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Cyclic adenosine 3′,5′-monophosphate (cAMP)–elevating agents, such as β 2 -adrenergic receptor (β 2 -AR) agonists and phosphodiesterase (PDE) inhibitors, remain a mainstay in the treatment of obstructive respiratory diseases, conditions characterized by airway constriction, inflammation, and mucus hypersecretion. However, their clinical use is limited by unwanted side effects because of unrestricted cAMP elevation in the airways and in distant organs. Here, we identified the A-kinase anchoring protein phosphoinositide 3-kinase γ (PI3Kγ) as a critical regulator of a discrete cAMP signaling microdomain activated by β 2 -ARs in airway structural and inflammatory cells. Displacement of the PI3Kγ-anchored pool of protein kinase A (PKA) by an inhaled, cell-permeable, PI3Kγ mimetic peptide (PI3Kγ MP) inhibited a pool of subcortical PDE4B and PDE4D and safely increased cAMP in the lungs, leading to airway smooth muscle relaxation and reduced neutrophil infiltration in a murine model of asthma. In human bronchial epithelial cells, PI3Kγ MP induced unexpected cAMP and PKA elevations restricted to the vicinity of the cystic fibrosis transmembrane conductance regulator (CFTR), the ion channel controlling mucus hydration that is mutated in cystic fibrosis (CF). PI3Kγ MP promoted the phosphorylation of wild-type CFTR on serine-737, triggering channel gating, and rescued the function of F508del-CFTR, the most prevalent CF mutant, by enhancing the effects of existing CFTR modulators. These results unveil PI3Kγ as the regulator of a β 2 -AR/cAMP microdomain central to smooth muscle contraction, immune cell activation, and epithelial fluid secretion in the airways, suggesting the use of a PI3Kγ MP for compartment-restricted, therapeutic cAMP elevation in chronic obstructive respiratory diseases.

Cystic fibrosis (CF) is an inherited disorder caused by mutations in the gene encoding for the cystic fibrosis transmembrane conductance regulator (CFTR) protein, an ATP-gated chloride channel expressed on the apical surface of airway epithelial cells. CFTR absence/dysfunction results in defective ion transport and subsequent airway surface liquid dehydration that severely compromise the airway microenvironment. Noxious agents and pathogens are entrapped inside the abnormally thick mucus layer and establish a highly inflammatory environment, ultimately leading to lung damage. Since chronic airway inflammation plays a crucial role in CF pathophysiology, several studies have investigated the mechanisms responsible for the altered inflammatory/immune response that, in turn, exacerbates the epithelial dysfunction and infection susceptibility in CF patients. In this review, we address the evidence for a critical role of dysfunctional inflammation in lung damage in CF and discuss current therapeutic approaches targeting this condition, as well as potential new treatments that have been developed recently. Traditional therapeutic strategies have shown several limitations and limited clinical benefits. Therefore, many efforts have been made to develop alternative treatments and novel therapeutic approaches, and recent findings have identified new molecules as potential anti-inflammatory agents that may exert beneficial effects in CF patients. Furthermore, the potential anti-inflammatory properties of CFTR modulators, a class of drugs that directly target the molecular defect of CF, also will be critically reviewed. Finally, we also will discuss the possible impact of SARS-CoV-2 infection on CF patients, with a major focus on the consequences that the viral infection could have on the persistent inflammation in these patients.

Cystic fibrosis is a genetic disease caused by mutations in the CFTR gene, whereas chronic obstructive pulmonary disease (COPD) is mainly caused by environmental factors (mostly cigarette smoking) on a genetically susceptible background. Although the etiology and pathogenesis of these diseases are different, both are associated with progressive airflow obstruction, airway neutrophilic inflammation, and recurrent exacerbations, suggesting common mechanisms. The airway epithelium plays a crucial role in maintaining normal airway functions. Major molecular and morphologic changes occur in the airway epithelium in both CF and COPD, and growing evidence suggests that airway epithelial dysfunction is involved in disease initiation and progression in both diseases. Structural and functional abnormalities in both airway and alveolar epithelium have a relevant impact on alteration of host defences, immune/inflammatory response, and the repair process leading to progressive lung damage and impaired lung function. In this review, we address the evidence for a critical role of dysfunctional airway epithelial cells in chronic airway inflammation and remodelling in CF and COPD, highlighting the common mechanisms involved in the epithelial dysfunction as well as the similarities and differences of the two diseases.

