Presence of cytochrome b-558 in guinea-pig alveolar macrophages-subcellular localization and relationship with NADPH oxidase.
The assignment of cytochrome b-558 as a component of the O2- (H2O2) -generating enzyme in guinea-pig alveolar macrophages was investigated. Guinea pig alveolar macrophages contained 76 pmol cytochrome b-558/mg protein, a value very similar to that of neutrophils. The rate of myristic acid-stimulated O2- generation by alveolar macrophages, calculated per cytochrome b-558, was only one-fourth that of neutrophils. An analysis of Percoll density gradient centrifugation profiles showed that the H2O2-generating activity of myristic acid-activated alveolar macrophages was concentrated in a single peak which was consistently associated with 5'-nucleotidase activity, a plasma membrane marker enzyme. A little H2O2-generating activity was seen with unactivated alveolar macrophages. Furthermore, the cytochrome b-558 of both myristic acid-activated and unactivated alveolar macrophages was also predominantly associated with 5'-nucleotidase activity and was found in trace amounts in a peak containing lysozyme activity, a marker of lysosome granules. Only about 6% of the cytochrome b-558 in plasma membranes from myristic acid-activated guinea-pig alveolar macrophages was anaerobically reduced by 0.5 mM NADPH, while under the same conditions about 30% of the heme protein of myristic acid-activated neutrophils was reduced. These results suggest two conclusions: firstly, that in both activated and unactivated alveolar macrophages, cytochrome b-558 is located in the plasma membrane, and the translocation of cytochrome b-558 does not occur during the activation of NADPH oxidase; and secondly, that a smaller part of cytochrome b-558 is associated with the activated NADPH oxidase of guinea pig alveolar macrophages compared with neutrophils.
Yamaguchi
T
T
National Institute of Hygienic Sciences, Tokyo, Japan.
Kaneda
M
M
D000818:Animals; D002458:Cell Fractionation; D002462:Cell Membrane / Q000032:analysis / Q000378:metabolism; D003573:Cytochrome b Group / Q000032:analysis* / Q000378:metabolism; D006168:Guinea Pigs; D006861:Hydrogen Peroxide / Q000378:metabolism; D008262:Macrophage Activation / Q000187:drug effects; D008264:Macrophages / Q000032:analysis* / Q000378:metabolism / Q000648:ultrastructure; D008297:Male; D019814:Myristic Acid; D009227:Myristic Acids / Q000494:pharmacology; D009247:NADH, NADPH Oxidoreductases / Q000032:analysis* / Q000378:metabolism; D009249:NADP / Q000494:pharmacology; D019255:NADPH Oxidases; D011650:Pulmonary Alveoli / Q000166:cytology* / Q000378:metabolism; D013053:Spectrophotometry; D013481:Superoxides / Q000378:metabolism
D003573:Cytochrome b Group; D009227:Myristic Acids; D019814:Myristic Acid; D013481:Superoxides; D009249:NADP; C036888:cytochrome b558; D006861:Hydrogen Peroxide; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases