Regulation of tumor necrosis factor cytotoxicity by calcineurin

Cyclosporin (CsA) inhibits mitochondrial death signaling and opposes tumor necrosis factor (TNF)-induced apoptosis in vitro. However, CsA is also a potent inhibitor of calcineurin, a phosphatase that may participate in cell death. Therefore, we tested the hypothesis that calcineurin regulates TNF cytotoxicity in rat hepatoma cells (FTO2B). TNF-treated FTO2B cells appeared apoptotic by DNA fragmentation, nuclear condensation, annexin V binding, and caspase activation. We studied two calcineurin inhibitors, CsA and FK506, and found that each potently inhibited TNF cytotoxicity. Western blot demonstrated calcineurin in FTO2B homogenates. In a model of mitochondrial permeability transition (MPT), we found that CsA prevented MPT and cytochrome c release, while FK506 inhibited neither. In summary, we present evidence that calcineurin participates in an apoptotic death pathway activated by TNF. CsA may oppose programmed cell death by inhibiting calcineurin activity and/or inhibiting mitochondrial signaling.     

Induction by mycoplasmas of tumor necrosis factor-alpha from macrophages

Several species of mycoplasmas including M. pneumoniae, the causative agent of human respiratory infection, were investigated for tumor necrosis factor-alpha (TNF-alpha) induction. The cytotoxic activity to Meth A cells of peritoneal macrophages purified from BALB/c mice was enhanced markedly when cultured with either viable or nonviable mycoplasmas. The supernatant of macrophage culture mixed with mycoplasmas, M. pneumoniae or A. laidlawii, showed a potent cytotoxic activity to TNF-alpha-sensitive but not to TNA-alpha-insensitive L cells. Addition of anti-TNA-alpha antiserum inhibited completely the cytotoxic activity of the supernatant, indicating that the cytotoxic activity is due mostly to TNF-alpha. These results strongly suggest that mycoplasmas possess an activity to induce TNF-alpha, which enhances the cytotoxic activity of macrophages and prevent infection with mycoplasmas in vivo.     

Protection from tumor necrosis factor cytotoxicity by protease inhibitors

Tumor necrosis factor (TNF) is cytocidal for human and murine cells when protein synthesis is inhibited by cycloheximide, but some protease inhibitors completely protect these cells from TNF cytotoxicity. Inhibitors of chymotrypsin-like proteases are active at lower concentrations than inhibitors of trypsin-like proteases. Both irreversible inhibitors, such as alkylating compounds, and reversible inhibitors, such as substrates of proteases, protect cells from the cytocidal activity of TNF. This protection is most effective when the cells are pretreated with these inhibitors before addition of TNF. When the protease inhibitors are removed, the cells gradually lose resistance to TNF cytotoxicity. The inhibitors do not interfere with the functioning of TNF-receptor complexes, since SK-MEL-109 melanoma cells treated with a protease inhibitor synthesize a TNF-induced protein. These findings suggest that a protease in involved in the cytocidal action of TNF.     

Induction by Staphylococcus aureus L-form of tumor necrosis factor-alpha from macrophages

Induction of tumor necrosis factor-alpha (TNF-alpha) by Staphylococcus aureus L-form was investigated. The supernatant of a macrophage culture mixed with S. aureus L-form showed a potent cytotoxic activity to L cells. Addition of anti TNF-alpha antibody inhibited completely the cytotoxic activity of the supernatant, indicating that the activity might be due mostly to TNF-alpha. To investigate localization of TNF-alpha production, the membranes of hypotonicity treated L-form were layered on a step-gradient composed of an upper and lower layers of 35% and 50% sucrose, respectively. The membranes were banded at the interface of 35% and 50% of sucrose. The activity of TNF-alpha production of the membrane fraction was 10-times higher than that of the soluble fraction.     

