Tumor cytotoxicity and interleukin 1 production of blood monocytes of lung cancer patients

Summary The effects of lung cancer on the abilities of blood monocytes to produce interleukin-1 and to mediate antitumor activity were examined. The functional integrity of blood monocytes was determined by their capacity to respond in vitro to a variety of activating agents and become tumoricidal, as assessed by a radioactive release assay and ability to produce interleukin-1 in vitro. The results show that the presence of lung cancer significantly increased the number of harvested blood monocytes and that the spontaneous tumoricidal activity of these monocytes was slightly high as compared to monocytes obtained from healthy donors. The production of interleukin-1 by monocytes of healthy donors and lung cancer patients was similar. Blood monocytes obtained from lung cancer patients were less cytotoxic against allogeneic A375 melanoma cells as compared with those of healthy donors subsequent to incubation with a soluble muramyl dipeptide analog or lipopolysaccharide, but were as tumoricidal as those from healthy donors when activated with lipophilic muramyl tripeptide (MTP-PE) entrapped in multilamellar liposomes. The finding that monocytes of patients with lung cancer can respond to MTP-PE encapsulated in liposomes, recommends the use of these liposomes in therapy of human lung cancer.     

Monocyte-mediated augmentation of human natural killer cell activity: conditions, monocyte and effector cell characteristics

The characteristics of the effector cells and monocytes, and conditions required for the monocyte-mediated augmentation of human natural killer (NK) cell activity were investigated. Enriched null cell populations were further fractionated by Percoll centrifugation and used as effector cells. The LGL-enriched fraction was less susceptible than either the unfractionated cells or the other Percoll fractions to the monocyte augmentation when mixed with monocytes in the chromium-release assay and when precultured with monocytes for 12 hr, retrieved by carbonyl iron treatment, and tested for NK activity against K562. This differential susceptibility was reflected at the single cell level. The LGL-enriched Percoll fraction did not display the increase in target-binding cells with lytic activity that was exhibited by the other effector cell preparations after culture with monocytes. No differences in Leu-7 and Leu-11 phenotypes were detected between enriched null cells that had been cultured with and without monocytes for 12 hr. At the monocyte level, it was shown that pretreatment of the monocytes with LPS did not alter their NK-augmenting activity appreciably. Glutaraldehyde-fixed monocytes were not effective, and actinomycin D-treated monocytes were less effective than untreated or irradiated monocytes when mixed with enriched null cells in the assay. Actinomycin D-treated monocytes did not augment and possibly suppressed NK activity tested after 12-hr culture, and irradiated monocytes were less effective for augmenting NK activity than untreated cells. Monocyte-mediated augmentation could be detected when the medium used for null cell-monocyte coculture was supplemented with a) different lots of fetal bovine serum, b) human AB serum, c) autologous serum, or d) no serum. Polymyxin B and indomethacin did not alter the monocyte effect. Finally, the monocyte-mediated augmentation of human NK was not MHC restricted, since allogeneic combinations were also effective. These results suggest that 1) lymphocytes other than LGL participate in the monocyte-mediated augmentation of NK activity, 2) the augmentation is probably activational rather than maturational, 3) the monocytes must be viable to be effective when mixed with null cells during the assay, 4) de novo RNA and/or protein synthesis by the monocytes is required for the monocytes to induce augmented activity in null cells after 12-hr coculture, 5) prostaglandin synthesis and endotoxin are probably not involved in the augmentation, 6) the phenomenon is not MHC restricted, and 7) monocytes may express augmentative and suppressive activities concurrently.     

