bluuebunny/crossref_metadata_2025 huggingface dataset:
Characterization of Purified Cryopreserved Human Monocyte Function in Assays of Superoxide Production, Accessory Cell Function, Chemotaxis, and in Fluorescent Cell Sorter Analysis
Authors: Stevenson Henry C , Bonvini Ezio , Favilla Teresa , Miller Paul , Akiyama Yukio , Hoffman Thomas , Oldham Robert , Kanapa Dorothy
Published: January 2018

The ability to use highly purified cryopreserved human monocytes in various in vitro assays has a number of practical and theoretical advantages, including convenience and the potential for enhanced reproducibility. Large numbers (up to 1 × 109) human monocytes can be isolated from a single donor in a purified suspension state, by a combination of leukapheresis and counter- current centrifugal elutriation (CCE) technologies. Following short- and long- term periods of cryopreservation, the viability and phagocytic function of these CCE-purified monocytes was unimpaired. Cryopreserved monocytes were similar to fresh cells in their ability to release superoxide anion (O2), although unstimulated and stimulated Of release values tended to increase slightly following weeks to months of cryopreservation. In contrast, even short-term cryopreservation diminished both the random migration and chemotactic responses of human monocytes; however, cryopreserved monocytes could be employed in this assay provided the calculated chemotactic ratio (chemotactic migration/spontaneous migration) was used. Cryopreserved monocytes demonstrate 70% of fresh monocyte accessory cell function in pokeweed mitogen- induced lymphocyte proliferation assays. When the binding of OKT3, OKM1, anti-DR, and fluoresceinated pokeweed mitogen to monocytes was analyzed in the fluorescence activator cell sorter (FACS), cryopreserved and fresh monocytes displayed a similar pattern of membrane reactivity.
The Iron-Hydrogen Peroxide-Iodide System Is Fungicidal: Activity Against the Yeast Phase of Blastomyces dermatitidis
Authors: Sugar Alan M , Chahal Resham S , Brummer Elmer , Stevens David A
Published: January 2018

A series of experiments show the potency of a newly described microbicidal system, involving iron, H2O2, and halide, in killing a fungus (Blastomyces dermatitidis). B dermatitidis has previously been shown susceptible to the myeloperoxidase-H2O2 halide system. The present studies show killing of either of two strains in 1 hour if Fe+ + at 5 × 10−5M, H2O2 at 5 × 10−M and Kl at 5 × 10−4M are all present (P < 0.001). EDTA, a Fe++ chelator, abrogates killing. The mechanism presumably utilizes hydroxyl radical, since an inhibitor, ethanol, also neutralizes the system. The bactericidal and fungicidal system is of great potential importance in vivo.
Deficient Kupffer Cell Phagocytosis and Lysosomal Enzymes in the Endotoxin-Low- Responsive C3H/HeJ Mouse
Authors: McCuskey Robert S , Urbaschek Renate , McCuskey Patricia A , Sacco Nancy , Stauber William T , Pinkstaff Carlin A , Urbaschek Bernhard
Published: January 2018

Various substances, including lysosomal enzymes, are produced by Kupffer cells and other macrophages; their release has been implicated in the toxic response to endotoxins. C3H/HeJ mice exhibit little or no response to doses of endotoxin that are lethal in syngeneic C3HeB/FeJ mice. To explore the nature of this deficient response, the Kupffer cells of these mice were studied using in vivo microscopic as well as histochemical and electron microscopical methods. In vivo, the rate of phagocytosis of single 0.8 μ m latex particles was measured in individual Kupffer cells as was the number of phagocytic cells per microscopic field. Frozen sections of livers were stained for a variety of lysosomal enzymes and liver specimens also were processed for electron microscopy. In comparison to the endotoxin-sensitive C3HeB/FeJ mice, the livers of the C3H/HeJ mice contained 60% fewer Kupffer cells that phagocytosed latex. However, the rate of phagocytosis by these cells was not statistically different and ranged from 19–26 sec. The volume density of acid-phosphatase-positive Kupffer cells was 40% less in the C3H/HeJ mice. Similar differences were observed with other lysosomal enzymes including cathepsins B and H and dipeptidyl peptidases I and II. However, light and electron microscopy revealed a relatively normal number of Kupffer cells in livers stained for peroxidase, a nonlysosomal enzyme. The results suggest that the insensitivity of C3H/HeJ mice to endotoxin may be related in part to a lysosomal enzyme deficiency and a paucity of phagocytic Kupffer cells in these animals.
Effect of Lidocaine on Production of Migration Inhibitory Factor and on Macrophage Motility: In Vitro Exposure of Guinea Pig Lymphocytes and Macrophages
Authors: Dickstein R A , Kiremidjian-Schumacher L , Stotzky G
Published: January 2018

Exposure of lymphocytes from guinea pigs sensitized to keyhole limpet hemo- cyanin and of macrophages from nonsensitized animals to noncytotoxic doses of lidocaine (10−4 to 10−6 M) resulted in the inhibition of the production of macrophage migration inhibitory factor and of macrophage motility. The inhibition of both processes was related to the concentration of lidocaine in the medium. The effects of lidocaine, a membrane-stabilizing drug, were apparently related to its ability to interact with the cell surface and cause changes in the surface ionic configuration of the cells, as determined by cell electrophoresis. The drug conferred permanent changes in the surface of lymphocytes at all concentrations tested, but the changes in the surface of macrophages induced in the presence of 10−5 and 10−6 M of the drug were reversible. The presence of noncytotoxic doses of lidocaine in the cellular environment resulted in significant changes in cellular functions that appeared to be related to the ability of the drug to interact with cell membranes in a manner determined by the specific surface properties of the cell.
Fluorescent Opsonization Assay: Binding of Plasma Fibronectin to Fibrin-Derivatized Fluorescent Particles Does Not Enhance Their Uptake by Macrophages
Authors: Falcone Domenick J
Published: January 2018