Reduction in tumor necrosis factor binding and cytotoxicity by hydrogen peroxide

The regulatory effect of H2O2 on both the cytotoxic activity and the specific binding of TNF-alpha was studied by using TNF-alpha-sensitized murine L929 cells. When these cells were exposed simultaneously to TNF-alpha and H2O2 (100 to 500 microM), the cytotoxic activity of TNF-alpha was inhibited by up to 66.6%. This inhibition was also effective when the cells were pretreated by H2O2, but not when TNF-alpha alone was preexposed to H2O2. These data suggest that H2O2 altered the cell sensitivity to TNF-alpha, without modifying the activity of the TNF-alpha molecule. Maximum loss of cell sensitivity to TNF-alpha occurred after 30-min preexposure to 500 microM H2O2. Complete restoration of TNF-alpha sensitivity was obtained within 12 h after H2O2 removal. It required protein synthesis as demonstrated by the suppressive effect of actinomycin D. The inhibitory effect of H2O2 was suppressed by catalase, but was unaffected by the scavengers of hydroxyl radical and hypochlorous acid, suggesting that H2O2 but not one of its metabolites was responsible for this inhibition. H2O2 inhibitory effect did not implicate any change in prostaglandin production or in PKC activity. In contrast, H2O2 effect was associated with an about 50% loss of the density of cell membrane 125I-TNF-alpha receptors (2949 vs 5620 binding sites per cell), without change in their affinity (3.9 vs 3.4 nM). Moreover H2O2 did not affect the rate of degradation of TNF-alpha, and only slightly increased the degree of internalization of 125I-TNF-alpha receptors. These findings indicate that H2O2 can down-regulate the cellular response to TNF-alpha, possibly by reducing the TNF-alpha-binding capacity.     

Reduction in tumor necrosis factor receptor affinity and cytotoxicity by glucocorticoids

Micromolar concentrations of glucocorticoids rendered L-M cells (a murine tumorigenic fibroblast line) less sensitive to the cytotoxic activity of murine TNF. The potency of different steroids paralleled their known anti-inflammatory potency, and pretreatment was more effective than post treatment. Sex steroids and mineralocorticoids were ineffective. Dexamethasone also decreased the sensitivity of MCF-7 (a human mammary carcinoma line) to the cytotoxic activity of human recombinant TNF. Pretreatment of both cell lines reduced the affinity of specific cell surface receptors for the binding of their species 125I-TNF about 3-fold while retaining the same number of binding sites. The decrease in sensitivity was not due solely to the inhibition of early TNF-induced events (such as binding, internalization or signal transduction). Dexamethasone modestly enhanced inhibition beyond that of neutralizing antiserum alone when both were added midway in the L-M killing reaction (after receptor down regulation but before the onset of complete cell death).     

Tumor necrosis factor-mediated cytotoxicity by tumor-associated macrophages

We have directly demonstrated that macrophages present within solid EMT6 mammary tumors (of BALB/c origin) produce TNF-alpha (TNF). These tumor-associated macrophages lysed WEHI-164, a TNF-sensitive cell line, very efficiently. This cytotoxicity was abrogated in the presence of anti-TNF antisera. In contrast, EMT6 cells, the tumor from which the macrophages were obtained, were not effectively lysed by the macrophages and were 100-fold less sensitive to lysis by recombinant mouse TNF. Thus, marked heterogeneity exists among tumors regarding sensitivity to TNF-mediated cytotoxicity. Similarly, macrophages which infiltrate into EMT6 multicellular spheroids implanted into the peritoneal cavity as well as free cells within the cavity exhibited TNF-mediated cytotoxicity of WEHI-164 cells, but failed to lyse EMT6 cells. The kinetics of lysis by these cells was similar to that of recombinant mouse TNF.     

Meconium enhances platelet-activating factor and tumor necrosis factor production by rat alveolar macrophages

Meconium aspiration syndrome (MAS) frequently results in inactivation of surfactant, persistent pulmonary hypertension (PPHN) and respiratory failure among newborn infants. Inflammation and inflammatory mediators play an important role in MAS. Since alveolar macrophages are thought to be very important cells in the pathogenesis of various inflammatory diseases, we evaluated whether meconium could stimulate rat alveolar macrophages to generate platelet-activating factor (PAF) and tumor necrosis factor (TNF)-alpha in vitro. We also examined the response to A23187 (calcium ionophore), 1-0-Hexadecyl-2-acetyl-sn-glycero-3-phosphocholine (synthetic PAF) and dexamethasone on meconium-induced release of PAF and TNF-alpha. PAF and TNF-alpha concentrations from supernatant fluid were measured after high-performance liquid chromatography purification by specific radioimmunoassay, and TNF-alpha concentrations were determined by using an enzyme-linked immunosorbent assay. Our results showed that alveolar macrophages exposed to meconium could enhance PAF and TNF-alpha production in a dose (0.1, 1, 5 and 10%, P<0.01)-dependent way. In the presence of A23187, the capability of meconium to stimulate PAF production was further enhanced in the supernatant fluids. Furthermore, treatment with synthetic PAF significantly increased the generation of TNF-alpha in response to meconium. On the other hand, dexamethasone effectively inhibited both PAF and TNF-alpha production stimulated by 5% meconium (P<0.01, P<0.01; respectively). We suggest that alveolar macrophages and PAF, TNF-alpha play an important role in the pathogenesis of lung injury and severe complications in MAS. Furthermore, the protective effect of glucocorticoids in MAS could be due, at least in part, to a suppression of PAF and TNF-alpha generation.     