Glucose and glutamine utilization by rat lymphocytes, monocytes and neutrophils in culture: a comparative study

Glucose and glutamine utilization and production of glutamate and lactate were determined for up to 48 h in lymphocytes, monocytes and neutrophils cultured in medium rich in metabolites and vitamins. Glucose was utilized by the three cell types in culture in the following order: neutrophils > monocytes > lymphocytes, whereas lactate was produced in the order: monocytes > neutrophils > lymphocytes. The consumption of glucose followed the activity of glucose-6-phosphate dehydrogenase but it was not related to hexokinase activity. Glutamine was consumed by the three leukocyte types in culture as follows: neutrophils > lymphocytes > or = monocytes. The consumption of glutamine was not fully related to the activity of phosphate-dependent glutaminase. The production of glutamate was not remarkably different among the three cell types. For comparison, glutamine and glucose utilization and glutamate and lactate production were also evaluated using 1-h incubated leukocytes. Under this condition, only glucose or glutamine was added to the medium. Glucose was utilized as follows: neutrophils > monocytes > lymphocytes, whereas lactate was produced in the following order: monocytes > or = neutrophils > lymphocytes. Glutamine was consumed as follows: neutrophils > lymphocytes > monocytes, whereas glutamate was produced as follows: neutrophils > or = monocytes = lymphocytes. The ratio of the amount of glucose/glutamine consumed by 1-h incubated cells was 0.5 for neutrophils, 1.5 for monocytes, and 0.3 for lymphocytes. However, the three cell types cultured for 48 h utilized glucose to a much higher degree than glutamine. The ratio of the amount of glucose/glutamine utilized by the cultured cells was 8.9 for neutrophils, 16.4 for monocytes, and 6.7 for lymphocytes. These observations support the proposition that glutamine is required in much higher amounts than glucose to accomplish the total metabolic requirement of leukocytes. Under conditions closer to physiological when the availability of a variety of metabolites and vitamins is not restricted, glucose is the preferred substrate for lymphocytes, monocytes and neutrophils.     

Lipopolysaccharide (LPS) modulation of human monocyte accessory cell function in promoting T-cell colonies: Inability of LPS and IL-1 to abrogate the need for monocytes with high HLA-DR expression

The abilities of human monocytes differentially expressing HLA-DR and of lipopolysaccharide (LPS) to influence T-cell colony responses were investigated. Optimal T-cell colony responses stimulated by soluble Staph protein A were crucially dependent on monocytes. Also, monocyte facilitation of colony responses was markedly inhibited by 10 μg/ml LPS and the addition of indomethacin reversed this inhibition. In contrast the inhibition of T-cell colony responses with 100 μg/ml LPS was not reversed with indomethacin and preincubation experiments with high concentrations of LPS showed the inhibition could be mediated through T cells by mechanisms other than prostaglandins. The treatment of monocytes with a monoclonal anti-HLA-DR reagent + C reduced the frequencies of monocytes expressing high levels of HLA-DR ~ fivefold and the resulting monocytes which expressed low levels of HLA-DR also poorly functioned in the promotion of colony responses compared to controls. LPS in the presence of indomethacin improved the ability of monocytes expressing low levels of HLA-DR to promote colony responses. However, these monocytes consistently failed to augment colony responses to those levels observed with untreated monocytes and their failure was not secondary to deficient interleukin 1 release. These results indicate that although LPS can somewhat potentiate the accessory cell function of certain human monocytes, it cannot abrogate an additional requirement for those monocytes expressing high levels of HLA-DR.     