A simple method for assessing the opsonic properties of plasma fibronectin is described. The method is based on the uptake of protein-derivatized fluorescent particles by mouse peritoneal macrophages. With this system, fibronectin stimulated the uptake of gelatin conjugated to carboxylated fluorescent particles. In contrast to the gelatinized particles, neither the covalent nor noncovalent interaction of fibronectin with fibrin particles stimulated their uptake by macrophages. These results provide additional evidence that fibronectin is not a conventional opsonin and that enhanced phagocytosis is not an obligatory response by macrophages to fibronectin-coated particles.
Phenylhydrazine-Induced Leukocytosis in the Rat
Authors: Dornfest Burton S , Lapin David M , Naughton Brian A , Adu Samuel , Korn Larry , Gordon Albert S
Published: January 2018

Rats were injected with phenylhydrazine (PHZ) for periods of up to 6 months, during which time a marked leukocytosis was induced. The highest leukocyte counts occurred within 4–5 days following injection. An initial injection of 4 mg/100 gm body weight evoked a mean total leukocyte count of 129 × 103 cells/μl. Successive weekly injections of 2 mg/100 gm resulted in a mean total leukocyte count of 70 × 103 cells/μl compared to a mean total leukocyte count of 12.5 × 103 cells/μl in saline-injected rats. Lymphocytes and monocytes accounted for approximately 75% of the total cell counts in both the PHZ-treated and control rats. The presence of increased numbers of mononuclear cells was confirmed by Percoll gradient separation and by phase-contrast microscopy. Although a leukocytosis was evident when using the automated Coulter electronic cell counter, it was not discernible when blood samples were counted manually in a hemocytometer by light microscopy. Histological examination of the thymus, lymph nodes, and spleen of the PHZ-treated rats indicated that lymphocytes and monocytes were mobilized from these sites. Lymphocyte depletion was evident, and germinal centers were found in all these lymphoid organs, indicating that PHZ induced a lymphopoietic response. A possible autoimmune etiology for PHZ-induced red blood cell destruction is discussed.
The Cytokinetic Behavior of Pulmonary Alveolar Macrophages in Monocytopenic Mice
Authors: Sawyer Richard T
Published: January 2018

The cytokinetic behavior of pulmonary alveolar macrophages (PAM) was studied by pulse labeling with 3HTdR in mice made monocytopenic by a single intravenous injection of the bone-seeking isotope strontium-89 (89Sr). In the presence or absence of blood monocytes, PAM population size was unchanged for up to 1 month of chronic, severe monocytopenia. Pulse-labeling studies performed during monocytopenia show that in control mice PAM population half-times were 17.8 days with a potential doubling time of 38 days, whereas T½ was 14.8 days with a 28.5 day population doubling time for PAM in 89Sr-treated mice. Analysis of the halving times of the PAM mean grain count and the halving times of the most highly pulse-labeled cohorts suggested that PAM cell cycle times (Tc) were 5.1 days with a PAM rate of disappearance of 10.8%/day in 88Sr-treated mice and Tc of 6.6 days with a PAM rate of disappearance of 11.4%/day in 89Sr-treated mice. As measured by 3HTdR-labeling techniques, these cytokinetic values are in close approximation to each other, suggesting that 89Sr treatment did not significantly alter either PAM population size or cytokinetic behavior. Employing experimental values it was possible to construct a simple model of PAM population growth that supports the concept that the PAM population is self-renewing in the adult mouse. Taken together, the data show that a major portion of the resident PAM need not depend on the daily influx of peripheral blood monocytes to maintain themselves in a kinetically steady state.
Binding of the Adhesive Protein Complex (LFA-1/Mac-1/p150,95) to Concanavalin A
Authors: Schmalstieg Frank C , Rudloff Helen E , Anderson Donald C
Published: January 2018

At least 30 proteins from human PMNL plasma membranes capable of binding concanavalin A (Con A), can be identified after surface labeling with 125I and subsequent sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Immunoprecipitation of the labeled proteins after solubilization in non-ionic detergents, with monoclonal antibodies (MAbs) directed against a family of leukocyte membrane proteins (LFA-1, Mac-1, p150,95, and the beta subunit these glycoproteins share), indicated that Mac-1 and p150,95 were bound by Con A. Dissociation of the alpha and beta subunits with sodium dodecyl sulfate, electrophoresis, transfer to nitrocellulose paper, and subsequent binding of these proteins by Con A demonstrated Con A retention by Mac-1-α, p150,95-α, and the common beta subunit. Affinity of Con A for LFA-1-α from human peripheral blood PMNL could not be confirmed by direct binding or electroblotting. Similar experiments in a patient deficient in LFA-1, Mac-1, p150,95, and the beta subunit confirmed that Mac-1-α and the beta subunit were important Con A-binding proteins.
Human Monocyte Production of Interleukin-1: Parameters of the Induction of Interleukin-1 Secretion by Lipopolysaccharides
Authors: Newton R C
Published: January 2018

