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Background: The management of chronic diseases, particularly in children, requires an integrated physical and psychological approach to both sick children and their family. This is the case of Cystic Fibrosis (CF), a complex genetic chronic disease, where, a comprehensive evaluation of the emotional impact and an effective multidimensional approach are indicated. Aim: This study investigates on parenting stress in children and adolescents with CF and its determinants related to parents, children and the disease severity. Methods: The study involved 34.04% adult males and 65.96% adult females (range 21-55 years) and 47 children with CF, 54.35% males and 45.65% females (range 1-17 years). The data were obtained through a Parenting Stress Index - Short Form (PSI-SF) questionnaire. According to the PSI-SF scoring system, three types of stress were detected: a typical stress pattern (normal), a high stress pattern (increased) and a defensive response, which may be considered as a high stress feature in children which requires monitoring and clinical evaluation. Results: This study shows a significant presence of stress in females (60.23%), of subject married (84.62%), unemployed (69.23%) and with education level such as "middle School" (61.54%). Concerning children of parents with high stress, it resulted most frequent children with one sibling (53.85%). Finally, by univariate analysis, it resulted a significant positive correlation between parenting stress and disease degree of children. Instead by multivariate analysis, we found that the variables: Number of siblings and Birth order were a significant positive and negative predictor of parenting stress respectively. Conclusion: An increased stress level was detected in less than one third of parents of subjects with CF. These data may be related to the psychological support which is part of the routine management of CF care team. However, as children's features seem to act as a determinant of stress more than parental ones, the parental-child dysfunction should be the target for further integrated interventions. Background: The management of chronic diseases, particularly in children, requires an integrated physical and psychological approach to both sick children and their family. This is the case of Cystic Fibrosis (CF), a complex genetic chronic disease, where, a comprehensive evaluation of the emotional impact and an effective multidimensional approach are indicated. Aim: This study investigates on parenting stress in children and adolescents with CF and its determinants related to parents, children and the disease severity. Methods: The study involved 34.04% adult males and 65.96% adult females (range 21-55 years) and 47 children with CF, 54.35% males and 45.65% females (range 1-17 years). The data were obtained through a Parenting Stress Index – Short Form (PSI-SF) questionnaire. According to the PSI-SF scoring system, three types of stress were detected: a typical stress pattern (normal), a high stress pattern (increased) and a defensive response, which may be considered as a high stress feature in children which requires monitoring and clinical evaluation. Results: This study shows a significant presence of stress in females (60.23%), of subject married (84.62%), unemployed (69.23%) and with education level such as “middle School” (61.54%). Concerning children of parents with high stress, it resulted most frequent children with one sibling (53.85%). Finally, by univariate analysis, it resulted a significant positive correlation between parenting stress and disease degree of children. Instead by multivariate analysis, we found that the variables: Number of siblings and Birth order were a significant positive and negative predictor of parenting stress respectively. Conclusion: An increased stress level was detected in less than one third of parents of subjects with CF. These data may be related to the psychological support which is part of the routine management of CF care team. However, as children’s features seem to act as a determinant of stress more than parental ones, the parental-child dysfunction should be the target for further integrated interventions.

Since the identification of Cystic Fibrosis (CF) as a disease in 1938 until 2012, only therapies to treat symptoms rather than etiological therapies have been used to treat the disease. Over the last few years, new technologies have been developed, and gene editing strategies are now moving toward a one-time cure. This review will summarize recent advances in etiological therapies that target the basic defect in the CF Transmembrane Receptor (CFTR), the protein that is mutated in CF. We will discuss how newly identified compounds can directly target mutated CFTR to improve its function. Moreover, we will discuss how proteostasis regulators can modify the environment in which the mutant CFTR protein is synthesized and decayed, thus restoring CFTR function. The future of CF therapies lies in combinatory therapies that may be personalized for each CF patient.

Background Recent standards of care mention chest radiography (CR) but not chest computed tomography (CT) in routine annual follow-up of children with cystic fibrosis (CF). To minimise radiation risk, CT or CR should only be performed if they impact clinical decision making. We investigated whether in addition to a wide range of commonly used clinical parameters, chest CT and/or CR in routine follow-up of CF patients influence clinical decisions. Methods 36 web based clinical vignettes (i.e. case simulations) were designed using clinical data from patients aged 8–18 years, randomly selected from two CF centres in The Netherlands. In a randomized cross-over design, clinicians assessed eight vignettes and suggested therapeutic/diagnostic management on two occasions, with a ten-week interval. Radiological information (CT or CR) was included at only one of the two assessments, in random order. Any differences in management could be attributed to information from CT or CR, and were compared by McNemar analysis. Results 44 European and Australian clinicians completed a total of 143 CT vignette pairs and 167 CR vignette pairs. CT was associated with a significant increase in antifungal treatment (Risk Ratio (RR) 2.8 (1.3–6.0, p = .02)), bronchoscopies (RR 1.6 (1.1–2.5, p = .04)), mycobacterial cultures (RR 1.3 (1.0–1.5, p = .02)), and ‘need for hospitalization’ (i.e. intravenous antibiotics and/or bronchoscopy) (RR 1.4 (1.0–1.9, p = .03)). CR led to a significant increase in inhaled antibiotics only (RR 1.3 (1.0–1.6, p = .04)). Conclusions CT but not CR, at routine biennial follow-up was associated with several changes in treatment and/or diagnostic testing, including the need for hospitalization.

