Proteasome and immunoproteasome function in cigarette smoke-mediated chronic lung disease

摘要

Chronic obstructive pulmonary disease (COPD) is projected to be the third leading cause of death by 2020 with cigarette smoke exposure being the main risk factor. Cigarette smoke leads to oxidative stress in the lung, resulting in protein damage and adaptive immune responses. Also, smokers and COPD patients are more susceptible to viral infections often followed by acute exacerbations of COPD pathogenesis. Lungs of COPD patients exhibit increased numbers of innate and adaptive immune cells, among these CD8+ T cells, whose abundance correlates with disease severity. The proteasome degrades more than 90 % of intracellular proteins - including damaged ones - into small peptides and is important to protect the cell from proteotoxic stress. Furthermore, the immunoproteasome, a specialized proteasome subtype which is expressed by default in antigen presenting cells and induced during infection, is involved in shaping adaptive immune responses by enhancing antigen presentation via major histocompatibility complex (MHC) I to cytotoxic CD8+ T cells. The effects of cigarette smoke on (immuno-)proteasome function have not been investigated so far. udThe first publication included in this thesis (van Rijt et al. 2012) explored the effects of acute cigarette smoke exposure on proteasome expression and activity. We observed that short-term exposure of cells to extracts of cigarette smoke directly impaired proteasome activity, while proteasomal protein expression was not altered. Oxidatively modified and polyubiquitinated proteins accumulated, suggesting augmentation of oxidative stress in cigarette smoke-treated cells. In lungs of mice acutely exposed to cigarette smoke, a similar effect could be observed: one of the three proteasome activities was significantly reduced, and ubiquitinated substrates for the proteasome were found to be accumulated, while proteasome expression levels were not changed.udThe second publication in this thesis (Keller et al. 2015) shows for the first time the cell-specific expression of immunoproteasomes in the lung and their induction by interferon-γ in vitro and by murid herpesvirus 68 (MHV-68) infection in vivo. Within these experiments, activity-based probes were used to clearly define the kinetics of standard and immunoproteasome subunit incorporation. In human lungs from controls or early-stage COPD patients, immunoproteasome expression was not changed. Immunoproteasomes localized mainly to alveolar macrophages, but not to parenchymal cells in both donors and end-stage COPD.udResults from recent experiments were accepted for publication in the meantime (Kammerl et al. 2016): we investigated MHC I antigen presentation in cigarette smoke extract-treated primary immune cells and bronchoalveolar lavage (BAL) cells from mice exposed to cigarette smoke for ten days. In vitro treatment of primary immune cells with cigarette smoke extract led to a decrease in the presentation of an immunoproteasome-dependent “self”-epitope. With the help of activity-based probes, we observed a shift from immuno- to standard proteasome activity in isolated alveolar macrophages from smoke exposed mice. This shift, however, was not sufficient to impact antigen presentation of an immunoproteasome-dependent epitope. The altered ratio of standard and immunoproteasome might be explained by transcriptional downregulation of immuno-, but not standard proteasomes by cigarette smoke in isolated alveolar macrophages of smoke-exposed mice, which was also observed in total BAL cells of early-stage COPD patients. In the lungs of end-stage COPD patients, activities of both standard and immunoproteasome subunits were significantly decreased, while total proteasome protein levels were not changed. udTaken together, we show that cigarette smoke directly impairs proteasome function in vitro and in vivo, which may exacerbate oxidative stress resolution in response to cigarette smoke, since the degradation of oxidatively modified and misfolded proteins is impaired. In addition, we observed alterations in immunoproteasome-dependent MHC I antigen presentation, which may contribute to increased susceptibility to virus-induced exacerbations, prolonged infection and possibly result in autoimmune responses.