Obstructive lung diseases such as cystic fibrosis (CF) and chronic obstructive pulmonary disease (COPD) are causes of high morbidity and mortality worldwide. CF is a multiorgan genetic disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene and is characterized by progressive chronic obstructive lung disease. Most cases of COPD are a result of noxious particles, mainly cigarette smoke but also other environmental pollutants. Areas covered: Although the pathogenesis and pathophysiology of CF and COPD differ, they do share key phenotypic features and because of these similarities there is great interest in exploring common mechanisms and/or factors affected by CFTR mutations and environmental insults involved in COPD. Various molecular, cellular and clinical studies have confirmed that CFTR protein dysfunction is common in both the CF and COPD airways. This review provides an update of our understanding of the role of dysfunctional CFTR in both respiratory diseases. Expert commentary: Drugs developed for people with CF to improve mutant CFTR function and enhance CFTR ion channel activity might also be beneficial in patients with COPD. A move toward personalized therapy using, for example, microRNA modulators in conjunction with CFTR potentiators or correctors, could enhance treatment of both diseases.

Introduction: Immunoglobulin (Ig) A protects mucosal surfaces from inhaled pathogens and antigens. While serum and sputum IgA are increased, it remains unknown whether polymeric immunoglobulin receptor (pIgR)-mediated transepithelial transport of IgA is affected in cystic fibrosis (CF). Aims: We wondered whether broncho-epithelial pIgR-mediated transport of IgA is altered and could contribute to lung infection by Pseudomonas aeruginosa (PA). Methods: Lung tissue from 19 CF patients, as compared to 17 controls, was assayed for pIgR immunostaining. Sputum samples from 49 CF patients and 46 controls were assessed for IgA, secretory (S)-IgA, IgM, S-IgM, secretory component (SC) and PA-specific IgA and IgM, as well as neutrophil elastase (NE) activity. IgA and IgM. Total and PA-specific IgA and IgM were also measured in serum. Results: Increased epithelial pIgR immunostaining was observed in CF compared to controls, in large (56% versus 8, mean, p Conclusion: We report that, in contrast with COPD, broncho-epithelial pIgR expression is enhanced in CF and associates with an increased production of sputum IgA and IgM, including PA-specific antibodies, indicating that humoral immunity is upregulated in the CF lung.

Vascular remodelling plays a central role in asthma and chronic obstructive pulmonary disease (COPD). Bradykinin (BK) is a vasoactive proinflammatory peptide mediating acute responses in asthma. We investigated the role of angiogenic factors in relation to BK receptors in asthma and COPD.Bronchial biopsies from 33 patients with COPD, 24 old (≥50 years) patients with (≥50 years) asthma, 18 old control smokers, 11 old control non-smokers, 15 young (≤40yrs) patients with (≤40 years) asthma and 10 young control non-smokers were immunostained for CD31, vascular endothelial growth factor-A (VEGF-A), angiogenin and BK receptors (B2R and B1R). Fibroblast and endothelial co-localisation of relevant molecules were performed by immunofluorescence. BK-induced VEGF-A and angiogenin release was studied (ELISA) in bronchial fibroblasts from subjects with asthma and COPD.In bronchial lamina propria of old patients with asthma, CD31 and VEGF-A(+) cell numbers were higher than old control non-smokers (p0.05). Angiogenin(+), B2R(+) and B1R(+) cell numbers in old patients with asthma were higher than in old control non-smokers, control smokers and patients with COPD (p0.01). Angiogenin(+) cell numbers were higher in patients with COPD than both old control groups (p0.05). In all patients with asthma the number of B2R(+) cells was positively related to the numbers of B1R(+) (rs=0.43), angiogenin(+) (rs=0.42) and CD31 cells (rs=0.46) (p0.01). Angiogenin(+) cell numbers were negatively related to forced expiratory volume in 1 s (rs=-0.415, p=0.008). Double immunofluorescence revealed that CD31 cells of capillary vessels coexpressed B2R and that fibroblasts coexpressed B2R, VEGF-A and angiogenin. BK (10(-6)M) induced significant angiogenin release in fibroblasts from asthma and to a lesser extent in COPD.Unlike COPD, this study suggests the involvement of BK receptors in bronchial vascular remodelling in asthma.