Enhancing effects of monocytes on modulation of a lymphocyte membrane antigen

Redistribution, or modulation, of some cell surface antigens occurs in the presence of specific antibody. The phenomenon of antigenic modulation may therefore affect the use of antibodies as therapeutic agents. This study was undertaken to investigate modulation of the 65,000 dalton T65 antigen, present on normal and malignant T cells and some malignant B cells, which is recognized by the monoclonal antibody T101. To induce cell surface antigenic modulation, normal or leukemic lymphoid cells were cultured in the presence of monoclonal antibody T101 for 3-hr periods. Removal of monocytes from mononuclear cell preparations resulted in significantly lower degrees of T65 antigenic modulation. The degree of antigenic modulation could be increased by adding monocytes back to monocyte-depleted lymphocyte suspensions. Furthermore, maximal modulation occurred in the presence of monocytes at T101 concentrations that were 3 logs lower than in the absence of monocytes. The enhancing effect of monocytes was dependent on the Fc portion of the T101 antibody molecule, and presumably was mediated by cross-linking of antigen-antibody complexes on the surface membrane of the modulating cell by Fc receptors present on monocytes. Further experiments performed to examine the characteristics of this enhancement of antigenic modulation by monocytes indicated that autologous as well as allogeneic monocytes were effective, indicating that the enhancing phenomenon was not dependent upon recognition of major histocompatibility antigens. Viable monocytes were required, but pretreatment of monocytes with sodium azide to inhibit energy production, or indomethacin to inhibit prostaglandin synthesis had no effect on this phenomenon. Polymorphonuclear leukocytes did not mediate similar enhancement, although monocytic and myeloid cell lines U937, THP-1, and HL-60 did. Spent culture medium from modulated cultures and preparations containing IL 1 activity did not enhance modulation of the T65 surface antigen on lymphocytes, suggesting that direct contact between lymphocytes and monocytes is required to mediate the effect. The finding that leukemic cells from patients with CLL undergo modulation of the T65 antigen to a much lower degree in vitro than observed in vivo, and that this difference can be overcome by the addition of monocytes, suggests that monocytes or the reticuloendothelial system may augment antigenic modulation in vivo.     

M2b monocytes predominated in peripheral blood of severely burned patients

Severely burned patients were shown to be carriers of M2 monocytes, and all of the monocytes isolated from peripheral blood of severely burned patients (19 of 19 patients) were demonstrated as M2b monocytes (IL-12(-)IL-10(+)CCL1(+) monocytes). Low levels of M2a (IL-12(-)IL-10(+)CCL17(+) monocytes) and M2c monocytes (IL-12(-)IL-10(+)CXCL13(+) monocytes) were demonstrated in peripheral blood of severely burned patients (M2a, 2 of 19 patients; M2c, 5 of 19 patients). M2b, M2a, and M2c monocytes were not detected in peripheral blood of healthy donors. However, M2b monocytes appeared when healthy donor monocytes were cultured in media supplemented with burn patient serum (15%). CCL2 was detected in sera of all burn patients, and M2b monocytes were not generated from healthy donor monocytes cultured with media containing 15% burn patient sera that were previously treated with anti-CCL2 mAb. In addition, M2b monocytes were generated from healthy donor monocytes in cultures supplemented with rCCL2. These results indicate that M2b monocytes are predominant in peripheral blood of severely burned patients who are carriers of CCL2 that functions to stimulate monocyte conversion from resident monocytes to M2b monocytes.     

Leptin stimulates endogenous cholesterol synthesis in human monocytes: New role of an old player in atherosclerotic plaque formation. Leptin-induced increase in cholesterol synthesis

The role of leptin in the pathomechanism of atherosclerosis, through its free radical generating ability is established. Its effect however, on the regulation of intracellular cholesterol synthesis has not been studied. The aim of the present study was to elucidate whether leptin influences endogenous cholesterol synthesis in monocytes. Furthermore, leptin signaling to HMG CoA reductase in control and hypercholesterolemic monocytes were compared. The in vitro effect of leptin was studied on freshly isolated human monocytes obtained from healthy control volunteers and patients with hypercholesterolemia. Our results can be summarized as follows: (1) Leptin is able to increase endogenous cholesterol synthesis in human monocytes in vitro. (2) The cholesterol synthesis increasing effect of the hormone is more pronounced in hypercholesterolemic monocytes with high basal cholesterol biosynthesis. (3) The leptin-induced Ca(2+) signal was involved in the enhancement of HMG CoA reductase activation in monocytes from both controls and hypercholesterolemic patients. (4) In control monocytes the Ca(2+) signal originated from intracellular pools, whereas in patients, Ca(2+)-influx and protein kinase C activation were found to be responsible for the leptin-effect. Mevalonate cycle inhibiting fluvastatin and 25-hydroxycholesterol decreased cholesterol production in leptin-stimulated monocytes. Our present study provides the first proof of the cholesterol synthesis enhancing effect of leptin through a statin-sensitive pathway in circulating monocytes. Furthermore our results suggest that leptin can be involved in the pathomechanism of atherosclerotic plaque formation also through its effect on cholesterol biosynthesis in monocytes.     

