Predictive role of natural killer cells in bronchoalveolar lavage fluid of patients with sarcoidosis.

Sarcoidosis is a rare granulomatous disease of unknown aetiology belonging to the wide group of interstitial lung diseases.). Although the limitlessness of BAL fluid is debated, it remains one of the best matrices for studying the pathogenesis of sarcoidosis. Natural killer (NK) cells have been described in BAL fluid from sarcoidosis patients. Elevated NK cells in BAL fluid from sarcoidosis patients have been found to be associated with poor outcomes. In this study, NK cells were evaluated in BAL samples from sarcoidosis patients at the time of diagnosis and associated with clinical characteristics in order to evaluate their prognostic role. Of the 276 patients suspected to have sarcoidosis on the basis of clinical and radiological findings, 248 had a final diagnosis of sarcoidosis. Clinical parameters, Scadding stage, and extrapulmonary localization were collected in a database. It resulted in fibrotic sarcoidosis patients being associated with an increase in lymphocyte percentages in BAL samples, particularly NK cells when compared with other groups. From ROC analysis, NK cell percentages in BAL samples resulted as being the best predictive markers in discriminating stage 4 of sarcoidosis from other RX stages (AUC=0.85, p<0.0001). Furthermore, after the stratification of patients on the basis of the number of extrapulmonary localizations, patients with an higher number of extrapulmonary localizations also showed higher percentages of NK cells in BAL fluid. In conclusion, NK cell percentages in BAL fluid can be considered a good prognostic marker of fibrotic phenotypes of sarcoidosis and involvement of other organs, although their diagnostic utility was poor.

Collagen Type IV Alpha 5 Chain in Bronchiolitis Obliterans Syndrome After Lung Transplant: The First Evidence.

INTRODUCTION: Bronchiolitis obliterans syndrome (BOS) is the most common form of CLAD and is characterized by airflow limitation and an obstructive spirometry pattern without parenchymal opacities. The protein signature of BOS lesions concerns extracellular matrix organization and aberrant basement membrane composition. In this pilot study, we investigated the presence of COL4A5 in the serum of patients with BOS. METHODS: 41 patients who had undergone LTX were enrolled. Of these, 27 developed BOS and 14 (control group) were considered stable at the time of serum sampling. Of BOS patients, serum samples were analysed at the time of BOS diagnosis and before the clinical diagnosis (pre-BOS). COL4A5 levels were detected through the ELISA kit. RESULTS: Serum concentrations of COL4A5 were higher in pre-BOS than in stable patients (40.5 ± 13.9 and 24.8 ± 11.4, respectively, p = 0.048). This protein is not influenced by comorbidities, such as acute rejection or infections, or by therapies. Survival analysis also reveals that a higher level of COL4A5 was also associated with less probability of survival. Our data showed a correlation between concentrations of COL4A5 and FEV1 at the time of diagnosis of BOS. CONCLUSION: Serum concentrations of COL4A5 can be considered a good prognostic marker due to their association with survival and correlation with functional parameters.

Influence of allergic status and nasal polyposis on long-term Benralizumab response in eosinophilic severe asthma.

Objective: It is unclear whether Benralizumab effectiveness in severe eosinophilic asthma can be influenced by nasal polyposis (NP) or allergic status associations. We evaluated whether Benralizumab long-term efficacy in asthma outcomes could be different in subjects with atopy (SAEA) compared to the effectiveness in those without allergies (SNAEA) and in individuals with NP compared to those without NP. Methods: This observational retrospective study considered 95 consecutive patients divided into allergic (SAEA; n:65[68.4%]; skin prick tests positive [SPT] and/or IgE values ≥100 UI/mL), and non-allergic (SNAEA; n:30[31.6%], SPT negative and normal IgE levels<100 UI/mL). Overall population was also divided into two groups according to NP presence (NP+:39[41%] and NP-:56[59%]). Benralizumab treatment mean was19.7±7.2 months (range 12-35). Results: No differences in Benralizumab effectiveness were found in asthma outcomes in patients with/without NP. SNOT-22 improvement was higher in NP+ (-22±24) compared to NP- groups (6.33±15.5;p=0.055). FEV1 (16.33±19.22%), ACT(7.45±3.95) increases and frequency of SABA use (3.37±4.99) reduction were higher in SAEA compared to what obtained in non-allergic subjects (FEV1:8.15±15.6%,p=0.043; ACT:4.89±3.57,p=0.005; SABA use:-1.16±1.84;p=0.015). 93.8% of SAEA patients whereas only 72.2% of SNAEA individuals reduced OC doses at least half after Benralizumab (p=0.035). These results were partially confirmed by linear regression models showing associations between allergic status and FEV1, ACT and SABA use changes (ß=8.37;p=0.048, ß=2.056;p=0.033 and ß=-2.184;p=0.042 respectively). Conclusion: Benralizumab effectiveness in asthma appears to be independent of NP presence. The allergic eosinophilic disease, compared to just eosinophilic asthma, may be a more severe phenotype. Benralizumab may have greater efficacy in SAEA on some outcomes.

