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Flagellates of the genus Leishmania are obligate intracellular parasites of vertebrates including man. The microorganisms reside and multiply inside the phagolysosomes of cells of the mononuclear phagocyte lineage. We here report on the spontaneous leishmanicidal activity exerted extracellularly by immature cells of the mononuclear phagocyte lineage. Highly purified, bone marrow-derived macrophage precursor cells displayed a strong spontaneous leishmanicidal activity already at very low effector/target rations (3:1, 6:1). This leishmanicidal activity was effective against both promastigotes and amastigotes as targets. The cytotoxic effect was evident within 4 h and maximal after 12 h of effector-target organism cocultivation, as determined by a radiolabel-release assay. An intimate cell-cell contact seemed necessary for the parasites to be killed.

In an open randomised trial 3 regimen of prophylaxis for deep vein thrombosis were compared in 105 patients undergoing elective hip arthroplasty. One part of the patients received one injection daily of a low molecular weight heparin, another part was treated with one injection of a combination containing a different type of low molecular weight heparin + dihydroergotamine per day and a third part of the patients received 3 applications of 5000 IU unfractionated heparin per day. The frequency of DVT determined by the 125iodine-labelled fibrinogen uptake test and phlebography was 22.9% in patients receiving the combination and 50.0% in those receiving the low-molecular weight heparin alone. The difference was statistically significant with 2 p = 0.03. The incidence of DVT in patients receiving unfractionated heparin was 30.6%. No difference in the development of bleeding complications was found between the three treatment groups.

The potential of the resident murine Kupffer cell to be cytotoxic in extra- and intracellular killing systems in vitro was investigated. Kupffer cells exerted no spontaneous cytotoxicity but were readily susceptible to activation with lymphokines. Such activated Kupffer cells very efficiently killed extracellular P815 cells and intracellular Leishmania spp. parasites. Kupffer cells could be induced to proliferate in vitro under the influence of colony-stimulating factor (D.-M. Chen, H.-S. Lin, P. Stahl, and R. Stanley, Exp. Cell Res. 121:103-109, 1979). Kupffer cell-derived macrophages cultured in vitro were identical to their liver-derived progenitors in terms of macrophage-specific surface antigens and with respect to extra- and intracellular cytotoxicity. These results demonstrated that Kupffer cells have a strong self-renewing potential and that essential Kupffer cell properties like antigenic determinants and cytotoxic potential remained stable throughout the replicative process. The possibility of Kupffer cell self-renewal in the intact organism is also discussed.

Purified polysaccharides (EPS) prepared from the plant Echinacea purpurea are shown to strongly activate macrophages. Macrophages activated with these substances develop pronounced extracellular cytotoxicity against tumor targets. The activation is brought about by EPS alone and is independent of any cooperative effect with lymphocytes. Also the production and secretion of oxygen radicals and interleukin 1 by macrophages is increased after activation with EPS. Cells of the macrophages lineage seem to be the main target for the action of these polysaccharides. EPS has no effect on T lymphocytes. B lymphocytes show a comparatively modest proliferation after incubation with E. purpurea EPS. Thus, these compounds, which are at least in tissue culture completely nontoxic, may be suited to activate in vivo cells of the macrophage system to cytotoxicity. They may therefore be of relevance in tumor and infectious systems.

Bacteriophage phi X174 viral strand DNA molecules shorter than genome length found late in the infectious cycle in Escherichia coli were 5' end labeled with 32P. Hybridization of the 32P-labeled molecules to restriction enzyme fragments of phi X replicative form DNA revealed an excess of phi X molecules whose 5' ends mapped in HaeIII fragments Z3 and Z4 in comparison with fragments Z1 and Z2. This suggests that initiation of phi X174 viral strand DNA synthesis may occur at internal sites on the complementary strand. There are several appropriately located sequences that might serve as n' (factor Y) recognition sequences and thereby facilitate discontinuous synthesis of the viral strand.