The role of monocytes in human lymphocyte activation by mitogens.

Studies were performed to determine the role of monocytes in human lymphocyte activation by mitogens. Velocity sedimentation at 1 x G in a new apparatus was utilized to obtain highly purified lymphocyte fractions (LF) nearly free of monocytes (0.02 to 0.4%) and a fraction (MF) enriched for monocytes (64 to 92%). The average peak responses of the lymphocyte fractions to phytohemagglutinin, concanavalin A, and pokeweed mitogen were 19, 10, and 9% of the responses achieved with unfractionated lymphocyte cultures containing approximately 20% monocytes. These changes were not attributable to altered dose requirements. When mitomycin-C-treated MF cells were used to reconstitute LF cultures, it was found that 4% monocytes fully restored the response to phytohemagglutinin whereas 8 to 16% monocytes were required for a normal response to the other mitogens. Higher numbers of MF cells produced supranormal responses, with 35 to 50% monocytes resulting in the optimal stimulation. Allogeneic monocytes were able to fully reconstitute the response of LF, and 2-mercaptoethanol (50 microM) was only slightly effective. In exploring possible mechanisms by which monocytes potentiate the mitogenic activity of lymphocytes, it was found that the supernatants of MF cultures could partially, but not completely, reconstitute LF responses, suggesting that contact with MF may be required for optimal effectiveness. Addition of graded numbers of monocytes to LF altered both the kinetics of the response and the peak level of proliferation. Monocyte depletion also resulted in markedly decreased survival of cultured unstimulated LF. These observations suggest a variety of possible effects of monocytes in potentiating mitogenic responses, including contact-mediated interactions with lymphocytes (possibly to present the mitogen optimally); enhancement of proliferation kinetics and the size of the responding subpopulation, and maintenance of a requisite growth factor(s) in the culture. Small differences in the monocyte content of cultured lymphocyte preparations may thus account for many of the often observed variations in mitogen responsiveness.     

Response of nitric oxide production to CpG oligodeoxynucleotides in turkey and chicken peripheral blood monocytes

We evaluated the innate immune response to various synthetic CpG-containing oligodeoxynucleotides (CpG ODNs) by measuring nitric oxide production in the peripheral blood monocytes from turkey poults. The results indicate that the presence of the CpG dinucleotide in ODNs was a prerequisite for activation of turkey monocytes and induction of nitric oxide (NO) synthesis. CpG motifs and sequence structure of the ODNs were also found to influence stimulatory activity greatly. The most potent CpG ODN to induce NO synthesis in turkey monocytes was human-specific CpG ODN M362, followed by CpG ODN 2006 (human), CpG ODN#17 (chicken) and CpG ODN 1826 (mouse). The optimal CpG motif for NO induction was GTCGTT. Phosphorothioate modification of CpG ODNs also significantly increased stimulatory activity. Compared with chicken monocytes, turkey monocytes appeared to be less sensitive to CpG motif variation, whereas chicken monocytes were found to respond more strictly to human-specific CpG ODNs or ODNs that contain GTCGTT motifs.     