Involvement of tumor necrosis factor in cytotoxicity mediated by human monocytes

Human monocytes cultured in a specially prepared medium free of lipopolysaccharide (LPS) constitutively produced a small, though significant, amount of tumor necrosis factor (TNF). Upon addition of LPS, the amount produced remained constant until the LPS concentration reached 1-10 ng/ml, whereupon the production of TNF dramatically increased, eventually becoming 100-fold greater than when the LPS concentration was below 1 ng/ml. Priming the monocytes with recombinant interferon-gamma (rIFN-gamma) before LPS exposure resulted in a 2- to 10-fold increase in TNF production, the highest relative increase being obtained at lower LPS concentrations and in the absence of LPS. Monocyte-produced TNF appears to be the effector molecule in monocyte-mediated killing of some target cell types, since antiserum against recombinant TNF inhibited killing of both actinomycin D-treated and untreated WEHI 164 cells by human monocytes. However, it also appears that TNF may not in all cases be an effector molecule in monocyte-mediated killing, since cytolysis of K562 cells mediated by IFN-gamma/LPS-activated monocytes was not inhibited by antiserum against recombinant TNF. Antiserum which was raised against a monocyte-derived cytotoxic factor and which neutralized recombinant TNF did, however, inhibit monocyte-mediated cytolysis of K562 cells, suggesting that an extracellular factor, perhaps related to TNF, was also involved in monocyte-mediated killing of K562 cells. A TNF-like activity was associated with the monocyte surface membrane, since paraformaldehyde-fixed monocytes expressed cytotoxic activity which was neutralized by antiserum against recombinant TNF. Fixed monocytes activated with rIFN-gamma in addition to LPS before fixation were generally more cytotoxic than those exposed to LPS alone, and those exposed to LPS were much more cytotoxic than those not exposed to LPS. Thus it is possible that high local TNF concentrations may be generated near the target cell upon direct contact between effector and target cells, and that also monocyte-associated TNF may in this way be involved in monocyte-mediated cytotoxicity.     

Induction of the synthesis of tumor necrosis factor receptors by interferon-gamma

Human HT-29 colon carcinoma and HeLa D98/AH2 and SK-MEL-109 melanoma cells were sensitive to synergistic growth inhibition by concentrations of recombinant human tumor necrosis factor (rTNF) and interferon-gamma (rIFN-gamma) which individually were only slightly inhibitory. We investigated whether this synergism could be explained by the presence of an increased number of TNF receptors in cells treated with rIFN-gamma. These receptors were measured by incubating cells resuspended from monolayers with 125I-rTNF. HT-29 cells treated for a few hours with rIFN-gamma could bind more 125I-rTNF than control untreated cells, but this binding returned to the level of control cells after 24 hr. The treatment with rIFN-gamma did not change the binding affinity of TNF receptors, but increased their number to 1800 per cell from a basal level of about 800 per cell. Inhibitors of RNA synthesis prevented this increase. HT-29 cells were significantly more growth-inhibited when treated first for 6 to 12 hr with rIFN-gamma and then with rTNF, than when treated first with rTNF and then with rIFN-gamma. Untreated HeLa D98/AH2 and SK-MEL-109 cells had 2400 and 9000 receptors per cell, with a KD similar to that of HT-29 cells (approximately 2 X 10(-10)M). A significant increase in TNF receptors after treatment with rIFN-gamma was observed in HeLa D98/AH2, but not in SK-MEL-109 cells. No increase in TNF receptors was detected in cells treated with rIFN-alpha 2. These results indicate that the synergism between rTNF and rIFN-gamma may be due, at least in part, to a transient induction of the synthesis of TNF receptors by rIFN-gamma in cells with a relatively low number of these receptors.     

