, immediately after the isolation of lymphocytes from human peripheral blood; mononuclear leukocytes, infected with measles virus in vitro, served as autologous target cells. Virus-specific cytotoxicity required the presence of both IgG antibodies against measles virus and of effector lymphocytes. The effector lymphocytes had Fc receptors and were mainly present in a fraction of non-T lymphocytes. Monocytes were not cytotoxic but rather inhibitory. These results indicate that lysis of virus-infected cells in this direct assay is due to antibody-dependent cellular cytotoxicity (ADCC), caused by K cells. Control experiments showed that the virus-infected target cells were sensitive to incubation with human serum or IgG, resulting in a nonspecific increase of ⁵¹Cr release; however, this did not affect the results of K-cell cytotoxicity. Maximal virus-specific lysis by ADCC did not reach the level obtained by complement-dependent cytotoxicity. Possible explanations for this difference are discussed.     

Human monocyte-macrophage-mediated antibody-dependent cytotoxicity to herpes simplex virus-infected cells.

Human peripheral blood mononuclear cells which mediate antibody-dependent cellular cytotoxicity (ADCC) against herpes simplex virus- (HSV) infected target cells consist of both adherent (MA) and nonadherent (MNA) effector cell populations. These two cell populations can be distinguished by their different phagocytic properties and morphologic appearance, their requirement for antibody in the ADCC reaction, and the rapidity with which they lyse target cells in the presence of immune serum. The MA cells are predominantly phagocytic and have the morphologic characteristics of monocyte-macrophages, whereas the MNA cells are nonphagocytic and appear to be small to medium-sized lymphocytes. Optimal expression of ADCC by MA cells requires higher concentrations of immune serum than does MNA cell-mediated ADCC. MA-mediated cell killing is first detectable by 8 hr and reaches completion after 24 hr of incubation. In contrast, MNA-mediated ADCC produces target cell damage by 2 hr and reaches completion at 8 hr of incubation. Unlike MNA effector cells, the MA effector cells are profoundly inhibited after preincubation with either latex or silica particles. The HSV immune status of the donor had no effect on the ability of either cell population to mediate ADCC. These data demonstrate the participation of both nonadherent mononuclear cells, presumably K cells, and monocyte-macrophages, in ADCC directed against HSV-infected target cells.     

Differential effects of ouabain on human cell-mediated cytotoxicity. I. Inhibition of mitogen-induced cellular cytotoxicity and antibody-dependent cellular cytotoxicity against chicken red cell targets.

The effects of ouabain, a known inhibitor of lymphoproliferation, were studied in relation to the cytotoxic effector function of human peripheral blood mononuclear leukocytes (MNL) against chicken red blood cell (CRC) targets. MNL effectors lysed ⁵¹Cr-labeled CRC targets in the presence of PHA (mitogen-induced cellular cytotoxicity—MICC) or rabbit anti-CRC antibody (antibody-dependent cellular cytotoxicity—ADCC) in the absence of ouabain. The addition of ouabain to the cytotoxic reaction caused profound diminution of MICC with greater than 90% suppression of killing at ouabain concentrations of 5 × 10^?4M; ADCC was much more resistant to the effects of ouabain with only 60 to 70% inhibition of killing at similar ouabain concentrations (P < 0.01). Similar ouabain inhibition of MICC occurred whether the effector cell populations were unseparated MNL, depleted of monocytes, enriched for T cells, or depleted of T cells, suggesting a generalized activity by ouabain against all effector cells active in MICC. Ouabain inhibition of MICC could be overcome by increasing PHA concentrations, indicating that ouabain inhibition was not due to irreversible toxic effects on effector cells. Increasing the concentration of anti-CRC antibody resulted in increased killing in this ADCC system and, paradoxically, ADCC cultures with the highest antibody concentrations were more completely inhibited by ouabain. This enhanced inhibitory effect of ouabain on ADCC cultures with the highest antibody concentrations was not observed when the effector cell population was first depleted of phagocytic cells, suggesting a preferential inhibitory action by ouabain against monocyte effectors in ADCC. Thus, the differential inhibitory effects of ouabain on MICC and ADCC against CRC targets may be in part explained by the differing ouabain sensitivities of the various effector cell subpopulations involved in these cell-mediated cytotoxic events.     

