Lung transplant (LTX) patients are at high risk of cytomegalovirus (CMV) infection, which is often associated with high mortality and morbidity. Reactivation of CMV causes cell injury due to the cytopathic effect of viral replication and triggering of T cell immunity. The aim of this study was to compare expression of immune checkpoints (ICs) (PD-1, CTLA-4, LAG-3 and TIGIT) in CD4, CD8 and CD56 and activation markers CD137, CD154 and CD69 of end-stage patients awaiting lung transplant. Eighteen pre-LTX positive for anti-CMV IgG titres and 18 healthy subjects were enrolled. IC and activation markers have been evaluated through flow cytometric analysis in HC and pre-LTX patients. Reactive (QF+) and unreactive (QF-) patients were stratified according to QuantiFERON-CMV assays. ICs' and activation markers' expression were determined before and after in vitro stimulation with pp-65 and IE-1 antigens. Lower expression of PD-1 was observed in CD4 and CD8 cells of pre-LTX patients than controls, whereas CTLA4 appeared upregulated in CD56 and CD8 cells. TIGIT is increased on the surface of CD4, CD8 and NK cells after peptide stimulation in QF-negative patients and PD-1 is only downregulated after stimulation in the QF-positive patients. This study provides new evidence of immune dysregulation in patients with end-stage lung disorders, particularly in relation to immune checkpoint cell biology. The change in QF+ mostly happens on cytotoxic cells NK and CD8, while the changes in QF- were observed in adaptive immune cells, including CD4 and CD8.

Background: Fractional exhaled nitric oxide (FeNO) is a biomarker of airway inflammation associated with airway hyper-responsiveness and type-2 inflammation. Its role in the management of severe asthmatic patients undergoing biologic treatment, as well as FeNO dynamics during biologic treatment, is largely unexplored. Purpose: The aim was to evaluate published data contributing to the following areas: (1) FeNO as a predictive biomarker of response to biologic treatment; (2) the influence of biologic treatment in FeNO values; (3) FeNO as a biomarker for the prediction of exacerbations in patients treated with biologics. Methods: The systematic search was conducted on the Medline database through the Pubmed search engine, including all studies from 2009 to the present. Results: Higher baseline values of FeNO are associated with better clinical control in patients treated with omalizumab, dupilumab, and tezepelumab. FeNO dynamics during biologic treatment highlights a clear reduction in FeNO values in patients treated with anti-IL4/13 and anti-IL13, as well as in patients treated with tezepelumab. During the treatment, FeNO may help to predict clinical worsening and to differentiate eosinophilic from non-eosinophilic exacerbations. Conclusions: Higher baseline FeNO levels appear to be associated with a greater benefit in terms of clinical control and reduction of exacerbation rate, while FeNO dynamics during biologic treatment remains a largely unexplored issue since few studies have investigated it as a primary outcome. FeNO remains detectable during biologic treatment, but its potential utility as a biomarker of clinical control is still unclear and represents an interesting research area to be developed.

Obstructive sleep apnea (OSA) is often observed in subjects with interstitial lung disease (ILD). It may have a negative impact on the course of ILD, but its prognostic significance in relation to other known indicators of poor outcome is unclear. After a detailed work-up, including overnight unattended type III polygraphy, all subjects newly diagnosed with ILDs referred to our clinics were followed-up for at least 1.5 years or until death or progression of disease [> 10% decline in forced vital capacity (FVC) below baseline]. We analyzed relationships between some prespecified variables of interest, including sleeping results, to establish parameters predictive of progressive course. Our population consisted of 46 subjects (mean age 59.6 years; males 61%); 23.9% and 41% had idiopathic pulmonary fibrosis and ILD associated with systemic diseases, respectively. Mean baseline forced vital capacity and diffusion capacity of carbon monoxide were 83% and 57% of predicted, respectively. Mean (± SE) Apnea–Hypopnea Index (AHI) was 17 (± 3) events/h. AHI in the ranges 5–14.9, 15–29.9, and ≥ 30 was recorded in 14 (31%), 6 (13%), and 9 (20%) subjects, respectively. Mean distance covered in the 6-MWG walk test (6MWT) was 302 (± 19) m and 26 subjects (57%) showed exertional oxyhemoglobin desaturation. The median follow-up was about 18 months. Multivariate logistic regression analysis showed that exertional desaturation (HR 8.2; 1.8–36.5 95% CI; p = 0.006) and AHI ≥ 30, namely the threshold of severe OSA (HR 7.5; 1.8–30.6; p = 0.005), were the only independent variables related to progressive disease course. We conclude that exertional desaturation and elevated AHI had independent negative prognostic significance in our ILD population.