Induction of tumor necrosis factor expression by a lectin fromViscum album

Summary A purified lectin (MLI) fromViscum album was used to test whether peripheral monocytes from human blood can be activated for the production of tumour necrosis factor (TNF). Cytotoxic activity was detected in the supernatant of MLI-stimulated monocyte cultures. This cytotoxic activity was completely inhibited by monoclonal antibodies to TNF. Small amounts of soluble TNF protein were measured in a TNF-specific enzyme-linked immunospecific assay system. Strong expression of TNF mRNA was induced in human monocytes as well as in macrophage cultures from C3H/HeJ mice having a low response to endotoxin after 2 h of stimulation. Both chains of the MLI were found to induce TNF mRNA equally well in human monocytes. In macrophages of endotoxin-low-responder mice the toxic A chain was a better inducer of TNF mRNA than the galactose-specific lectin B chain. Thus, MLI has immunomodulating effects in activating monocytes/macrophages for inflammatory responses.     

Production of interleukin-1 and tumor necrosis factor by cisplatin-treated murine peritoneal macrophages

Supernatants collected from cisplatin-treated macrophages demonstrated enhanced cytotoxicity against actinomycin-D-treated L929 cells and also enhanced the thymocyte proliferation in response to concanavalin A, showing that cisplatin-treated macrophages release interleukin-1 (IL-1) and tumor necrosis factor (TNF) into the culture supernatant. The supernatant collected from untreated macrophages showed little TNF and IL-1 activity. The release of TNF and IL-1 was observed to be dependent on the dose and duration of cisplatin treatment. Medium alone containing cisplatin did not enhance thymocyte proliferation and had little cytotoxic effect on actinomycin-D-treated L929 cells. Cisplatin-treated macrophage culture supernatants were chromatographed over a Superose 12 column on an FPLC system. TNF activity eluted in two major peaks with apparent molecular weights of 50-55 and 15-20 kilodaltons, respectively. The kinetics of IL-1 release was also studied. Maximum production and release of IL-1 were observed up to 24 h after cisplatin treatment and then gradually declined. Freeze-thaw lysates of cisplatin-treated macrophages also showed enhanced IL-1 activity. Paraformaldehyde (PFA)-fixed cisplatin-treated macrophages showed significantly enhanced cytotoxic activity against L929 cells as compared to PFA-fixed untreated macrophages. PFA-fixed cisplatin-treated macrophages also enhanced thymocyte proliferation. These results suggest that cisplatin treatment of murine macrophages also results in increased expression of membrane-associated IL-1 and TNF activity.     

Modulation of colony-stimulating factor-1 receptors on macrophages by tumor necrosis factor

The effect of murine rTNF-alpha on the binding of human 125I-rCSF-1 to murine thioglycolate-elicited peritoneal exudate macrophages (PEM) was investigated. At 4 degrees C, 125I-CSF-1 binding to PEM was inhibited by preincubation with human rCSF-1, but not by other cytokines. When PEM were incubated with various cytokines at 37 degrees C, murine rTNF-alpha caused greater than 90% decrease in 125I-CSF-1 binding. This decrease was time, temperature and TNF dose dependent, and was not affected by preincubation with cycloheximide. The reduction in CSF-1-binding activity was reversed by prolonged incubation at 37 degrees C even in the presence of TNF. However, PEM preincubated with TNF subsequently washing free of residual TNF resulted in a rapid recovery of CSF-1 binding. This recovery of CSF-1-binding activity required protein synthesis. Binding studies suggested that the decrease in 125I-CSF-1 binding was most likely caused by a reduction in the number of CSF-1 receptors. In addition, preincubation with TNF at 37 degrees C inhibited 125I-CSF-1 binding on mononuclear phagocytes, including the macrophage cell line J774, bone marrow-derived macrophages, and nonelicited macrophages from three different strains of mice. In contrast, 125I-murine rTNF-alpha binding to PEM was not inhibited by preincubation with CSF-1 at 4 degrees C or 37 degrees C. These data suggest that TNF may play a role in the modulation of receptor expression on blood cells, and may point to a role for this pleiotropic cytokine in the regulation of hemopoiesis.     

