Human macrophages play a key role in atherogenesis and are believed to be the progenitors of the cholesteryl ester (CE)-laden foam cells present in early atherosclerotic lesions. Several mechanisms by which macrophages accumulate CE have been recently described. One involves a perturbation in LDL metabolism subsequent to macrophage activation. Thus, we decided to study the effect of macrophage activation by immune complexes on N-LDL metabolism. Initially, LDL-containing immune complexes (LDL-IC) were chosen, since increased plasma levels of these IC have been reported in patients with coronary heart disease. Human macrophages stimulated for 22 h with LDL-IC (250 micrograms/ml) and incubated afterwards for 20 h with 10 micrograms/ml 125I-N-LDL showed a six- and fourfold increase in the accumulation and degradation, respectively, of 125I-N-LDL over the values observed in nonstimulated cells. Scatchard analysis of 125I-N-LDL-specific binding suggests an increase (20-fold) in the number of LDL receptors in macrophages stimulated with LDL-IC. We studied other immune complexes varying in size and antigen composition. Some of the IC were able to stimulate, although to a lesser degree, the uptake of N-LDL by macrophages. Lipoprotein IC are more efficient and have the greatest capacity to increase N-LDL uptake and CE accumulation. We conclude that human macrophage activation by LDL-IC leads to an increase in LDL receptor activity and promotes in vitro foam cell formation.

Antigen-Antibody Complex↗

PubMed1988

Ex vivo activation of killer monocytes (AKM) and their application to the treatment of human cancer.

Human blood monocytes activated by gamma interferon have been shown to be highly tumoricidal against colon cancer cells in vitro. Monocytes from patients with peritoneal colorectal carcinomatosis (PCC) were purified by a combination of cytapheresis and elutriation procedures, followed by in vitro incubation with gamma interferon for 18 hours. After debulking surgery, activated killer monocytes (AKM) were reinfused into patients' peritoneal cavities weekly for 16 weeks. To date, six patients have completed the entire protocol and three have completed maintenance therapy. All have tolerated the therapy well with acceptable toxicity. Midway through the protocol, we analyzed the trafficking pattern of the AKM by prelabeling them with 111In. Distribution was relatively homogeneous throughout the peritoneum; at the second staging celiotomy, three of the seven PCC patients were found to have very small amounts of recurrent disease in places to which the AKM were felt to have had limited access (other areas remained disease-free); these areas of recurrent disease were surgically resectable. AKM have also been infused systemically into five cancer patients. First, the patients were infused with unactivated 111In-labeled monocytes; 1 month later each patient received 111In-labeled gamma interferon-activated AKM. Trafficking studies indicated that both forms of monocytes migrated to sites in the reticuloendothelial system. We have seen virtually no complications from intravenous infusions of either unactivated or gamma interferon-activated AKM.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1988

Effects of adherence, activation and distinct serum proteins on the in vitro human monocyte maturation process.

Elutriator-purified human monocytes were cultured in a serum-free (SF) medium, and various serum proteins and functional activating agents were assessed for their effects on the in vitro maturation of human monocytes to macrophages. Following 3 days of suspension culture in Teflon labware, 60% of the monocytes were easily recovered. When varying concentrations of human AB serum (HuAB) were employed, human monocyte maturation progressed rapidly; the kinetics of this maturation process during cell suspension culture were very similar to the pattern observed following adherence culture. In contrast, when SF medium was employed, a marked retardation of the monocyte maturation process was observed; this could not be attributed to any changes in cell recovery and/or viability. Thus, cells could be maintained in their monocytoid form for 3 days when cultured in SF medium. When HuAB was added after 3 days of culture, human monocyte maturation into macrophages proceeded at a normal rate. We attempted to characterize certain of the serum protein(s) found in HuAB which promoted the monocyte maturation process. Human immunoglobulin G (IgG) was found to be the most potent serum protein in increasing 5'-N activity and decreasing peroxidase activity of suspension cultured monocytes. Immunoglobulin M (IgM) and albumin (Alb) were shown not to have significant monocyte maturation activity. Heat-treated human gamma globulin and IgG purified by high-performance liquid chromatography (HPLC) were shown to have patterns identical with that of untreated HGG and IgG with regard to promoting monocyte maturation; F(ab')2 was not an active maturation promoter, indicating the need for an intact Fc portion of the IgG molecule. Fibrinogen and fibronectin also had maturation promoting activity. Finally, addition of the potent monocyte functional activators, muramyl dipeptide (MDP), polyriboinosinic:polyribocytidilic acid (Poly I:C), and lipopolysaccharide (LPS) had no effect on the monocyte maturation process. Thus, neither cell adherence or activation appear to be critical for the monocyte to macrophage maturation process. Instead, we hypothesize that in addition to proper nutritional support, a group of serum proteins (unified mainly by their ability to interact with monocyte membrane receptors) appear to be the principal promoters of this process.

