Do Circulating Extracellular Vesicles Strictly Reflect Bronchoalveolar Lavage Extracellular Vesicles in COPD?

Cell-derived extracellular vesicles (EVs) found in the circulation and body fluids contain biomolecules that could be used as biomarkers for lung and other diseases. EVs from bronchoalveolar lavage (BAL) might be more informative of lung abnormalities than EVs from blood, where informa- tion might be diluted. To compare EVs’ characteristics in BAL and blood in smokers with and without COPD. Same-day BAL and blood samples were obtained in 9 nonsmokers (NS), 11 smokers w/o COPD (S), and 9 with COPD (SCOPD) (FEV1: 59 ± 3% pred). After differential centrifugation, EVs (200–500 nm diameter) were identified by flow cytometry and labeled with cell-type specific antigens: CD14 for macrophage-derived EVs, CD326 for epithelial-derived EVs, CD146 for endothelial-derived EVs, and CD62E for activated-endothelial-derived EVs. In BAL, CD14-EVs were increased in S compared to NS [384 (56–567) vs. 172 (115–282) events/μL; p = 0.007] and further increased in SCOPD [619 (224–888)] compared to both S (p = 0.04) and NS (p < 0.001). CD326-EVs were increased in S [760 (48–2856) events/μL, p < 0.001] and in SCOPD [1055 (194–11,491), p < 0.001] when compared to NS [15 (0–68)]. CD146-EVs and CD62E-EVs were similar in the three groups. In BAL, significant differences in macrophage and epithelial-derived EVs can be clearly detected between NS, S and SCOPD, while these differences were not found in plasma. This suggests that BAL is a better medium than blood to study EVs in lung diseases

Pretreatment rate of decay in forced vital capacity predicts long-term response to pirfenidone in patients with idiopathic pulmonary fibrosis.

Pirfenidone reduces functional decline and disease progression in patients with Idiopathic Pulmonary Fibrosis (IPF). However, response to treatment is highly heterogeneous. In this study, we evaluated whether response to pirfenidone is influenced by the pre-treatment rate of forced vital capacity (FVC) decline. Fifty-seven IPF patients were categorized as rapid (RP) or slow progressors (SP) based on whether their FVC decline in the year preceding pirfenidone treatment was > or <10% predicted. Patients were followed-up every 6 months and up to 24 months following institution of pirfenidone treatment. In the entire population, pirfenidone reduced significantly the rate of FVC decline from 222 ml/yr to 68 ml/yr at 12 month (p<0.01) and 86 ml/yr at 24 month (p=0.04) follow-up. In RP, the reduction of FVC decline was evident at 6 months (706 ml/yr pre-treatment vs 35 ml/yr; p<0.01) and maintained, though to a lesser degree, at 12 (105 ml/yr; p< 0.01) and 24 months (125 ml/yr; p<0.02). Conversely, among SP the reduction in FVC decline was not significant at any of the time points analyzed. Pirfenidone reduces significantly the rate of FVC decline in patients with IPF. However, the beneficial effect is more pronounced and long-lasting in patients with rapidly progressive disease

Dual polarization of human alveolar macrophages progressively increases with smoking and COPD severity

BACKGROUND: It is known that tissue macrophages derive not only from blood monocytes but also from yolk sac or fetal liver, and the tissue of residence guides their function. When isolated, they lose tissue specific signatures, hence studies of human macrophages should be ideally done directly in the tissue. The aim of this study was to investigate directly in human lung tissue the polarization of alveolar macrophage (AM), classic (M1) or alternative (M2), in health and disease, using COPD as a model. METHODS: Surgical lungs from 53 subjects were studied: 36 smokers whose FEV(1) varied from normal to severe COPD, 11 non-smokers and 6 normal donors. iNOS and CD206 immunohistochemistry was used to quantify the percentage of AM polarized as M1 or M2 in lung sections. RESULTS AND DISCUSSION: The percentage of M1 and M2 increased progressively with smoking and COPD severity, from 26% to 84% for M1 and from 7% to 78% for M2. In donors 74% of AM were negative for M1 and 93% for M2. Confocal microscopy showed co-localization of M1 and M2 in the same AM in severe COPD. CONCLUSION: In normal lungs alveolar macrophages were mostly non-polarized. With smoking and COPD severity, M1 and M2 polarization increased significantly and so did the co-expression of M1 and M2 in the same alveolar macrophage. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12931-017-0522-0) contains supplementary material, which is available to authorized users