Fibrotic hypersensitivity pneumonitis (fHP) is a frequently misdiagnosed fibrosing interstitial pneumonia, which often remains undiagnosed due to the lack of uniformity of diagnostic criteria. Its features are similar to those of other ILDs, especially idiopathic pulmonary fibrosis (IPF), and biomarkers with potential clinical value have been proposed. We reviewed the recent literature on serum and BAL biomarkers, focusing on their clinical role in the diagnosis and management of fHP. We searched Medline/Pubmed results from 2005 until April 2020. The manuscripts of interest selected by our search were limited in number and proposed different clinical biomarkers in serum (IgG antibodies, macrophage inflammatory proteins-1, epithelial cell proteins) and BAL (lymphocytes, T-cell mediators). This is the first review to summarize all the serum and BAL biomarkers for fHP proposed in the literature. This review summarized the main biomarkers investigated in fibrotic hypersensitivity pneumonitis because an urgent aim of subsequent research will be to validate and standardize them for diagnostic purposes.

Background: Nintedanib is an oral multitarget tyrosine kinase inhibitor approved for the treatment of patients with idiopathic pulmonary fibrosis (IPF). Recent evidence demonstrated that nintedanib reduced functional disease progression also in subjects with non-IPF progressive fibrosing interstitial lung disease (PF-ILD). However, real-life data on the effectiveness of nintedanib in PF-ILD and familial pulmonary fibrosis (FPF) are lacking. Methods: this retrospective monocentric study enrolled 197 patients affected with IPF, PF-ILD and FPF treated with nintedanib at the Referral Centre of Siena from 2014 to 2021. Pulmonary functional tests and survival data were collected throughout the observation period for the evaluation of mortality and disease progression outcomes. Results: nintedanib treatment significantly reduced the FVC decline rate in IPF and PF-ILD subgroups, but not in FPF subjects. No significant differences were observed among the subgroups in terms of survival, which appeared to be influenced by gender and impaired lung function (FVC < 70% of predicted value). Concerning disease progression rate, a diagnosis of FPF is associated with more pronounced FVC decline despite nintedanib treatment. Conclusions: our research studies the effectiveness and safety of nintedanib in reducing functional disease progression of IPF and PF-ILD. FPF appeared to be less responsive to nintedanib, even though no differences were observed in terms of survival.

Background Cytomegalovirus (CMV) is the major and most common opportunistic infection complicating lung transplant (LTX). The aim of this study was to analyse the epidemiological aspects of CMV infection in lung transplant patients subject to a pre-emptive anti-CMV approach and to study the impact of this infection on lung transplant outcome, in terms of onset of chronic lung allograft dysfunction (CLAD). Methods This single-centre retrospective study enrolled 87 LTX patients (median age 55.81 years; 41 females, 23 single LTX, 64 bilateral LTX). All patients were managed with a pre-emptive anti-CMV approach. The incidences of the first episode of CMV infection, 1, 3, 6 and 12 months after LTX, were 12.64%, 44.26%, 50.77% and 56.14%. A median interval of 41 days elapsed between LTX and the first episode of CMV infection. The median blood load of CMV-DNA at diagnosis was 20,385 cp/ml; in 67.64% of cases, it was also the peak value. Patients who had at least one episode had shorter CLAD-free survival. Patients who had three or more episodes of CMV infection had the worst outcome. Results CMV infection was confirmed to be a common event in lung transplant patients, particularly in the first three months after transplant. It had a negative impact on transplant outcome, being a major risk factor for CLAD. The hypothesis that lower viral replication thresholds may increase the risk of CLAD is interesting and deserves further investigation.