Splenic leukocytes from chickens injected with Salmonella pullorum antigen stimulate production of corticosteroids by isolated adrenal cells

Corticosteroid levels in the blood serum of White Rock pullets were significantly increased within 1 hr after an iv injection of heat-killed Salmonella pullorum antigen (SP-Ag), and reached levels 7 to 10 times that of PBS-injected controls within 2 hr after SP-Ag injection. Incubations with isolated adrenal cells indicated that serum from SP-Ag-injected birds had the ability to stimulate the synthesis or release of corticosteroids twice that of serum PBS-injected birds. Stripping the serum from SP-Ag-injected birds with activated charcoal and precipitated silica (Quso G-32) removed the corticosteroids and the adrenal-stimulating ability. A 2-hr incubation of isolated adrenal cells with leukocytes from spleens removed from chickens 1 hr after injection with SP-Ag, using stripped serum as the medium, stimulated a two- to fivefold increase in corticosteroid as compared to splenic leukocytes from PBS-injected chickens incubated in the same medium. The results indicate that an "ACTH-like" substance was produced by the S. pullorum antigen-stimulated splenic leukocytes.     

Virus-induced enhancement of lymphocyte-mediated antibody-dependent cytotoxicity (ADCC) in vitro

The effect of Parotis virus on antibody-dependent cellular cytotoxicity in vitro (ADCC) of human lymphocytes was investigated in a 51Cr-release assay and, at the effector cell level, in an ADCC plaque assay. Target cells were bovine or chicken erythrocytes, which are not susceptible to natural cytotoxicity (NK) of human lymphocytes. They were not killed when incubated with virus-treated lymphocytes in the absence of antibodies. Treatment of the lymphocytes or the target cells with small amounts of virus, however, resulted in a very significant enhancement of ADCC. The same results were obtained with live or UV-inactivated virus, suggesting that enhancement was a passive phenomenon not requiring infection. Enhancement was already significant after 3 hr of incubation, indicating that it was independent of endogenously released interferon. Enhancement of ADCC by virus was due to effector cell recruitment rather than due to the increase of the cytotoxic potential of the individual K cell. The highest frequency of effector cells was present in Percoll fractions enriched in large granular lymphocytes (LGL). Virus treatment resulted in recruitment of effector cells carrying T cell markers such as the T3 antigen (OKT3+), receptors for sheep erythrocytes, or Fc receptors for IgM. In contrast, the absolute number of K cells carrying the HNK-1 marker (Leu-7) or receptors for C3 fragments was not changed by the virus. It is concluded that Parotis virus enhances ADCC by improving effector cell-target cell contacts, resulting in recruitment of effector cells with T cell characteristics. Recruitment is accompanied by a significant reduction of the antibody concentration needed for ADCC induction. This virus-mediated enhancement of ADCC may be of importance for protection of the host in the early phases of a virus infection in which the amounts of anti-viral IgG antibodies capable of inducing cellular cytotoxicity may yet be very small.     

Effects of chicken intestinal antimicrobial peptides on humoral immunity of chickens and antibody titres after vaccination with infectious bursal disease virus vaccine in chicken

Sixty chickens were randomly divided into two groups (30 chickens in each group) to determine the effect of oral administration of chicken intestinal antimicrobial peptides (CIAMP) on the humoral immune response. Chickens of both groups were fed the same diet. In the treatment group chickens received drinking water supplemented with CIAMP (1 microg/ml) right after hatching. Samples of blood, bursa of Fabricus, spleen and intestine were taken at day 1, 4, 7, 10 and 17 of experiment. CIAMP supplementation enhanced the content of IgG and IgM in serum from day 4-10 and day 10-17, respectively, (p < 0.05), IgM-forming cells in bursa of Fabricus and spleen at the age of 7 days (p < 0.05) and IgG-forming cells in bursa of Fabricus at the age of 4 days (p < 0.05). In addition, CIAMP enhanced the IgA-forming cells in caecal tonsils diffuse area at day 4 (p < 0.05). Furthermore, CIAMP enhanced the antibody response to infectious bursal disease virus vaccine (IBDV) in chickens 21 days following IBDV vaccine administration (p < 0.05). These results suggested that CIAMP could modulate the humoral immune response of chickens and increased the antibody titres of infectious bursal disease virus vaccine.     

Polyadnylic acid sequences in RNA able to transfer specific delayed type hypersensitivity in vitro.