Immune inflammation and disease progression in idiopathic pulmonary fibrosis

The clinical course in idiopathic pulmonary fibrosis (IPF) is highly heterogeneous, with some patients having a slow progression and others an accelerated clinical and functional decline. This study aims to clinically characterize the type of progression in IPF and to investigate the pathological basis that might account for the observed differences in disease behavior. Clinical and functional data were analyzed in 73 IPF patients, followed long-time as candidates for lung transplantation. The forced vital capacity (FVC) change/year (< or 6510% predicted) was used to define "slow" or "rapid" disease progression. Pathological abnormalities were quantified in the explanted lung of 41 out of 73 patients undergoing lung transplantation. At diagnosis, slow progressors (n = 48) showed longer duration of symptoms and lower FVC than rapid progressors (n = 25). Eleven slow and 3 rapid progressors developed an acute exacerbation (AE) during follow-up. Quantitative lung pathology showed a severe innate and adaptive inflammatory infiltrate in rapid progressors, markedly increased compared to slow progressors and similar to that observed in patients experiencing AE. The extent of inflammation was correlated with the yearly FVC decline (r = 0.52, p = 0.005). In conclusion an innate and adaptive inflammation appears to be a prominent feature in the lung of patients with IPF and could contribute to determining of the rate of disease progression

From cigarette smoking to Chronic Obstructive Pulmonary Disease or Idiopathic Pulmonary Fibrosis. Why?

Chronic Obstructive Pulmonary Disease (COPD) and Idiopathic Pulmonary Fibrosis (IPF) are two chronic lung diseases with distinct clinical, pathological and epidemiologic features. In both disorders cigarette smoking represents a major risk factor; indeed, it causes a repetitive alveolar damage that elicit inflammatory response and may enhance the establishment of the disease. During my PhD, we investigated the role of innate and adaptive immune response in COPD and IPF, and how it may be related to different clinical outcomes. In COPD, the role of inflammation is consolidated; alveolar macrophages (AM), which are one of the main actors of innate immune response, were firstly recognized to play a key role in the pathogenesis of the disease, by releasing protease that damage lung parenchyma. In our studies, we investigated how AM may be involved in the development of COPD with alternative mechanisms; for example, we analyzed AM polarization, classic (M1) and alternative (M2), in lung tissue and how it may be influenced by smoking and disease severity. The model of COPD as a disease caused only by an antiprotease-protease imbalance has been overtaken; indeed, not all subjects with α1-Antitrypsin (AAT) deficiency (AATD) develop disease, meaning that other immune regulatory factors are involved. In fact, we investigated α1AT polymerization in AM as a pro-inflammatory trigger, due to its abnormal accumulation in AM endoplasmatic reticulum. Moreover, we performed a genome wide analysis in siblings with AATD, with extreme phenotypes of the disease (emphysema/no emphysema), searching for genes variants that may modulate or promote inflammatory response, possibly explain these opposite manifestations. Since the study of biomarkers is becoming of great interest in the field, we focused our research on adaptive immune cells in the blood, examining blood eosinophils (BE) and lymphocytes (BL), to investigate whether they can be predictor of clinical outcomes, and if they reflect what is happening in the lung. In addition, we have started to unravel the possible role of the peripheral BL in the development or prevention of cancer. Then we moved from peripheral blood to the lung tissue, examining how lymphocyte and eosinophil counts in the lung may change after the introduction of an anti-inflammatory treatment with Roflumilast, adding some clues on its mechanisms of action. Indeed, the study of lung infiltrate is mandatory to better understand the pathogenesis of the disease, and we are currently developing new techniques to deeply analyze the immune cells in COPD lung, and the presence of markers of immunomodulation. In IPF, the role of inflammation is more debated than COPD. Our idea that an inflammatory reaction could be an important part of IPF has lead us to investigate the possible role of innate and adaptive immune response in the pathogenesis of the disease, not only in explanted lungs of end-stage IPF, but also in the surgical lung biopsies of patients with mild disease. The pathological evidences we found were then correlated to clinical outcomes, as disease progression, to study if they may account for the variability in lung function decline. Moreover, we also explored whether different lung tissue inflammatory profiles may impact on the response to Pirfenidone treatment, which exerts also an anti-inflammatory effect. Finally, we investigated HRCT as a possible non-invasive predictor of disease behavior and treatment response in IPF. The results of the studies we have conducted so far, highlights how inflammatory response, and especially the immune regulation, is pivotal in COPD and IPF, adding novel findings to previous knowledge in the areas of innate and adaptive inflammation, and putting the basis for future research in these smoking-induced diseases