Objective We evaluated the prevalence, Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) gene profile, clinical data, management and outcome for infants with a CFTR-related metabolic syndrome/CF Screen Positive, Inconclusive Diagnosis (CRMS/CFSPID) designation from six Italian centres. Methods All newborn bloodspot screening (NBS) positive infants born from January 2011 to August 2018 with a CF diagnosis or a CRMS/CFSPID designation were enrolled. Data on sweat testing, genetics, clinical course and management were collected. Results We enrolled 257 CF patientsand 336 infants with a CRMS/CFSPID designation (CF: CRMS/CFSPID ratio of 1:1.30).Blood immuno-reactive trypsinogen (IRT) was significantly lower in CRMS/CFSPID infants and the F508del variant accounted for only 20% of alleles. Children with CRMS/CFSPID showed a milder clinical course, pancreatic sufficiency compared to CF infants. Varied practice across centres was identified regarding sweat testing, chest radiograph (8-100%) and salt supplementation (11-90%). Eighteen (5.3%) CRMS/CFSPID infants converted or were reclassified to diagnosis of CF. Four infants (1.3%) developed a clinical feature consistent with a CFTR-related disorder (1.2%). Twenty-seven were re-classified as healthy carriers (8.0%) and 16 as healthy infants (4.8%). Conclusions We have identified considerable variability in the evaluation and management of infants with an inconclusive diagnosis following NBS across six Italian centres. CRMS/CFSPID is more regularly seen in this population compared to countries with higher prevalence of F508del.Conversion to a CF diagnosis was recorded in 18 (5.3%) of CRMS/CFSPID infants and in 16 was as a result of increasing sweat chloride concentration.

Background: The goal of mucoactive therapies in cystic fibrosis (CF) is to enhance sputum clearance and to reduce a progressive decline in lung function over the patient’s lifetime. We aimed to investigate the level of consensus among specialists from Italian CF Centers on appropriateness of therapeutic use of dornase alfa (rhDNase) for CF patients. Method: A consensus on appropriate prescribing in CF mucoactive agents was appraised by an online Delphi method, based on a panel of 27 pulmonologists, coordinated by a Scientific Committee of six experts in medical care of patients with CF. Results: Full or very high consensus was reached on several issues related to therapeutic use of dornase alfa for CF patients in clinical practice. Conclusions: Modified Delphi method was used to define the most appropriate use of dornase alfa in routine CF to improve lung function and long-term outcomes in patients, in agreement with international guidelines on CF management.

OBJECTIVE: To verify whether growth hormone receptor (GHR) gene expression plays a role in growth of children with cystic fibrosis (CF), as a consequence of the chronic inflammatory condition and malnutrition. DESIGN: We enrolled 49 prepubertal patients (24 males and 25 females) affected by CF in a stable clinical condition, 19 of whom had been diagnosed through newborn screening and 30 following presentation of symptoms. Patients had no significant comorbidity affecting growth or cystic fibrosis transmembrane conductance regulator (CFTR)-related diabetes requiring insulin therapy. Blood was collected during two follow-up visits to measure insulin-like growth factor (IGF-I), growth hormone-binding protein (GHBP), and GHR gene expression. Recruited as a control group were 52 healthy children, sex- and age-matched, were recruited as a control group. METHODS: We compared body mass index (BMI), height, weight, IGF-I, GHBP, and GHR gene expression values (evaluated by Chemiluminescent Immunometric assay; ELISA and real-time PCR, respectively) in CF patients diagnosed through newborn screening (NBS) or by symptoms (late diagnosis [LD]) and in healthy controls. RESULTS: BMI increased significantly in patients between the time of diagnosis and check-up (P

Cystic fibrosis (CF) is a disease caused by loss-of-function mutations affecting the CF transmembrane conductance regulator (CFTR), a chloride channel. Recent evidence indicates that CFTR is inhibited by a gluten/gliadin-derived peptide (P31-43), causing an acquired state of CFTR inhibition within the gut that contributes to the pathogenesis of celiac disease (CD). Of note, CFTR inhibition does not only cause intra- and extracellular ion imbalances but also affects proteostasis by activating transglutaminase-2 (TGM2) and by disabling autophagy. These three phenomena (CFTR inhibition, TGM2 activation, and autophagy impairment) engage in multiple self-amplifying circuitries, thus forming an “infernal trio”. The trio hinders enterocytes from returning to homeostasis and instead locks them in an irreversible pro-inflammatory state that ultimately facilitates T lymphocyte-mediated immune responses against another gluten/gliadin-derived peptide (P57–68), which,upon deamidation by activated TGM2, becomes fully antigenic. Hence, the pathogenic protein gliadin exemplifies a food constituent the exceptional immunogenicity of which arises from a combination of antigenicity (conferred by deaminated P57–68) and adjuvanticity (conferred by P31-43). CF can be treated by agents targeting the “infernal trio” including CFTR potentiators, TGM2 inhibitors, and autophagy enhancers. We speculate that such agents may also be used for CD therapy and indeed could constitute close-to-etiological treatments of this enteropathy.

BackgroundThe sweat chloride test (ST) is the gold standard for cystic fibrosis (CF) diagnosis in symptomatic patients, within the newborn screening and in the follow-up of CF patients during molecular therapies. However, false positives have been reported in patients with different diseases. We describe and discuss 4 cases due to different clinical conditions in which we recorded false positive ST, and the test remained altered for a period of varying length.Cases presentationCase 1: Eight months old female child suffering from constipation, recurrent vomiting and failure to thrive, family history of recurrent pancreatitis without mutations in thePRSS1andSPINK1genes. Both ST and fecal elastase were altered although noCFTRgene mutations were found. Due to rapid clinical deterioration, celiac disease was suspected and diagnosed by laboratory tests and intestinal biopsy. After 2 weeks of gluten-free diet ST and fecal elastase normalized.Case 2: 14 months old male suffering from bilateral renal dysplasia, episodes of metabolic alkalosis, recurrent respiratory infections and recurrent vomiting. The child had more ST positives, but noCFTRmutations were found. During follow-up, he developed sensorineural hearing loss and an atrial septic defect was found. Finally, a diagnosis of Klinefelter was made, but the ST normalized several years later.Case 3 and 4: Two boys with stubborn constipation and fecal occlusion treated with Poly Ethylene Glycol (PEG) with salts showed pathological ST. The test returned normal a few days after stopping treatment.ConclusionsWe hypotesized the possible causes of ST alteration in these conditions: in celiac disease it could be due to a transient dysregulation of the aquaporins, rapidly reversed by the diet; in Klinefelter, it may be due to stable pubertal hypoandrogenism; while, the PEG formulation itself contains salts that can temporarily alter ST.

