Effects of human peripheral blood monocytes,monocyte-derived macrophages,and spleen mononuclear phagocytes on Toxoplasma gondii

In vitro effects of human peripheral blood monocytes, peripheral blood monocyte-derived macrophages, and spleen mononuclear phagocytes on Toxoplasma gondii were studied. In almost all instances, over 80% of human monocytes and monocyte-derived macrophages infected with Toxoplasma in vitro destroyed the organism. Degeneration of intracellular Toxoplasma was not due to decreased viability of organisms in the challenge inoculum. Human monocytes did not elaborate into the culture medium substances which altered the capacity of Toxoplasma to survive and replicate within mouse macrophages. The early reduction in intracellular Toxoplasma was not affected by inhibitors of various intracellular processes or by diseases associated with altered cellular immunity (sarcoidosis, infectious mononucleosis, or lymphoma.) The Toxoplasma that remained after 6 hr within human monocytes and macrophages multiplied. This multiplication was observed both microscopically and in a radioassay which detects uptake of [³H]uracil or [³H]deoxyuridine into nucleic acids of intracellular Toxoplasma. Intracellular Toxoplasma in monocytes cultured with poly(I:C) or in monocyte-derived macrophages cultured with lymphokines showed decreased uptake of radiolabeled precursors into nucleic acids of intracellular Toxoplasma. Treatment of monocytes with endotoxin did not alter nucleic acid synthesis of surviving intracellular Toxoplasma. These results suggest that human mononuclear phagocytes in peripheral blood and in tissue (spleen) have the capacity to eliminate a large percentage of the Toxoplasma that they ingest or that invade them. The inhibition of nucleic acid synthesis of remaining Toxoplasma by exposure of monocyte-derived macrophages to lymphokines suggests that lymphocyte products may be important for elimination of the Toxoplasma that remain and multiply within a small proportion of mononuclear phagocytes.     

Mouse mononuclear cell chemotaxis. I. Differential response of monocytes and macrophages.

The chemotactic responses of peripheral blood monocytes and peritoneal macrophages to lymphocyte- and complement-derived stimuli were studied in vitro. Monocyte and macrophage chemotactic responses to these stimuli were different in both dose-response and time course. These differences provide a useful functional marker between these two cell populations.     

Purification and partial characterization of elastase activity from rat alveolar and peritoneal macrophages

Macrophage elastase was purified from conditioned media from alveolar and thioglycollate-elicited peritoneal macrophages. The enzyme was purified to apparent electrophoretic homogeneity by preparative isoelectric focusing after a purification step consisting of low ionic strength dialysis and sequential batch fractionation on DEAE-Sephadex A-50. The proteinase activities isolated from alveolar and peritoneal macrophages showed the same physical and biochemical properties. This fact suggests that the same enzyme activity is present in rat macrophages of two different anatomical sites. The molecular weight and isoelectric point of the enzyme were estimated to be 22,500 and 8.3, respectively. The enzyme, characterized as a metallo proteinase, had elastolytic activity, as well as activity toward Suc-(Ala)3-NA. It is inhibited by o-phenanthroline, chicken ovoinhibitor, and EDTA, but not by phenylmethylsulfonyl fluoride or soybean trypsin inhibitor. The macrophage enzyme possesses biochemical and biophysical properties different from the rat pancreatic and granulocyte elastases (which are serine proteinases), and from the metallo proteinase with elastolytic activity isolated from rat platelets.     

Ultrastructural immunocytochemical localization of lysozyme in human monocytes and macrophages

Summary In order to investigate the mechanism of synthesis and secretion of lysozyme (LZ) by human mononuclear phagocytes, the ultrastructural localization of LZ was studied by a pre-embedding direct immunoperoxidase method. Blood monocytes showed a reaction product for LZ in cytoplasmic granules, whereas cultured monocytes showed the reaction product in phagosomes as well as granules at 5 h of culture and in numerous large granules at 3 days of culture. In Kupffer cells, LZ was present in cytoplasmic granules, vacuoles and phagosomes. Some Kupffer cells showed a positive reaction for LZ in the rough endoplasmic reticulum, perinuclear cisterna and Golgi apparatus. Macrophages in the lymph nodes contained LZ in cytoplasmic granules. Bone marrow macrophages contained numerous phagosomes with electron-dense degradation products of erythrocytes, but the reaction product for LZ could not be clearly identified. The present study demonstrated that LZ is present in the granules of human mononuclear phagocytes and released into phagosomes. An in-vitro culture study, furthermore, demonstrated that macrophages produce LZ-containing large granules distinct from those of monocytes. However, findings that indicate the synthesis and secretion of LZ by cultured monocytes, as suggested previously by other investigators, were not observed in this study.     