Summary Background Long-term complications of cystic fibrosis include osteoporosis and fragility fractures, but few data are available about effective treatment strategies, especially in young patients. We investigated treatment of low bone mineral density in children, adolescents, and young adults with cystic fibrosis. Methods We did a multicentre trial in two phases. We enrolled patients aged 5–30 years with cystic fibrosis and low bone mineral density, from ten cystic fibrosis regional centres in Italy. The first phase was an open-label, 12-month observational study of the effect of adequate calcium intake plus calcifediol. The second phase was a 12-month, double-blind, randomised, placebo-controlled, parallel group study of the efficacy and safety of oral alendronate in patients whose bone mineral apparent density had not increased by 5% or more by the end of the observational phase. Patients were randomly assigned to either alendronate or placebo. Both patients and investigators were masked to treatment assignment. We used dual x-ray absorptiometry at baseline and every 6 months thereafter, corrected for body size, to assess lumbar spine bone mineral apparent density. We assessed bone turnover markers and other laboratory parameters every 3–6 months. The primary endpoint was mean increase of lumbar spine bone mineral apparent density, assessed in the intention-to-treat population. This study is registered with ClinicalTrials.gov, number NCT01812551. Findings We screened 540 patients and enrolled 171 (mean age 13·8 years, SD 5·9, range 5–30). In the observational phase, treatment with calcium and calcifediol increased bone mineral apparent density by 5% or more in 43 patients (25%). 128 patients entered the randomised phase. Bone mineral apparent density increased by 16·3% in the alendronate group (n=65) versus 3·1% in the placebo group (n=63; p=0·0010). 19 of 57 young people (33·3%) receiving alendronate attained a normal-for-age bone mineral apparent density Z score. In the observational phase, five patients had moderate episodes of hypercalciuria, which resolved after short interruption of calcifediol treatment. During the randomised phase, one patient taking alendronate had mild fever versus none in the placebo group; treatment groups did not differ significantly for other adverse events. Interpretation Correct calcium intake plus calcifediol can improve bone mineral density in some young patients with cystic fibrosis. In those who do not respond to calcium and calcifediol alone, alendronate can safely and effectively increase bone mineral density. Funding Telethon Foundation (Italy).

Bradykinin-induced interleukin (IL)-8 release should potentially activate neutrophils releasing myeloperoxidase (MPO) and subsequently generating "nitrosative stress". We studied bradykinin-induced expression of bradykinin B(2) receptor and bradykinin- and lipopolysaccharide (LPS)-induced IL-8 release, MPO (marker of neutrophil activation) and 3-nitrotyrosine (3-NT; marker of "nitrosative stress") production in human bronchial epithelial cells BEAS-2B alone or in co-culture with human neutrophils. We evaluated B(2) receptor protein expression in BEAS-2B cells by immunostainings and Western blot analysis, and measured respectively bradykinin- or LPS-induced IL-8 release in BEAS-2B cells and bradykinin- and/or LPS-induced MPO and 3-NT production in BEAS-2B cells co-cultured with human neutrophils by ELISA. In addition, we evaluated bradykinin- and/or LPS-induced 3-NT formation in BEAS-2B cells co-cultured with human neutrophils by immunocytochemistry. Bradykinin up-regulates B(2) receptor expression (P

Purpose. Neutrophil-dominated airway inflammation is a key feature of progressive lung damage in cystic fibrosis (CF). Thus, reducing airway inflammation is a major goal to prevent lung damage in CF. However, current anti-inflammatory drugs have shown several limits. PI3Kγplays a pivotal role in leukocyte recruitment and activation; in the present study we determined the effects of genetic deletion and pharmacologic inhibition of PI3Kγon airway inflammation and structural lung damage in a mouse model of CF lung disease.Methods.βENaC overexpressing mice (βENaC-Tg) were backcrossed with PI3Kγ-deficient (PI3KγKO) mice. Tissue damage was assessed by histology and morphometry and inflammatory cell number was evaluated in bronchoalveolar lavage fluid (BALF). Furthermore, we assessed the effect of a specific PI3Kγinhibitor (AS-605240) on inflammatory cell number in BALF.Results. Genetic deletion of PI3Kγdecreased neutrophil numbers in BALF ofPI3KγKO/βENaC-Tg mice, and this was associated with reduced emphysematous changes. Treatment with the PI3Kγinhibitor AS-605240 decreased the number of neutrophils in BALF ofβENaC-Tg mice, reproducing the effect observed with genetic deletion of the enzyme.Conclusions. These results demonstrate the biological efficacy of both genetic deletion and pharmacological inhibition of PI3Kγin reducing chronic neutrophilic inflammation in CF-like lung diseasein vivo.