The Role of Bronchoscopy in the Diagnosis and Management of Patients with SARS-Cov-2 Infection

Bronchoscopy has several major diagnostic and therapeutic indications in pulmonology. However, it is an aerosol-generating procedure that places healthcare providers at an increased risk of infection. Now more than ever, during the spread of the coronavirus disease 2019 (COVID-19) pandemic, the infectious risk during bronchoscopy is significantly raised, and for this reason its role in diagnostic management is debated. In this review, we summarized current evidence regarding the indications for bronchoscopy and the measures that should be applied to decrease risk exposure. Indeed, seeing the long-lasting period of the pandemic, resuming standard of care for all patients is required

From cigarette smoking to Chronic Obstructive Pulmonary Disease or Idiopathic Pulmonary Fibrosis. Why?

Chronic Obstructive Pulmonary Disease (COPD) and Idiopathic Pulmonary Fibrosis (IPF) are two chronic lung diseases with distinct clinical, pathological and epidemiologic features. In both disorders cigarette smoking represents a major risk factor; indeed, it causes a repetitive alveolar damage that elicit inflammatory response and may enhance the establishment of the disease. During my PhD, we investigated the role of innate and adaptive immune response in COPD and IPF, and how it may be related to different clinical outcomes. In COPD, the role of inflammation is consolidated; alveolar macrophages (AM), which are one of the main actors of innate immune response, were firstly recognized to play a key role in the pathogenesis of the disease, by releasing protease that damage lung parenchyma. In our studies, we investigated how AM may be involved in the development of COPD with alternative mechanisms; for example, we analyzed AM polarization, classic (M1) and alternative (M2), in lung tissue and how it may be influenced by smoking and disease severity. The model of COPD as a disease caused only by an antiprotease-protease imbalance has been overtaken; indeed, not all subjects with α1-Antitrypsin (AAT) deficiency (AATD) develop disease, meaning that other immune regulatory factors are involved. In fact, we investigated α1AT polymerization in AM as a pro-inflammatory trigger, due to its abnormal accumulation in AM endoplasmatic reticulum. Moreover, we performed a genome wide analysis in siblings with AATD, with extreme phenotypes of the disease (emphysema/no emphysema), searching for genes variants that may modulate or promote inflammatory response, possibly explain these opposite manifestations. Since the study of biomarkers is becoming of great interest in the field, we focused our research on adaptive immune cells in the blood, examining blood eosinophils (BE) and lymphocytes (BL), to investigate whether they can be predictor of clinical outcomes, and if they reflect what is happening in the lung. In addition, we have started to unravel the possible role of the peripheral BL in the development or prevention of cancer. Then we moved from peripheral blood to the lung tissue, examining how lymphocyte and eosinophil counts in the lung may change after the introduction of an anti-inflammatory treatment with Roflumilast, adding some clues on its mechanisms of action. Indeed, the study of lung infiltrate is mandatory to better understand the pathogenesis of the disease, and we are currently developing new techniques to deeply analyze the immune cells in COPD lung, and the presence of markers of immunomodulation. In IPF, the role of inflammation is more debated than COPD. Our idea that an inflammatory reaction could be an important part of IPF has lead us to investigate the possible role of innate and adaptive immune response in the pathogenesis of the disease, not only in explanted lungs of end-stage IPF, but also in the surgical lung biopsies of patients with mild disease. The pathological evidences we found were then correlated to clinical outcomes, as disease progression, to study if they may account for the variability in lung function decline. Moreover, we also explored whether different lung tissue inflammatory profiles may impact on the response to Pirfenidone treatment, which exerts also an anti-inflammatory effect. Finally, we investigated HRCT as a possible non-invasive predictor of disease behavior and treatment response in IPF. The results of the studies we have conducted so far, highlights how inflammatory response, and especially the immune regulation, is pivotal in COPD and IPF, adding novel findings to previous knowledge in the areas of innate and adaptive inflammation, and putting the basis for future research in these smoking-induced diseases

