Effect of lung surfactant on the release of factor increasing monocytopoiesis by macrophages

Upon ingestion of particulate and soluble material, at the site of an inflammation macrophages release the factor increasing monocytopoiesis (FIM), which accelerates the rate of division of the monoblasts and promonocytes in the bone marrow. It is not known, however, whether FIM is released by macrophages present at noninflamed sites. Since FIM is secreted only during phagocytosis and alveolar macrophages ingest surfactant in vivo, the present study was performed to find out whether surfactant induces the release of FIM by alveolar macrophages. Resident alveolar macrophages were found to contain FIM and secrete this factor in vitro in the absence of an introduced phagocytable particle. Resident peritoneal macrophages also contained FIM and released this factor after exposure to surfactant. These findings suggest that in the absence of an inflammatory stimulus in vivo, alveolar macrophages that have ingested surfactant release FIM to maintain the normal production of monocytes in the bone marrow.     

Synergistic myelopoietic actions in vivo after administration to mice of combinations of purified natural murine colony-stimulating factor 1, recombinant murine interleukin 3, and recombinant murine granulocyte/macrophage colony-stimulating factor.

Combinations of low dosages of purified murine hematopoietic colony-stimulating factors (CSFs)--L-cell CSF type 1 (CSF-1), recombinant interleukin 3 (IL-3), and recombinant granulocyte/macrophage CSF (GM-CSF)--were compared with single CSFs for their influence on the cycling rates and numbers of bone marrow granulocyte/macrophage, erythroid, and multipotential progenitor cells in vivo in mice pretreated with human lactoferrin. Lactoferrin was used to enhance detection of the stimulating effects of exogenously administered CSFs. Concentrations of CSFs that were not active in vivo when given alone were active when administered together with other types of CSF. The concentrations of CSF-1, IL-3, and GM-CSF needed to increase progenitor cell cycling rates were reduced by factors of 40-200, 10-50, and 40- greater than 400, respectively; the concentrations needed to increase progenitor cell numbers were reduced by factors of 40-500 (CSF-1), 20-80 (IL-3), and greater than 40- greater than 200 (GM-CSF) when these forms of CSFs were administered in combination with low dosages of one of the other forms of CSFs. The results demonstrate that different CSFs can synergize when administered in vivo to increase the cycling rates and numbers of marrow hematopoietic progenitor cells. These findings may be of relevance physiologically to the regulation of myeloid blood cell production by CSFs.     

Roles of factor increasing monocytopoiesis (FIM) and macrophage activation in host resistance to listeria monocytogenes

The present contribution concerns two aspects of host resistance in overcoming an infection withListeria monocytogenes. One of these aspects is the regulation of monocyte production by the factor increasing monocytopoiesis (FIM), a macrophage-derived factor. Listeria-resistant C57BL/10 mice and Listeria-sensitive CBA mice produce and secrete similar amounts of FIM in response to an inflammation induced by soluble Listeria antigen. However, monocyte precursors in the bone marrow of Listeria-resistant mice react to an injection of serum containing FIM by increased monocyte production, whereas Listeria-sensitive mice are unable to react to this stimulus. The other aspect of host resistance toL. monocytogenes is the activation of macrophages leading to increased bactericidal activity. Macrophages of both mouse strains stimulated first with live BCG and then with PPD, killed ingested Listeria faster than resident peritoneal macrophages did. However, recombinant interferon-, thought to be the most important macrophage-activating factor, did not induce increased listericidal activity in macrophages.Der vorliegende Beitrag befaßt sich mit zwei Aspekten der Abwehrmechanismen des Wirtsorganismus in der Bewältigung von Infektionen durchListeria monocytogenes. Einer der Aspekte ist die Regulation der Monozytenproduktion durch einen Faktor (FIM), der die Monozytopoese erhöht und von Makrophagen gebildet wird. Als Reaktion auf eine durch lösliches Listeria-Antigen induzierte Entzündung bilden und sezernieren Listeria-resistente C57BL/10-Mäuse und Listeria-empfindliche CBA-Mäuse ähnliche Mengen an FIM. Die Monozyten-Vorstufen im Knochenmark Listeria-resistenter Mäuse reagieren hingegen auf eine Injektion von Serum, das FIM enthält, mit einer verstärkten Monozytenproduktion, während Listeria-empfindliche Mäuse nicht in der Lage sind, auf diesen Stimulus zu reagieren. Der andere Aspekt der Abwehr des Wirtsorganismus gegenL. monocytogenes ist die Makrophagenaktivierung, die zu einer Erhöhung der bakteriziden Aktivität führt. Bei beiden Mäusestämmen konnten Makrophagen, die erst mit lebendem BCG und anschließend mit PPD stimuliert wurden, phagozytierte Listerien rascher abtöten als residente Peritonealmakrophagen. Dagegen gelang es mit rekombinantem Gamma-Interferon, das als der wichtigste Makrophagenaktivierungsfaktor angesehen wird, nicht, eine Induktion erhöhter listerizider Aktivität in Makrophagen auszulösen.     