Induction of monocyte antitumor response by human cancer cells transduced with TNF-GFP fusion gene: possible implications for immunotherapy of cancer

This study was undertaken to determine how human pancreatic cancer (HPC-4) cells transduced with the TNF-GFP fusion gene (TLG) alter the antitumor response of human monocytes in vitro and whether they could act as an antitumor vaccine. In our model, HPC-4 cells were transduced with retroviral vector harboring TLG gene and designated as HPC-4(TLG). The TLG protein expression was confirmed by Western blot and flow cytometry analysis. Monocytes were co-cultured with transduced and control HPC-4 cells. The secretion of TNF, IL-10 and IL-12 was measured by ELISA. The cytotoxicity of monocytes against HPC-4 cells was determined by MTT test. The results show that the HPC-4(TLG) cells expressed membrane-bound, intracellular and secretory TLG protein. When cultured with HPC-4(TLG) cells, monocytes released a higher amount of TNF, but IL-10 and IL-12 secretion was inhibited. The pre-exposure of monocytes to HPC-4(TLG), but not to HPC-4, cells did not decrease TNF nor increase IL-10 production, thus not leading to monocyte deactivation. Also, the antitumor cytotoxicity of monocytes stimulated with HPC-4(TLG) was not downregulated, which occurred when non-transduced HPC-4 cells were used. In conclusion, compared to parental HPC-4 cells, TLG gene transduced HPC-4 cells induced stronger antitumor response of monocytes in vitro and prevented deactivation of monocytes.     

Erythromyeloid tumor cells (K562) induce PGE synthesis in human peripheral blood monocytes

Human peripheral blood monocytes were found to spontaneously produce prostaglandin of the E series (PGE) in culture medium (0.5 ng to 3.0 ng/7.5 X 10(5) cells), and the addition of K562 tumor cells enhanced the production by five- to 15-fold after 18 hr of incubation. PGE2 (10(-6) M) inhibited the cytolytic activity of freshly isolated peripheral blood monocytes against K562 target cells by 50%. The PGE production was inhibited by inhibitors of cyclo-oxygenase (indomethacin, aspirin, and ETYA) when present during the incubation. However, pretreatment of monocytes with these cyclo-oxygenase inhibitors was ineffective in preventing PGE production. Kinetic experiments showed that appreciable stimulation of PGE production occurred only after 6 hr of co-culture. Other human tumor cell lines (HSB, SB, and CEM) enhanced PGE production upon co-culture with monocytes but to a lesser extent (twofold to threefold). Monocytes treated with 0.4% formaldehyde or heat (56 degrees C) were not capable of producing PGE when cultured alone or with K526 tumor cells. In contrast, formaldehyde-treated, but not heat-treated, K562 tumor cells were able to induce monocytes to produce PGE. By using a single cell conjugation assay, K562 tumor cells were found to bind equally well to treated or untreated monocytes. In contrast, the lytic activity of treated monocytes against K562 target cells was abolished. The presence of protein synthesis inhibitor, cycloheximide, was found to inhibit PGE production by monocytes cultured alone or with K562 tumor cells. Supernatants from K562 tumor cell cultures were also capable of inducing monocytes to produce PGE, and their effect on PGE production from monocytes was suppressed by cycloheximide. In addition, pretreatment of either K562 tumor cells or monocytes with an irreversible protein synthesis inhibitor, emetine, also suppressed the production of PGE upon co-culture with the untreated counterpart. The production of PGE by monocytes in response to exposure to tumor cells may represent a mechanism whereby tumor cells subvert host immune defense against them.     

Isolation of functional subsets of human peripheral blood monocytes.

Monocytes were isolated by counterflow centrifugation of Ficoll-Hypaque separated peripheral blood mononuclear cells. The monocytes formed a bimodal volume distribution of "large" and "small" phagocytic esterase-positive, peroxidase-positive cells with peaks at 470 and 410 mu3, respectively. The large monocytes were predominately Fc receptor positive, and were able to lyse both sensitized human and chicken erythrocyte targets in ADCC assays, whereas the small monocytes were largely FcR negative and were inactive against sensitized human erythrocyte targets. However, ADCC against chicken erythrocyte targets was seen in some fractions containing small monocytes and was probably due to FcR+ lymphocytes (K cells) in those fractions. These experiments establish that monocytes are effectors of ADCC against both human and chicken erythrocyte targets and that the peripheral blood monocyte is heterogeneous in size, function, and surface receptor distribution.     