Exposure to gluten, a protein present in wheat rye and barley, is the major inducer for human Celiac Disease (CD), a chronic autoimmune enteropathy. CD occurs in about 1% worldwide population, in genetically predisposed individuals bearing human leukocyte antigen (HLA) DQ2/DQ8. Gut epithelial cell stress and the innate immune activation are responsible for the breaking oral tolerance to gliadin, a gluten component. To date, the only treatment available for CD is a long-term gluten-free diet. Several studies have shown that an altered composition of the intestinal microbiota (dysbiosis) could play a key role in the pathogenesis of CD through the modulation of intestinal permeability and the regulation of the immune system. Here, we show that gliadin induces a chronic endoplasmic reticulum (ER) stress condition in the small intestine of a gluten-sensitive mouse model and that the coadministration of probiotics efficiently attenuates both the unfolded protein response (UPR) and gut inflammation. Moreover, the composition of probiotics formulations might differ in their activity at molecular level, especially toward the three axes of the UPR. Therefore, probiotics administration might potentially represent a new valuable strategy to treat gluten-sensitive patients, such as those affected by CD.

BACKGROUND: Cystic fibrosis (CF) is a multi-organ genetic disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene which encodes the CFTR protein. CF-associated liver disease (CFLD) is a common complication; diagnosis is based on clinical, laboratory findings and abdominal imaging. However, non-invasive diagnostic approaches are needed to early detect CFLD, its progression and severity. Recent studies demonstrate a possible role of point shear wave elastography (p-SWE) with liver stiffness measurement (LSM) as a tool for CFLD diagnosis also in children. This non-invasive technique measures liver stiffness to assess liver fibrosis and is suggested to be less operator-dependent compared to ultrasonography. Aim of our prospective observational study is to investigate the role of p-SWE with LSM for CFLD diagnosis in children and adolescents with CF and to compare this finding with aspartate aminotransferase to platelet ratio index (APRI), fibrosis index based on four factors (FIB-4) and gamma-glutamyl-transpeptidase to platelet ratio (GPR) indices. METHODS: Fifty-nine children with CF, who had routinely undergone abdominal imaging, were consecutively enrolled. Laboratory findings and clinical data were recorded, as abdominal ultrasound and shear wave elastography at baseline. The cases were divided into two groups based on collected data and classified as CFLD and CFnoLD (without liver disease) according to Debray criteria. APRI, FIB-4 and GPR fibrosis indices were also evaluated. RESULTS: Twenty-four/59 (40.7%) were defined as CFLD. LSM test is superior to the APRI (P

Objective We aimed to assess the order of severity of the defects of 3 direct determinants of glucose regulation—beta-cell function, insulin clearance, and insulin sensitivity—in patients with cystic fibrosis (CF), categorized according their glucose tolerance status, including early elevation of mid-level oral glucose tolerance test (OGTT) glucose values (>140 and Methods A total of 232 CF patients aged 10 to 25 years underwent OGTT. Beta-cell function and insulin clearance were estimated by OGTT mathematical modeling and OGTT-derived biomarkers of insulin secretion and sensitivity were calculated. The association between glucometabolic variables and 5 glucose tolerance stages (normal glucose tolerance [NGT], AGT140, indeterminate glucose tolerance [INDET], impaired glucose tolerance [IGT], cystic fibrosis–related diabetes CFRD]) was assessed with a general linear model. Results Beta-cell function and insulin sensitivity progressively worsened across glucose tolerance stages (P Conclusions In CF patients, each of the 5 glucose tolerance stages shows a unique pattern of defects of the direct determinants of glucose regulation, with AGT140 patients significantly differing from NGT and being similar to IGT. These findings suggest that AGT140 should be recognized as a distinct glucose tolerance stage and that reconsideration of the grade of glucometabolic deterioration across glucose tolerance stages in CF is warranted.

Background Cystic Fibrosis Related Diabetes (CFRD) is a frequent comorbidity of patients with Cystic Fibrosis (CF). A worsening of clinical conditions appears before CFRD. It has been demonstrated a decline in pulmonary function and nutritional status also in patients with prediabetes. Few trials show that insulin may be beneficial in prediabetic CF patients, to date guidelines do not recommend for this condition. Case presentation We report a case of a patient treated with insulin glargine at 13 years, due to glycemic intolerance, and with Lumacaftor/Ivacaftor at 15 years. A reduction of pulmonary exacerbations was observed after glargine therapy, also confirmed after the starting of Lumacaftor/ Ivacaftor in this patient. Pulmonary function improved only after the first year of glargine therapy, then a deterioration appeared due to the natural history of CF lung damage. During the COVID-19 lockdown, poor adherence to care contributed to diabetes mellitus onset needing high insulin requirements. After two weeks the patient returned to prediabetic condition and his previous dose of glargine. Conclusions our case highlights firstly that insulin glargine has contributed to preserve him from further clinical worsening due to prediabetes in the years before pandemic, secondly the negative impact of COVID-19 lockdown on the clinical course of a chronic disease as CF.