The role of lipopolysaccharide (LPS) as an activator of human monocyte interleukin-1 (IL-1) synthesis and secretion has been examined in this study. The results demonstrate that when blood monocytes are prepared under low endotoxin conditions, they do not spontaneously secrete IL-1 activity. When cells are exposed to LPS extracted from different bacterial species, there is variation seen in the potency, with LPS from Salmonella species being the most potent in inducing IL-1 activity from human monocytes. This material is tenfold more potent than LPS obtained from three different strains of Escherichia coli and 10,000-fold more potent than material obtained from two other bacterial species. Detoxified endotoxins are inefficient activators for IL-1 secretion. When monocytes are exposed to LPS, there is a rapid rise in the level of IL-1 activity detected. Activity can be detected in cell lysates after 1 hr with appreciable accumulation seen over the first 6 hr of culture. This is accompanied by IL-1 release into the surrounding medium after 2 hr of culture with subsequent accumulation. Monocyte synthesis of IL-1 activity appears to be sensitive to fg/ml levels of Salmonella minnesota LPS, while appreciable secretion of this activity by monocytes requires pg/ml levels.
Cyclic AMP Levels and Cellular Kinetics During Maturation of Human Promyelocytic Leukemia Cells
Authors: Chaplinski Thomas J , Niedel James E
Published: January 2018

Differentiation of human promyelocytic leukemia cells (HL-60) in response to several classes of inducing agents is characterized by the sequential appearance of granulocytic or monocytic markers. Compounds that increase intracellular cAMP in HL-60 cells induce a program of maturation in which cells demonstrate early functional phagocytic properties.

Cyclic nucleotide metabolism was studied during monocytic and granulocytic differentiation of the HL-60 cell line. In synchronous and nonsynchronous cell cultures, cyclic AMP levels were raised 300-fold or 25-fold in response to N6, O2-dibutyryl adenosine 3′:5′-cyclic monophosphate or the combination of prostaglandin E2 and theophylline, respectively. No reproducible changes in intracellular adenosine 3′:5′-cyclic monophosphate levels occurred in response to retinoic acid or dimethyl sulfoxide, suggesting that changes in adenosine 3′:5′-cyclic monophosphate levels alone do not mediate cell maturation induced by these compounds. Agents that increased intracellular cyclic AMP in HL-60 cells slowed the progress through the S and G2-M phase of the cell cycle, whereas other agents such as dimethyl sulfoxide stimulated cells through this same period.
A Quantitative Lymphocyte Localization Assay
Authors: Borgs P , Hay J B
Published: January 2018

Multiple lesions (up to 100 sites) were induced in the skin of sheep using either allogeneic lymphocytes or, in BCG-sensitized animals, tuberculin. Cells recovered from an indwelling lymph catheter draining a prescapular lymph node were labeled with 111-indium, returned to venous blood, and allowed to circulate for 3 hours. Sheep were killed, and the skin lesions and lymph nodes were removed and counted in a gamma spectrometer. High levels of radioactivity (up to 38,000 cpm/lesion) were recovered from lesions, and only a few hundred cpm were recovered from comparable normal skin sites. Dose-response relationships and time kinetics were demonstrated for these lesions, and the radioactivity on blood and lymph cells was measured. The contribution of cell-free radioactivity was negligible. Using replicate injection sites, analytical, internally controlled studies can now be initiated to study the induction, promotion, and suppression of lymphocyte traffic.
The Role of Langerhans Cells and Keratinocytes in Epidermal Immunity
Authors: Lee Choi Ken , Sauder Daniel N
Published: January 2018

The immunology of the epidermis has received considerable study over recent years. After the antigen-presenting capacity of epidermal Langerhans cells was confirmed, subsequent studies suggested that keratinocytes could modulate certain immunologic events through production of a cytokine, epidermal cell-derived thymocyte-activating factor (ETAF). Most recently, a murine epidermal cell population, the dendritic Thy-1-positive cell, has been shown to possess natural killer-cell-like activity. In this review, the biology of these cell types are discussed. A discussion of allergic contact hypersensitivity and its alteration by ultraviolet light is used to illustrate some of the complex control mechanisms that continue to be the subject of ongoing study.
Pulmonary Hemodynamic Effects of Antisheep Serum-Induced Leukopenia
Authors: Heath Carole A , Lai Linda , Bizios Rena , Malik Asrar B
Published: January 2018

Antisheep antileukocyte serum (ALS) was produced in rabbits, purified, and adsorbed against erythrocytes and platelets. The ALS was infused intra-arterially over a 3-hour period into anesthetized sheep (n = 6) prepared with lung lymph fistulas. Pulmonary vascular resistance (PVR) and pulmonary lymph flow (Qlym) increased two- to threefold while the lymph-to-plasma protein concentration ratio (L/P) did not change from baseline, suggesting an increase in pulmonary vascular permeability to proteins. The arterial leukocyte count decreased from 4,935 ± 840 to 1,385 ± 325 cells/μl, the neutrophil count decreased from 1,045 ± 265 to 340 ± 130 cells/μl, and the platelet count decreased from 2.8 × 105 ± 0.2 × 105 to 0.65 × 105 ± 0.12 × 105 cells/μl. ALS induced neutrophil aggregation in vitro, but not platelet aggregation. In the present study, we also examined the effects of ALS-induced leukopenia on the increase in pulmonary vascular permeability after intravenous α-thrombin challenge to assess the role of leukocytes in mediating the increased permeability response. Sheep (n = 5) were made leukopenic by an intramuscular injection of ALS; Qlym was in the normal range 4 to 5 hours after the ALS administration. In the leukopenic group, thrombin challenge caused an increase in Qlym from 7.8 ± 0.7 to 12.9 ± 1.9 ml/hr (a 64% increase) while L/P decreased from 0.86 ± 0.04 to 0.70 ± 0.05 (a 19% decrease). In contrast, thrombin-induced intravascular coagulation in control sheep (n = 5) produced a 250% increase in Qlym with an increase in L/P. The results indicate that leukocytes are required for the increase in lung vascular permeability after thrombin-induced intravascular coagulation.
In Vitro Effects of Platelet Factor 4 on Normal Human Neutrophil Functions
Authors: Bebawy Sam T , Gorka John , Hyers Thomas M , Webster Robert O
Published: January 2018