A rejuvenation effect of the antifibrotic therapy correlates with lung function improvement in IPF patients

Idiopathic pulmonary fibrosis (IPF) is an irreversible and fatal lung disease destined to increase as a sequel of the Severe Acute Respiratory Syndrome Coronavirus-2. Several studies implicate telomere shortening as hallmark in the pathomechanism of IPF. Our understanding of disease pathogenesis remains, however, incomplete, which hampers also the development of more effective drugs. Aims of this study are: 1) to analyze telomere length in blood leukocytes (LTL) in well phenotypically characterized IPF patients at diagnosis, before treatment with antifibrotic drugs, Pirfenidone and Nintedanib (T0); 2) to investigate in follow up (T1) a possible change in the rate of LTL by combining data on clinic, hematochemical tests, respiratory function and occupational exposure. We examined a group of 24 IPF patients that underwent to a therapy follow up (T1) (mean±SD 297±124 days). We observed an increase in LTL at T1 compared to LTL at T0 (mean±SD LTL (T/S), T1 vs T0, 1.29 ± 0.26 vs 1.19 ± 0.27; p=0.051). Multiple linear regression analysis reveals that the LTL increase, at T1, is significantly related to the time of treatment (p=0.002), LTL at T0 (p<0.0001) and to a slowing in lung function decline (FVC%pred) (p=0.054). The other variables considered including occupational exposure, pack-years, occupational risk factor, gender and body mass index are not significantly related. Our research would indicate a rejuvenation effect of the antifibrotic therapy by measuring the LTL that correlates with lung function improvement. This suggests that targeting fundamental mechanisms of cellular aging has the potential to interfere with the severity of the disease

Extracellular Vesicles in Lung Cancer: Bystanders or Main Characters?

Lung cancer still represents the main cause of cancer death worldwide. The poor survival is mainly related to the diagnosis which is often obtained in advanced stages when the disease is unresectable and characterized by the worst prognosis. Only in the last decades have great discoveries led to the development of new therapies targeted to oncogenes and to boost the host immune response against the tumor. Tumor identification and molecular/immunological characterization rely on bioptic samples which represent the gold standard for diagnosis. Nonetheless, less invasive procedures providing small samples will be more and more common in the future. Extracellular vesicles (EV), submicron particles released by any cell type, are candidates for diagnostic and prognostic biomarkers. EV are mediators of intercellular communication and can convey cytokines, miRNAs, antigens, and many other factors of tumorigenesis. This review summarizes the most appealing findings on lung-cancer-related EV, debating the evidence on circulating versus airway EV as potential biomarkers in disease management and the main studies on the role of these particles on lung cancer pathogenesis. Overall, the available results point toward a wide range of possible applications, supported by the promising achievements of genotyping on BAL fluid EV and proteomic analysis on pleural effusion EV. Nonetheless, the study of lung EV is still affected by remarkable methodological issues, especially when in vitro evidence is translated into humans. Whether EV still represent an “information fog” or can be useful in lung cancer management will be discussed, with possible hints on how to improve their usage