P13 Analysis of the mechanisms of increased severity of influenza-related pneumonia in mice

to endotoxin shock or polymicrobial sepsis. Endotoxin shock was induced by intraperitoneal injection of LPS (60mg/kg). Sepsis was initiated by cecal ligation and puncture (CLP) (6 punctures with a 21gauge needle). Results: Clarithromycinmarkedly expanded splenic and lung CD11b+ Gr−1+cells in normal mice. Pretreatment with clarithromycin improved mortality in mouse models of LPS-induced shock and sepsis. Since we confirmed that more than 90% of Gr−1+ cells are CD11b+Gr−1+ cells, we substitute anti-Gr−1 antibody for depleting CD11b+Gr−1+ cells. Improved mortality by clarithromycin was cancelled by depleting Gr−1+ cells both in endotoxin shock and sepsis models. Furthermore, adoptive transfer of sorted CD11b+Gr−1+cells obtained from clarithromycin-treated mice protected against LPSand CLP-induced lethality, which were associated with decrease of pro-inflammatory cytokines (TNF-a, IL-6, and IFN-g) and increase of anti-inflammatory cytokine IL-10 in serum and bronchoalveolar lavage fluids. In vitro study, CD11b+Gr−1+cells obtained from clarithromycin-treated mice prohibited the proliferation of CD3+ T cells in the presence of anti-CD3 and anti-CD28, as compared to the control CD11b+Gr−1+ cells. Consistently, microarray analysis demonstrated that expressions of arginase-1, a key enzyme of inhibiting T cell proliferation, are much higher in CD11b+Gr−1+cells obtained from clarithromycin-treated mice compared to the control CD11b+Gr-1+cells. In addition, production of pro-inflammatory cytokines (TNF-a, IL-6, and IFN-g) were significantly suppressed, while anti-inflammatory cytokine IL-10 was increased when bone marrowderived macrophages or thiogllycolate-elicited macrophages were cocultured with CD11b+Gr−1+ cells obtained from clarithromycintreated mice. Conclusion: We have identified for the first time the unique population of immunosuppressive CD11b+Gr−1+ cells expanded by clarithromycin, which are essential for the anti-inflammatory properties of clarithromycin.