Platelet factor (PF4) prepared from human outdated platelets by heparinagarose affinity chromatography was confirmed to be chemotactic for human neutrophils and in a concentration-dependent fashion caused significant release of lysosomal enzymes (myeloperoxidase, lysozyme, beta-glucuronidase) from human neutrophils treated with cytochalasin B. Lysosomal enzyme release from PF4-stimulated neutrophils was rapid and reached a plateau by 1–3 min. PF4 did not cause release of the cytoplasmic enzyme lactate dehydrogenase which indicates that exocytosis of granule-containing lysosomal enzymes did not result from cytolysis. In contrast, superoxide anion generation from human neutrophils stimulated with PF4 was undetectable even at the highest PF4 concentration tested (2 × 10−5 M). Pretreatment of neutrophils with PF4 caused significant increased adherence of neutrophils to plastic surfaces and cultured pulmonary artery endothelial cells. The concentration of PF4 that elicited neutrophil chemotaxis, lysosomal enzyme release and increased adherence is slightly higher than those concentrations found in normal human sera. However, the results suggest that PF4 may be an important mediator in neutrophil-platelet interactions and the induction of acute inflammation especially at sites of platelet microthrombi where the concentration of PF4 would be elevated.
Effects of Selenium In Vitro on Human T-Lymphocyte Functions and K-562 Tumor Cell Growth
Authors: Watson Ronald Ross , Moriguchi Satoru , McRae Bart , Tobin Lucia , Mayberry J Carter , Lucas David
Published: January 2018

In vitro E-rosette formation, lymphocyte mitogenesis, and natural killer (NK) cell activity of human blood lymphocytes were strongly inhibited by high concentrations (10−4 M) of sodium selenite, sodium selenate, and selenium dioxide. Lower concentrations (10−5 or 10−7 M) also inhibited E-rosette formation and natural killer cell activity against K-562 tumor cells. Lymphocyte transformation induced by concanavalin A (con A) or pokeweed mitogen (PWM) was also inhibited by all selenium compounds tested, but only at the highest concentrations (10−5 and 10−4 M). There was depression of the total number of viable lymphocytes following incubation with selenium dioxide only at a high concentration (10−4 M). Interferon production was enhanced at lower levels (10−9 to 10−6M) of selenium dioxide while a higher concentration (10−5 and 10−4M) appeared to inhibit its production. The mechanism of inhibition by selenium compounds (10−4 M) is due, in part, to the decrease of viable lymphocytes. It is unclear how other and lower concentrations (10−7 or 10−9 M) of selenium compounds inhibit E-rosette formation, NK activity, or K-562 tumor cell growth.
Induction of Macrophage Growth by Effete Cells
Authors: Yui Satoru , Yamazaki Masatoshi
Published: January 2018

When mouse peritoneal cells in early phase-inflammatory exudates were cultured in vitro, the number of polymorphonuclear leukocytes (PMNs) rapidly decreased, whereas the number of macrophages gradually increased during culture. Macrophage growth was also induced by addition of effete PMNs or their debris to macrophage monolayers. This phenomenon was not restricted to senescent PMNs, because effete spleen cells or effete tumor cells also induced macrophage growth. The activity of these cell debris was stable on heating at 100° and was partially recovered in the lipid fraction. These data suggest that tissue macrophages may proliferate when they scavenge effete PMNs or other host cells and that these cell debris may be important mediators in inducing growth of peripheral macrophages in inflammation and tumors or the normal steady state.
Prostaglandin E and Thromboxane B2 Release by PMA-Induced U937 Cells in Response to C3b
Authors: Schenkein Harvey A
Published: January 2018

The human monocytic cell lines HL-60 and U937 were examined for their ability to release PGE and TxB2 in response to the C3 cleavage fragment C3b. It was found that U937 responds to C3b by releasing both eicosanoids only after a 48 h induction period in the presence of PMA. HL-60 failed to respond. The response of U937 was further studied and characterized. It was found that the continual presence of PMA in culture was not required for the response to C3b, and that by replenishing the culture medium every 48 hr, the response could be maintained for up to 12 days. This contrasts with the previously observed behavior of human monocytes, which fail to respond after 24 hr in culture. Further studies demonstrated that maximal levels of PGE and TxB2 were elicited by 25–50 μg/ml of C3b, and that the kinetics of appearance of these eicosanoids in culture supernatants was very similar to that previously observed for monocytes. Maximal levels of PGE and TxB2 occurred following 24–48 hr in culture. In addition, a monoclonal a-CR1 antibody stimulated PG release by these cells. It is expected that this cell line will provide a model for study of the influence of cell differentiation on the response of human mononuclear phagocytes to C3b.
Cellular Immune Functions of Adults Treated With a Daily, Long-Term, Low Dose of 13-cis Retinoic Acid
Authors: Watson Ronald Ross , Jackson J C , Alberts David S , Hicks Mary Jane
Published: January 2018