This narrative paper reviews the current knowledge of Home Oxygen Therapy (HOT) in stable Chronic Obstructive Pulmonary Disease (COPD) and Interstitial Lung Disease (ILD), two major causes of Long-Term Oxygen Therapy (LTOT) prescription. There is evidence that LTOT improves survival in COPD subjects with chronic severe respiratory failure. HOT is also used to contrast exercise and sleeping hypoxemia and to improve Quality of Life (QoL) and symptoms. Ambulatory Oxygen Therapy (AOT) did not assure generalized improvements in symptoms and Quality of Life (QoL) of COPD subjects. There is short-term evidence in a real-life study that AOT may improve QoL in ILD subjects with Exercise Oxygen Desaturation (EOD) and exertional dyspnea. There are some differences between guidelines and practices, which translate into variations in characteristics and rates of ILD and COPD subjects admitted to LTOT and AOT. Indications on titration of oxygen flow and the best oxygen delivery device for optimal management of AOT in COPD and ILD subjects are often vague or lacking. More work is needed for optimizing and customizing HOT in COPD and ILD subjects.

Background: Idiopathic pulmonary fibrosis (IPF) is a chronic progressive interstitial lung disease limited to the lungs. Immunological dysregulation may significantly participate in the pathophysiology of IPF. The immunological responses to nintedanib therapy in IPF patients were investigated for the first time in this study. Materials and methods: Fifty IPF patients (median age (IQR) 69 (65-75) years; 38 males), were selected retrospectively. Flowcytometry analysis were performed to phenotype immunological biomarkers in peripheral blood from IPF patients after 1 year of antifibrotic therapy and a group of healthy volunteers. Results: Before starting antifibrotic treatment, IPF patients showed increased CD1d+CD5+ (p = 0.0460), Treg (p = 0.0354), T effector (CD25(high)CD127(high)) (p = 0.0336), central cells (CD4(+)CD45RA(-)) (p = 0.0354), effector cells (CD4(+)CD45RA(+)) (p = 0.0249) and follicular cell percentages (p = 0.0006), notably Tfh1 (p = 0.0412) and Tfh17 (p = 0.0051) cell percentages, in respect with healthy controls (HC). After nintedanib therapy, Breg (p = 0.0302), T effector (p = 0.0468), Th17.1 (p = 0.0146) and follicular cells (p = 0.0006), notably Tfh1 (p = 0.0006) and Tfh17 (p = 0.0182) cell percentages, were significantly decreased. In the logistic regression, Tfh panel showed a significant area under the receiver operating characteristics curve (AUROC) to distinguish IPF than HC (90.5%), as well as t0 and t1 (99.3%). Conclusion: In conclusion, the immunological results obtained in this study demonstrate that nintedanib significantly helps to restore immunological responses in IPF patients. These findings will be useful in the search for biomarkers predictive of response to antifibrotic treatment.

Background: Severe acute respiratory syndrome caused by novel coronavirus 2 (SARS-CoV-2) emerged in Wuhan (China) in December 2019. Here we evaluated a panel of biomarkers to phenotype patients and to define the role of immuno-inflammatory mediators as biomarkers of severity. Materials and methods: Serum samples were obtained from 24 COVID-19 patients on admission to hospital, before any treatment or infusion of intravenous steroids or invasive ventilation. KL-6 IL-6 and C-peptide were measured by chemiluminescent enzyme immunoassay. IL-6 assay was validated for accuracy and precision. The validity of variables used to distinguish severe from mild-to-moderate patients was assessed by areas under curves (AUC) of the receiver operating characteristic (ROC) and logistic regression was performed to combine parameters of the two groups. Results: In the severe group, IL-6, CRP and KL-6 concentrations were significantly higher than in mild-to moderate patients. KL-6, IL-6 and CRP concentrations were directly correlated with each other. ROC curve analysis of the logistic regression model including IL-6, KL-6 and CRP showed the best performance with an AUC of 0.95. Conclusions: Besides corroborating previous reports of over-expression of IL-6 in severe COVID-19 patients requiring mechanical ventilation, analytical determination of other mediators showed that IL-6 concentrations were correlated with those of KL-6 and CRP. The combination of these three prognostic bioindicators made it possible to distinguish severe COVID-19 patients with poor prognosis from mild-to-moderate patients.

