15-Lipoxygenase expression and 15 (S)-hydroxyeicoisatetraenoic acid release and reincorporation in induced sputum of asthmatic subjects

Background: Recent evidence shows that 15(S)-hydroxyeicoisatetraenoic acid (15[S]-HETE) can be released and rapidly reincorporated into cellular lipids. These mechanisms exert several immunoregulatory functions that may be relevant in airway inflammation. Objective: Our purpose was to evaluate the levels of both soluble and cell-associated 15(S)-HETE and to examine 15- lipoxygenase (15-LO) messenger RNA (mRNA) expression in sputum samples obtained from 10 control and 18 asthmatic subjects. Methods: Levels of 15(S)- HETE were measured by reverse-phase HPLC separation followed by RIA in supernatants and in cell membrane-extracted phospholipids after acid hydrolysis. 15-LO mRNA was evaluated by primed in situ hybridization (PRINS). Combined immunocytochemistry and PRINS was used to identify the phenotype of cells bearing 15-LO transcripts. Results: Levels of both soluble and cell- associated 15(S)-HETE were higher in asthmatic than in control subjects (P < .0001). The percentage of cells expressing 15-LO mRNA was higher in asthmatic than in control subjects (P < .01). On double staining for specific cell-type markers and 15-LO mRNA, macrophages were the major source for 15-LO. Conclusion: This study shows that the induced sputum technique allows the evaluation of 15-LO activity and that soluble, cell-associated 15(S)-HETE and 15-LO levels are higher in asthmatic than in control subjects. In addition, this study indicates that, in induced sputum, airway macrophages are the major source of 15(S)-HETE in asthma.