The effects of long-term consumption of 13-cis retinoic acid (13-cRA) on cellular immune functions were measured in young, adult volunteers. The retinoid was administered for 9 months at about 0.13 mg/kg/day. The mean 8AM concentrations of 13-cRA ranged between 30 and 60 ng/ml of serum throughout the study. Corticosteroid levels in plasma decreased significantly throughout treatment, declining from 15.2 ug/dL to 9.1 mg/dL (p < 0.05). T-cell mitogenesis stimulated by PHA or A Con A was not significantly affected, although this parameter was slightly depressed during the first 2 months of treatment. The percentage of B-lymphocytes tended to decrease during treatment and returned to normal after cessation of 13-cRA (p < 0.05), while the percentage of T-cells as measured by E-rosette and by fluorescent antibody tagging of surface antigens did not change. The percentage of non T-cells tended to increase slightly during treatment.
Increased Leukocyte Diversity and Responsiveness to B-Cell and T-Cell Mitogens in Cell Suspensions Prepared by Enzymatically Dissociating Murine Lymph Nodes
Authors: Monfalcone Alan P , Szakal Andras K , Tew John G
Published: January 2018

The isolation of viable follicular dendritic cells (FDCs) from murine lymph nodes requires that the nodes be enzymatically dissociated with collagenase and the protease dispase. The present study was undertaken to compare leukocyte populations derived from enzymatically dissociated and mechanically disrupted lymph nodes. We examined cell viability, cell number, cell types, and the proliferative response to the mitogens LPS, PHA, and Con A. Cells were prepared by taking the inguinal, brachial, axillary, and popliteal lymph nodes from one side of the animals and mechanically disrupting them. The corresponding lymph nodes from the other side of the same animals were digested with enzymes. The enzyme-treated lymph nodes yielded substantially more cells and had a higher percentage of viable cells. Over 40% more viable cells were available for cell culture using the enzyme dissociation method. The numbers of FDCs, macrophages, plasma cells, and fibroblasts were clearly increased. The cells dispersed by enzymatic means responded better than the mechanically dispersed cells to both B-cell and T-cell mitogens. This was especially striking in cultures at lower cell densities. We conclude that the method of cell dissociation has a marked effect on the types of viable cells released and available for culture, as well as on the ability of the cells to respond. We believe the cell types released by enzymatic dissociation and their response in culture more accurately reflect conditions in vivo.
Partial Characterization of Protein Kinase C and Inhibitor Activity of Protein Kinase C in Rabbit Peritoneal Neutrophils
Authors: Huang Chi-Kuang , Oshana Scott C
Published: January 2018

Analysis of the cytosol fraction containing protein kinase C activity from rabbit peritoneal neutrophils by DEAE-cellulose chromatography identified protein kinase C activity in the fractions eluted with 0.08 M–0.14 M NaCl and protein kinase C inhibitor activity in the fraction eluted with 0.16 M–0.5 M NaCl. On further analysis by Sephadex G-150 gel filtration, one peak of protein kinase C and two peaks of inhibitor activity were identified. The peak of protein kinase C activity eluted at Ve/Vo 1.6 corresponding to a Stokes radius of 35 å. The first peak of the inhibitor activity eluted at Ve/Vo 1.4 and the second peak of the inhibitor activity eluted at Ve/Vo 2.5. The peak of phosphoprotein phosphatase activity does not coincide with the peaks of inhibitor activity of protein kinase C.
Tetrahydrocannabinol-Induced Suppression of Macrophage Spreading and Phagocytic Activity In Vitro
Authors: Lopez-Cepero Mayra , Friedman Michelle , Klein Thomas , Friedman Herman
Published: January 2018

The effects of tetrahydrocannabinol (THC) on several parameters of macrophage function in vitro were assessed. Delta 9 THC added to cultures of normal mouse peritoneal cells in vitro affected the ability of the cells to spread on glass surfaces and also had some effect on their ability to phagocytize yeast. These effects were dose related. A concentration of 20 μg of THC almost completely inhibited macrophage spreading, but it also decreased viability and the total number of these cells. Doses of 10 or 5 μg of THC also inhibited spreading but had little effect on cell viability or number. A dose of 1.0 μg of THC had some inhibitory effect on spreading and the lowest dose affecting spreading appeared to be about 0.05 μg per culture. Higher doses of THC were necessary to inhibit phagocytosis of yeast particles as determined by direct microscopic examination or use of radiolabeled yeast as the test particles. These results indicate that several readily measured functions of macrophages may be suppressed by THC.
Modulation of Murine Macrophage Metabolism by Glycolipids: Inhibition of LPS-Induced Metabolism by Specific Gangliosides
Authors: Ryan John L , Gobran Laurice , Morrison David C
Published: January 2018

We have investigated the ability of gangliosides isolated from murine brain to modulate macrophage metabolism. LPS elicits a profound increase in glucose consumption by cultured resident macrophages. When highly purified mouse brain gangliosides are added to macrophage cultures, a modest inhibition of baseline glucose utilization occurs. Both disialogangliosides and trisialogangliosides mediate this effect. In the absence of serum, these gangliosides may be toxic to cultured macrophages. GT1b is the only brain ganglioside tested that specifically inhibited LPS-induced macrophage metabolism. Sialic acid appears to be a necessary component of the gangliosides, although it is not sufficient to induce inhibition. Asialoganglioside and free sialic acid have no effect on macrophage metabolism in comparison with intact gangliosides. The inhibition of LPS-induced metabolism may be explained by the ability of LPS to form stable molecular complexes with both disialogangliosides and trisialogangliosides. These experiments also suggest that the ganglioside content of macrophages may influence functional responses of macrophages to exogenous stimuli.
Proliferative Activity of Macrophages in the Greater Omentum of the Mouse in Relation to the Early Postnatal Development of the Vascular Structures
Authors: Dux Kazimierz
Published: January 2018

The rudimental omentum in newborn mice presents itself as a thin, poorly vascularized membrane covering the stomach. Imprints and whole mounted preparations of the omental membrane of mice from seven different inbred strains, aged 1–10 days, were made and sequential morphological changes were described. It was found that omentum of suckling mice contains numerous foci or colonies of mononuclear phagocytes (omental macrophages).

