Effect of roflumilast on inflammatory cells in the lungs of cigarette smoke-exposed mice

BACKGROUND: We reported that roflumilast, a phosphodiesterase 4 inhibitor, given orally at 5 mg/kg to mice prevented the development of emphysema in a chronic model of cigarette smoke exposure, while at 1 mg/kg was ineffective. Here we investigated the effects of roflumilast on the volume density (V(V)) of the inflammatory cells present in the lungs after chronic cigarette smoke exposure. METHODS: Slides were obtained from blocks of the previous study and V(V )was assessed immunohistochemically and by point counting using a grid with 48 points, a 20× objective and a computer screen for a final magnification of 580×. Neutrophils were marked with myeloperoxidase antibody, macrophages with Mac-3, dendritic cells with fascin, B-lymphocytes with B220, CD4+ T-cells with CD4+ antibody, and CD8+T-cells with CD8-α. The significance of the differences was calculated using one-way analysis of variance. RESULTS: Chronic smoke exposure increased neutrophil V(V )by 97%, macrophage by 107%, dendritic cell by 217%, B-lymphocyte by 436%, CD4+ by 524%, and CD8+ by 417%. The higher dose of roflumilast prevented the increase in neutrophil V(V )by 78%, macrophage by 82%, dendritic cell by 48%, B-lymphocyte by 100%, CD4+ by 98% and CD8+ V(V )by 88%. The lower dose of roflumilast did not prevent the increase in neutrophil, macrophage and B-cell V(V )but prevented dendritic cells by 42%, CD4+ by 55%, and CD8+ by 91%. CONCLUSION: These results indicate (i) chronic exposure to cigarette smoke in mice results in a significant recruitment into the lung of inflammatory cells of both the innate and adaptive immune system; (ii) roflumilast at the higher dose exerts a protective effect against the recruitment of all these cells and at the lower dose against the recruitment of dendritic cells and T-lymphocytes; (iii) these findings underline the role of innate immunity in the development of pulmonary emphysema and (iiii) support previous results indicating that the inflammatory cells of the adaptive immune system do not play a central role in the development of cigarette smoke induced emphysema in mice.