5'-Nucleotidase↗

PubMed1988

Adoptive cancer immunotherapy utilizing lymphokine activated killer cells and gamma interferon activated killer monocytes.

Explore the source record for details and available documents.

Animals↗

PubMed1988

The treatment of cancer with activated cytotoxic leukocyte subsets.

Cancer biotherapy focuses on stimulating natural defense mechanisms designed to control or eradicate cancer. Many distinct cancer biotherapy strategies have been formulated over the past two decades; one of the most promising involves the removal of cytotoxic leukocyte subsets from cancer patients, augmenting the cytotoxic function of these cells ex vivo and re-administering these activated cells with additional function-enhancing biologicals. This approach is termed adoptive cellular immunotherapy and is best represented at present by the activated killer monocyte and the lymphokinc activated killer cell therapies. The activation of cytotoxic monocytes with gamma interferon and the activation of cytotoxic lymphocytes with interleukin-2 are prototypic examples of adoptive immunotherapy research activity which is likely to expand in the near future.

Drug Evaluation↗

PubMed1988

An artificial lymph gland: a new approach to cancer therapy.

We are witnessing the initial stages of an era of extracorporeal blood cell manipulation aimed at amplifying, modifying, modulating, and regulating immune function. In some respects, this manipulation reproduces lymph organ functions. The basic technology is blood cell separation by continuous flow centrifugation resulting in isolation of mononuclear cells. These cells are activated in-vivo and ex-vivo with biological response modifiers. These modifiers appear to have applicability in the adoptive immunotherapy of cancer as shown in preliminary data obtained with lymphokine-activated killer (LAK) cells used in the treatment of advanced cancer in humans. The potential utilization of lymphokines as activators of tumoricidal activity by immunologic effector cells harvested from the cancer patient by cytapheresis is the subject of active research.

Cell Separation↗

PubMed1988

Glycosylation of low-density lipoprotein enhances cholesteryl ester synthesis in human monocyte-derived macrophages.

Glucose can react with the lysine residues of low-density lipoproteins (LDLs) and convert the lipoprotein to a form with a receptor-mediated uptake by cultured cells that is impaired. However, in contrast to other modified lipoproteins taken up by both murine and human macrophages via the scavenger-receptor pathway that may induce the formation of foam cells, glycosylated LDL is not recognized by murine macrophages, and thus far, it has not been shown to lead to marked intracellular accumulation of cholesterol in human macrophages. This study illustrates that glycosylated LDL incubated with human monocyte-derived macrophages, at a concentration of 100 micrograms LDL/ml medium, stimulates significantly more cholesteryl ester (CE) synthesis than does control LDL (10.65 +/- 1.5 vs. 4.8 +/- 0.13 nmol.mg-1 cell protein.20 h-1; P less than .05). At LDL concentrations similar to those of plasma, the rate of CE synthesis in macrophages incubated with glycosylated LDL is more markedly enhanced than that observed in cells incubated with control LDL (3-fold increase). The marked stimulation of CE synthesis in human macrophages exposed to glycosylated LDL is paralleled by a significant increase in CE accumulation in these cells (P less than .001). The increase in CE synthesis and accumulation seem to be mediated by an increase in the degradation of glycosylated LDL by human macrophages. Glycosylated LDL enters the macrophages and is degraded by the classic LDL-receptor pathway in slightly smaller amounts than control LDL, but its degradation by pathways other than the classic LDL receptor or scavenger receptor is markedly enhanced.(ABSTRACT TRUNCATED AT 250 WORDS)

Cholesterol Esters↗

PubMed1987

Fate of gamma-interferon-activated killer blood monocytes adoptively transferred into the abdominal cavity of patients with peritoneal carcinomatosis.