Background: Krebs von den Lungen-6 (KL-6) was suggested as ILD biomarker including idiopathic pulmonary fibrosis (IPF). Lung cancer is one of the most severe comorbidity of IPF patients. This study aims to serially analyze KL-6 in IPF patients after 24 months of Nintedanib and to first investigate the biomarker behavior in IPF associated with adenocarcinoma. Materials and methods: One hundred and forty-two ILD patients (median (IQR), 69 (63–75) years; 86 males) were retrospectively enrolled. Serial serum samples were collected from IPF patients before starting antifibrotic therapy and after 12 months. Serum KL-6 levels were measured by KL-6 reagent assay (Fujirebio Europe, UK). Results: Increased KL-6 concentrations were identified in IPF-LC patients than IPF, fibrotic hypersensitivity pneumonitis, and pulmonary fibrosis associated with autoimmune disease groups. A cut-off value was calculated to distinguish IPF and IPF-LC patients. IPF patients monitored for 24 months with Nintedanib showed persisted increased levels of KL-6 with a progressive decline of FVC percentages. Conclusion: This preliminary study offers a first demonstration that very high serum concentrations of KL-6 in IPF-LC patients are associated with poor prognosis. Moreover, serial evaluation of serum KL-6 in IPF patients over 24 months of Nintedanib treatment revealed that most patients experienced a stabilization of lung function parameters and of serum concentrations of KL-6.

Seven species of coronavirus cause acute respiratory illness in humans. Coronavirus HKU 1 (CoV HKU 1) was first described in 2005 in an adult patient with pneumonia in Hong Kong. Although it is a well-known respiratory tract pathogen, there is not much information about its role in hospitalized adults, especially in southern Europe. Here, we describe a case of radiologically demonstrated CoV HKU 1-related bronchiolitis with acute respiratory failure in an adult female without significant comorbidities except obesity.

Sarcoidosis is a multi-systemic granulomatous disease of unknown origin. Recent research has focused upon the role of autoimmunity in its development and progression. This study aimed to determine and define the disturbance and distribution of T and B cell subsets in the alveolar and peripheral compartments. Thirteen patients were selected for the study [median age, interquartile range (IQR) = 57 years (48-59); 23% were male]. Twelve healthy controls [median age, IQR = 53 years (52-65); 16% male] were also enrolled into the study. Cellular and cytokine patterns were measured using the cytofluorimetric approach. Peripheral CD8 percentages were higher in sarcoidosis patients (SP) than healthy controls (HC) (p = 0.0293), while CD4 percentages were lower (p = 0.0305). SP showed low bronchoalveolar lavage (BAL) percentages of CD19 (p = 0.0004) and CD8 (p = 0.0035), while CD19(+)CD5(+)CD27(-) percentages were higher (p = 0.0213); the same was found for CD4 (p = 0.0396), follicular regulatory T cells (T-reg) (p = 0.0078) and T-reg (p < 0.0001) cells. Low T helper type 17 (Th17) percentages were observed in BAL (p = 0.0063) of SP. Peripheral CD4(+) C-X-C chemokine receptor (CXCR)5(+)CD45RA(-)) percentages and follicular T helper cells (Tfh)-like Th1 (Tfh1) percentages (p = 0.0493 and p = 0.0305, respectively) were higher in the SP than HC. Tfh1 percentages and Tfh-like Th2 percentages were lower in BAL than in peripheral blood (p = 0.0370 and p = 0.0078, respectively), while CD4(+) C-X-C motif CXCR5(+)CD45RA(-) percentages were higher (p = 0.0011). This is the first study, to our knowledge, to demonstrate a link between an imbalance in circulating and alveolar Tfh cells, especially CCR4-, CXCR3- and CXCR5-expressing Tfh subsets in the development of sarcoidosis. These findings raise questions about the pathogenesis of sarcoidosis and may provide new directions for future clinical studies and treatment strategies.

