scholarly journals Modulation of phagosome-lysosome fusion in mouse macrophages.

1981 ◽  
Vol 153 (4) ◽  
pp. 1015-1020 ◽  
Author(s):  
M C Kielian ◽  
Z A Cohn
Keyword(s):  
Concanavalin A ◽  
Immune Serum ◽  
Cross Linking ◽  
Activated Macrophages ◽  
Enzymatic Modification ◽  
Inflammatory Agent ◽  
Proteose Peptone ◽  
Mouse Macrophages ◽  
Control Fusion ◽  
Rate Limiting

A previously described fluorescence assay has been used to characterize factors that modulate phagosome-lysosome (P-L) fusion in mouse macrophages. Fusion was not affected by enzymatic modification or by concanavalin A cross-linking of the plasma membrane or by coating the phagocytic particle with concanavalin A or immune serum. Pretreatment of cells with 10-5-10-4 M colchicine, or treatment immediately after ingestion with 1-10 microgram/ml cytochalasin did not alter P-L fusion; implying that the cytoskeleton does not control fusion in a rate-limiting way. Fusion was strikingly elevated in 5-h cultures of activated macrophages from immune-boosted mice. A lower enhancement was seen in cells activated by proteose-peptone, a nonspecific inflammatory agent.

scholarly journals Extracellular cytolysis by activated macrophages and granulocytes. I. Pharmacologic triggering of effector cells and the release of hydrogen peroxide.

1979 ◽  
Vol 149 (1) ◽  
pp. 84-99 ◽  
Author(s):  
C F Nathan ◽  
L H Brukner ◽  
S C Silverstein ◽  
Z A Cohn
Keyword(s):  
Trypan Blue ◽  
Response Curve ◽  
Target Cells ◽  
Activated Macrophages ◽  
Lymphoma Cells ◽  
Effector Cells ◽  
Proteose Peptone ◽  
Standard Assay ◽  
Different Types ◽  
J774 Cells

Lymphoma cells were rapidly lysed by activated macrophages and granulocytes in the presence of PMA. Release of 51Cr from lymphoma cells correlated closely with their destruction as viewed by scanning electron microscopy, and with reduction in the number of trypan blue-excluding cells. The standard assay involved 51 Cr release measured at 4.5 h, but injury appeared to be complete in 1 h. Of eight different types of effector cells tested, only those releasing abundant H2O2 in response to PMA were effective, that, is BCG-, C. parvum-, or casein-activated macrophages, or thioglycollate-elicited granulocytes. Normal macrophages, J774 cells, or macrophages elicited with thioglycollate broth or proteose-peptone were ineffective. BCG-activated macrophages and granulocytes caused 50% specific release of 51Cr from P388 lymphoma cells at E:T ratios between 1.4 and 4.5, and from mouse erythrocytes at E:T ratios of 0.017 to 0.025. 10 types of target cells varied widely in their susceptibility to lysis by reagent H2O2, with one-half maximal lysis occurring at H2O2 concentrations ranging from 3.63 X 10(-6) M to 3.85 X 10(-5) M. Effector cells were expected to generate approximately that much H2O2 during the period of injury. Susceptibility of the target cells to lysis by PMA-triggered granulocytes correlated closely with their sensitivity to H2O2 (r = 0.98). The membrane-active agents LPS and digitonin, which did not trigger H2O2 release, did not trigger cytotoxicity. The dose-response curve for triggering of H2O2 release by PMA was identical to that for triggering cytotoxicity. These results provided strong circumstantial evidence for the importance of H2O2 in extracellular cytolysis by activated macrophages and granulocytes when pharmacologically triggered.