Background: Newborn screening (NBS) early-identifies cystic fibrosis (CF), but in CF-screening positive inconclusive diagnosis (CF-SPID) the results of immunoreactive trypsinogen (IRT), molecular analysis and sweat test (ST) are discordant. A percentage of CF-SPID evolves to CF, but data on long-term monitoring are lacking. We describe the follow-up of all CF and CF-SPID identified between 2008 and 2019. Methods: NBS was performed by IRT followed by molecular analysis and ST between 2008 and 2014, double IRT followed by molecular analysis and ST after 2014. Results: NBS revealed 47 CF and 99 CF-SPID newborn, a ratio 1:2.1&mdash, the highest reported so far. This depends on the identification by gene sequencing of the second variant with undefined effect in 40 CF-SPID that otherwise would have been defined as carriers. Clinical complications and pulmonary infections occurred more frequently among CF patients than among CF-SPID. Two CF-SPID cases evolved to CF (at two years), while eight evolved to CFTR-related disorders (CFTR-RD), between one and eight years, with bronchiectasis (two), recurrent pneumonia (four, two with sinonasal complications), recurrent pancreatitis (two). No clinical, biochemical or imaging data predicted the evolution. Conclusion: Gene sequencing within the NBS reveals a higher number of CF-SPID and we first describe an approach to early identify CFTR-RD, with relevant impact on their outcome.

About 50% of patients with cystic fibrosis (CF) have sinonasal complications, which include inferior turbinate hypertrophy (NTH) and/or nasal polyposis (NP), and different degrees of lung disease, which represents the main cause of mortality. Monitoring of sinonasal disease requires complex instrumental procedures, while monitoring of lung inflammation requires invasive collection of bronchoalveolar lavage fluid. The aim of this study was to investigate the associations between salivary cytokines levels and CF-related airway diseases. Salivary biochemical parameters and cytokines, i.e., interleukin-6 (IL-6), IL-8, and tumor necrosis factor alpha (TNF-&alpha, ), were analyzed in resting saliva from healthy subjects and patients with CF. Patients with CF showed significantly higher levels of salivary chloride, IL-6, IL-8, and TNF-&alpha, and lower calcium levels than healthy subjects. Among patients with CF, IL-6 and IL-8 were significantly higher in patients with NTH, while TNF-&alpha, was significantly lower in patients with NP. A decreasing trend of TNF-&alpha, in patients with severe lung disease was also observed. On the other hand, we did not find significant correlation between cytokine levels and Pseudomonas aeruginosa or Stenotrophomonas maltophilia colonization. These preliminary results suggest that salivary IL-6 and IL-8 levels increase during the acute phase of sinonasal disease (i.e., NTH), while the end stages of pulmonary disease and sinonasal disease (i.e., NP) show decreased TNF-&alpha, levels

The clinical manifestation of cystic fibrosis (CF) is heterogeneous also in patients with the same cystic fibrosis transmembrane regulator (CFTR) genotype and in affected sibling pairs. Other genes, inherited independently of CFTR, may modulate the clinical manifestation and complications of patients with CF, including the severity of chronic sinonasal disease and the occurrence of chronic Pseudomonas aeruginosa colonization. The T2R38 gene encodes a taste receptor and recently its functionality was related to the occurrence of sinonasal diseases and upper respiratory infections. We assessed the T2R38 genotype in 210 patients with CF and in 95 controls, relating the genotype to the severity of sinonasal disease and to the occurrence of P. aeruginosa pulmonary colonization. The frequency of the PAV allele i.e., the allele associated with the high functionality of the T2R38 protein, was significantly lower in i) CF patients with nasal polyposis requiring surgery, especially in patients who developed the complication before 14 years of age; and ii) in CF patients with chronic pulmonary colonization by P. aeruginosa, especially in patients who were colonized before 14 years of age, than in control subjects. These data suggest a role for T2R38 as a novel modifier gene of sinonasal disease severity and of pulmonary P. aeruginosa colonization in patients with CF.

Patients with cystic fibrosis (CF) have low cholesterol absorption and, despite enhanced endogenous biosynthesis, low serum cholesterol. Herein, we investigated cholesterol metabolism in a murine CF model in comparison towild type(WT) testing serum and liver surrogate biomarkers together with the hepatic expression of genes involved in cholesterol metabolism. CF mice display lower sterols absorption and increased endogenous biosynthesis. Subsequently, we evaluated the effects of a cholesterol-supplemented diet on cholesterol metabolism in CF and WT mice. The supplementation in WT mice determines biochemical changes similar to humans. Instead, CF mice with supplementation did not show significant changes, except for serum phytosterols (−50%), liver cholesterol (+35%) and TNFα mRNA expression, that resulted 5-fold higher than in CF without supplementation. However, liver cholesterol in CF mice with supplementation resulted significantly lower compared to WT supplemented mice. This study shows that in CF mice there is a vicious circle in which the altered bile salts synthesis/secretion contribute to reduce cholesterol digestion/absorption. The consequence is the enhanced liver cholesterol biosynthesis that accumulates in the cell triggering inflammation.

Introduction: Cystic Fibrosis (CF) is an inherited, lethal and expensive rare disease affecting more than 85,000 people worldwide. CF is caused by more than 2000 loss-of-function mutations in the gene coding for the cystic fibrosis transmembrane conductance regulator (CFTR). Personalized interventions on individual CF patients probably constitute the future of CF management.Areas covered: The molecular genetics exploration of CF has led in the last years to the development of mutation-specific therapies that aim at directly targeting the mutant CFTR to reverse disease phenotype. Here, we will summarize the strategies of drug repositioning, i.e. the use of ‘old drugs’ for ‘new purposes’ in the perspective of a personalized approach to CF treatment.Expert opinion: Rare diseases like CF may well enter the era of precision medicine. A personalized/repositioning medicine in CF could help to provide the right drug to the right patient and potentially reduce the costs of developing disease-modifying drugs.