Five patients with colorectal cancer widely metastatic to peritoneal surfaces have been treated i.p. with infusions of autologous blood monocytes made cytotoxic by in vitro incubation with human gamma-interferon. The monocytes were purified by a combination of cytapheresis and counter-current centrifugal elutriation procedures; each week approximately 350 million activated monocytes were given to patients as adoptive immunotherapy by a single i.p. instillation. On the eighth cycle of treatment the trafficking of i.p. infused blood monocytes was studied in two patients by prelabeling the cells with 111In. These activated cells became distributed widely within the peritoneal cavity. Two and 5 days after infusion their position within the peritoneum had not changed. When peritoneal specimens were obtained 36 h after 111In-labeled monocyte infusion, labeled monocytes were demonstrated to be associated with the serosal surfaces by autoradiographic analysis. Scintiscanning structures outside the abdominal cavity revealed that 111In-labeled monocytes infused i.p. did not traffic to other organs during the 5 days of the study. We conclude that i.p. adoptive transfer of autologous killer blood monocytes is an effective way of delivering these cytotoxic cells to sites of tumor burden on peritoneal surfaces in these cancer patients.

Colonic Neoplasms↗

PubMed1987

Reaction of T lymphocytes with anti-T3 induces translocation of C-kinase activity to the membrane and specific substrate phosphorylation.

Reaction of the T cell membrane with monoclonal antibodies to T3 can initiate cellular activation, and this is associated with increased intracellular Ca2+ and inositol-trisphosphate (IP3) release. We therefore studied the possible involvement of Ca2+/phospholipid-dependent kinase (C-kinase) in these phenomena. Quantitative assays of exogenous substrate phosphorylation in unstimulated cells showed Ca2+/phospholipid-dependent kinase activity in the cytosol, but no comparable activity in the particulate fractions corresponding to membrane and cytoskeleton material. At concentrations of soluble anti-T3 that partially activate T cells in the absence of macrophages, there was a 50 to 60% decrease in C-kinase activity in the cytosol, with a comparable increase in activity in the membrane fraction. A similar transfer of activity was also induced with the known C-kinase activator, 12-O-tetradecanoyl-phorbol-13-acetate, although redistribution was more rapid in onset, more complete, and more sustained. Redistribution of enzyme activity was additionally confirmed by qualitative assays of endogenous substrate phosphorylation. Labeling of intact cells followed by immunoprecipitation analysis with anti-T3 indicated signal-dependent phosphorylation of two components of the T3 complex and an unidentified 94,000 substrate that was resistant to reduction and alkylation. These findings are consistent with an important role for C-kinase in transduction of membrane events by the T3-Ti complex.

Antibodies↗

PubMed1987

Stimulation of cholesteryl ester synthesis in human monocyte-derived macrophages by low-density lipoproteins from type 1 (insulin-dependent) diabetic patients: the influence of non-enzymatic glycosylation of low-density lipoproteins.

Diabetes mellitus is an independent risk factor in the development of atherosclerosis. In this study we aimed to demonstrate whether there is an abnormal interaction between low-density lipoproteins from diabetic patients and human macrophages. We measured cholesteryl ester synthesis and cholesteryl ester accumulation in human monocyte-derived macrophages (obtained from non-diabetic donors) incubated with low density lipoproteins from Type 1 (insulin-dependent) diabetic patients in good or fair glycaemic control. Low density lipoproteins from the diabetic patients stimulated more cholesteryl ester synthesis than low density lipoproteins from non-diabetic control subjects (7.19 +/- 1.19 vs 6.11 +/- 0.94 nmol/mg cell protein/20 h, mean +/- SEM, p less than 0.05). The stimulation of cholesteryl ester synthesis by low density lipoproteins isolated from diabetic patients was paralleled by a significant increase in intracellular cholesteryl ester accumulation (p less than 0.02). There were no significant differences in the lipid composition of low density lipoproteins between the diabetic and control groups. Non-enzymatic glycosylation of low density lipoproteins was higher in the diabetic group (p less than 0.01) and correlated significantly with cholesteryl ester synthesis (r = 0.58). Similarly, low-density lipoproteins obtained from non-diabetic subjects and glycosylated in vitro stimulated more cholesteryl ester synthesis in macrophages than control low density lipoproteins. The increase in cholesteryl ester synthesis and accumulation by cells exposed to low density lipoproteins from diabetic patients seems to be mediated by an increased uptake of these lipoproteins by macrophages.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1987

Expression and secretion of type beta transforming growth factor by activated human macrophages.