Stimulation by autologous serum preheated at 66 °C of the incorporation of [3H]uridine by cultured lymphocytes: comparison with stimulation by concanavalin A

1977 ◽  
Vol 55 (3) ◽  
pp. 215-222 ◽  
Author(s):  
C. M. David ◽  
D. R. Forsdyke
Keyword(s):  
Concanavalin A ◽  
Maximum Velocity ◽  
Autologous Serum ◽  
Rabbit Serum ◽  
Con A ◽  
Rate Limiting Step ◽  
Inhibitory Molecule ◽  
Lymph Node Cells ◽  
Stimulatory Factor ◽  
Rate Limiting

Rabbit serum, preheated at 66 °C for 30 min, produced a stimulation of the incorporation of [3H]uridine by cultured autologous lymph-node cells similar to that caused by concanavalin A (Con-A). However, whereas the percentage stimulation by Con-A increased with time, that by serum preheated at 66 °C (66 °C-serum) reached a peak after 3–4 h and then declined. The decline was greater at higher cell concentrations. Isotope-dilution studies showed that stimulation at 3 h by 66 °C-serum or by Con-A reflected an increase in the maximum velocity of the rate-limiting step for incorporation of [3H]uridine and not a decrease in the pool of uridine and (or) uridine competitors. Experiments in which serum concentration and the relative proportions of serum preheated at 66 °C and serum preheated at 37 °C were varied, suggested that preheating serum at 66 °C removes an inhibitory factor and exposes a stimulatory factor. The stimulatory activity of 66 °C-serum was not dialysable. The results are compatible with a model which requires that lectins activate cultured lymphocytes by influencing the distribution of an inhibitory molecule (perhaps α2-macroglobulin) between the cell surface and the culture medium.

Interaction of Alveolar Macrophages with Nocardia asteroides : Immunological Enhancement of Phagocytosis, Phagosome-Lysosome Fusion, and Microbicidal Activity

1980 ◽  
Vol 30 (2) ◽  
pp. 578-587
Author(s):  
Carole Davis-Scibienski ◽  
Blaine L. Beaman
Keyword(s):  
Lymph Node ◽  
Alveolar Macrophages ◽  
Cell Damage ◽  
Immune Serum ◽  
Nocardia Asteroides ◽  
Bacterial Cells ◽  
Activated Macrophages ◽  
Additional Increase ◽  
Lining Material ◽  
Lymph Node Cells

Normal and specifically activated rabbit alveolar macrophages were infected in vitro with Nocardia asteroides GUH-2. In the presence of serum from normal rabbits, no significant differences were noted between normal and activated alveolar macrophages with respect to phagocytosis, incidence of phagosomelysosome fusion, or nocardicidal activity. However, all of these macrophage functions were enhanced by various immunological components. Serum from immunized rabbits enhanced phagocytosis of nocardial cells by activated macrophages, and there was an additional increase in phagocytosis observed when alveolar lining material was present. Complement had no effect on the ability of the macrophages to phagocytize nocardial cells. The greatest percentage of organisms phagocytized was observed when specifically primed lymph node cells, alveolar lining material, and serum from immunized rabbits were present in the incubation medium. N. asteroides GUH-2 inhibited phagosome-lysosome fusion in normal macrophages in the presence of serum from normal rabbits. However, addition of serum from immunized rabbits or the addition of specifically primed lymphocytes increased the amount of phagosome-lysosome fusion, whereas complement had no effect on this fusion process. Nocardial viability was not reduced when either normal or activated macrophages were infected with bacteria in the presence of normal serum, immune serum, or alveolar lining material. However, specifically activated macrophages incubated with primed lymph node cells obtained from immunized rabbits were able to both decrease the number of viable organisms recovered and to increase the incidence and extent of bacterial cell damage. The greatest number of organisms were killed by specifically activated macrophages when the bacterial cells were incubated with primed lymph node cells suspended in immune serum and alveolar lining material. These results indicate that activated macrophages alone are not sufficient to kill ingested N. asteroides GUH-2 and that specifically primed lymphocytes are important in host resistance to nocardial infections.

scholarly journals Concanavalin A binding to human erythrocytes leads to alterations in properties of the membrane skeleton