These cells reveal a high proliferative activity when examined 1 hr after exposure of tritiated thymidine. The labeling index in the population of omental macrophages is variable, ranging in individual mice from less than 5% to 25%. This wide variation is a nonrandom phenomenon. It was found in all seven strains of mice and depends mainly on the age of the mouse. Two possible interpretations are as follows: 1) Some interrelation exists between the intensity of the proliferative activity of omental macrophages and the development of vascular and lymphoid structures that are characteristic for the omentum of adult mice. 2) Omental macrophages originate from local precursors that are not dependent on the bone marrow.
Expression of Class II-MHC Antigens by Tumor-Associated and Peritoneal Macrophages: Systemic Induction During Tumor Growth and Tumor Rejection
Authors: Evans Robert , Blake Sharon S , Saffer Jeffrey D
Published: January 2018

It was shown previously that combination therapy of tumor-bearing mice resulted in amplification of antitumor responses at the site of tumor rejection and involved some of the classical components of the immunological network. As a continuation of this work, we now show that amplification of immune responses involves an increase in class II-MHC antigen (Ia) expression both at the tumor site and peripherally in the peritoneal cavity. Of further interest, however, was the observation that there was also an increase in la expression by tumor-associated macrophages (TAM) and peritoneal macrophages (PMs) in mice bearing progressing tumors. In these experiments, Ia expression by TAM and PMs was assessed in C57BL/6J mice bearing the progressing syngeneic MCA/76-9 sarcoma as well as in mice that had received combination therapy consisting of an intraperitoneal injection of cyclophosphamide (CY) and an intravenous injection of tumor-sensitized T lymphocytes (immune cells). When progressing tumors were analyzed, it was seen that by the fourth day after tumor cell implantation, 45–60% TAM expressed la, a level that was sustained throughout the course of tumor growth. The treatment of tumor-bearers with CY had no effect on la expression by TAM. However, the number of la-expressing TAM increased significantly after combination therapy and, as the tumors regressed, reached a peak of up to 100% in the second week after therapy. TAM were shown by Northern blot hybridization to contain mRNA encoding for the laβ chain. When la expression by PMs was assessed, it was seen that during tumor progression there was an increased expression of la over background (from < 10 to about 40%), beginning 9–12 days after tumor cell implantation and continuing for the duration of the experiments. This was not influenced by CY injection. Combination therapy significantly increased the number of PMs expressing la (up to 80%). la expression by PMs could be induced rapidly in vivo by injecting normal B6 mice intraperitoneally (ip) with T cells isolated from tumors induced to regress by combination therapy. PMs isolated up to 15 days after injection were shown to express Ia. Moreover, the ip injection of a mixture of specific MCA/76-9 tumor cells and these tumor-associated lymphocytes resulted in a more rapid rate and a higher level of Ia expression by PMs compared with the level induced by either treatment alone. In contrast to these findings, it was shown that there was little la expression by TAM and PMs from mice bearing the relatively nonimmunogenic sarcoma MCA/77-23. Overall, the data suggest that antitumor immune responses result in a widespread systemic increase in la expression by macrophages.
Phorbol Ester Circumvents the Need for Macrophages as Well as for Mitogenic Lectins in the Stimulation of Lymphocytes With Wheat Germ Agglutinin or the Calcium Ionophores A23187 or Ionomycin
Authors: Grier Charles E. , Mastro Andrea M
Published: January 2018

Numerous biochemical events precede the proliferation of primary lymphocytes stimulated by mitogenic lectins in the presence of macrophages. Various compounds can activate parts of this response. Specifically the tumor-promoting phorbol ester, 12-O-tetradecanoylphorbol-13-acetate, (TPA), can replace the requirement for macrophages, apparently by mimicking the macrophage product interleukin 1 (IL1). Wheat germ agglutinin (WGA), itself a non-mitogenic lectin, is reported to cause a calcium flux, phosphatidylinositol turnover, and enhance interleukin 2 (IL2) synthesis. In spite of these positive responses, WGA inhibits DNA synthesis caused by mitogenic lectins. Nevertheless, in this study, we tested the possibility that together TPA and WGA could complement and bring about DNA synthesis. This prediction turned out to be true. The combination of two non-mitogenic compounds resulted in lymphocyte proliferation. The TPA overcame the inhibitory effects of WGA. Moreover, macrophages were not required. The TPA also synergized with the calcium ionophores A23187 or ionomycin to cause lymphocyte proliferation in the absence of macrophages. WGA and the ionophores together did not cause proliferation, a finding which suggested that they fulfill the same roles. These observations led us to conclude that at least two signals were required for lymphocyte stimulation. One signal caused the mobilization of calcium and the other signal circumvented the need for macrophages or macrophage products possibly by mimicking diacylglycerol, the activator of protein kinase C.
Variations in Macrophage Antigen Phenotype: A Correlation Between Ia Antigen Reduction and Immune Dysfunction During Tumor Growth
Authors: Garner Ronald E , Malick Adrian P , Elgert Klaus D
Published: January 2018