Alveolar macrophages activated with concanavalin A and peripheral blood monocytes activated with lipopolysaccharide secrete type beta transforming growth factor (TGF-beta). There is minimal TGF-beta secretion in unactivated monocytes, even though TGF-beta mRNA is expressed in these cells at a level similar to that in activated, lipopolysaccharide-treated cultures. U937 lymphoma cells, which have monocytic characteristics, also express mRNA for TGF-beta. Freshly isolated monocytes, both control and lipopolysaccharide-treated, secrete an acid-labile binding protein that inhibits TGF-beta action. We conclude the following: (i) that expression of TGF-beta mRNA is unrelated to monocyte activation, (ii) that secretion of TGF-beta is induced by monocyte activation, and (iii) that cosecretion of TGF-beta and its monocyte/macrophage-derived binding protein may modulate growth factor action. In contrast, monocytic expression of other growth factor genes, such as the B chain of platelet-derived growth factor, is not constitutive and requires activation.

Cell Line↗

PubMed1987

Halothane anesthesia decreases human monocyte hydrogen peroxide generation. Protection of monocytes by activation with gamma interferon.

In an effort to determine the impact of halothane anesthesia on certain human cell-mediated immune functions, normal, purified human monocytes and lymphocytes were exposed to halothane in vitro at varying concentrations for up to 8 hours. Subsequently, these human effector cells were analyzed for their ability to function in several cell-mediated immunologic assays. Natural killer cell activity against K-562 was unaffected by halothane in most of the donors tested. Similarly, the ability of purified monocytes to inhibit MBL-2 tumor cell growth was unchanged. Halothane appeared to decrease the proliferative response of lymphocytes to phytohemagglutinin (PHA) in approximately 50% of the normal donors tested. In contrast, the ability of monocytes to lyse antibody-coated red cell targets (ADCC) was unaffected by even maximal exposure to halothane. Of interest was the finding that human monocytes exposed to as low as 2% halothane anesthesia for 4 hours displayed a dramatic down-regulation of hydrogen peroxide (H2O2) release. Since it is known that hydrogen peroxide and other incompletely reduced forms of oxygen secreted by monocytes can play a major role in the antimicrobial, antitumor, and inflammatory functions of these cells, this finding may help explain the enhanced susceptibility of post-operative patients to infections.

Adult↗

PubMed1986

Synergistic action of adenosine and fMet-Leu-Phe in raising cAMP content of purified human monocytes.

The content of cAMP was measured in monocytes treated with fMet-Leu-Phe and adenosine, either singly or in combination. Adenosine caused a small and variable rise in cAMP, which was considerably less than that caused by fMet-Leu-Phe. The rise induced by peptide plus adenosine was twice the sum of the increases caused by each agent alone. An inhibitor of phosphodiesterase also enhanced the adenosine-induced rise in cAMP. The data suggest that the increase in cAMP by adenosine-induced cyclase activation is limited by the activity of phosphodiesterase, and that the latter can be inhibited by fMet-Leu-Phe.

4-(3-Butoxy-4-methoxybenzyl)-2-imidazolidinone↗

PubMed1986

The effect of anesthetic agents on human immune system function. I. Design of a system to deliver inhalational anesthetic agents to leukocyte cultures in vitro.

A novel system for exposing purified human mononuclear leukocyte subsets or any cultured cell to inhalational anesthetic agents has been devised. Monocytes and lymphocytes are purified by counter-current centrifugal elutriation and put into culture vessels with and without appropriate functional activators. The culture vessels are placed into one of four anesthetic agent exposure chambers, each containing a different concentration of the anesthetic agent to be tested. The system that delivers the anesthetic agent to the culture vessels is essentially the same as the one used in the operating room; the actual levels of anesthetic agent delivered are monitored. In this report, we present evidence that halothane can be successfully tested using the above-cited experimental design; this agent substantially inhibits the secretion of interferon by human mononuclear cells. The ability of monocytes to secrete alpha interferon in response to polyinosinic: polycytidylic acid (poly I:C) was significantly depressed following 4 h in vitro exposure to increasing concentrations of halothane; the secretion of gamma interferon by lymphocytes in response to phytohemagglutinin (PHA) was also depressed, although to a lesser degree. The system presented should allow for the in vitro exploration of the effects of inhalational agents on purified human leukocyte subset functions as well as for the analysis of the effect of these agents on monocyte/lymphocyte interactions.

Halothane↗