Severe asthma and long-term Benralizumab effectiveness in real-life.

OBJECTIVE: Long-term efficacy of Benralizumab in real life is not clearly known. We assessed the long-term effectiveness persistence to anti-IL-5R treatment in a group of severe eosinophilic asthmatics. PATIENTS AND METHODS: We retrospectively analyzed 95 individuals affected by severe asthma (36 males  ̶ 37.9%; mean age 58.1 ± 12.2) treated with Benralizumab (mean time 19.7 ± 7.2 months, range 12-35). Outcomes were evaluated at the beginning and at the end of patients' treatment periods. RESULTS: Mean baseline blood eosinophils were 897.5 ± 720.1 cells/µL (11 ± 5.6%) decreasing to 7.4 ± 20.6 cells/µL (0.97 ± 0.26%; p < 0.0001) after Benralizumab. FENO likewise decreased from 63.9 ± 68.4 to 28.4 ± 23.6 ppb, while FEV1% significantly improved (p < 0.0001). Mean FEF25-75 also increased from 45.8 ± 24.6% to 60.7 ± 24.6%, whereas RAW dropped from 202.15 ± 109.6% to 135.2 ± 54.75% (p < 0.0001). Also, lung volumes greatly decreased. ACT/ACQ significantly improved, while exacerbations number fell from 4.1 ± 2.4, before anti-IL-5R, to 0.33 ± 0.77, after treatment (p < 0.0001). Rhinitis severity levels and SNOT-22 also changed favorably. Patients that took long-term OCs were 71.6% before treatment, decreasing to 23.2% after Benralizumab (p < 0.0001), with an OCs dose reduction from 14.8 ± 8.9 to 1.45 ± 2.8 mg/day (p < 0.0001). 51.6% of subjects used SABA as needed before Benralizumab, falling to 4.2% after treatment. Several patients showed a reduction of ICS doses, SABA use and maintenance therapy step-down. Clinical/biological response with anti-IL-5R remained constant or even improved in terms of exacerbations or maintenance therapy reductions over time. On the contrary, FEF25-75% improvement slowed down in the long-term. No relationship was found between baseline blood eosinophil number and therapeutic response. CONCLUSIONS: Long-term Benralizumab effectiveness persistence in all outcomes in real life was confirmed.

Differential redox proteomic profiles of serum from severe asthma patients after one month of benralizumab and mepolizumab treatment.

Mepolizumab and Benralizumab are biological drugs for severe asthma patients able to reduce moderate-to-severe exacerbation rate (peripheral eosinophilial % mepolizumab 1.6 ± 1.2; benralizumab 0; p < 0.0001), improving the quality of life and lung function parameters (FEV1%: mepolizumab 87.1 ± 21.5; benralizumab 89.7 ± 15, p < 0.04). Here we report a preliminary redox proteomic study highlighting the level of oxidative burst present in serum from patients before and after one month of both treatments. Our results highlighted apolipoprotein A1 oxidation after Mepolizumab treatment, that could be related to HDL functionality and could represent a potential biomarker for the treatment. On the other hand, after one month of Benralizumab we detected higher oxidation levels of ceruloplasmin and transthyretin, considered an important oxidative stress biomarker which action help to maintain redox homeostasis.

Prognostic bioindicators in severe COVID-19 patients.

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.

Incidence and risk factors for respiratory tract bacterial colonization and infection in lung transplant recipients.

