8,DandE). inAdipo/mice F4/80+CD11c+cells also expressed IL-6, suggesting that IL-6 is regulated by adiponectin in these alveolar macrophages. Transcriptomic analysis identified serum amyloid A3 (Saa3), which promotes IL-17A expression, as the gene most differentially augmented by ozone inAdipo/vs. wild-type mice. After ozone,Saa3mRNA expression was markedly greater inAdipo/vs. wild-type mice but reduced inAdipo//IL-6/vs.Adipo/mice. In conclusion, our data support a pivotal role of IL-6 in the hyperinflammatory condition observed inAdipo/mice after ozone exposure and suggest that this role of IL-6 involves its ability to induce Saa3, IL-17A, and G-CSF. Keywords:neutrophils, IL-17A, macrophages, T cells, serum amyloid A3, CCL20 ozone is an air pollutantproduced by reactions between automobile exhaust and sunlight. Once inhaled, ozone oxidizes lipids, proteins, and other species present in the lung lining fluid (18,34,53). Products of these reactions damage epithelial cells, cause the generation of cytokines and chemokines, and induce pulmonary inflammation characterized by BML-275 (Dorsomorphin) neutrophil and macrophage influx into the lungs. Ozone also causes pulmonary injury leading to increased permeability of the alveolar/capillary barrier (20,31). Adiponectin is an insulin-sensitizing hormone secreted almost exclusively by adipocytes. Serum adiponectin is reduced in obese humans and animals and is elevated upon weight loss (2,17,23,65). Adiponectin has anti-inflammatory properties in vitro and in vivo (51,60,63). Indeed, mice lacking adiponectin (Adipo/mice) have augmented neutrophilic influx into the lungs upon subacute exposure to ozone (0.3 ppm for 2472 h) (28,29). This augmented neutrophilia is dependent on IL-17A (28): ozone causes greaterIl17amRNA abundance inAdipo/vs. wild-type (WT) mice, and anti-IL-17A mAb treatment prior to ozone exposure reduces bronchoalveolar lavage (BAL) neutrophils inAdipo/mice to levels close to those observed in WT mice. We also demonstrated increased numbers of IL-17A+cells (mainly interstitial macrophages/monocytes and T cells) in the lungs after ozone exposure. These IL-17A+cells were augmented inAdipo/vs. WT mice (28). Anti-IL-17 mAb treatment also resulted in reduced granulocyte colony-stimulating factor (G-CSF) inAdipo/mice exposed to ozone, whereas other IL-17A-dependent neutrophil chemotactic factors were not affected (28). IL-6 is a pleiotropic proinflammatory cytokine induced by infectious agents (22), in autoimmunity (47), and upon exposure to environmental toxicants, including ozone BML-275 (Dorsomorphin) (3,27). Indeed, the pulmonary neutrophil recruitment induced by ozone (0.3 ppm for 72 h) is partly IL-6 dependent (27,50). Compared with WT mice,Adipo/mice not only have augmented neutrophil influx but also have increased BAL IL-6 after ozone exposure (28,29). IL-6 is important in polarizing T cells toward a Th17 phenotype (62). Hence, the purpose of this study was to examine the hypothesis that IL-6 is required for the increases in IL-17A expression and subsequent neutrophil recruitment observed inAdipo/mice exposed to ozone. To test this hypothesis, we exposed Keratin 10 antibody WT,IL-6/, andAdipo/mice as well as mice deficient in both adiponectin and IL-6 (Adipo//IL-6/mice) to ozone (0.3 ppm) or to room air for 72 h and assessed pulmonary inflammation and IL-17A expression. We also conducted a microarray analysis of lungs ofAdipo/and WT mice exposed to ozone or air. We found that serum amyloid A3 (SAA3), an acute-phase protein induced by inflammation and capable of inducing IL-17A expression in the lungs (1), was higher inAdipo/mice vs. WT mice exposed to ozone and that IL-6 deficiency virtually ablated this effect of adiponectin deficiency. == METHODS == == == == Animals. == This study was approved by the Harvard Medical BML-275 (Dorsomorphin) School Standing Committee on Animals (IACUC). Adiponectin-deficient (Adipo/) mice BML-275 (Dorsomorphin) were originally obtained from Dr. Y. Matsuzawa (41). IL-6-deficient (IL-6/) mice were obtained from The Jackson Laboratory (Bar Harbor, ME).Adipo/andIL-6/mice were crossbred to generate double-deficient mice (Adipo//IL-6/). Sex- and age (1113 wk old)-matched WT C57BL/6J mice were obtained from The Jackson Laboratory. == Protocol. == Mice were exposed to either ozone (0.3 ppm) or ambient air for 24, 48, or 72 h, as previously described (28). Immediately after exposure, mice BML-275 (Dorsomorphin) were euthanized with an intraperitoneal overdose of pentobarbital. A tracheostomy was performed and the trachea was cannulated. BAL was performed and the lungs were harvested and used to extract RNA for RT-PCR. In another cohort, after BAL, blood was flushed from the lungs.