BACKGROUND:Pirfenidone and nintedanib are the sole pharmacological therapies currently approved for idiopathic pulmonary fibrosis (IPF). Limited comparison data is available in literature, despite they are both prescribed for mild-to-moderate disease. Here, we describe our almost 10 years real-life experience with antifibrotic treatment to investigate potential differences in terms of efficacy.POPULATION AND METHODS:We retrospectively recruited patients diagnosed with IPF and treated with pirfenidone or nintedanib at Siena Referral Center. Clinical, functional, safety and radiological data was collected at baseline and during the follow-up, according to our Center protocol.RESULTS:We retrospectively recruited 263 IPF patients (139 treated with pirfenidone and 124 with nintedanib) in the study. After 885.3 ± 559.5 days of observation, the median survival was 1224 days. No significant differences were found between pirfenidone and nintedanib in terms of survival and time to decline of forced vital capacity >10% (p = 0.8786 and p = 0.1677, respectively). A smaller lung diffusion for carbon monoxide (DL CO ) decrease was found after 1 year of therapy with nintedanib in respect to pirfenidone (p = 0.0167). Overall, 21 patients permanently discontinued antifibrotic treatment due to side effects (14 with pirfenidone, 7 with nintedanib); no fatal adverse events were recorded.DISCUSSION:Our results showed a similar effectiveness between pirfenidone and nintedanib in terms of mortality and functional disease progression. Both drugs confirmed their good tolerability profile and no new safety alerts were observed. Nintedanib was associated with a smaller reduction of DL CO after 1 year of follow-up compared with pirfenidone, maybe due to its antiangiogenic properties.

Background: Idiopathic pulmonary fibrosis (IPF) is a severe progressive interstitial lung disease. At 5-year follow-up, 15% of IPF patients develop lung cancer, which significantly reduces the survival rate. Here we review the literature on the clinical role of oncomarkers in IPF progression, and describe the trend of routine oncomarkers in IPF patients over the longest follow-up yet reported. Materials and methods: A systematic search of the literature in PubMed was performed to find relevant studies published up to 24 September 2020. The most common oncomarkers were chosen to select papers related to pulmonary fibrosis. Then, 24 IPF patients and 25 non-IPF patients, followed at Careggi ILD Referral Centre and Siena Regional Referral Centre for ILD, were enrolled consecutively. Results: A few studies reported an association between serum oncomarkers and severity of IPF. NSE, CEA, Ca19.9, and Ca125 were higher in the IPF, than in the non-IPF, group at every follow-up (p < 0.05). Ca15.3 concentrations were higher in the IPF, than the non-IPF, group at t3 (p = 0.0080) and t4 (p = 0.0168). To improve the specificity and sensitivity of Ca15.3, a panel of biomarkers was analyzed, with the IPF group as dependent variable, and chitotriosidase, Cyfra 21.1, Ca15.3, Ca125, and Ca19.9 as independent variables. Conclusions: This study focused on the discovery of multiple biomarker signatures, such as combinations of oncomarkers, that are widely and routinely available in biochemistry laboratories. The combination of clinical parameters and biological markers could help achieve more accurate results regarding prognosis and response to treatment in IPF. Our results could pave the way for a more “personalized” medical approach to patients affected by IPF.