Summary Candida albicans is implicated in intestinal diseases. Identifying host signatures that discriminate between the pathogenic versus commensal nature of this human commensal is clinically relevant. In the present study, we identify IL-9 and mast cells (MCs) as key players of Candida commensalism and pathogenicity. By inducing TGF-β in stromal MCs, IL-9 pivotally contributes to mucosal immune tolerance via the indoleamine 2,3-dioxygenase enzyme. However, Candida-driven IL-9 and mucosal MCs also contribute to barrier function loss, dissemination, and inflammation in experimental leaky gut models and are upregulated in patients with celiac disease. Inflammatory dysbiosis occurs with IL-9 and MC deficiency, indicating that the activity of IL-9 and MCs may go beyond host immunity to include regulation of the microbiota. Thus, the output of the IL-9/MC axis is highly contextual during Candida colonization and reveals how host immunity and the microbiota finely tune Candida behavior in the gut., Graphical Abstract, Highlights • IL-9/IL-9R signaling affects MC function in mucosal candidiasis • IL-9 and mucosal MCs contribute to barrier function loss in leaky gut models • IL-9 and stromal MCs induce local protective tolerance in infection via IDO1 • IL-9 and mucosal MCs expand and IDO1 decreases in human celiac disease, Deciphering the mechanisms by which Candida albicans promotes either pathology or protective tolerance in the gut could be clinically relevant. Renga et al. show a key role for IL-9 and mast cells in promoting either inflammatory dysbiosis and pathology or tolerance in leaky gut models and human celiac disease.

Diabetes mellitus is the most common comorbidity in cystic fibrosis (CF). Recently, more attention has been paid to early glucose metabolism derangements (GMDs). The subject of this report is a female patient, affected by CF since 3 months of age. She presented with intermittent diabetes during early childhood. At the age of 10 years, oral glucose tolerance test (OGTT) was performed and showed glucose intolerance (IGT) status; glargine insulin therapy was started. At the age of 13 years, CF-related diabetes with fasting hyperglycemia occurred, so rapid insulin at meals was added. During the following year, clinical and nutritional status improved. Stable clinical conditions were observed in the following 3 years. This is the first case of very long-term follow-up concerning a CF patient with GMDs. Our case confirms the importance of paying attention to early GMDs in very young CF patients and seems to suggest that earlier therapy could ameliorate CF natural history.

Background Primary ciliary dyskinesia (PCD) and cystic fibrosis (CF) are increasingly compared. There are no chest magnetic resonance imaging (MRI) comparative studies of PCD and CF. We assessed clinical, functional, microbiological and MRI findings in PCD and mild CF patients in order to evaluate different expression of lung disease. Methods Twenty PCD (15.1 years) and 20 CF subjects with mild respiratory impairment (16 years, 70% with pancreatic insufficiency) underwent MRI, spirometry, and sputum cultures when clinically stable. MRI was scored using the modified Helbich system. Results PCD was diagnosed later than CF (9.9 versus 0.6 years, p = 0.03), despite earlier symptoms (0.1 versus 0.6 years, p = 0.02). In the year preceding the study, patients from both groups underwent two systemic antibiotic courses (p = 0.48). MRI total scores were 11.6 ± 0.7 and 9.1 ± 1 in PCD and CF, respectively. FEV1 and FVC Z-scores were −1.75 (range, −4.6–0.7) and −0.6 (−3.9–1.8) in PCD, and −0.9 (range, −5.4–2.3) and −0.3 (−3.4–2.5) in CF, respectively. No difference was found between lung function or structure, despite a higher MRI subscore of collapse/consolidation in PCD versus CF (1.6 ± 0.1 and 0.6 ± 0.2, p

Cystic fibrosis (CF) is an inherited, prematurely lethal rare disease affecting more than 85,000 people worldwide. CF is caused by more than 2000 loss-of-function mutations in the gene coding for the cystic fibrosis transmembrane conductance regulator (CFTR). This review summarizes recent advances in the etiological therapies of CF that aim at repairing the functional defect of CFTR by means of CFTR modulators. We will discuss the state of art of the mutation-specific treatments that are designed to target different steps of the CFTR biogenesis perturbed by mutations in CFTR gene. Moreover, we will discuss how drug repositioning, namely the use of drugs already approved for the treatment of other human diseases, may be repurposed in CF patients to circumvent CFTR dysfunction. Finally, we highlight how the combined use of two or more compounds acting on different disease mechanisms is required to achieve clinical benefit in CF population.

Structural lung disease begins very early in children with cystic fibrosis (CF), often in the first three months of life. Inhaled medications represent an attractive therapeutic approach in CF that are routinely used as early intervention strategies. Two aerosolized solutions, hypertonic saline and dornase alfa, have significant potential benefits by improving mucociliary clearance, with minimal associated side-effects. In particular, they favor rehydration of airway surface liquid and cleavage of extracellular DNA in the airways, respectively, consequently reducing rate of pulmonary disease exacerbations. Indirect anti-inflammatory effects have been documented for both drugs, addressing each of the three interrelated elements in the vicious cycle of lung disease in CF: airway obstruction, inflammation and infection. This short review aimed to summarize the main papers that support potential clinical impact of inhaled solutions on pulmonary disease in CF.