Lung macrophages serve as obligatory intermediate between blood monocytes and alveolar macrophages

Alveolar macrophages are a unique type of mononuclear phagocytes that populate the external surface of the lung cavity. Early studies have suggested that alveolar macrophages originate from tissue-resident, local precursors, whereas others reported their derivation from blood-borne cells. However, the role of circulating monocytes as precursors of alveolar macrophages was never directly tested. In this study, we show through the combined use of conditional cell ablation and adoptive cell transfer that alveolar macrophages originate in vivo from blood monocytes. Interestingly, this process requires an obligate intermediate stage, the differentiation of blood monocytes into parenchymal lung macrophages, which subsequently migrate into the alveolar space. We also provide direct evidence for the ability of both lung and alveolar macrophages to proliferate.     

Cadmium-induced depression of the respiratory burst in mouse pulmonary alveolar macrophages, peritoneal macrophages and polymorphonuclear neutrophils.

No profound alteration in the resting O₂ consumption of mouse pulmonary alveolar macrophages, polymorphonuclear neutrophils or peritoneal macrophages incubated in media containing either cadmium chloride or cadmium acetate was observed. However, when heat-killed $\text{P. aeruginosa}$, opsonized in autologous serum, were added to the cell suspension a significant depression in the respiratory burst accompanying the phagocytic event was manifested. The suppression of the respiratory burst appeared to be related to the concentration of cadmium. The possible alteration in the relationship between macrophage microtubule assembly and endocytosis is discussed.     

Demonstration and quantitative analysis of mononuclear phagocytes (monocytes and tissue macrophages) in the sinusoid capillaries of the rat adrenal cortex

Mononuclear phagocytes of the Rat adrenal cortex were identified by light microscopy after injection of Chinese ink or colloidal iron (Imferon 200). They may be found as endothelial, periendothelial or intracapillary cells. The number of mononuclear phagocytes was much greater in the adult than in the young Rat, with a significant peak 3 hrs. after injection of Chinese ink in the young Rat and two peaks, 4 and 12 hrs. after injection in the adult.     

Quantitative studies of phagocytosis by alveolar macrophages

The initial rate of phagocytosis by rabbit alveolar macrophages of paraffin oil emulsions stabilized with albumin was markedly increased by prior incubation of the emulsion with serum. The active component(s) of serum was non-dialyzable and heat-labile and was absent from zymosan-treated serum. Magnesium and calcium were both required for the maximal rate of phagocytosis. At 4 °C or in the presence of 1 mM N-ethylmaleimide, the rate of phagocytosis was less than 2% of the control (37° C) rate. The initial rates of phagocytosis of this emulsion by alveolar macrophages from rabbits injected with Freund's adjuvant were not demonstrably different from those observed with macrophages from normal rabbits. Per of mg of cell protein, polymorphonuclear leukocytes ingested serum-treated emulsion more rapidly than did macrophages, but per cell the rates were not different.     

Albumin catabolism in vitro by cultured peritoneal and pulmonary mononuclear phagocytes

1.

1. Because of claims that albumin is degraded by cells of the reticuloendothelial system the appearance of non-protein-bound ¹²⁵I was measured during incubation of biologically screened ¹²⁵I-labeled rat albumin or heat-denatured ¹²⁵I-labeled rat albumin in cultures containing rat peritoneal or pulmonary mononuclear phagocytes.     

Detection of Fc receptors for IgA on rat alveolar and peritoneal macrophages.