1987 ◽  
Vol 241 (2) ◽  
pp. 521-525 ◽  
Author(s):  
S M Gokhale ◽  
N G Mehta
Keyword(s):  
Concanavalin A ◽  
Membrane Skeleton ◽  
Cross Linking ◽  
Dependent Manner ◽  
Con A ◽  
Normal Cells ◽  
Skeletal Protein ◽  
Low Ionic Strength ◽  
Dose Dependent ◽  
Resistance To Heat

Three properties related to the erythrocyte membrane skeleton are found to be altered after the binding of concanavalin A (Con A) to erythrocytes or their isolated membranes. Con A binding to normal erythrocytes imparts resistance to heat (49 degrees C)-induced fragmentation of the cells. The fragmentation, due to denaturation of spectrin at 49 degrees C, is prevented by Con A in a dose-dependent manner, but levels off at concentrations of Con A in excess of 100 micrograms/ml. The binding of Con A to ghosts isolated from normal, trypsin- or Pronase-treated cells prevents (completely or substantially) the elution of the skeletal protein complex when the membranes are extracted under low-ionic-strength conditions in the cold. The Con A-agglutinated membranes of trypsin- and Pronase-treated, but not normal, cells show cross-linking of skeletal proteins and band 3 with dimethyl adipimidate, a 0.86 nm (8.6 A)-span bifunctional reagent. The extent of cross-linking is greater in the Pronase-treated membrane than in the less-agglutinable trypsin-treated membranes. The results show that, after Con A has bound, rearrangements occur in the membrane that alter properties of the skeletal proteins. Additionally, redistribution of the skeletal proteins and the Con A receptor occurs in the lectin-agglutinated membranes.

scholarly journals Differential Use of Chondroitin Sulfate to Regulate Hyaluronan Binding by Receptor CD44 in Inflammatory and Interleukin 4-activated Macrophages

2011 ◽  
Vol 286 (22) ◽  
pp. 19179-19190 ◽  
Author(s):  
Brian Ruffell ◽  
Grace F. T. Poon ◽  
Sally S. M. Lee ◽  
Kelly L. Brown ◽  
Sie-Lung Tjew ◽  
...  
Keyword(s):  
Chondroitin Sulfate ◽  
Cell Surface Receptor ◽  
Interleukin 4 ◽  
Mouse Bone Marrow ◽  
Activated Macrophages ◽  
Cd44 Expression ◽  
Mouse Macrophages ◽  
Inflammatory Stimuli ◽  
Anti Inflammatory ◽  
Hyaluronan Binding

CD44 is a cell surface receptor for the extracellular matrix glycosaminoglycan hyaluronan and is involved in processes ranging from leukocyte recruitment to wound healing. In the immune system, the binding of hyaluronan to CD44 is tightly regulated, and exposure of human peripheral blood monocytes to inflammatory stimuli increases CD44 expression and induces hyaluronan binding. Here we sought to understand how mouse macrophages regulate hyaluronan binding upon inflammatory and anti-inflammatory stimuli. Mouse bone marrow-derived macrophages stimulated with tumor necrosis factor α or lipopolysaccharide and interferon-γ (LPS/IFNγ) induced hyaluronan binding by up-regulating CD44 and down-regulating chondroitin sulfation on CD44. Hyaluronan binding was induced to a lesser extent in interleukin-4 (IL-4)-activated macrophages despite increased CD44 expression, and this was attributable to increased chondroitin sulfation on CD44, as treatment with β-d-xyloside to prevent chondroitin sulfate addition significantly enhanced hyaluronan binding. These changes in the chondroitin sulfation of CD44 were associated with changes in mRNA expression of two chondroitin sulfotransferases, CHST3 and CHST7, which were decreased in LPS/IFNγ-stimulated macrophages and increased in IL-4-stimulated macrophages. Thus, inflammatory and anti-inflammatory stimuli differentially regulate the chondroitin sulfation of CD44, which is a dynamic physiological regulator of hyaluronan binding by CD44 in mouse macrophages.