It has been suggested that genes other than CFTR could modulate the severity of lung disease in cystic fibrosis (CF). Neutrophil Fcgamma receptor II (FcgammaRII) is involved in host defense against microorganisms and in inflammatory response. We evaluated the association between genetic variability of this gene and both airway infection with Pseudomonas aeruginosa and severity of lung disease in patients with CF. We studied 167 Italian unrelated patients with CF and 50 control subjects. The distribution of FcgammaRIIA genotypes in CF patients was compared with that in control subjects and the different genotypes were related with the presence or absence of P. aeruginosa infection and markers of disease severity in CF patients. The distribution of FcgammaRIIA genotypes was not significantly different between CF patients and controls. We observed that in CF patients with the same CFTR genotype (DeltaF508/DeltaF508), those carrying the R allele of FcgammaRIIA had an increased risk of acquiring chronic P. aeruginosa infection (P=0.042, R.R.: 4.38; 95% CI: 1.17/22.4). Moreover, the frequency of R/R genotype in patients with chronic P. aeruginosa infection seems to be higher than that of control subjects and patients without chronic infection. The observation that CF patients carrying the R allele of FcgammaRIIA are at higher risk of acquiring chronic P. aeruginosa infection suggests that the FcgammaRII loci genetic variation is contributing to this infection susceptibility.

Experimental guinea pig asthma is a reliable and clinically relevant facsimile of human disease. The guinea pig is the preferred choice for use as a model of allergic bronchial asthma in the evaluation of anti-asthmatic drugs, since the airway anatomy and the response to inflammatory mediators is similar to humans. Further, the great strength of this model is the direct anaphylactic bronchoconstriction upon antigen challenge. Under certain conditions a late asthmatic response can be measured and airway hyperresponsiveness is observed in vitro and in vivo. Moreover, the inflammatory response is comparable with the human situation. More recent studies describe a chronic model for asthma in which airway remodeling is induced as can be observed in the asthmatic patients. The focus here is to demonstrate that guinea pig asthma models are useful for testing novel therapeutics.

Airway wall inflammation, IL-4, and mucus hypersecretion are thought to be associated.To quantify bronchial inflammatory cells in smokers with chronic bronchitis (CB) with and without airflow obstruction (AO), determining the cells expressing IL-4 and IL-5 and their association with submucosal gland mucin.We applied immunohistochemistry to identify, and double-labeling to colocalize, IL-4 and IL-5 to distinct inflammatory cells in resected bronchi from (1) 11 asymptomatic smokers (AS), (2) 11 smokers with CB, and (3) 10 smokers with CB and AO.There were greater numbers of mucosal and gland CD45(+) leukocytes in CB (epithelium, 673/mm(2); subepithelium, 698/mm(2); gland, 517/mm(2)) than in AS (331, 237, and 178/mm(2), respectively; p0.01 for all) or CB + AO (375, 243, and 215/mm(2), respectively; p0.05 for all). There were greater numbers of subepithelial and submucosal gland plasma cells in CB (subepithelium, 110/mm(2); gland, 213/mm(2)) compared with AS (38 and 41/mm(2), respectively; p0.01 for both), and more subepithelial mast cells in CB (204/mm(2)) than in AS (65/mm(2); p0.01) or CB + AO (115/mm(2); p0.01). In CB, the percentage of gland occupied by mucin was positively correlated with the numbers of interstitial CD45(+) cells, plasma cells, and IL-4 protein(+) cells. In CB, 69 and 62% of gland-associated plasma cells expressed IL-4 and IL-5, respectively.Inflammatory cells are increased in bronchial submucosal glands and mucosa of large airways in smokers with CB. Gland-associated plasma cells express IL-4, and these likely promote mucus hypersecretion.

The defect in interferon-γ (IFN-γ) production that results in a T helper cell type 2-dominated response may be responsible for a decrease in the apoptosis of allergen-activated T cells in asthma. We investigated the effect of recombinant IFN-γ on proliferation, Fas/Fas ligand (FasL) expression, and apoptosis in allergen-stimulated peripheral blood mononuclear cells obtained from atopic, asthmatic patients and nonatopic, control subjects. The addition of IFN-γ at the start of cultures markedly inhibited the proliferative response to a specific allergen in cells from all asthmatic patients, whereas no change was observed in cells from nonatopic, control subjects. IFN-γ induced an increase in the expression of Fas and FasL by allergen-stimulated CD4+ T cells from asthmatic patients and caused the apoptosis of these cells. A Fas-blocking monoclonal antibody prevented the inhibitory effect of IFN-γ on allergen-induced proliferation. These results suggest that IFN-γ inhibits the proliferation of allergen-stimulated CD4+ T cells from atopic, asthmatic patients by inducing the surface expression of Fas and FasL, which in turn triggers their apoptotic program. The defect in IFN-γ production involved in the allergic, immune response may therefore be responsible for a decrease in apoptosis of allergen-activated T lymphocytes in the airways of atopic, asthmatic patients.