Antibody-depedent cell-mediated cytotoxicity (ADCC) could be initiated without protein synthesis [human peripheral blood lymphocytes as effector cells incubated with 10^?3M cycloheximide, (Cy)], although the reaction did not achieve its full lytic ability. This partial inhibition of ADCC was dependent on the dose of Cy. Both ADCC and protein synthesis returned to normal values after removal of the inhibitor. The kinetics of the reaction carried out by Cy-treated effector cells for short periods was similar to that of controls. After this time, the percentage of lysed target cells increased continuously in controls, while the cytotoxiciy of Cy-treated effector cells reached a plateau. When effector cells carried out ADCC in the presence of Cy, their lytic mechanism was “wasted,” and it could be recovered only by removal of the inhibitor. Our results indicate that effector cells have a preformed lytic mechanism operative in ADCC. This lytic mechanism is consumed during the reaction and its recovery requires protein synthesis.     

Gamma delta T cells from HIV+ donors can be expanded in vitro by zoledronate/interleukin-2 to become cytotoxic effectors for antibody-dependent cellular cytotoxicity

Background aimsImmunotherapy using γδ T cells capable of mediating antibody-dependent cellular cytotoxicity (ADCC) is a promising anti-human immunodeficiency virus (HIV) strategy. Approved aminobispohsphonate drugs, for example zoledronate (Zometa), stimulate γδ T cells in cancer patients, where they may promote direct tumor killing. Knowing that γδ T cells are modulated during HIV disease, documenting their responses and potential for controlling HIV is important. We investigated whether zoledronate/interleukin (IL)-2 could expand cytotoxic Vδ2 cells from HIV⁺ donors and whether these cells functioned in ADCC.MethodsPeripheral blood mononuclear cells from uninfected controls and HIV⁺ individuals receiving anti-retroviral therapy were treated with isopentenyl pyrophosphate (IPP) or zoledronate plus IL-2 to expand the Vδ2⁺ subset. Immunophenotyping and functional analyzes (cytotoxicity or cytokine expression) allowed us to compare cell properties from individual donors and to compare the responses to each stimulating agent.ResultsZoledronate stimulated a greater expansion of Vδ2 cells in HIV⁺ individuals compared with phosphoantigen IPP, and these cells expressed CD16. CD56 expression (a marker for cytotoxic cells) was lower on zoledronate-expanded cells, consistent with significantly lower cytotoxicity against the Daudi tumor cell line. Cells expanded with either zoledronate or IPP were active in ADCC, were similar in terms of interferon (IFN)-γ and tumor necrosis factor (TNF)-α expression, and degranulated in response to Fc receptor cross-linking.ConclusionsZoledronate causes ex vivo expansion of Vδ2 cells from HIV⁺ individuals. Despite lower expression of CD56 and decreased direct cytotoxicity, these effectors were potent in ADCC. Zoledronate/IL-2- expanded cells have potential for immunotherapy to activate Vδ2 cells in HIV patients and enhance ADCC.     

Spontaneous rosette-forming lymphocytes in sheep lymph. A marker for antigen-binding cells

Type O Rh positive human red blood cells (HRBC), native or treated with one of three enzymes (papain, trypsin, or neuraminidase), were labeled with ⁵¹Cr and then sensitized with anti-Rh immune globulin. These cells served as targets in antibody-dependent cellular cytotoxicity (ADCC) for unfractionated human mononuclear cells (MC), MC depleted of monocytes by adhesion to plastic, and MC enriched for monocytes. Enzyme-treated HRBC were lysed with greater efficiency in ADCC than native HRBC. This was explained by the finding that the enzyme modified HRBC were lysed both by lymphocytes and monocytes, whereas native HRBC were lysed only by monocytes. The lysis of native HRBC was strongly inhibited by small amounts of human serum or free IgG. In contrast, the lysis of enzyme-treated HRBC was considerably more resistant to inhibition by human serum or free IgG. The enhanced lysis of enzyme-treated HRBC could not be the result of increased binding of antibody to the target cells, since augmented lysis was observed both for HRBC sensitized before neuraminidase treatment as well as for HRBC sensitized after neuraminidase treatment. These results suggest that the surface charge on target cells plays a critical role in determining which classes of leukocytic effector cells are active in ADCC systems.     

