The contribution of interstitial macrophages (IM) to lung defense, homeostasis, and pathophysiology is for the most part unknown. Studies on this cell type are difficult because they are not readily accessible in large numbers or in high purity. In the present work, various nonenzymatic and enzymatic methods were compared with the aim of isolating and characterizing pure populations of lung IM. The results of our studies demonstrate that most procedures currently used to isolate IM yield subpopulations of alveolar macrophages (AM) or IM highly contaminated by AM and granulocytes. We found that lavage of the lung yielded only one half of the total AM present in the tissue. The remainder of the AM could only be obtained by extensive washing of cut and disaggregated lung tissue, which is considered by some investigators to be an effective procedure for IM isolation. According to our results, cells recovered by lavage and washing of cut and disaggregated lung tissue were morphologically and histochemically identical, were strongly positive for nonspecific esterase, highly phagocytic, and appeared to represent subpopulations of AM with apparently varying degrees of adherence to the alveolar walls. We also found that IM could be obtained in high purity by sequential digestion of the remaining lung tissue with 60 and 175 IU/ml of collagenase followed by selective adherence. Digestion of the tissue with 60 IU/ml of collagenase resulted in a highly enriched population of granulocytes and also reduced their contamination in the IM population. The resulting IM were distinct from AM by morphology and histochemistry. Like AM, these cells displayed Fc receptor-mediated phagocytosis.(ABSTRACT TRUNCATED AT 250 WORDS)

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Nitric oxide is a highly reactive mediator released in the liver by hepatocytes, Kupffer cells and endothelial cells during endotoxin-induced inflammation. In this study we determined whether Ito cells also produce nitric oxide after exposure to endotoxin. For induction of endotoxemia, rats were injected intravenously with Escherichia coli lipopolysaccharide (2.5 mg/kg). Ito cells were isolated from the animals 48 hr later by means of in situ perfusion of the liver with protease and collagenase followed by purification on an arabinogalactan gradient. Ito cells from untreated and endotoxemic rats were found to produce low levels of nitric oxide in response to interferon-gamma. In both cell types, this response depended on L-arginine and was blocked by NG-monomethyl-L-arginine, a specific nitric oxide synthase inhibitor. Cells from rats treated with endotoxin produced significantly more nitric oxide than did cells from untreated animals; this was due, at least in part, to increased expression of protein for an inducible form of nitric oxide synthase. These cells also responded to stimulation with lipopolysaccharide in vitro, as well as the combination of interferon-gamma and lipopolysaccharide, which was synergistic in stimulating nitric oxide production. Tumor necrosis factor-alpha and macrophage colony-stimulating factor were also found to stimulate nitric oxide production by Ito cells from endotoxemic rats. In addition, in these cells, tumor necrosis factor-alpha synergized with interferon-gamma in inducing nitric oxide production. The combination of interferon-gamma and lipopolysaccharide was also found to inhibit Ito cell DNA synthesis, as measured on the basis of [3H]-thymidine uptake.(ABSTRACT TRUNCATED AT 250 WORDS)

The production of the immunotransmitter, interleukin 1 (IL-1), by peritoneal exudate cells (PEC) was examined in iron-deficient and control rats. Three groups of weanling male Sprague-Dawley rats were fed a purified diet containing 6, 12, or 35 ppm iron for 6 wk to produce severe iron deficiency, moderate iron deficiency, or adequate iron status. Crude IL-1 samples were prepared from acute PEC and assayed for activity. IL-1 preparations from severely and moderately iron-deficient rats enhanced mouse thymocyte proliferation in vitro less than half as much as IL-1 preparations from control rats. In a rabbit bioassay, injection of IL-1 prepared with PEC from either group of iron-deficient rats had little effect on body temperature or plasma minerals, while IL-1 from iron-adequate source PEC produced a febrile response and markedly lowered plasma iron and zinc in recipient rabbits. Severe or moderate iron deficiency, then, clearly impairs IL-1 production by rat PEC.

Copper, iron and zinc concentrations were measured in tissues of young (2 mo), mature (14 mo) and aged (26 mo) male Fischer rats fed either a normal protein (16% casein) or high protein (32% casein) diet for 30 d. Spleen copper concentrations decreased with maturity but were not affected by dietary protein level. Age, dietary protein and age X protein interaction affected spleen iron concentrations. Splenic iron was increased significantly only in mature and aged rats fed the normal protein diet. High protein-fed aged rats had decreased splenic zinc. High protein feeding increased renal zinc in the young and aged rats compared to normal protein feeding. At both protein levels, liver iron increased in the mature rats. Upon aging, zinc levels in the heart increased in the normal protein group and decreased in the high protein group. A significant interaction between age and protein was observed on heart zinc. Thus, the concentrations of tissue trace minerals are affected by age, dietary protein and protein X age interaction in young, mature and aged male rats.