Regulation of tumor necrosis factor gene expression and protein synthesis in murine macrophages treated with recombinant tumor necrosis factor

The effect of murine rTNF-alpha on c-fos and TNF mRNA accumulation and protein synthesis was investigated in bone marrow-derived macrophages to examine the mechanism(s) by which TNF modulates macrophage activity. A rapid and transient expression of the c-fos gene was induced by murine rTNF. This was blocked by 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine, an inhibitor of protein kinase C, suggesting that the murine rTNF stimulated a protein kinase C-dependent signal transduction pathway. Although LPS induced the accumulation of one TNF mRNA species, murine rTNF induced the synthesis of two distinct TNF mRNA species. Both LPS- and murine rTNF-induced TNF mRNA accumulation was equally enhanced by pretreatment with mouse rIFN-gamma. In contrast, cycloheximide pretreatment had little effect on murine rTNF-induced TNF mRNA accumulation, whereas this treatment increased LPS-induced TNF mRNA by sevenfold. These results argue that TNF mRNA accumulation can be modulated in macrophages by distinct mechanisms. As assessed by Western blot and immunoprecipitation analysis, LPS stimulated the synthesis of both cell-associated and secreted forms of TNF protein. In comparison, newly synthesized TNF protein was not detected when macrophages were treated with murine rTNF alone or in combination with murine rIFN-gamma. This demonstrates that although murine rTNF stimulated the synthesis of two distinct TNF mRNA species, additional signal(s) are necessary for their translation into protein and that such signals are present after LPS stimulation.     

Induction of tumour necrosis factor receptor-expressing macrophages by interleukin-10 and macrophage colony-stimulating factor in rheumatoid arthritis

Despite its potent ability to inhibit proinflammatory cytokine synthesis, interleukin (IL)-10 has a marginal clinical effect in rheumatoid arthritis (RA) patients. Recent evidence suggests that IL-10 induces monocyte/macrophage maturation in cooperation with macrophage-colony stimulating factor (M-CSF). In the present study, we found that the inducible subunit of the IL-10 receptor (IL-10R), type 1 IL-10R (IL-10R1), was expressed at higher levels on monocytes in RA than in healthy controls, in association with disease activity, while their expression of both type 1 and 2 tumour necrosis factor receptors (TNFR1/2) was not increased. The expression of IL-10R1 but not IL-10R2 was augmented on monocytes cultured in the presence of RA synovial tissue (ST) cell culture supernatants. Cell surface expression of TNFR1/2 expression on monocytes was induced by IL-10, and more efficiently in combination with M-CSF. Two-color immunofluorescence labeling of RA ST samples showed an intensive coexpression of IL-10R1, TNFR1/2, and M-CSF receptor in CD68⁺ lining macrophages. Adhered monocytes, after 3-day preincubation with IL-10 and M-CSF, could produce more IL-1β and IL-6 in response to TNF-α in the presence of dibutyryl cAMP, as compared with the cells preincubated with or without IL-10 or M-CSF alone. Microarray analysis of gene expression revealed that IL-10 activated various genes essential for macrophage functions, including other members of the TNFR superfamily, receptors for chemokines and growth factors, Toll-like receptors, and TNFR-associated signaling molecules. These results suggest that IL-10 may contribute to the inflammatory process by facilitating monocyte differentiation into TNF-α-responsive macrophages in the presence of M-CSF in RA.     

Peroxidases enhance macrophage-mediated cytotoxicity via induction of tumor necrosis factor

Tumor necrosis factor (TNF) is a monokine which is involved in macrophage-mediated cytotoxicity (MMC). We have previously reported that peroxidases can activate thioglycollate-induced macrophages to the tumoricidal state in vitro. The present study was undertaken in an attempt to correlate peroxidase-induced MMC with production of TNF. Horseradish peroxidase (HRP) was used as the principal model for these studies. Resident and thioglycollate-induced macrophages exposed to peroxidases were examined for both MMC against 3T12 cells and production of TNF. Thioglycollate-induced macrophages exposed to HRP, bovine lactoperoxidase, or human myeloperoxidase demonstrated enhanced secretion of TNF. When exposed to HRP, both resident and thioglycollate-induced macrophages secreted significant amounts of TNF and acquired the ability to lyse 3T12 cells. However, resident macrophages were considerably less efficient in both their cytotoxic activity and TNF secretion. Macrophage-mediated cytotoxicity was eliminated by the addition of specific antisera to TNF. In addition, replacement of culture supernatants within 24 hr after exposure of the macrophages to HRP increased tumor cell killing in the absence of additional detectable TNF production, suggesting that other factors may be involved in peroxidase-induced MMC. These results indicate that TNF is intimately associated with peroxidase-induced MMC and suggest a possible role for peroxidases as immunomodulators via augmentation of macrophage capacities and functions.     

