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Enhanced superoxide anion generation but reduced leukotriene B4 productivity in thioglycollate-elicited peritoneal
1Research Institute for Disease of the Chest, Medical School, Kyushu University, Fukuoka, Japan.
Abstract:
Lipoxygenase metabolism of arachidonic acid was compared between peritoneal macrophages from untreated rats and those from rats on day 7 after intraperitoneal injection of thioglycollate broth (TG). Resident macrophages (M phi) from untreated rats produced mainly LTB4 (303 +/- 25 pmol/5 x 10(6) cells) and 5-HETE (431 +/- 56 pmol/5 x 10(6) cells) when stimulated with 5 micrograms/ml calcium ionophore A23187 for 20 min at 37 degrees C. On the other hand, TG-elicited M phi generated less amounts of lipoxygenase metabolites (157 +/- 10 pmol LTB4 and 319 +/- 19 pmol 5-HETE/5 x 10(6) cells) with the same stimulus. Then, leukotriene productivity was examined by using subcellular fractions of each M phi lysate and an unstable epoxide intermediate, leukotriene A4. LTA4 hydrolase activity was mainly contained in soluble fractions from the both groups of M phi. The cytosol fraction from the resident M phi exhibited the following specific and total activity; 2.2 +/- 0.1 nmol LTB4/mg protein/5 min and 12.2 +/- 0.5 nmol LTB4/5 min per 10(8) cells. On the contrary, the cytosol fraction from the TG-elicited M phi showed 1.9 +/- 0.1 nmol LTB4/mg protein/5 min and 9.6 +/- 0.3 nmol LTB4/5 min per 10(8) cells. The resident M phi, however, generated 0.14 +/- 0.04 nmol O2-/min/4 x 10(5) cells whereas the TG-elicited M phi did 0.49 +/- 0.13 nmol O2-/min/4 x 10(5) cells when stimulated with wheat germ lectin. These results suggest that the TG-elicited macrophages show enhanced superoxide production but generate less lipoxygenase metabolites.(ABSTRACT TRUNCATED AT 250 WORDS)
Insights
Thioglycollate-elicited macrophages exhibit increased superoxide production but reduced lipoxygenase metabolism of arachidonic acid compared to resident macrophages. This indicates a shift in inflammatory mediator generation following thioglycollate broth stimulation.
Area of Science:
- Biochemistry
- Immunology
- Cell Biology
Background:
- Macrophages play a crucial role in inflammatory responses.
- Lipoxygenase pathway metabolizes arachidonic acid into inflammatory mediators like leukotrienes.
- Thioglycollate broth is used to elicit peritoneal macrophages, altering their phenotype.
Purpose of the Study:
- To compare the lipoxygenase metabolism of arachidonic acid in resident and thioglycollate-elicited (TG-elicited) rat peritoneal macrophages.
- To investigate the activity of LTA4 hydrolase in subcellular fractions of both macrophage types.
- To assess and compare superoxide production in resident and TG-elicited macrophages.
Main Methods:
- Peritoneal macrophages were obtained from untreated rats (resident) and rats 7 days post-thioglycollate broth injection (TG-elicited).
- Lipoxygenase metabolites (LTB4, 5-HETE) were quantified after stimulation with calcium ionophore A23187.
- LTA4 hydrolase activity was measured in soluble and cytosol fractions.
- Superoxide production was assessed upon stimulation with wheat germ lectin.
Main Results:
- TG-elicited macrophages produced significantly less LTB4 and 5-HETE compared to resident macrophages.
- LTA4 hydrolase activity per milligram of protein and per cell was slightly lower in TG-elicited macrophages.
- TG-elicited macrophages demonstrated markedly enhanced superoxide production when stimulated with wheat germ lectin.
Conclusions:
- Thioglycollate-elicited macrophages exhibit a functional shift towards enhanced superoxide production.
- These elicited macrophages show diminished capacity for lipoxygenase metabolism of arachidonic acid.
- The findings suggest differential inflammatory mediator generation profiles between resident and activated macrophage populations.