Involvement of ras p21 protein in signal-transduction pathways from interleukin 2, interleukin 3, and granulocyte/macrophage colony-stimulating factor, but not from interleukin 4.

The protooncogene ras acts as a component of signal-transduction networks in many kinds of cells. The ras gene product (p21) is a GTP-binding protein, and the activity of the protein is regulated by bound GDP/GTP. Recent studies have shown that a certain class of growth factors stimulates the formation of active p21-GTP complexes in fibroblasts and that oncogene products with enhanced tyrosine kinase activities have a similar effect on ras p21. We have measured the ratio of active GTP-bound p21 to total p21 in several lymphoid and myeloid cell lines in order to understand the role of ras in the proliferation of these cells. Interleukin 2 (IL-2), IL-3, and granulocyte/macrophage colony-stimulating factor (GM-CSF) enhance the formation of the active p21.GTP, whereas IL-4 has no effect on p21-bound GDP/GTP. These results strongly suggest that ras p21 acts as a transducer of signals from IL-2, IL-3, and GM-CSF, but not from IL-4.     

Effect of macrophage colony-stimulating factor on in vitro osteoclast generation and bone resorption.

To investigate the role of recombinant human macrophage colony-stimulating factor (rhM-CSF) on in vitro bone resorption, two bone explants, each at a different developmental stage, were adopted, namely 1) radii and 2) metatarsals of 17-day-old embryonic mice. At this stage of gestation, bone resorption in the radii is exclusively dependent upon fusion of osteoclast precursors and activation of mature osteoclasts, whereas in metatarsals it is dependent upon the generation of new osteoclasts. rhM-CSF showed no effect in radii, but stimulated 45Ca release in metatarsals, when they were either intact or periosteum stripped in coculture with embryonal liver as a source of hemopoietic progenitors. The rhM-CSF-induced increase in 45Ca release was paralleled by a higher number of osteoclasts. The stimulating effect was found to be in a concentration range between 250-500 U/ml M-CSF. The action of rhM-CSF was blocked by irradiation, indicating that it is dependent upon cell proliferation. These results, thus, show that M-CSF stimulates bone resorption only when it is dependent on generation of new osteoclasts. M-CSF does not appear to have any effect on the activity of mature osteoclasts. The mechanism of action might be direct on osteoclast precursors or indirect on accessory cells influencing osteoclast generation.     

Production of colony-stimulating factor by leukemic leukocytes.

The production of colony-stimulating factor (CSF) by the peripheral blood cells of untreated patients with acute myeloid leukemia (AML) was measured in the agar culture system using normal human bone marrow as the source of colony-forming units (CFUc). CSF production was found to be variable and was related to the morphologic subtype of AML--cells from patients with monocytic leukemia produced normal or large quantities of CSF, while (with one exception) those from patients with myeloblastic leukemia produced little or no CSF. There was a general relationship between CSF production and serum lysozyme levels. Attempts to demonstrate a consistent inhibitory effect exerted by leukemic peripheral blood cells on normal leukopoiesis in vitro were negative. Results instead suggested that the addition to the feeder layer of cells from patients with monocytic leukemia could raise CSF levels above those obtained with normal peripheral blood leukocytes alone, possibly by recruiting additional CFUc from normal marrow.     

Effects of interleukin 3, interleukin 6, and granulocyte colony-stimulating factor on sorted murine splenic progenitor cells.

In order to examine the effect of recombinant growth factors on hemopoietic stem cells, these cells were enriched using wheat germ agglutinin (WGA) and monoclonal antibodies for lineage markers (Lin) such as B220, L3T4, Lyt-2, asialo GM1, Mac-1, and AL-21. Spleen colony-forming units (CFU-S) and in vitro colony-forming units were highly enriched in the fraction of WGA+Lin- spleen cells. To eliminate committed progenitor cells, spleen cells of 5-fluorouracil (5-FU)-treated mice were used. By this treatment, day-8 CFU-S disappeared but day-14 CFU-S were preserved. Day-14 CFU-S were also contained in the fraction of WGA+Lin- cells, which made up about 0.5% of total nucleated spleen cells. Moreover, this fraction contained primitive stem cells that could reconstitute the hemopoiesis of irradiated mice. Sorted WGA+Lin- spleen cells obtained from male 5-FU-treated mice were injected into lethally irradiated female mice. Southern hybridization using a mouse Y chromosome-specific probe showed that the bone marrow, spleen, and thymus of the recipients was reconstituted by male mouse-derived cells. When sorted WGA+Lin- spleen cells of the 5-FU-treated mice were cultured in vitro in the presence of recombinant interleukin 3 (IL-3), interleukin 6 (IL-6), and granulocyte colony-stimulating factor (G-CSF), colony formation was observed only in wells with IL-3, whereas unfractionated spleen cells formed colonies in the presence of IL-3, IL-6, or G-CSF. However, IL-6 but not G-CSF acted synergistically on enriched hemopoietic stem cells in the presence of IL-3. These data suggest that G-CSF or IL-6 did not affect primitive stem cells independently but showed the effect on these cells indirectly or synergistically with IL-3.     