Virus DNA replication and protein synthesis in Amsacta moorei entomopoxvirus-infected Estigmene acrea cells

Amsacta moorei entomopoxvirus DNA synthesis was detected in Estigmene acrea cells by [³H]thymidine incorporation 12 hr after virus inoculation. Hybridization of ³²P-labeled Amsacta entomopoxvirus DNA to the DNA from virus-infected cells indicated that viral-specific DNA synthesis was initiated between 6 and 12 hr after virus inoculation. A rapid increase in the rate of virus DNA synthesis was detected from 12 to 24 hr after virus inoculation. Amsacta entomopoxvirus protein biosynthesis in E. acrea cells was studied by [su35S]methionine incorporation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Extracellular virus and virus-containing occlusion bodies were first detected in virus-infected cell cultures 18 hr after virus inoculation. Thirty-seven virus structural proteins, ranging in molecular weight from 13,000 to 208,000 were detected in both occluded and nonoccluded forms of the virus. The biosynthesis of virus structural proteins increased rapidly from 18 to 34 hr after infection. A major viral-induced protein corresponding in molecular weight to viral occlusion body protein (110,000) was detected approximately 24 hr after virus inoculation.     

Enhancement of the antibody-dependent cellular cytotoxicity of human peripheral blood lymphocytes with interleukin-2 and interferon α

Antibody-dependent cellular cytotoxicity (ADCC) is regarded as an important mechanism by which monoclonal antibodies (mAb) can exert an antitumour effect in vivo. It may be possible, therefore, to enhance the therapeutic efficacy of mAb by cytokines that are able to enhance the ADCC of human CD3^–, CD56⁺, CD16⁺ natural killer (NK) cells. We investigated in vitro the effects of recombinant interferon (rIFN) and recombinant interleukin 2 (rIL-2), alone or in combination, on the ADCC of human peripheral blood NK cells. Both cytokines enhanced the ADCC of the human effector cells. rIFN induced a maximally increased ADCC after an exposure of human effector cells to 20 IU/ml for 15–30 min, while rIL-2 induced optimal ADCC after incubation of the cells for 2 days in 20–50 U/ml. We now show that activation of the NK cells with a combination of rIL-2and rIFN induced significantly higher levels of ADCC than either cytokine alone. The highest ADCC was induced if the cells were first exposed to rIL-2 before rIFN was added to the culture. Culture of NK cells in medium or rIL-2 decreased the expression of FcRIII (CD16), indicating that intensity of CD16 expression and level of ADCC are not directly correlated, although blocking experiments with a mAb directed against CD16 showed that this FcR was essential for ADCC of the human effector cells.Supported by a grant from the Dutch Cancer Society (grant NKI-84-14)     

Detection of Amsacta moorei entomopoxvirus and vaccinia virus proteins in cell cultures restrictive for poxvirus multiplication

The titer of Amsacta entomopoxvirus (EPV) protein detected in murine L-929 cells by enzyme-linked immunosorbent assay (ELISA) decreased to within preimmune serum levels by 24 hr after inoculation of the virus which indicates that Amsacta EPV structural protein biosynthesis does not occur in the vertebrate cell line. A viral-induced protein of approximately 100,000 M_r was detected by [³⁵S]methionine incorporation 4 hr after inoculation of Tn-368 cells with Amsacta EPV. Biosynthesis of protein which reacted with vaccina antiserum was detected in Estigmene acrea (BTI-EAA) cells by ELISA 10 hr after inoculation with 10 PFU of virus per cell. The amount of putative vaccinia structural protein detected in BTI-EAA cells increased approximately twofold by 70 hr after virus inoculation. No increase in vaccinia structural protein biosynthesis was detected in BTI-EAA cells inoculated with vaccinia virus previously inactivated by heat and UV light.     