Variable la antigen expression by macrophages (Mφ) was examined during tumor growth by measuring: i) la antigen masking and immunofluorescence by anti-la antibody, ii) accessory cell function in concanavalin A (Con A) and mixed lymphocyte reaction (MLR)-induced T cell proliferation, and iii) Mφ stimulatory function in the MLR. Tumor-induced progressive loss of la antigen expression was shown by immunofluorescence and corroborated by anti-la blockade of MLR stimulatory activity of normal but not tumor-bearing hosts (TBH) splenic Mφ. The TBH splenic Mφ supported Con A-induced proliferation of syngeneic T cells (la antigen-independent) but did not support syngeneic T cell proliferation in the MLR (la antigen-dependent). Irrespective of tissue source, normal and TBH Mφ differed in their MLR stimulatory capabilities. In general, splenic Mφ preparations were better stimulators of allogeneic T cell blastogenesis in the MLR than thioglycollate-elicited peritoneal Mφ. Kinetic studies with TBH Mφ showed a significant progressive loss in MLR stimulatory activity, which was especially pronounced with peritoneal Mφ. Expression of la antigens by normal but not TBH Mφ were diminished by 24-h in vitro plating of the peritoneal Mφ. Indomethacin treatment showed Prostaglandin E2 was not a direct in vitro factor in la antigen-mediated reduction of splenic Mφ MLR stimulatory activity. Taken together, these data delineate a loss of Mφ la antigen expression, resulting in a decrease in la antigen-mediated functional activities during tumor growth.
Changes in Splenic Macrophage Mac Antigen Expression During Tumor Growth: A Kinetic Study of Accessory Cell Function and Antigen-Defined Phenotypes
Authors: Garner Ronald E , Elgert Klaus D
Published: January 2018

Three monoclonal antibodies, anti-Mac-1, -2, and -3, were used to modify accessory cell activity of whole spleen cell (WSC) or splenic adherent cell (SAC) preparations from normal or tumor-bearing hosts (TBH). Ligand activation by anti-Mac-1, -2 and -3 modified normal and TBH WSC lectin responses differentially; the most pronounced effect was with anti-Mac 1, which augmented normal WSC responsiveness, whereas anti-Mac-2 and -3 augmented TBH WSC mitogenesis. Ligand interaction with Mac-2 epitopes resulted in significantly suppressed normal host WSC responsiveness. With complement, anti-Mac-1 and -3 each reduced normal and TBH WSC proliferation. Reconstitution of blastogenesis was obtained by combining Mac-2-depleted with Macs-depleted normal host WSC or by combining Mac-1 -depleted with Mac-2-depleted TBH WSC. To evaluate the role of different types of splenic adherent cells in T cell lectin responsiveness, adherent cells were collected and depleted by antibody plus complement treatment and added back to normal T cells. Removal of TBH SAC indicated the number of Mac-1+ SAC susceptible to lysis increased during tumor growth, whereas those susceptible to anti-Mac-2 and-3 treatment decreased. Removal of Mac-1 + normal host SAC stimulated the supportive accessory function of the remaining SAC. Enhancing accessory cell function diminished after removal of Mac-2+ or Mac-1 + normal and TBH SAC, respectively. T cell responsiveness was increased by adding back combinations of normal host SAC, Mac-1-depleted with Mac-3-depleted SAC or Mac-1-depleted with Mac-2-depleted SAC but not by Mac-2-depleted with Macs-depleted SAC. In contrast, none of the TBH SAC combinations were completely restored in accessory activity. In summary, SAC from normal host demonstrated an accesssory cell function corresponding to a Mac-1- phenotype, which was either replaced or obscured by the predominance of a Mac-1+ phenotype in TBH.
Differences in Fc Receptor Expression Between Small and Large Guinea Pig Monocytes
Authors: Noga Stephen J , Normann Sigurd J , Weiner Roy S
Published: January 2018

Guinea pig monocytes have been separated into subpopulations differing in size, morphology, and cytochemistry. We report now that these peripheral blood monocyte subpopulations differ also in native Fc receptor expression. All freshly isolated small monocytes formed Fc rosettes with sensitized sheep erythrocytes, whereas Fc receptor expression was not detectable on large monocytes. However, Fc receptor expression developed rapidly in culture such that by 48 hours all monocytes were positive. This phenomenon was inhibited by 2-deoxy-D-glucose and cycloheximide, suggesting that Fc receptor expression depended upon glycolysis and protein synthesis. Large monocytes participated in antibody-dependent lysis of sheep erythrocyte targets. That large monocytes lacking Fc receptors were cytolytic in this assay was concordant with expression of Fc receptors during the 16-hour test period. We believe that the large number of lymphocytes present in the Fc receptor-positive small monocyte fraction interfered with their measurement of antibody-dependent cytotoxicity. We conclude that Fc receptor expression on guinea pig monocytes is heterogeneous within the peripheral circulation. These differences quickly diminish in vitro with the expression of Fc receptors by all monocytes. Additionally, antibody-dependent lysis of sensitized erythrocytes by guinea pig mononuclear cells is due to monocytes and not lymphocytes or Kurloff cells.
A New Monoclonal Antibody, TRPM-3, Binds Specifically to Certain Rat Macrophage Populations: Immunohistochemical and Immunoelectron Microscopic Analysis
Authors: Takeya Motohiro , Hsiao Lily , Takahashi Kiyoshi
Published: January 2018