15-lipoxygenase (15-LO) has been implicated in the inflammation of chronic bronchitis (CB), but it is unclear which of its isoforms, 15-LOa or 15-LOb, is primarily involved. To detect 15-LO gene (mRNA) and protein expression, we have applied in situ hybridization (ISH) and immunohistochemistry (IHC), respectively, to bronchial biopsies obtained from 7 healthy nonsmokers (HNS), 5 healthy smokers (HS), and 8 smokers with CB, and additionally include the airways of lungs resected from 11 asymptomatic smokers (AS) and 11 smokers with CB. Compared with HNS, biopsies in CB demonstrated increased numbers of 15-LOa mRNA+ cells (median: HNS = 31.3/mm(2) versus CB = 84.9/mm(2), P < 0.01) and protein+ cells (HNS = 2.9/mm(2) versus CB = 32.1/mm(2), P < 0.01). The HS group also showed a significant increase in protein+ cells (HNS = 2.9/mm(2) versus HS = 14/mm(2), P < 0.05). In the resected airways, 15-LOa protein+ cells in the submucosal glands of the CB group were more numerous than in the AS group (AS = 33/mm(2) versus CB = 208/mm(2); P < 0.001). 15-LOa mRNA+ and protein+ cells consistently outnumbered 15-LOb by approximately 7- and 5-fold, respectively (P < 0.01). Quantitative reverse transcriptase polymerase chain reaction of complementary biopsies confirmed the increased levels of 15-LOa in CB compared with that in either HNS or HS (P < 0.05). There was no difference between the subject groups with respect to 15-LOb expression. The numbers of cells expressing mRNA for 15-LOa in CB showed a positive association with those expressing interleukin (IL)-4 mRNA (r = 0.80; P < 0.01). We conclude that the upregulation of 15-LO activity in the airways of HS and of smokers with CB primarily involves the 15-LOa isoform: the functional consequences of its association the upregulation of IL-4 in chronic bronchitis requires further study.

We wished to determine if the inflammatory cells surrounding the airway mucus-secreting glands in chronic bronchitis (CB) were associated with interleukin (IL)-4 and IL-5 mRNA expression and whether the CD8 T cell population expressed these cytokines. Digoxigenin-labeled IL-4 and IL-5 antisense RNA probes were used to detect gene expression in 11 asymptomic smokers (AS), 11 smokers with CB alone with normal lung function, and 10 smokers with chronic bronchitis and coexisting chronic obstructive pulmonary disease (CB+COPD; FEV(1)% of predicted of 43-77% and FEV(1)/ FVC of 51-68%). There were approximately three times as many IL-4 than IL-5 mRNA(+) cells. The highest number of IL-4 mRNA(+) cells were in the submucosal glands of the CB group with normal lung function (216/mm(2)), significantly higher than the values in either the AS (63/mm(2)) or the CB+COPD (87/mm(2)) groups, respectively (p < 0.01). There were similar group differences when the total numbers of inflammatory cells were compared. Accordingly, there was a positive correlation between the number of IL-4 mRNA(+) cells and the total number of inflammatory cells in both the subepithelium and glandular compartments (r = 0.60; p = 0.01 and r = 0.70; p = 0.02, respectively). There were no significant associations between the numbers of CD8(+) and IL-4 or IL-5 mRNA(+) cells. Of 1328 IL-4(+) and 1404 CD8(+) cells counted none was double labeled. Of 727 IL-5(+) and 1569 CD8(+) cells, none was double labeled. In contrast, as a positive control, 34% of tumor necrosis factor (TNF)-alpha(+) cells were also CD8(+) and 15% of CD8(+) cells were TNF-alpha positive. Thus, cells other than the CD8(+) phenotype produce IL-4 and IL-5 in CB. We conclude that there is increased inflammation and IL-4 gene expression in the mucus-secreting glands and the airway mucosa of smokers with bronchitis: both are lower in those with CB and coexisting COPD suggesting that airway inflammation in CB is reduced when airway obstruction develops.

The efficiency of some components of the host defense system has been evaluated in 32 atopic patients with rhinitis, asthma or asthma and rhinitis. No alterations in components of complement (C3 and C4), serum immunoglobulins (IgG, IgA, and IgM) and neutrophil functional parameters were found. As expected, serum IgE values were higher than in controls. A decreased percentage of total circulating T lymphocytes, an increased percentage of Ia1-positive cells, but normal values of OKT4- and OKT8-positive T lymphocytes were demonstrated. However, a greater heterogeneity in the distribution of the suppressor-cytotoxic subset was shown in atopic patients.