BACKGROUND: The effect of complex alleles in cystic fibrosis (CF) is poorly defined for the lack of functional studies. OBJECTIVES: To describe the genotype-phenotype correlation and the results of either in vitro and ex vivo studies performed on nasal epithelial cells (NEC) in a cohort of patients with CF carrying cystic fibrosis transmembrane conductance regulator (CFTR) complex alleles. METHODS: We studied 70 homozygous, compound heterozygous or heterozygous for CFTR mutations: p.[Arg74Trp;Val201Met;Asp1270Asn], n=8; p.[Ile148Thr;Ile1023_Val1024del], n=5; p.[Arg117Leu;Leu997Phe], n=6; c.[1210-34TG[12];1210-12T[5];2930C>T], n=3; p.[Arg74Trp;Asp1270Asn], n=4; p.Asp1270Asn, n=2; p.Ile148Thr, n=6; p.Leu997Phe, n=36. In 39 patients, we analysed the CFTR gating activity on NEC in comparison with patients with CF (n=8) and carriers (n=4). Finally, we analysed in vitro the p.[Arg74Trp;Val201Met;Asp1270Asn] complex allele. RESULTS: The p.[Ile148Thr;Ile1023_Val1024del] caused severe CF in five compound heterozygous with a class I-II mutation. Their CFTR activity on NEC was comparable with patients with two class I-II mutations (mean 7.3% vs 6.9%). The p.[Arg74Trp;Asp1270Asn] and the p.Asp1270Asn have scarce functional effects, while p.[Arg74Trp;Val201Met;Asp1270Asn] caused mild CF in four of five subjects carrying a class I-II mutation in trans, or CFTR-related disorders (CFTR-RD) in three having in trans a class IV-V mutation. The p.[Arg74Trp;Val201Met;Asp1270Asn] causes significantly (pT] and a class I-II mutation had mild CF or CFTR-RD (gating activity: 18.5-19.0%). CONCLUSIONS: The effect of complex alleles partially depends on the mutation in trans. Although larger studies are necessary, the CFTR activity on NEC is a rapid contributory tool to classify patients with CFTR dysfunction.

(1) Background: Diagnostic testing for cystic fibrosis (CF) is based on a sweat chloride test (SCT) considering the appropriate signs and symptoms of the disease and results of a gene mutation analysis. In 2014, the Istituto Superiore di Sanit&agrave, (ISS) established a pilot Italian external quality assessment program for CF SCT (Italian EQA-SCT), which is now a third party service carried out by the ISS. (2) Methods: The ongoing scheme is prospective, enrollment is voluntary, and the payment of a fee is required. Results are shared through a dedicated web-facility. Assessment covers the analysis, interpretation, and reporting of results. (3) Results: Thirteen, fifteen, sixteen, and fifteen different laboratories, respectively, participated from 2015 to 2016 and from 2018 to 2019 in the Italian EQA-SCT scheme. Eleven different laboratories participated each year in all four rounds of the Italian EQA-SCT. (4) Conclusions: The overall results obtained from the laboratories participating constantly clearly show that their qualitative and quantitative performance improved significantly. This is due to the opportunity&mdash, after receiving the EQA results&mdash, to constantly review their performance and address any inconsistencies. We firmly believe that participation in the EQA program will improve the quality of participating laboratories and that EQA participation should become mandatory as a fundamental requirement for laboratory accreditation.

Cystic fibrosis (CF) is a life shortening–autosomal recessive disease caused by loss–of–function mutations in the gene encoding the CF transmembrane conductance regulator (CFTR) that transports essential electrolytes in and out of specialized cells (1). The loss of CFTR causes anomalies in the epithelial ion transport, leading to dehydration of cell surfaces and resulting in the accumulation of sticky obstructions that eventually destroy the integrity and physiology of different organ systems (2).

Background An increasing number of patients have been described as having a number of Cystic Fibrosis Transmembrane conductance Regulator (CFTR) variants for which it lacks a clear genotype–phenotype correlation. We assesses the clinical features of patients bearing the S737F (p.Ser737Phe) CFTR missense variant and evaluated the residual function of CFTR protein on nasal epithelial cells (NEC). Methods A retrospective database was performed from individuals homozygous or compound heterozygous for the S737F variant followed in the Cystic Fibrosis (CF) Centre of Florence. We performed a nasal brushing in cooperating patients and compared the results with those of patients followed in the pediatric CF Centre of Naples. Results 9/295 (3%) subjects carrying at least S737F CFTR variant on one allele were identified. Patients were diagnosed in 7/9 cases by newborn screening and in two cases for dehydration with hypochloremic metabolic alkalosis; at diagnosis sweat chloride levels (SCL) were in the pathological range in only one case. After a mean follow up of 8,6 years (range 0,5–15,8), SCL were in the pathological range in 8/9 cases (mean age at CF diagnosis: 1,5 years), all patients were pancreatic sufficiency and respiratory function was normal. The gating activity on NEC was 15.6% and 12.7% in two patients compound heterozygous for W1282X and DelE22_24, while it was ranged between 6,2% and 9,8% in CF patients. Conclusions S737F is a CFTR mutation associated to hypochloremic alkalosis in childhood, mild CF phenotype in teenage years and a residual function of CFTR protein.

Background The sweat test is one of the main diagnostic tools used in newborn screening programs and as a confirmatory test, in case of suspect of Cystic Fibrosis (CF). Since sweat chloride (Cl) concentration is also considered an appropriate parameter to explore the efficacy of CFTR modulators in clinical trials, it is crucial to evaluate the biological variability of this test in healthy and pathological conditions. The aim of this pilot study was to determine the intra-individual biological variability of sweat Cl, both in healthy individuals and CF patients and to assess its correlation with diet, season, and menstrual cycle. Methods Thirty-five out of 36 selected subjects (6-18 years) were enrolled by 2 CF care centers and assigned to 3 cohorts: CF, CFTR-related disorder (CFTR-RD) and healthy volunteers. Each participant was subjected to eight sweat tests in different conditions and time of the year. Data were analyzed using linear mixed effects models for repeated measures, taking also into account intra-individual correlations. Results We observed a high intra-individual variability of sweat Cl, with the lowest mean CV% values among CF patients (20.21 in CF, 29.74 in CFTR-RD, and 31.15 in healthy subjects). Gender and diet had no influence on sweat Cl variability, nor had pubertal age and menstrual phase. Conclusion Results of this pilot study confirmed that sweat Cl variability is high in CF patients, although non-CF individuals displayed even higher mean CV% values. Season significantly influenced sweat test values only in CF patients, likely due to changes in their hydration status.