Metabolie Aspects in Mice Administered Live Salmonella typhimurium

The influence of intraperitoneal inoculation of live Salmonella typhimurium on carbohydrate and lipid metabolisms in mice was investigated at doses of 9.2 × 10⁷ cells, 1.9 × 10⁸ cells, and 3.8 × 10⁸ cells. The hepatic glycogen content in mice at 18 hr after the inoculation decreased in inverse proportion to an increase in the injection dose. The activities of hepatic phosphorylase and G-6-P_ase increased significantly after 2 hr, but after 18 hr the levels of both enzyme activities, especially G-6-P_ase, declined in inverse porportion to an increase in dose of viable cells administered to the mice. The levels of serum glucose and free fatty acids (FFA) in mice markedly decreased at doses of 1.9 × 10⁸ and 3.8 × 10⁸ cells after a transient rise at early stage (1 hr) after the injection. Marked hypertriglyceridemia was seen in infected mice. However, the activity in serum lipoprotein lipase (LPL) was reduced by an increase in the injection dose. The effect of intraperitoneal administration of viable cells on the serum triglyceride level was prevented in mice immunized with S. typhimurium endotoxin or administered with the anti-endotoxin serum. These results indicate that hypertriglyceridemia mainly results from the action of endotoxin in the pathogen. Serum lactate dehydrogenase (LDH) activity markedly increased at the dose of 3.8 × 10⁸ cells within 8–16 hr, and the infected mice exhibited a leakage of isozymes LDH-3 and 5 in the serum 16 hr post-inoculation.     

KINETICS OF ERYTHROPOIETIN STIMULATED PROLIFERATION OF ERYTHROID PROGENITORS IN THE SPLEEN

The effect of RBC transfusion and erythropoietin (EPO) on the proliferation of immature erythrocyte progenitors was studied in the spleens of RBC transfused, lethally irradiated mice injected with bone marrow. Transfusion decreased expansion of the progenitors and slowed their proliferation: the mean cycle time as measured by per cent labelled mitosis (PLM) on the third day after injection of bone marrow was 10.7 hr in transfused as compared to 5.6 hr in non-transfused mice. One injection of five units of erythropoietin on day 2 decreased the mean cycle time to 7.3 hr in transfused mice and increased expansion of the progenitor cells. The effects of erythropoietin on cell proliferation were prompt: a significant increase of incorporation of ³H-TdR into DNA occurred within 2 hr of injection. Erythroblasts were absent from the spleens of transfused, irradiated bone marrow injected mice; however, erythroblasts appeared by 72 hr and 48 hr following EPO injection either 2 days or 5 days after transplantation respectively. Increased uptake of radioactive iron in spleen after erythropoietin injection preceded the appearance of erythroblasts by 2 and 1 days when erythropoietin was injected either 2 or 5 days after marrow transplantation respectively. The increase in cellular proliferation induced by erythropoietin in transfused irradiated mice injected with bone marrow equivalent to 0.35 femoral shaft was manifested as an increase of the total DNA content in the spleen by 119 μg (11.9 × 10⁶ cells) within 48 hr of injection. The cellular increment produced by EPO injection on day 5 to mice given 0.05 femoral shaft consisted mainly of undifferentiated mononuclear cells, most of which were labelled, with erythroblasts comprising only one quarter of the increment. Erythropoietin inactivated by mild acid hydrolysis failed to increase cellular proliferation.     

Glycovariant anti-CD37 monospecific protein therapeutic exhibits enhanced effector cell-mediated cytotoxicity against chronic and acute B cell malignancies

TRU-016 is a SMIP^TM (monospecific protein therapeutic) molecule against the tetraspanin transmembrane family protein CD37 that is currently in Phase 2 trials in Chronic Lymphocytic Leukemia (CLL) and Non-Hodgkin Lymphoma (NHL). In an attempt to enhance the ADCC function of SMIP-016, the chimeric version of TRU-016, SMIP-016^GV was engineered with a modification in a glycosylation site in the Fc domain. The wild-type and glycovariant SMIP proteins mediate comparable Type I antibody-like direct cytotoxicity in the presence of anti-human Fc crosslinker and show a similar tyrosine phosphorylation pattern post-treatment. However, NK cells stimulated with the SMIP-016^GV exhibit enhanced activation and release 3-fold more interferon-γ compared with SMIP-016. SMIP-016^GV shows enhanced ADCC function against cells expressing CD37 with NK cell effectors derived from both normal and CLL-affected individuals. Enhanced ADCC is observed against CLL cells and is sustained at concentrations of SMIP-016^GV as low at 5E⁻⁶ µg/mL on cells expressing minimal CD37 antigen. In support of the biological relevance of this, SMIP-016^GV mediates effective ADCC against primary acute lymphoblastic leukemia (ALL) cells with low surface expression of CD37. Collectively, these data suggest potential use of the novel therapeutic agent SMIP-016^GV with enhanced effector function for B cell malignancies, including CLL and ALL therapy.     