Concanavalin A receptors and capping in control and activated macrophages

1983 ◽  
Vol 15 (1) ◽  
pp. 59-69 ◽  
Author(s):  
K. Donaldson ◽  
J. M. G. Davis ◽  
K. James
Keyword(s):  
Concanavalin A ◽  
Activated Macrophages ◽  
Concanavalin A Receptors

scholarly journals THE CYTOTOXIC EFFECT OF MOUSE MACROPHAGES ON SYNGENEIC AND ALLOGENEIC ERYTHROCYTES

1973 ◽  
Vol 137 (3) ◽  
pp. 807-820 ◽  
Author(s):  
H. Melsom ◽  
R. Seljelid
Keyword(s):  
Cytotoxic Effect ◽  
Peritoneal Macrophages ◽  
Target Cells ◽  
Activated Macrophages ◽  
Con A ◽  
Osmotic Effect ◽  
Prolonged Cultivation ◽  
Cytotoxic Reaction ◽  
Mouse Macrophages ◽  
Mouse Peritoneal Macrophages

A cytotoxic effect of mouse peritoneal macrophages against syngeneic and allogeneic erythrocytes was demonstrated by isotope release and release of hemoglobin. The cytotoxic effect was dependent on the contact between viable, activated macrophages and target cells. Activation was accomplished by prolonged cultivation of macrophages and by the presence of Zn++ and Con-A. Immunization did not prove necessary. Morphological observations as well as experiments with various salt concentrations indicate that the cytotoxic reaction may involve some kind of osmotic effect upon the target cells.

scholarly journals Kinetic evidence for a common mechanism of capping on lymphocytes

1982 ◽  
Vol 204 (1) ◽  
pp. 229-237 ◽  
Author(s):  
Anthony N. Corps ◽  
James C. Metcalfe ◽  
Tullio Pozzan
Keyword(s):  
Concanavalin A ◽  
Cytochalasin B ◽  
Specific Effect ◽  
Common Mechanism ◽  
Cross Linking ◽  
Detectable Effect ◽  
Membrane Flow ◽  
Surface Immunoglobulin ◽  
Fixed Times ◽  
Concanavalin A Receptors

1. Differences in the rates at which ligands cap various receptors on the same cells, and their sensitivity to various drugs, have been interpreted as evidence that there are distinct mechanisms for ‘fast’ and ‘slow’ cap formation. We have examined the factors which determine the rate of cap formation of three receptors on mouse splenic lymphocytes or thymocytes, and compared the effects of cytochalasin B or colchicine under conditions where the different receptors cap at similar rates. 2. When surface immunoglobulin, concanavalin A receptors, or θ antigen are induced to cap at their maximal rates by appropriate concentrations of one or more cross-linking ligands, the half-time for maximal capping of each receptor population is between 1.5 and 3.0min at 37°C. Slower rates of cap formation are obtained by using non-optimal concentrations of the cross-linking ligands. 3. When the three receptors were induced to cap at similar rates (either maximal or slower), 10μm-cytochalasin B caused a similar decrease in the rate of cap formation for each receptor, without affecting the eventual extent of capping. At comparable capping rates on control cells, colchicine (10μm) increased the rate of cap formation for surface immunoglobulin and concanavalin A receptors to a similar extent, without affecting the eventual extent of cap formation. In contrast, colchicine had no detectable effect on the capping of θ antigen. 4. From these results, we conclude that there are no intrinsic differences in the rates at which different receptors can be induced to cap that can be used to diagnose differences in their mechanisms of cap formation. The observation that ligand concentration and the drugs acting on the cytoskeleton generally affect the rate but not the extent of cap formation accounts for the wide variation in reported effects of the drugs on cap formation measured at fixed times. The receptor-specific effect of colchicine on surface immunoglobulin and concanavalin A receptors, but not θ antigen, is not readily compatible with models of cap formation which depend on lipid or membrane flow.

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