To evaluate incidence of and risk factors for respiratory bacterial colonization and infections within 30 days from lung transplantation (LT). We retrospectively analyzed microbiological and clinical data from 94 patients transplanted for indications other than cystic fibrosis, focusing on the occurrence of bacterial respiratory colonization or infection during 1 month of follow-up after LT. Thirty-three percent of patients developed lower respiratory bacterial colonization. Bilateral LT and chronic heart diseases were independently associated to a higher risk of overall bacterial colonization. Peptic diseases conferred a higher risk of multi-drug resistant (MDR) colonization, while longer duration of aerosol prophylaxis was associated with a lower risk. Overall, 35% of lung recipients developed bacterial pneumonia. COPD (when compared to idiopathic pulmonary fibrosis, IPF) and higher BMI were associated to a lower risk of bacterial infection. A higher risk of MDR infection was observed in IPF and in patients with pre-transplant colonization and infections. The risk of post-LT respiratory infections could be stratified by considering several factors (indication for LT, type of LT, presence of certain comorbidities, and microbiologic assessment before LT). A wider use of early nebulized therapies could be useful to prevent MDR colonization, thus potentially lowering infectious risk.

Right cardiac involvement in lung diseases: a multimodality approach from diagnosis to prognostication.

Lung diseases are amongst the main healthcare issues in the general population, having a high burden of morbidity and mortality. The cardiovascular system has a key role in patients affected by respiratory disorders. More specifically, the right ventricle (RV) enables the impaired lung function to be overcome in an initial stage of disease process, reducing the severity of dyspnoea. In addition, two of the main causes of death in this setting are RV failure and sudden cardiac death (SCD). Echocardiography is regarded as a useful and easily available tool in assessing RV function. Several noninvasive echocardiographic parameters of elevated pulmonary pressures and RV function have been proposed. The combination of different parameters and imaging methods is paramount and researches regarding RV impairment using these indices has been specifically addressed in relation to the chronic obstructive and restrictive lung disease in order to guide the clinicians in the management of these patients. Cardiac involvement in lung diseases is often observed, and RV changes are reported also in early stages of pulmonary diseases. The role of right ventricle in chronic respiratory disease patients has to be evaluated in detail to describe the response to therapy and the degree of disease progression through multimodality and advanced imaging techniques. The aim of this review is to describe the different pathophysiological mechanisms of cardiac impairment in primary lung disease (such as chronic obstructive pulmonary disease (COPD), idiopathic pulmonary fibrosis (IPF) and sarcoidosis) and to summarize the role of cardiac multimodality imaging in the diagnosis and the prognosis of these diseases.

Effects of rituximab therapy on B cell differentiation and depletion.

Rituximab is a human/murine chimeric anti-CD20 monoclonal antibody. It is largely used to treat B cell malignancies and has become standard in the management of B cell­mediated diseases such as rheumatoid arthritis and granulomatosis with polyangitis. The effects of rituximab need to be monitored by B cell phenotyping. Evaluate possible surface markers for monitoring B cell development in response to rituximab treatment. This review discusses the literature on the B cell surface markers analysed by flow cytometry in patients treated with rituximab. A panel of biomarkers of response to treatment to monitor by flow cytometry is also suggested. B cell phenotyping is useful to predict clinical relapses after rituximab treatment. The proposed panel of biomarkers includes CD38++CD24++IgD+/- immature B cells and IgD-CD38+/- memory B cells. In responders, Th1/Th2 balance and tolerance cells (CD4+CD25+CD127-/low Treg cells and CD19+CD24hiCD38hi Breg cells) tend to be restored after rituximab therapy. Furthermore, in responder patients, indirect depletion of CD19+/-CD27++CD38++ preplasma cells can be proposed as a predictor of response. Flow cytometric analysis of samples from patients treated with rituximab is a useful strategy to stratify patients according to response to treatment. Identification of B cell differentiation stages by means of a specific flow cytometry panel could improve monitoring of rituximab effects and enable non-responders to be distinguished from good responders.

Regulatory T Cells in Severe Persistent Asthma in the Era of Monoclonal Antibodies Target Therapies.

Asthma is an immunoinflammatory disease characterized by bronchial hyper-reactivity to different external stimuli. New monoclonal target treatments have been developed, but few studies have investigated the role of regulatory T cells in severe asthma and the modulatory effect of biological therapy on regulatory T cell functions. Their dysfunction may contribute to the development and exacerbation of asthma. Here we review the recent literature on the potential immunological role of regulatory T cells in the pathogenesis of severe asthma. The analysis of the role of regulatory T cells was performed in terms of functions and their possible interactions with mechanisms of action of the novel treatment for severe asthma. In an era of biological therapies for severe asthma, little data is available on the potential effects of what could be a new therapy: monoclonal antibody targeting of regulatory T cell numbers and functions.

Omalizumab treatment in Samter's triad: case series and review of the literature.