Pulmonary hypertension (PH) is defined as an elevated mean pulmonary artery pressure at rest (mPAP ≥ 25 mmHg), evaluated by right heart catheterization (RHC). The aim of the present study was to evaluate HRCT findings in relation to transthoracic echocardiographic data to better characterize PH in IPF patients and to identify a non-invasive composite index with high predictive value for PH in these patients. 37 IPF patients were enrolled in this retrospective study. All patients underwent a complete assessment for PH, including transthoracic Doppler echocardiography, HRCT scan and right heart catheterization. Right heart catheterization was done in 19 patients (51.3%) as pre-lung transplant assessment and in 18 patients (48.6%) to confirm PH, suspected on the basis of echocardiography. Twenty out of 37 patients (54%) were confirmed to have PH by RHC. Multivariate regression showed that the combination of sPAP, PA area measured by HRCT and the ratio of the diameter of the segmental artery to that of the adjacent bronchus in the apicoposterior segment of the left upper lobe was strongly correlated with mPAP (R2 = 0.53; p = 0.0009). The ROC analysis showed that 931.6 was the ULN for PA area, with 86% sensitivity and 61% specificity (0.839 AUC); 20.34 was the ULN for the ratio of PA area to ascending aorta diameter, with 100% sensitivity and 50% specificity (0.804 AUC). The composite index proposed in the present study could help early detection of IPF patients suspected of PH requiring confirmation by RHC (if deemed clinically necessary).

Adipokines (APN) are mainly secreted by adipocytes, macrophages and various other cells, along with their role in the regulation and mediation of inflammatory responses. APN is almost exclusively synthesized by adipocytes and regulated by peroxisome proliferator-activated receptor γ (PPARγ) that is involved in the epithelial-mesenchymal transition, linked lung fibrosis. Leptin is involved in acute lung injury with a role in lung fibrogenesis. Little is known about the relationship between APN/leptin and idiopathic pulmonary fibrosis (IPF) and the few studies available in the literature used ELISA to detect these lipid mediators. Our study is also the first to measure adipokines by the new multiplex assay and for the first time were performed in bronchoalveolar lavage (BAL) from IPF patients. This preliminary study suggests that APN levels in serum could be useful for predicting the prognosis of IPF, as they are inversely correlated with DLco percentages and BMI. Moreover, this first analysis of APN in BAL from IPF patients by a new method demonstrated an inverse correlation between these levels and BMI values and a direct correlation with eosinophil percentages, both of which are negative prognostic factors of IPF.

After publication of our article [1] the authors have notified us that one of the names has been incorrectly tagged.

Benralizumab and mepolizumab are new therapies for severe eosinophilic asthma. Theyare both humanized IgG antibodies, targeting the IL‐5 receptor and IL‐5, respectively, suppressing the corresponding pathways. No specific biomarkers have been proposed to evaluate treatment response to benralizumab or mepolizumab. The aim of this proteomic study was to compare serum protein profiles of patients with severe eosinophilic asthma before and after anti‐IL5 or anti‐IL5R therapies. Proteomic analysis highlighted 22 differently abundant spots. Among the proteins identified, CAYP1, A1AT and A2M expression was significantly modified in both groups of patients after therapies while ceruloplasmin showed a significant modification in the group of benralizumab treatment. These differentially expressed proteins could be potential biomarkers of response to mepolizumab and benralizumab treatments and need further evaluation.

Background: Calcium methabolism alterations are quite common in sarcoidosis and have been correlated with disease activity. Aims and Objectives: The aim of the study is to investigate the clinical significance of calcium methabolism alterations in patients with chronic sarcoidosis. We paid particular attention for associations with specific disease phenotypes and chitotriosidase expression. Methods: 212 chronic sarcoidosis patients (97 males, 56.07±12 years old), followed at Referral Centre of Siena, were retrospectively recruited in the study. Demographic, clinical, functional, radiological data and serum-urinary calcium methabolism were entered in a electronical database for analysis. Chitotriosidase and ACE levels were measured too as well as bone mineral density and lung function tests. Results: Hypercalciuria and hypercalcemia were observed in 18.8% and 1.8% of patients, respectively. Urinary calcium levels correlated with chitotriosidase activity (r=0.33, p=0.0042). Patients with worsening persistent disease showed the highest levels of urinary calcium. DLco percentages correlated inversely with urinary calcium (r=0.1482; p=0.0397). Conclusions: Calcium metabolism alterations, particularly hypercalciuria, was observed in a significant percentage of patients of sarcoidosis. Urinary calcium correlated with clinical status, DLCO and serum chitotriosidase activity, suggesting its potential role as biomarker of activity and severity of sarcoidosis.