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Comparative studies on alveolar macrophages and polymorphonuclear leukocytes. I. H2O2 and O2- generation by rabbit
Abstract:
The oxidative metabolism of rabbit alveolar macrophages (A-MO) was compared with that of rabbit polymorphonuclear leukocytes (PMN) with respect to H2O2 generation by intact cells or subcellular fractions. Rabbit PMN exhibited an increase in the oxygen uptake and a marked release of H2O2 upon addition of heat-killed E. coli in the presence and absence of opsonin. However, rabbit A-MO exhibited an increase in the oxygen uptake upon addition of E. coli only in the presence of anti-E. coli serum as an opsonin, whereas a very small amount of H2O2 release was observed during ingestion of the opsonized E. coli. The generation of O2- and H2O2 by a granule-rich fraction isolated from phagocytosing PMN was larger than that by a similar fraction isolated from resting PMN. However, there was no significant difference in O2- and H2O2 generation by the granule fractions between phagocytosing and resting A-MO in the presence of either NADH or NADPH. In contrast to the granule fraction of rabbit PMN, the O2- and H2O2 generating activities in the A-MO granule fraction were higher in the presence of NADH than in the presence of NADPH. The rates of NADH and NADPH oxidation by both A-MO and PMN granule fractions were measured with and without addition of Mn2+ to the assay medium. The effect of Mn2+ on the NAD(P)H oxidase was found to differ between rabbit A-MO and PMN.
Insights
Rabbit polymorphonuclear leukocytes (PMN) generate more hydrogen peroxide (H2O2) than alveolar macrophages (A-MO) during bacterial phagocytosis. This difference in oxidative metabolism impacts immune response.
Area of Science:
- Cellular immunology
- Oxidative metabolism
- Phagocytosis
Background:
- Alveolar macrophages (A-MO) and polymorphonuclear leukocytes (PMN) are key phagocytic cells in the innate immune system.
- Oxidative metabolism, particularly hydrogen peroxide (H2O2) generation, is crucial for microbial killing by phagocytes.
- Understanding the differences in H2O2 production between A-MO and PMN is important for elucidating their distinct roles in host defense.
Purpose of the Study:
- To compare the oxidative metabolism and H2O2 generation capacity of rabbit alveolar macrophages (A-MO) and polymorphonuclear leukocytes (PMN).
- To investigate H2O2 production by intact cells and subcellular fractions of A-MO and PMN during phagocytosis.
- To analyze the activity of NAD(P)H oxidase in granule fractions and the effect of Mn2+ on enzyme activity.
Main Methods:
- Measurement of oxygen uptake and H2O2 release by intact rabbit A-MO and PMN upon stimulation with heat-killed E. coli, with and without opsonin.
- Isolation of granule-rich fractions from phagocytosing and resting A-MO and PMN.
- Assay of superoxide anion (O2-) and H2O2 generation by granule fractions using NADH or NADPH as substrates.
- Measurement of NADH and NADPH oxidation rates in granule fractions with and without Mn2+.
Main Results:
- Rabbit PMN showed increased oxygen uptake and significant H2O2 release upon E. coli addition, regardless of opsonin presence.
- Rabbit A-MO exhibited increased oxygen uptake only with opsonized E. coli and released very little H2O2.
- Granule fractions from phagocytosing PMN generated more O2- and H2O2 than resting PMN fractions.
- A-MO granule fractions showed no significant difference in O2- and H2O2 generation between phagocytosing and resting states.
- A-MO granule fractions utilized NADH more effectively than NADPH for H2O2 generation, unlike PMN fractions.
- The effect of Mn2+ on NAD(P)H oxidase activity differed between rabbit A-MO and PMN granule fractions.
Conclusions:
- Rabbit PMN are more efficient at generating H2O2 during bacterial phagocytosis compared to rabbit A-MO.
- Differences in oxidative metabolism and substrate utilization (NADH vs. NADPH) exist between A-MO and PMN granule fractions.
- The modulation of NAD(P)H oxidase by Mn2+ is distinct in A-MO and PMN, suggesting differential regulatory mechanisms.