Background Recurrent (RP) and chronic pancreatitis (CP) may complicate Cystic Fibrosis (CF). It is still unknown if mutations in genes involved in the intrapancreatic activation of trypsin (IPAT) or in the pancreatic secretion pathway (PSP) may enhance the risk for RP/CP in patients with CF. Methods We enrolled: 48 patients affected by CF complicated by RP/CP and, as controls 35 patients with CF without pancreatitis and 80 unrelated healthy subjects. We tested a panel of 8 genes involved in the IPAT, i.e. PRSS1, PRSS2, SPINK1, CTRC, CASR, CFTR, CTSB and KRT8 and 23 additional genes implicated in the PSP. Results We found 14/48 patients (29.2%) with mutations in genes involved in IPAT in the group of CF patients with RP/CP, while mutations in such genes were found in 2/35 (5.7%) patients with CF without pancreatitis and in 3/80 (3.8%) healthy subjects (p

Background A clinical heterogeneity was reported in patients with Cystic Fibrosis (CF) with the same CFTR genotype and between siblings with CF. Methods We investigated all clinical aspects in a cohort of 101 pairs of siblings with CF (including 6 triplets) followed since diagnosis. Results Severe lung disease had a 22.2% concordance in sib-pairs, occurred early and the FEV1% at 12 years was predictive of the severity of lung disease in the adulthood. Similarly, CF liver disease occurred early (median: 15 years) and showed a concordance of 27.8% in sib-pairs suggesting a scarce contribution of genetic factors; in fact, only 2/15 patients with liver disease in discordant sib-pairs had a deficiency of alpha-1-antitrypsin (a known modifier gene of CF liver phenotype). CF related diabetes was found in 22 pairs (in 6 in both the siblings). It occurred later (median: 32.5 years) and is strongly associated with liver disease. Colonization by P. aeruginosa and nasal polyposis that required surgery had a concordance > 50% in sib-pairs and were poorly correlated to other clinical parameters. The pancreatic status was highly concordant in pairs of siblings (i.e., 95.1%) but a different pancreatic status was observed in patients with the same CFTR mutations. This suggests a close relationship of the pancreatic status with the “whole” CFTR genotype, including mutations in regulatory regions that may modulate the levels of CFTR expression. Finally, a severe course of CF was evident in a number of patients with pancreatic sufficiency. Conclusions Physicians involved in care of patients with CF and in genetic counseling must be aware of the clinical heterogeneity of CF even in sib-pairs that, at the state of the art, is difficult to explain. Electronic supplementary material The online version of this article (10.1186/s12890-018-0766-6) contains supplementary material, which is available to authorized users.

Candida albicans is implicated in intestinal diseases. Identifying host signatures that discriminate between the pathogenic vs commensal nature of this human commensal is clinically relevant. In the present study we have identified IL-9 and mast cells (MC) as key players of Candida commensalism and pathogenicity. By inducing TGF-β in stromal MC, IL-9 pivotally contributed to mucosal immune tolerance via the indoleamine 2,3-dioxygenase enzyme. However, Candida-driven IL-9 and mucosal MC also contributed to barrier function loss, dissemination and inflammation in experimental leaky gut models and were upregulated in patients with celiac disease. Inflammatory dysbiosis occurred in IL-9 and MC deficiency, a finding indicating that the activity of IL-9 and MC may go beyond host immunity to include regulation of the microbiota. Thus, the output of the IL9/MC axis is highly contextual during Candida colonization and reveals how host immunity and the microbiota finely tune Candida behavior in the gut.

Diabetes mellitus is the most common comorbidity in cystic fibrosis (CF), occurring in a variable number of children and adolescents. Glucose metabolism derangements (GMDs) are responsible for a negative impact on the general health status of CF patients. Screening of GMDs is important since the youngest age and should be performed by means of OGTT, including its intermediate times, that could detect other non-traditional GMDs. Insulin treatment, administered before overt diabetes, could be beneficial in reducing the number of pulmonary infections, in improving both pulmonary function and nutritional status. Early screening of GMDs in pediatric age can exert an important preventing role regarding all aspects of health status of patients with CF.

Background In cystic fibrosis (CF) the defective CF transmembrane conductance regulator protein may be responsible for the impaired transport of glutathione (GSH), the first line defense of the lung against oxidative stress. The aim of this single-blind, randomized, placebo-controlled trial was to evaluate the effect of inhaled GSH in patients with CF. Methods 54 adult and 51 pediatric patients were randomized to receive inhaled GSH or placebo twice daily for 12months. Results Twelve month treatment with inhaled GSH did not achieve our predetermined primary outcome measure of 15% improvement in FEV 1 %. Only in patients with moderate lung disease, 3, 6 and 9months therapy with GSH resulted in a statistically significant increase of FEV 1 values from the baseline. Moreover GSH therapy improved 6-minute walking test in pediatric population. GSH was well tolerated by all patients. Conclusions Inhaled GSH has slight positive effects in CF patients with moderate lung disease warranting further study. Trial registry ClinicalTrials.gov; No.: NCT01450267; URL: www.clinicaltrialsgov.