The presence of Fc receptors for IgA on alveolar macrophages was determined by rosette assay and immunogold labeling. IgA-mediated phagocytosis by alveolar macrophages was observed. Results of these assays were compared between rats receiving no treatment and those receiving long-term cortisone administration. Sheep erythrocytes coated with dextran and an IgA monoclonal antibody specific for the alpha 1,3 linkages of dextran bound to 16% of alveolar macrophages. However, peritoneal macrophages did not form rosettes with dextran-IgA-coated erythrocytes. Immunogold labeling by transmission electron microscopy revealed that most Fc receptors for IgA were found on the membrane of pseudopodia of activated alveolar macrophages. Long-term cortisone administration diminished the phagocytosis and phagocytic index of alveolar macrophages, thereby contributing to decreased host resistance to infection (e.g., Pneumocystis carinii pneumonia).     

Sodium-dependent nucleoside transport in mouse lymphocytes, human monocytes, and hamster macrophages and peritoneal exudate cells.

Mouse splenocytes and hamster peritoneal exudate cells (PEC), including macrophages, were shown to contain a predominantly Na(+)-dependent and inhibitor (6-[(4-nitrobenzyl)-mercapto]purine ribonucleoside, NBMPR)-resistant transport system for adenosine and other nucleosides. Adenosine (1 microM) was transported about equally in mouse thymocytes and human monocytes from peripheral blood by a Na(+)-dependent system and the NBMPR-sensitive facilitated diffusion system. Hamster PEC also transported inosine, tubercidin, formycin B, uridine, and thymidine in a NBMPR-insensitive manner. With the exception of formycin B, all nucleosides were phosphorylated intracellularly to varying degree, adenosine being almost fully phosphorylated. During the time course of routine experiments (30 s) formycin B was concentrated twofold over external medium levels (1 microM) without any drop-off in the transport rate. On the basis of metabolic studies it was estimated that uridine and tubercidin were also transported against a concentration gradient. Inosine, guanosine, 2'-deoxyadenosine, tubercidin, formycin B, and the pyrimidines uridine, thymidine, and cytidine (all 100 microM) inhibited transport of adenosine and inosine about 50-100%, while 3'-deoxyinosine showed weak inhibitory action. Transport of thymidine was strongly inhibited by nucleosides except by 3'-deoxyinosine. The Na(+)-dependent, active, and concentration transport system appears to be a feature of many immune-type cells, and its presence offers particular conceptual possibilities for the therapy of infections located in these cells.     

Interaction of murine colony-stimulating factor (CSF-1) with alveolar mononuclear phagocytes

Colony-stimulating factor (CSF-1) was purified from serum-free L-cell-conditioned medium (LCM) and iodinated so that we could study its interaction with murine alveolar macrophages. At 0 °C, the binding of ¹²⁵ICSF-1 to alveolar macrophages reached a stable maximum within 16 h. Under this condition, the binding of ¹²⁵ICSF-1 at various concentrations was saturated at about 3 ng/ml. The binding sites of ¹²⁵ICSF-1 were sensitive to trypsin but not to DNase or RNase treatment. At 37 °C, the trypsin-treated cells regenerated more than 90% of their original binding sites within 12 h. Whereas more than 97% of these alveolar macrophages were phagocytic and esterase-positive, autoradiographic studies showed that only 10–31 % of them were capable of binding to ¹²⁵ICSF-1. These results indicate that the frequencies of CSF-1-binding cells and alveolar macrophage colony-forming cells (AL-CFC) are closely correlated, but no causal relationship has been established.     

Complement proteins and macrophages. 1. Quantitative estimation of factor B produced by mouse peritoneal macrophages

Cobra venom factor was used for the detection of factor B synthesized by mouse peritoneal macrophages. This method was shown to be specific for factor B assay by neutralization by antimouse factor B antibody. The amount of factor B in the culture supernatant, assessed by this method, was found to be dependent on the medium used for cultivation of macrophages. The addition of 25% L cell-conditioned medium to minimal essential medium (LCM-MEM) enhanced the production of factor B and also of lysozyme. Kinetic analysis in LCM-MEM showed that factor B produced by 6 x 10(4) cells/cm2 increased up to 72 hr and reached a plateau at 96 hr. The amounts of factor B and lysozyme produced in LCM-MEM depended upon the number of macrophages. Production of factor B was completely inhibited by 1 microgram of cycloheximide per ml and was restored by its removal.