Role of interleukin 1 in promoting human monocyte-mediated tumor cytotoxicity

Human peripheral blood monocytes from normal donors obtained by separation on a Percoll gradient showed considerable cytotoxicity against tumor cells when preincubated in vitro for 24 hr with human monocyte-derived interleukin 1 (IL 1). In contrast, monocytes after pretreatment in medium alone had low cytotoxic activity. All the IL 1 preparations, including IL 1 which was purified by high-performance liquid column chromatography (HPLC), as well as crude culture supernatant from human monocytes promoted monocyte-mediated cytotoxicity in the same dose-dependent manner as the thymocyte growth-promoting activity. There was no endotoxin or interferon (IFN) activity in the highly purified IL 1, suggesting that IL 1 itself was the active moiety. The effect of IL 1 on monocyte-mediated cytotoxicity was partially inhibited by indomethacin, whereas pretreatment of monocytes with prostaglandin (PG) E1 or E2 rather than IL 1 also resulted in substantial monocyte cytotoxicity. Thus, the effect of IL 1 on monocyte-mediated cytotoxicity is presumably mediated by PGE. Since fresh monocytes that were not preincubated exhibited levels of spontaneous cytotoxic activity similar to that of monocytes preincubated with IL 1, it seemed likely that the effect of IL 1 was to maintain the spontaneous level of activity rather than to induce cytotoxic activity. To elucidate this possibility, monocytes were first preincubated in medium alone for a longer period, and after losing their spontaneous activity they were further incubated with or without IL 1. Such "aged" monocytes did not develop cytotoxic activity in response to IL 1 but did in response to other agents known to induce macrophage cytotoxicity, such as endotoxin or lymphokine-containing supernatants. Therefore, the major effect of IL 1 actually seemed to prolong the cytotoxic state of monocytes. These results also suggest that IL 1 released by macrophages or monocytes may play a role in host defense against neoplastic cells by acting on monocytes as an autostimulating factor.     

Differential survival of Leishmania donovani amastigotes in human monocytes

Leishmania donovani is an important intracellular protozoal pathogen of man; it is found solely within macrophages in its amastigote stage in humans, and exists in its extracellular, flagellated promastigote stage in the sandfly, its arthropod vector. To determine if either stage of L. donovani was capable of surviving within monocytes--the oxidatively active precursors of tissue macrophages--interactions of the parasite with human monocytes were studied in vitro. Amastigotes and promastigotes were ingested to a comparable degree by monocytes; whereas 79% of promastigotes were killed within 48 hr, however, amastigotes survived and multiplied threefold over 5 days. Promastigotes, which have been shown to be sensitive to hydrogen peroxide-peroxidase-halide microbicidal mechanisms, elicited a phagocytic oxidative burst that was 49% of the response to serum-opsonized zymosan, as assessed by luminol-enhanced chemiluminescence. NBT was reduced to formazan in 71% of monocytes exposed to promastigotes. The death of promastigotes within monocytes could be attributed at least in part to oxidative microbicidal mechanisms because there was no significant decrease in the number of cell-associated parasites in monocytes from donors with chronic granulomatous disease of childhood. In contrast to promastigotes, amastigotes survived within monocytes, despite eliciting an oxidative response that was 27% of the response produced by serum-opsonized zymosan; this response was not significantly different from that produced by promastigotes. In a phagocyte-free system, amastigotes were found to be sevenfold more resistant than were promastigotes to the lethal effects of hydrogen peroxide. The survival of L. donovani in human monocytes is thus dependent on the parasite stage; promastigotes are ingested, they elicit an oxidative burst, and the majority are killed by oxidative microbicidal mechanisms, whereas amastigotes are ingested and survive to parasitize human monocytes successfully, despite eliciting a phagocytic oxidative burst.     