OBJECTIVE: Samter's triad is the combination of asthma, aspirin sensitization, and nasal polyposis. Few data are available on the use of omalizumab in this disease. The study aimed to describe the impact of omalizumab on clinical and functional parameters and the quality of life of a series of patients with Samter's triad. Moreover, we aimed to provide a review of the literature on this topic. PATIENTS AND METHODS: We retrospectively described four patients with Samter's triad undergoing omalizumab therapy. Clinical, functional, and immunological data of these patients were collected at baseline and follow-up. RESULTS: Reduction of asthma exacerbations and salbutamol rescue therapy were observed in all patients after anti-IgE treatment together with an improvement in the quality of life. A significant improvement in FEV1, FVC, and FEF25-75 was observed. No major side-effects were observed. A total of 14 studies regarding omalizumab in aspirin-exacerbated respiratory diseases were included in the review, comprising 78 patients. All studies reported a good efficacy in improving asthma control; restoration of aspirin tolerance was repeatedly reported. CONCLUSIONS: The results of our case series and review of the literature suggest that omalizumab effectively improves asthma control, lung function tests, and quality of life in patients with Samter's triad.

NK and NKT-like cells in granulomatous and fibrotic lung diseases.

ABTRACT: Background The pathogenetic and regulatory roles of natural killer (NK) and natural killer T-like cells in interstitial lung diseases (ILDs), fibrotic and granulomatous of unknown etiology are unclear. Objectives Here we investigated NK and NKT-like cells in peripheral blood (PB) and Bronchoalveolar lavage (BAL) from patients with ILDs. Method 190 patients (94 male mean age 61 ± 14.3 years) and 8 controls undergoing bronchoscopy for ILD diagnostic work-up were enrolled consecutively; 115 patients sarcoidosis, 24 chronic fibrotic hypersensitivity pneumonitis and 43 patients other ILDs [32 idiopathic pulmonary fibrosis (IPF) and 11 non-specific interstitial pneumonia (NSIP)]. PB and BAL were processed by flow cytometry using monoclonal antibodies to differentiate NK and NKT-like cells. Results NK% in BAL was significantly different among ILDs (p = 0.02). Lower NK% was observed in BAL from sarcoidosis than other ILDs (p < 0.05). Similar findings were observed for NKT-like, whereas no differences were found for PB NK%. Difference of NK% was observed between BAL and PB in all groups (p < 0.001). Sarcoidosis patients reported the best area under the curve for NKT-like (AUC = 0.678, p = 0.0015) and NK cells (AUC = 0.61, p = 0.001). In the IPF-NSIP subgroup, NK% cell was inversely correlated with FVC% (r = - 0.34, p = 0.03) and DLCO% (r = - 0.47, p = 0.0044). Conclusions NK and NKT-like were expressed differently in BAL from patients with different ILD and were significantly depleted in sarcoidosis respect to other ILDs. This suggests that these cells may play a protective role in the pathogenesis of sarcoidosis.

Pleuroparenchymal fibroelastosis (PPFE) associated with giant cell arteritis: A coincidence or a novel phenotype?

Pleuroparenchymal fibroelastosis (PPFE) is a rare interstitial lung disease characterized by the fibrotic thickening of subpleural and parenchymal areas of the upper lobes. It may be both idiopathic or secondary to infections, interstitial lung diseases and/or drug exposure. Often PPFE patients report recurrent lower respiratory tract infections, suggesting that repeated inflammatory alterations induced by pulmonary infections may contribute to the development/progression of PPFE. Here, we report for the first time the case of a patient affected by Giant cell Arteritis with histologically proven PPFE. The lung involvement in GCA is rare and interstitial lung diseases are usually reported as an uncommon clinical manifestation of GCA. Our patient is probably the first case presenting PPFE associated with GCA and we wonder if this is a real associative disease or a coincidence perhaps, secondary to drug effects.

Pirfenidone and Nintedanib in idiopathic pulmonary fibrosis: Real-life experience in an Italian referral centre.

