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Macrophages (M phi s) undergo a physiological response known as the macrophage disappearance reaction (MDR) in response to certain stimuli in the peritoneal compartment. The types of stimuli that can cause the MDR, the relationship of the MDR to the host immunological response, and the possible role of the MDR in M phi activation are reviewed. The data indicate that the MDR occurs in response to both acute nonspecific inflammatory and specific immune delayed hypersensitivity processes and that the MDR may play an important role in M phi activation.

Resident peritoneal macrophages (M phi) were labeled in situ by intraperitoneal (i.p.) injection of the green fluorescent cell tracking dye PKH-1. After immunofluorescence staining with M phi specific monoclonal antibodies (Mabs) and phycoerythrin (PE) second antibody, the resident M phi were labeled with both the green dye and red Mab label, while recruited M phi were labeled only with the red Mab tag. These populations were distinguished by two-color flow cytometry. PKH-1 labeled resident peritoneal M phi were followed for 1-49 days in mice that received no further treatment (steady state). Dye labeled M phi were still detectable after 49 days in vivo, although their green fluorescence intensity had decreased steadily over time. The decrease in dye intensity was limited to M phi, as the fluorescence intensity of PKH-1 labeled peritoneal lymphocytes did not change. Resident M phi populations were clearly separated from recruited M phi by the intensity of their staining with PKH-1 for up to 28 days. No decrease in the number of resident (dye labeled) peritoneal M phi was observed over 1-28 days. These data indicate that resident peritoneal M phi were not replaced by recruited blood monocytes in the steady state.

The effect of inhaled ammonium sulfate on benzo[a]pyrene carcinogenesis in the lungs of Syrian golden hamsters was studied. Exposure to ammonium sulfate at an airborne concentration 20 times average United States ambient levels resulted in a significant depression (p less than 0.05) of benzo[a]pyrene carcinogenesis in the first 6 mo of the study. However, at 2 yr, the termination of the study, there were no differences in cancer incidence between groups receiving benzo[a]pyrene and benzo[a]pyrene plus ammonium sulfate. In addition, at the concentration studied, inhaled ammonium sulfate did not significantly increase the incidence or severity of pneumonitis or pulmonary fibrosis in the hamster. However, this inhalation did increase the incidence of emphysema but not the severity. The decreased incidence of cancer during the first 6 mo of this study in animals receiving both benzo[a]pyrene and ammonium sulfate suggests that interaction between sulfate and benzo[a]pyrene does occur, but is insufficient to afford long-term protection against the development of cancer. No enhancement of carcinogenesis by benzo[a]pyrene occurs in the presence of inhaled sulfate.

Structural and functional characteristics of Fc receptors for IgG (Fc gamma) on human neutrophils were examined with two monoclonal antibody probes specific for the Fc gamma receptors, Leu 11b and 3G8. To determine the distribution, density, and membrane mobility of the Fc gamma receptor, we used immunogold staining techniques, flow cytometry analysis, and fluorescence microscopy. Both 3G8 and Leu 11b inhibited several cell functions, thereby depicting the regulatory role of the Fc gamma receptor in mediating neutrophil activities. Among the functions studied were release of lysosomal enzymes, release of superoxide anion (O2-), and Fc-dependent rosette formation and phagocytosis. The densities of Fc gamma determinants recognized by Leu 11b and 3G8 on cells from a patient with chronic myelogenous leukemia were less than the density of epitopes on neutrophils from a normal individual. Taken together, the detailed analysis of physical and functional aspects of the Fc gamma receptor on neutrophils described in this study serve as a model for further assessment of the use of Fc gamma phenotyping of cells as a diagnostic tool.

The kinetics of macrophage (M phi) recruitment to the peritoneum following the induction of acute inflammation by thioglycollate broth (TG) was evaluated after prelabeling resident M phi with the fluorescent cell tracking dye, PKH-1. Most of the PKH-1-labeled resident M phi disappeared from the recoverable peritoneal cell population within the first hour after injection of TG. This disappearance coincided with the inflammatory influx of neutrophils (PMNs) and was sustained for at least 5 days after administration of TG, although the PMN number had returned to resident levels by this time. PKH-1-labeled peritoneal M phi were observed again in most animals at 7 days after injection of TG. The number of labeled M phi recovered at 7 days was approximately twice the number of resident peritoneal M phi in control animals which did not receive the TG broth. These additional M phi may include progeny of either the resident M phi or other local M phi precursors, such as omental M phi, which were labeled by the PKH-1 injection.

Technical advancements in computer hardware and the associated computerized architectures allow the design and fabrication of dedicated, modestly expensive computerized workstations to aid humans in complex decision-making tasks. We refer to these as "Intelligent Workstations" because they accommodate the hypothesis-driven reasoning ability of the user into the system. Concomitant with the rapid developments in computers, diagnostic and therapeutic technologies related to cell analysis have undergone growth. Combining these technologies requires careful formulation and analysis of the diagnostic model involving image cytometry. Here we present a model of the intelligent workstation for image cytometry and illustrate its advantages in cell-surface phenotyping. Combining cell morphology with cell-surface phenotyping has clear diagnostic utility that no other widely available methods can supply. Although the reported results are based on samples of peripheral blood, cell suspensions of disaggregated cells can also be analyzed.

A novel method for labeling resident peritoneal macrophages (M phi) by injection of a dye into the peritoneal cavity is described. The dye, which fluoresces green, is selectively taken up by the resident M phi. Dye labeled cells can be further characterized by labeling of cell surface antigens with monoclonal antibodies (Mabs) and phycoerythrin conjugated second antibody. After such labeling with the Mabs F4/80 or Mac 1 the resident M phi were labeled by both the green dye and the red Mab markers, while recruited M phi or neutrophils were labeled with just the red Mab; the two populations of cells were readily distinguished by two-color flow cytometry. This technique enabled identification of resident and recruited M phi in each animal without the use of radioisotopes, irradiation, or bone marrow ablation. Sufficient numbers of cells can be analyzed from each animal so that individual animals could be evaluated. We found no adverse effects of this labeling technique on expression of cell surface antigens or M phi mediated cytotoxicity. We did find evidence that the i.p. injection induced a mild inflammation in the peritoneal cavities of animals injected with either the dye or the balanced salt solution vehicle. Examination of the intracellular staining pattern indicated that the label rapidly sequestered in the cytoplasm of the M phi, possibly in the lysosomes. Dye solubility studies showed that the dye was partially soluble at the concentration used for in vivo labeling. We hypothesize that the M phi labeling occurred by a combination of phagocytosis of dye aggregates and endocytosis of labeled plasma membrane.

An automated method of sulfate analysis is described, which can detect sulfate concentrations as low as 2.5 mug per ml water. The assay is based on the reaction of sodium rhodizonate and barium forming a colored complex. Sulfate quantitavely interferes with this reaction. The assay is reproducible in the range of 2.5 to 30 mug sulfate per ml water. This method conveniently and accurately measures water soluble sulfate filtered from the air, and is especially useful in assaying samples containing microgram quantities of sulfate from experimental inhalation apparatus.