Cilostazol reduces MCP-1-induced chemotaxis and adhesion of THP-1 monocytes by inhibiting CCR2 gene expression

The chemotaxis and adhesion of monocytes to the injured endothelium in the early atherosclerosis is important. Cilostazol, a specific phosphodiesterase type III inhibitor, is known to exhibit anti-atherosclerotic effects mediated by different mechanisms. This study aimed to investigate the modulating effect of cilostazol on the MCP-1-induced chemotaxis and adhesion of monocytes. The gene expression of CCR2, the major receptor of MCP-1 in THP-1 monocytes, was also analyzed. The chemotaxis of monocytes toward MCP-1 was investigated using the transwell filter assay. Cilostazol dose-dependently inhibited the MCP-1-induced chemotaxis of monocytes which was shown to be cAMP-dependent. Using western blot analysis and flow cytometry method, we demonstrated the decrease of CCR2 protein at the cell membrane of monocytes by cilostazol treatment. Results from RT/real-time PCR confirmed the decrease of CCR2 mRNA expression by cilostazol which was also mediated by cAMP. Similar inhibition was also noted in human peripheral monocytes. The post-CCR2 signaling pathways including p44/42 and p38 MAPK were examined by western blot analysis. Result confirmed the inhibitory effect of cilostazol on the phosphorylation of p44/42 and p38 MAPK after MCP-1 stimulation. The activation of monocytes after MCP-1 treatment exhibited enhanced adhesion to vascular endothelial cells which was dose-dependently suppressed by cilostazol. Together, cilostazol was demonstrated, for the first time, to inhibit the CCR2 gene expression and MCP-1-induced chemotaxis and adhesion of monocytes which might therefore reduce the infiltration of monocytes during the early atherosclerosis. The present study provides an additional molecular mechanism underlying the anti-atherosclerotic effects of cilostazol.     

Activation and immunoregulation of antigen-specific human b lymphocyte responses: multifaceted role of the monocyte

The multifaceted role of the monocyte in the induction and modulation of antigen-specific antibody responses by human B cells was delineated. Monocytes were absolutely required for the induction of specific antibody responses to both TT and KLH in an antigen-induced in vitro assay. Monocytes were also required for the PWM induction of specific antibody in immunized subjects. Pulsing monocytes with specific antigen or with PWM consistently stimulated proliferation of T cells in absence of added antigen and could also stimulate specific antibody synthesis although less consistently. Stimulation of specific antibody responses with antigen required fewer numbers of monocytes than did stimulation of specific antibody responses with PWM. Polyclonal antibody synthesis induced by PWM was also dependent on monocytes. However, polyclonal antibody synthesis induced by supraoptimal concentrations of antigen was usually optimal in the absence of monocytes and was actually suppressed when increased numbers of monocytes were added to monocyte-depleted cultures. Monocyte supernatants could not replace the absolute requirements for monocytes in the induction of specific antibody synthesis. However, monocyte supernatants could profoundly modulate the antigen-specific as well as the polyclonal Ig response of lymphocytes to either antigen or PWM stimulation in a manner closely resembling monocytes themselves. Thus, we demonstrated that monocytes and their products play a critical role in the activation and immunoregulation of antigen-specific antibody responses of human B cells.     

The cell-mediated lympholysis-inducing capacity of highly purified human monocytes and T lymphocytes in primary and secondary mixed leukocyte cultures

In this study the capacity of T cells and monocytes to induce cell-mediated lympholysis (CML) in primary and secondary MLC was investigated. The T lymphocytes were enriched by rosetting with sheep red blood cells (E) and further purified by sedimentation at unit gravity, which completely removed the contaminating monocytes. In addition, a highly purified monocyte population was obtained by 1 X G sedimentation of the non-E rosette-forming cells. These purified T cells have a poor CML-inducing capacity in primary and secondary MLC. In contrast, monocytes were very effective in inducing CML in both primary and secondary MLC. Induction of CML by monocytes in primary MLC was inhibited by heterologous anti-Ia-like antisera, indicating that the induction of CML by monocytes was related to the presence of HLA-DR (Ia-like) antigens on these cells.