An anti-macrophage monoclonal antibody designated TRPM-3 was produced using thioglycollate-elicited rat peritoneal macrophages as immunogen. By the immunoper-oxidase method, TRPM-3 was found to be specific for certain macrophage populations, such as marginal zone macrophages and marginal metallophils of the spleen, sinus macrophages of the lymph nodes, and omentum macrophages. No epithelial cells, peripheral blood monocytes, granulocytes, or lymphocytes had reacted with TRPM-3. Immunoelectron microscopically, reaction to TRPM-3 was clearly demonstrated on the plasma membrane of splenic marginal zone macrophages, lymphatic sinus macrophages, omentum macrophages, and peritoneal macrophages. Accordingly, TRPM-3 was considered to have recognized the particular membrane antigen expressed by restricted macrophage populations.
Experimental Histoplasmosis in the Beige Mouse
Authors: Patiño Maria Mercedes , Williams Dwight , Ahrens Joan , Graybill John R
Published: January 2018

Mice carrying the beige mutation (bg/bg) on a C57BI/6 background were challenged with Histoplasma capsulatum. bg/bg mice had higher mortality and higher lung tissue fungal counts in their lungs than either bg/ + or C57BI/6 mice challenged with equal inocula. Immunologic studies showed that bg/bg mice developed normal delayed-type hypersensitivity (DTH) reactions to histoplasmin, but had deficient NK cell cytotoxic activity against YAC-1 target cells. Studies of macrophage killing of H. capsulatum in vitro showed that T lymphocytes of either bg/ + or bg/bg mice were able to activate fungal killing by bg/ + but not by bg/bg macrophages. These studies, while not excluding a role for the NK cell, suggest that macrophage dysfunction may be critical in the greater susceptibility of the bglbg mouse and, by extension, that macrophage function is of major importance in host defense against H. capsulatum.
Neutrophils from burn patients are unable to increase the expression of CD11b/CD18 in response to inflammatory stimuli
Authors: Rodeberg David A , Bass Robert C , Alexander J Wesley , Warden Glenn D , Babcock George F
Published: January 2018

Neutrophils (PMNs) from patients with thermal injury are dysfunctional for the CD11b/CD18-dependent functions of diapedesis, chemotaxis, and phagocytosis. The expression of CD11b/CD18 on normal PMNs is increased after an inflammatory stimulus. We proposed that CD11b/CD18 expression on burn patient PMNs would respond abnormally to inflammatory stimuli. PMNs were obtained from nonseptic burn patients during the second week after thermal injury. PMNs from burn patients incubated with lipopolysaccharide (LPS), N-formyl-methionyl-leucyl-phenylalanine, phorbol myristate acetate, or A23187 did not increase the expression of CD11b/CD18 to the same degree exhibited by normal PMNs. This inability to increase CD11b/CD18 was not due to differences in CD14 receptor expression, LPS binding, or factors present in the serum of burn patients. The upregulation of CD35 also was decreased on burn patient PMNs. Western blot analysis revealed decreased quantities of CD11b protein in burn patient PMNs compared with normal control PMNs. The deficiency in CD11b/ CD18 expression after inflammatory stimuli may explain some of the abnormalities observed in burn PMN function.
Leukosialin (CD43) behavior during adhesion of human monocytic THP-1 cells to red blood cells
Authors: Soler M , Merant C , Servant C , Fraterno M , Allasia C , Lissitzky J C , Bongrand P , Foa C
Published: January 2018

To understand the modulation and the behavior of glycocalyx elements during adhesion, we explored one of its components, the CD43 molecule, on human monocytic THP-1 cells exposed to cytokine stimulation and its redistribution during heterotypic adhesion to opsonized erythrocytes. First we demonstrated by immunofluorescence and immunoprecipitation that CD43 is dys-sialylated in monocytic THP-1 cells stimulated by interferon-γ (IFN-γ) and tumor necrosis factor α (TNF-α) and stimulation increased correlated to heterotypic adhesion. CD43 anti-adhesive effect seemed to be related to sialic acid moeties because an increase in adhesion was also induced by sialidase treatment and by monoclonal antibodies recognizing sialic acid-dependent epitopes on CD43. Second, a redistribution of CD43 molecules was observed after adhesion, resulting in the exclusion of CD43 molecules from contact areas as demonstrated by immunofluorescence and by ultrastructural immunogold localization. We therefore demonstrated in monocytic THP-1 cells that some glycocalyx molecules can be modulated by cytokines and redistributed during adhesion. These results support the concept that CD43 can regulate cell interactions.
Effect of neutrophil serine proteinases and defensins on lung epithelial cells: modulation of cytotoxicity and IL-8 production
Authors: Van Watering Sandra , Mannesse-Lazeroms Sylvia P G , Dijkman Joop H , Hiemstra Pieter S
Published: January 2018

Neutrophil accumulation in the lung may contribute to tissue injury as observed in inflammatory diseases. Both oxidative and non-oxidative mechanisms are involved in neutrophil-mediated tissue injury. Non-oxidative mechanisms include the release of neutrophil granule proteins such as the serine proteinases elastase and cathepsin G, and the non-enzymatic defensins. Because stimulated neutrophils are thought to release their products simultaneously, we investigated possible interactions between purified defensins and serine proteinases with respect to induction of cellular injury and their ability to induce interleukin-8 (IL-8) synthesis in cells of the lung epithelial cell line A549. Whereas defensins induced cell lysis, elastase and cathepsin G induced detachment of A549 cells. Co-incubation of elastase and cathepsin G revealed an additive effect on detachment, whereas defensins inhibited serine proteinase-induced detachment. Vice versa, both serine proteinases reduced defensin-induced cell lysis. Furthermore, elastase and cathepsin G prevented defensin-induced IL-8 synthesis. In contrast, no inhibitory interaction between cathepsin G and defensins was observed with respect to their antibacterial activity. The results from this study indicate that, at sites of inflammation, neutrophil-mediated injury might be regulated by interactions between released defensins and serine proteinases.