We reported earlier that dephosphorylated nascent phi X174 viral strand DNA molecules were less extensively degraded from the 5' end by spleen exonuclease than were non-nascent molecules. Experiments described here revealed that the insensitivity to the 5'-OH end-specific nuclease was more evident among the longer molecules in the population than among the shorter, all of the molecules being less than unit length in size. The smallest molecules in the population were about as sensitive to the enzyme as the control molecules and hence must possess unblocked 5'-terminal nucleotides. Degradation of the nascent DNA with the 3' end-specific snake venom phosphodiesterase revealed only a small enrichment for [3H]thymidine near the 3' end, seemingly insufficient to account completely for the apparent insensitivity of the longer molecules to spleen exonuclease. When the nascent molecules were isolated without the use of proteolytic enzymes, some pronase-sensitive material was found associated with the DNA, particularly the longer molecules. We suggest that the resistance of the longer nascent (pronase-treated) molecules to spleen exonuclease occurs because they have remnants of the viral gene A or A* protein covalently bound to the 5' end.

Precursor cells for NK activity, present in the light fraction of fresh mouse bone marrow, were cultivated in vitro in the presence of either CSF-1, IL-2, or a combination of both factors. In the presence of only CSF-1, strong proliferation was induced. Cells quickly passed the macrophage precursor stage and matured to typical macrophages. Neither granula formation nor NK activity were induced. Under culture conditions with only IL-2 NK activity had developed after 3 d, however, no significant proliferation occurred. In the presence of both factors strong proliferation was induced, and concomitantly, granula formation and NK activity developed. Apparently, proliferation depended on CSF-1 and granula formation, and NK cytotoxicity was induced by IL-2. When proliferating cells with strong anti-YAC-1 activity from a culture in CSF-1 plus IL-2 were further cultivated in only IL-2, the content of granula further increased, whereas proliferation gradually stopped. In contrast, when these cells from CSF-1 plus IL-2 culture were further cultivated in only CSF-1, granula disappeared and NK activity was lost, whereas sustained proliferation and differentiation to macrophages occurred. Only under culture conditions with both factors were proliferation and NK activity both maintained. More than 90% of cells from a 3-d culture in CSF-1 plus IL-2 expressed the NK 1.1. marker, whereas F4/80 was only marginally detected by FACS analysis. After two further days in culture, 70% of the cells expressed F4/80 and 60% coexpressed NK 1.1. and F4/80. By setting the size scatter in order to gate for large granular cells, a population was obtained with 100% coexpression of NK1.1. and F4/80. The data indicate that early cells of the macrophage lineage can develop into different functional and morphological directions depending on the varying influence of IL-2 and CSF-1.

Mature mononuclear phagocytes have been receiving much attention as effectors of spontaneous candidacidal activity, although with controversial results due to differences in the effector populations and the methods used in different laboratories. We here systematically compare the fungistatic activity of immature and mature cells of the murine macrophage series. The results show that nonadherent, nonphagocytic precursor cells (isolated either [90% purity] from bone marrow liquid cultures or from the organs of mice in which inflammatory conditions had been elicited in vivo) exerted a strong extracellular candidastatic activity. In contrast, mature macrophages, either obtained from different anatomical areas (spleen, liver, lung, peritoneal cavity) or matured in vitro from the precursor populations, displayed striking heterogeneity in their ability to inhibit the growth of Candida albicans, depending on the anatomical site they were derived from. Lymphokine activation did not alter the fungistatic pattern of the untreated cells. The different macrophage populations behaved very differently also in the production of reactive oxygen intermediates (ROI) in response to phagocytosis of C. albicans. The amounts of ROI generated, however, showed no correlation with candidastatic ability. Low levels of candidastatic activity exerted by resident peritoneal macrophages (good ROI producers) were inhibited by catalase, whereas high levels of growth inhibition by Kupffer cells (poor ROI producers) after 8 h of assay were hardly influenced by the enzyme. Our data suggest the existence of two different effector mechanisms in macrophage-mediated C. albicans growth inhibition, a rather inefficient ROI-dependent one, and a second, very efficient oxygen-independent mechanism. The implications of these findings are discussed.