Effect of histamine and histamine antagonists on natural and antibody-dependent cellular cytotoxicity of human lymphocytes in vitro

The in vitro effect of histamine and its antagonists, cimetidine and clemastine fumarate, on natural killer (NK) and antibody-dependent cellular Cytotoxicity (ADCC) activities of human lymphocytes was investigated. The histamine 1 (H1) antagonist, clemastine fumarate, and the histamine 2 (H2) antagonist, cimetidine, but not histamine alone, inhibited the NK and ADCC activities of lymphocytes when added directly to the mixture of effector and target cells in a ⁵¹Cr-release assay. This inhibition was proportional to the concentration of drugs added and was observed at various effector to target ratios against several targets. H1 and H2 antagonists also inhibited NK activities of T cells as well as Percoll-separated, NK-enriched effector cells. The inhibition was significantly reversed by histamine. In target binding assays, clemastine fumarate and cimetidine also decreased the target binding capacity of effector lymphocytes. Further, PBL precultured with histamine (10^?3–10^?4M) for 24 hr showed a significant decrease in their NK and ADCC activities. In coculture experiments, PBL precultured with histamine suppressed the NK activity of normal autologous effector lymphocytes. PBL precultured with histamine showed an increased number of OKT8⁺ cells, as estimated using monoclonal antibodies. The suppression of Cytotoxicity was not due to either direct toxicity, steric hindrance, crowding, or cell death, but by functionally viable suppressor cells. An immunoregulatory role for histamine in NK and ADCC reactions is proposed.     

Cells of the J774 macrophage cell line are primed for antibody-dependent cell-mediated cytotoxicity following exposure to gamma-irradiation.

Activation of macrophages (M phi) for host defense against tumor cells follows a sequence of priming events followed by an initiating stimulus that results in production and release of cytotoxic molecules that mediate target cell killing. We have developed a model to study specific macrophage cytotoxicity in vitro utilizing a cultured murine M phi cell line, J774. Specific cytotoxicity of cultured human gastrointestinal tumor cells is achieved in the presence of murine IgG2a monoclonal antibody (mAb) 17-1-A. The ability of these cells to mediate antibody-dependent cell-mediated cytotoxicity (ADCC) is greatly enhanced following gamma-irradiation. ADCC can be demonstrated at mAb 17-1-A concentrations greater than or equal to 1 microgram/ml and effector/target cell ratios greater than or equal to 2. Exposure to doses greater than or equal to 10 Gy of gamma-irradiation increases ADCC threefold. Varying the duration from J774 M phi exposure to gamma-irradiation until addition of antibody-coated target cells showed that the primed state for ADCC is stable for at least 8 days but approximately 24 hr is required for complete development of the primed state. mAb-dependent target cell death begins 8 hr after addition of mAb and labeled target cells to primed effector cells and is complete by 24 hr. Incubation of unirradiated J774 M phi effector cells with recombinant murine interferon-gamma (rmIFN-gamma) also results in enhanced ADCC, but the extent of target cell killing achieved is less than that following priming by gamma-irradiation. Concomitant priming of gamma-irradiated J774 M phi with rmIFN-gamma increases the extent of ADCC. Further study of irradiated J774 cells may elucidate the molecular pathways utilized by M phi for achieving and maintaining the primed state for ADCC. Irradiated J774 cells will also provide a homogenous, stably primed cell type in which to examine the mechanism(s) of cytotoxicity employed by tumoricidal M phi.     

Suppression of the mitogen-stimulated blastogenic response during reticuloendotheliosis virus-induced tumorigenesis: investigations into the mechanism of action of the suppressor.

Splenic lymphocytes from chickens infected with reticuloendotheliosis virus (REV) are suppressed in their ability to undergo PHA-induced blastogenesis. This suppression can be detected within 72 hr after virus injection and requires active viral infection since i.p. or i.v. injection of UV-inactivated REV does not result in inhibition of the blastogenic response. Suppressor cells from the spleens of REV-infected birds severely inhibit the ability of spleen cells from uninfected chickens to respond to PHA at a ratio of 1:20, suggesting that each suppressor cell may be capable of suppressing more than one target cell. Contact between suppressor and target cells is required for the rapid inhibition of the normal PHA response. The suppressive mechanism is cytostatic in nature, and apparently of host origin since neither REV, nor REV-infected (or transformed) cells mediate the suppression. The ability of the suppressor cells to impair the blastogenic response of spleen cells is sensitive to trypsin, suggesting that an inhibitory protein is exposed on the surface of the suppressor cells.