Monocyte-conditioned medium and interleukin 1 induce granulocyte-macrophage colony-stimulating factor production in the adherent cell layer of murine bone marrow cultures

We have studied how production of colony-stimulating factors (CSF) can be induced in murine long-term bone marrow cultures (LTBMC). We found that the adherent cells, but not the nonadherent cells, of LTBMC synthesized and secreted large amounts of CSF upon stimulation with monocyte-conditioned medium (MCM) from the early phase of monocyte culturing. This CSF induced both granulocyte- and macrophage-containing colonies. Interleukin 1 (IL-1) also induced CSF production by the adherent cells, although not to the same extent as MCM. Medium conditioned by E-rosette-positive lymphocytes could not substitute for MCM. CSF production varied in long-term bone marrow cultures less than two weeks old, but thereafter the amount of CSF obtained was relatively independent of the age of the cultures (2-26 weeks). No correlation was found between the content of granulocyte-macrophage colony-forming cells (GM-CFC) in the nonadherent cell fraction of LTBMC and the ability of the adherent cell layer to produce CSF. These results suggest a two-stage process for CSF synthesis. Monocytes produce a factor(s) that, in a second step, leads to bioassayable levels of CSF in the supernatant of adherent cells in LTBMC.     

Decreased atherosclerosis in mice deficient in both macrophage colony-stimulating factor (op) and apolipoprotein E.

To develop a murine model system to test the role of monocyte-derived macrophage in atherosclerosis, the osteopetrotic (op) mutation in the macrophage colony-stimulating factor gene was bred onto the apolipoprotein E (apoE)-deficient background. The doubly mutant (op/apoE-deficient) mice fed a low-fat chow diet had significantly smaller proximal aortic lesions at an earlier stage of progression than their apoE-deficient control littermates. These lesions in the doubly mutant mice were composed of macrophage foam cells. The op/apoE-deficient mice also had decreased body weights, decreased blood monocyte differentials, and increased mean cholesterol levels of approximately 1300 mg/dl. Statistical analysis determined that atherosclerosis lesion area was significantly affected by the op genotype and gender. The confounding variables of body weight, plasma cholesterol, and monocyte differential, which were all affected by op genotype, had no significant additional effect on lesion area once they were adjusted for the effects of op genotype and gender. Unexpectedly, there was a significant inverse correlation between plasma cholesterol and lesion area, implying that each may be the result of a common effect of macrophage colony-stimulating factor levels. The data support the hypothesis that macrophage colony-stimulating factor and its effects on macrophage development and function play a key role in atherogenesis.     

Cloning, sequence, and expression of a human granulocyte/macrophage colony-stimulating factor.

Human granulocyte/macrophage colony-stimulating factor (GM-CSF) is a glycoprotein that is essential for the in vitro proliferation and differentiation of precursor cells into mature granulocytes and macrophages. In this report we have used a mouse GM-CSF cDNA clone to isolate human GM-CSF clones from libraries made from HUT-102 messenger RNA and mitogen-stimulated T-lymphocyte messenger RNA. The human cDNA clones contained a single open-reading frame encoding a protein of 144 amino acids with a predicted molecular mass of 16,293 daltons and showed 69% nucleotide homology and 54% amino acid homology to mouse GM-CSF. One of these cDNA clones was shown to direct the synthesis of biologically active GM-CSF using a yeast expression system. The gene for human GM-CSF appears to exist as a single-copy gene.     

Acetaldehyde induces granulocyte macrophage colony-stimulating factor production in human bronchi through activation of nuclear factor-kappa B.

Acetaldehyde, a metabolite of alcohol and primary mediator of alcohol-induced asthma, causes bronchoconstriction via histamine release from airway mast cells. Acetaldehyde also is found in cigarette smoke and may cause airway inflammation. The purpose of this study was to determine the effect of acetaldehyde on cytokine production and nuclear factor kappa B (NF-kappa B) activation in human bronchial tissues. Human bronchi were prepared from normal parts of lung tissues resected for lung cancer (n = 11). The bronchi were cultured in the presence of 5 x 10(-4) M of acetaldehyde for 24 hours and the concentrations of eotaxin, granulocyte macrophage colony-stimulating factor (GM-CSF), interleukin-5, interleukin-8, and regulated on activation, normal T cells expressed and secreted in cultured supernatants were determined by enzyme-linked immunosorbent assay. Tissues also were immunohistochemically stained for NF-kappa Bp65. Acetaldehyde significantly increased GM-CSF production from human bronchi and nuclear translocation of NF-kappa Bp65 in airway epithelium but had no effects on other cytokines. Our findings suggest that acetaldehyde potentially causes airway inflammation via increased GM-CSF production through nuclear translocation of NF-kappa B.