Chronic inflammation of the lung, as a consequence of persistent bacterial infections by several opportunistic pathogens represents the main cause of mortality and morbidity in cystic fibrosis (CF) patients. Mechanisms leading to increased susceptibility to bacterial infections in CF are not completely known, although the involvement of cystic fibrosis transmembrane conductance regulator (CFTR) in microbicidal functions of macrophages is emerging. Tissue macrophages differentiate in situ from infiltrating monocytes, additionally, mature macrophages from different tissues, although having a number of common activities, exhibit variation in some molecular and cellular functions. In order to highlight possible intrinsic macrophage defects due to CFTR dysfunction, we have focused our attention on in vitro differentiated macrophages from human peripheral blood monocytes. Here we report on the contribution of CFTR in the bactericidal activity against Pseudomonas aeruginosa of monocyte derived human macrophages. At first, by real time PCR, immunofluorescence and patch clamp recordings we demonstrated that CFTR is expressed and is mainly localized to surface plasma membranes of human monocyte derived macrophages (MDM) where it acts as a cAMP-dependent chloride channel. Next, we evaluated the bactericidal activity of P. aeruginosa infected macrophages from healthy donors and CF patients by antibiotic protection assays. Our results demonstrate that control and CF macrophages do not differ in the phagocytic activity when infected with P. aeruginosa. Rather, although a reduction of intracellular live bacteria was detected in both non-CF and CF cells, the percentage of surviving bacteria was significantly higher in CF cells. These findings further support the role of CFTR in the fundamental functions of innate immune cells including eradication of bacterial infections by macrophages.

Abstract-Endothelial cells express tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptors, but the function of TRAIL in endothelial cells is not completely understood. We explored the role of TRAIL in regulation of key intracellular signal pathways in endothelial cells. Key Words: tumor necrosis factor-related apoptosis-inducing ligand Ⅲ nitric oxide Ⅲ prostanoids T umor necrosis factor (TNF)-related apoptosisinducing ligand (TRAIL)/Apo-2L is a member of the TNF family of cytokines, which are structurally related proteins playing important roles in regulating cell death, immune response, and inflammation. 1 TRAIL is a type II membrane protein, which can be proteolytically cleaved to a soluble form, 2 as previously shown also for TNF-␣ and CD95 (Apo-1/Fas)L. The unique feature of TRAIL, compared with other members of the TNF family, is its ability to induce apoptosis in a variety of malignant cells both in vitro and in vivo, displaying minimal toxicity on normal cells and tissues. 3,4 TRAIL interacts with 4 high affinity transmembrane receptors belonging to the apoptosis-inducing TNFreceptor (R) family. TRAIL-R1 (DR4) and TRAIL-R2 (DR5) transduce apoptotic signals on binding of TRAIL, whereas TRAIL-R3 (DcR1) and TRAIL-R4 (DcR2) are homologous to DR4 and DR5 in their cysteine-rich extracellular domain, but they lack the intracellular death domain and apoptosis inducing capability. It has been proposed that TRAIL-R3 and TRAIL-R4 function as decoy receptors protecting normal cells, including endothelial cells, from apoptosis. 5,6 It has been shown that endothelial cells express TRAIL receptors, 6 and TRAIL protein is expressed in the medial smooth cell layer of the aorta and pulmonary arteries. 7 Whereas cleavage of Fas ligand from the cell surface requires the action of zinc-dependent metalloproteases, generation of soluble TRAIL involves the action of cysteine proteases. 2 Notably, the vessel wall is a rich source of cysteine proteases. 8 Using human umbilical vein endothelial cells (HUVECs) as a model system, the aim of this study was to investigate the ability of TRAIL to modulate intracellular pathways that play a key role in endothelial cell biology. In particular, we have analyzed whether TRAIL was able to modulate the production of nitric oxide (NO), which regulates vascular tone, promotes endothelial cell survival and migration, and inhibits platelet adhesion and aggregation, leukocyte adherence, and vascular smooth muscle cell proliferation, therefore providing antithrombotic and antiinflammatory activity. 9 -12 Moreover, we have investigated the expression and/or activity of cyclooxygenases (COX) in response to TRAIL, because also these enzymes, by catalyzing the rate-limiting step in the biosynthesis of prostanoids, 13 such as prostaglandin (PGE) 2 , prostacyclin (PGI) 2 , and thromboxane (TXA) 2 , have a profound influence on blood pressure, regional blood flow, vascular remodeling, and angiogenesis.

BackgroundStenotrophomonas maltophilia has recently gained considerable attention as an important emerging pathogen in cystic fibrosis (CF) patients. However, the role of this microorganism in the pathophysiology of CF lung disease remains largely unexplored. In the present study for the first time we assessed the ability of S. maltophilia CF isolates to adhere to and form biofilm in experimental infection experiments using the CF-derived bronchial epithelial IB3-1cell line. The role of flagella on the adhesiveness of S. maltophilia to IB3-1 cell monolayers was also assessed by using fliI mutant derivative strains.ResultsAll S. maltophilia CF isolates tested in the present study were able, although at different levels, to adhere to and form biofilm on IB3-1 cell monolayers. Scanning electron and confocal microscopy revealed S. maltophilia structures typical of biofilm formation on bronchial IB3-1 cells. The loss of flagella significantly (P < 0.001) decreased bacterial adhesiveness, if compared to that of their parental flagellated strains. S. maltophilia CF isolates were also able to invade IB3-1 cells, albeit at a very low level (internalization rate ranged from 0.01 to 4.94%). Pre-exposure of IB3-1 cells to P. aeruginosa PAO1 significantly increased S. maltophilia adhesiveness. Further, the presence of S. maltophilia negatively influenced P. aeruginosa PAO1 adhesiveness.ConclusionsThe main contribution of the present study is the finding that S. maltophilia is able to form biofilm on and invade CF-derived IB3-1 bronchial epithelial cells, thus posing a rationale for the persistence and the systemic spread of this opportunistic pathogen in CF patients. Experiments using in vivo models which more closely mimic CF pulmonary tissues will certainly be needed to validate the relevance of our results.