Enhancement of cell-mediated cytotoxicity by recombinant tumor necrosis factor (TNF alpha)

The effects of recombinant tumor necrosis factor (rTNF alpha) on the immune responses were investigated. A single iv injection of rTNF alpha (6 x 10(3) U) caused regression of sarcoma-180 transplanted into BALB/c nu/+ mice, but failed to regress this tumor in nu/nu mice. A higher dose of rTNF alpha (2 x 10(4) U) was necessary to induce antitumor effect in nu/nu mice. A host-related factor seemed to be involved in mediating tumor regression. Therefore, the effects of rTNF alpha on various T-dependent immune responses, including delayed footpad reaction (DFR), cell mediated cytolysis (CMC), and plaque-forming cells (PFC) were examined in BALB/c mice, immunized ip with chicken erythrocytes (CRBC). A single injection of rTNF alpha, at the time of the antigen administration, induced the augmentation of CMC to CRBC in a dose-dependent manner. DFR and PFC were not affected in optimal immunization procedures. The TNF alpha injection, at or after the time of antigen administration, was more effective in inducing augmentation of CMC. The increase in CMC by TNF alpha was mediated by nonadherent, Thy 1.2, Lyt 2.2 positive cells and neutralization of TNF alpha by the anti-TNF alpha monoclonal antibody abolished the effect on CMC. These results indicated that the human recombinant TNF alpha induced changes in the T-cell-mediated responses.     

Spontaneous cytotoxicity and tumor necrosis factor production by peripheral blood monocytes from AIDS patients

Peripheral blood monocytes (PBM) from AIDS patients have exhibited defects in some but not all of the immune functions yet tested. This study has examined the capacity of AIDS PBM to lyse tumor target cells as well as their ability to secrete TNF. Untreated PBM from AIDS patients were significantly cytotoxic to U937 target cells and responded to IFN-gamma pretreatment with augmented cytotoxicity. Both the spontaneous and IFN-gamma-stimulated cytotoxic activity was significantly (p less than 0.01) higher than that observed with normal PBM. The cytotoxic activity depended on the E:T ratio used and was higher in AIDS PBM at all ratios tested (10:1 to 40:1). Because TNF has been implicated in macrophage cell-mediated cytotoxicity, we examined whether the elevated cytotoxic activity of AIDS PBM was associated with an increase in TNF production. Supernatants from PBM cultured overnight with or without IFN-gamma were tested in a bioassay measuring cytotoxicity against U937 target cells as well as in an RIA specific for TNF. Supernatants derived from either unstimulated or IFN-gamma-treated AIDS PBM exhibited significantly higher levels of cytotoxicity than supernatants from normal macrophages. Both normal and AIDS PBM produced higher levels of cytotoxic factors in response to IFN-gamma. As determined by the RIA, AIDS PBM spontaneously released high levels of TNF whereas little TNF was produced by normal PBM. Treatment with IFN-gamma augmented the level of TNF production in both AIDS and normal PBM. These results demonstrate that PBM from AIDS patients have undergone in vivo activation as manifested by both cytotoxicity against tumor target cells and production of TNF. Target cell lysis by both AIDS PBM and their supernatants was inhibited by monoclonal anti-rTNF, suggesting that the increase in PBM cell-mediated cytotoxicity was caused by an increase in TNF production. The significance of these findings in the pathogenesis of the disease is discussed.     

Stimulation of production of tumor necrosis factor by murine macrophages when exposed in vio and in vitro to weak electromagnetic waves in the centimeter range

Whole-body microwave sinusoidal irradiation of male NMRI mice, exposure of macrophages in vitro, and preliminary irradiation of culture medium with 8.15-18 GHz (1 Hz within) at a power density of 1 microW/cm2 caused a significant enhancement of tumor necrosis factor production in peritoneal macrophages. The role of microwaves as a factor interfering with the process of cell immunity is discussed.