BACKGROUND: Idiopathic pulmonary fibrosis has a median survival time after diagnosis of 2-5 years. The main goal of treating IPF is to stabilize or reduce the rate of disease progression. Nintedanib and Pirfenidone have been a breakthrough in the management of IPF. Here we evaluated the effectiveness of Pirfenidone and Nintedanib in a population of IPF patients diagnosed in the last 12 months at Florence ILD Referral Centre. METHODS: In the last 12 months, 82 IPF patients (66 male, mean age 78.3±23.8 years) were diagnosed and started antifibrotic therapy with Pirfenidone or Nintedanib. Their clinical and functional details were analyzed retrospectively at time 0 and after 6 and 12 months of therapy. RESULTS: The median age of the patients treated with Nintedanib was higher than that of the Pirfenidone group (p<0.0001). The most common symptoms at disease onset were exertional dyspnoea and dry cough with no differences between the two groups (p<0.05). All IPF patients manifested bibasal crackles at the time of diagnosis. No significant differences in FVC, FEV1, TLC and DLCO were found at time 0 or after 6 months between patients treated with Pirfenidone and Nintedanib (p>0.05). After 1 year, lung function test parameters of patients treated with Pirfenidone had remained stable from baseline. DISCUSSION: This study emphasizes that both antifibrotic drugs appeared to be a good therapeutic choice in terms of functional stabilization, also in older patients.

Effects of omalizumab in severe asthmatics across ages: A real life Italian experience.

BACKGROUND: This retrospective study aimed at evaluating long-term effects of Omalizumab in elderly asthmatics in a real-life setting. METHODS: 105 consecutive severe asthmatics (GINA step 4-5; mean FEV1% predicted:66 ± 15.7) treated with Omalizumab for at least 1 year (treatment mean duration 35.1 ± 21.7 months) were divided into 3 groups according to their age at Omalizumab treatment onset: 18-39, 40-64 and ≥ 65 years. RESULTS: Comorbidities, number of overweight/obese subjects and patients with late-onset asthma were more frequent among older people. A similar reduction of inhaled corticosteroids dosage and SABA on-demand therapy was observed in all groups during Omalizumab treatment; a similar FEV1 increased was also observed. Asthma Control Test (ACT) improved significantly (p < 0.001) in the three groups, increasing from 15 [IQR:12-18] to 24 [IQR:22-25] in younger subjects, from 14 [IQR:10-16] to 21 [IQR:20-23] in the 40-64-year-group and from 15 [IQR:12-16] to 20 [IQR:18-22] in elderly patients where improvement was lower (p = 0.039) compared to younger people. Asthma exacerbations decreased significantly after Omalizumab but the percentage of exacerbation-free patients was higher in younger people (76.9%) compared to middle aged patients (49.2%) and the elderly (29%) (p = 0.049). After Omalizumab treatment, the risk for exacerbations was lower in subjects aged 40-64 (OR = 0.284 [CI95% = 0.098-0.826], p = 0.021) and 18-39 (OR = 0.133 [CI95% = 0.026-0.678], p = 0.015), compared to elderly asthmatics. Also, a significantly reduced ACT improvement (ß = -1.070; p = 0.046) passing from each age class was observed. CONCLUSION: Omalizumab improves all asthma outcomes independently of age, although the magnitude of the effects observed in the elderly seems to be lower than in the other age groups.

Comparative proteomic analysis of bronchoalveolar lavage of familial and sporadic cases of idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a chronic lung disease characterized by progressive deterioration of the alveolar integrity. Among IPF identified phenotypes, that of familial (f-)IPF is usually associated with several gene mutations which are seldom observed in sporadic (s-)IPF. This study aimed at investigating the molecular patterns and variability in f-IPF and s-IPF patients through a differential proteomic analysis. Protein patterns of bronchoalveolar lavage fluid (BALF) samples from 10 familial and 17 sporadic IPF patients were compared using 2D electrophoresis and mass spectrometry. Principal component analysis (PCA) was applied to proteomic data and an enrichment analysis was also performed to characterize specific pathogenic mechanisms and to identify potential biomarkers. BALF samples from f-IPF showed 87 protein spots differentially expressed than those from s-IPF samples; once identified, these spots revealed 22 unique proteins. The functional analysis showed that the endothelial reticulum stress probably plays a central pathogenetic role in f-IPF with an up-regulation of proteins involved in wounding and immune responses, coagulation system, and ion homeostasis. Up-regulated proteins in the s-IPF group were those involved in the oxidative stress response. PCA analysis of differentially expressed proteins clearly distinguished f-IPF from s-IPF patients, and in agreement with radiological and histological patterns, pointed out a higher heterogeneity in f-IPF than s-IPF samples. The 'Slit/Robo signaling', 'clathrin-coated vesicle' and 'cytoskeleton remodelling', were extrapolated by 'pathways analysis' and the results of 'diseases (by biomarkers)' highlighted a 'connective tissue and autoimmune disease', two aspects of increasing interest in IPF.