Introduction: primary ciliary dyskinesia (PCD) and cystic fibrosis (CF) are increasingly compared. There are no chest magnetic resonance imaging (MRI) comparative studies of PCD and CF. Aims and objectives: we assessed clinical, functional, microbiological and MRI findings in PCD and mild CF patients in order to evaluate different expression of lung disease. Methods: 20 PCD (15.1 years) and 20 CF subjects with mild respiratory impairment (16 years, 70% with pancreatic insufficiency) underwent MRI, spirometry, and sputum cultures when clinically stable. MRI was scored using the modified Helbich system. Results: PCD was diagnosed later than CF (9.9 versus 0.6 years, p=0.03), despite earlier symptoms (0.1 versus 0.6 years, p=0.02). In the year preceding the study, patients from both groups underwent two systemic antibiotic courses (p= 0.48). MRI total scores were 11.6±0.7 and 9.1±1 in PCD and CF, respectively. FEV1 and FVC Z- scores were -1.75 (range, -4.6-0.7) and -0.6 (-3.9-1.8) in PCD, and -0.9 (range, -5.4- 2.3) and -0.3 (-3.4-2.5) in CF, respectively. No difference was found between lung function or structure, despite a higher MRI subscore of collapse/consolidation in PCD versus CF (1.6±0.1 and 0.6±0.2, p Conclusions: MRI is a valuable radiation-free tool for comparative PCD and CF lung disease assessment. Patients with PCD may exhibit similar MRI and lung function changes as CF subjects with mild pulmonary disease. Delay in PCD diagnosis is unlikely the only determinant of similarities.

Cystic fibrosis (CF), the most common lethal monogenic disease in Caucasians, is characterized by recurrent bacterial infections and colonization, mainly by Pseudomonas aeruginosa, resulting in unresolved airway inflammation. CF is caused by mutations in the gene coding for the cystic fibrosis transmembrane conductance regulator (CFTR) protein, which functions as a chloride channel in epithelial cells, macrophages, and other cell types. Impaired bacterial handling by macrophages is a feature of CF airways, although it is still debated how defective CFTR impairs bacterial killing. Recent evidence indicates that a defective autophagy in CF macrophages leads to alterations of bacterial clearance upon infection. Here we use bone marrow-derived macrophages from transgenic mice to provide the genetic proof that defective CFTR compromises both uptake and clearance of internalized Pseudomonas aeruginosa. We demonstrate that the proteostasis regulator cysteamine, which rescues the function of the most common F508del-CFTR mutant and hence reduces lung inflammation in CF patients, can also repair the defects of CF macrophages, thus restoring both bacterial internalization and clearance through a process that involves upregulation of the pro-autophagic protein Beclin 1 and re-establishment of the autophagic pathway. Altogether these results indicate that cysteamine restores the function of several distinct cell types, including that of macrophages, which might contribute to its beneficial effects on CF.

Etiological therapies aim at repairing the underlying cause of cystic fibrosis (CF), which is the functional defect of the cystic fibrosis transmembrane conductance regulator (CFTR) protein owing to mutations in the CFTR gene. Among these, the F508del CFTR mutation accounts for more than two thirds of CF cases worldwide. Two somehow antinomic schools of thought conceive CFTR repair in a different manner. According to one vision, drugs should directly target the mutated CFTR protein to increase its plasma membrane expression (correctors) or improve its ion transport function (potentiators). An alternative strategy consists in modulating the cellular environment and proteostasis networks in which the mutated CFTR protein is synthesized, traffics to its final destination, the plasma membrane, and is turned over. We will analyze distinctive advantages and drawbacks of these strategies in terms of their scientific and clinical dimensions, and we will propose a global strategy for CF research and development based on a reconciliatory approach. Moreover, we will discuss the utility of preclinical biomarkers that may guide the personalized, patient-specific implementation of CF therapies.

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).

Cystic fibrosis (CF) is an autosomal recessive disease caused by mutations in the cystic fibrosis transmembrane regulator (CFTR) gene. About 2000 mutations have been described so far. We setup an ex vivo model of human nasal epithelial cells (HNECs) to study CF patients testing the effect of novel mutations and molecular therapies. We performed sampling (by brushing), followed by culture and analysis of HNECs using a series of molecular techniques. We performed 50 brushings from CF patients and controls. Using cultured cells, we: i) demonstrated the widely heterogeneous CFTR expression in patients and in controls; ii) defined the splicing effect of a CFTR mutation; iii) assessed the CFTR gating activity in patients bearing different mutations; iv) demonstrated that butyrate significantly enhances CFTR expression. Based on our data, we can conclude: 1) HNEC brushing is performed without anaesthesia and is well tolerated in all CF patients (children and adults); 2) HNECs can be preserved for up to 48 hours before culture allowings multicentre studies; 3) HNECs culture can be considered a suitable model to study the molecular effects of new CFTR gene mutations and/or uncertain meaning specific mutations of carriers; 4) an ex vivo model of HNECs may be used to evaluate, before human use, the effect of new drugs on patients' cells bearing specific CFTR mutations; 5) the methodology is adequate for a quantitative measurement, by fluorescence, of the CFTR gating activity of the HNECs from patients with different genotypes identifying: a) CF patients bearing two severe mutations with an activity10% (compared to controls - 100%); b) CF patients bearing at least a mild mutation with an activity of 10-20%; c) CF carriers (heterozygous subjects) with an activity between 40-70%.La fibrosi cistica (FC) è una malattia autosomica recessiva causata da mutazioni nel gene CFTR (Cystic Fibrosis Transmembrane conductance Regulator). Finora sono state descritte circa 2000 mutazioni, ma per la maggior parte di esse è difficile definirne l’effetto senza complesse procedure in vitro. Abbiamo effettuato il campionamento (mediante brushing), la cultura e l’analisi di cellule epiteliali nasali umane (HNEC) utilizzando una serie di tecniche che possono aiutare a testare l’effetto delle mutazioni CFTR. Abbiamo eseguito 50 brushing da pazienti FC e controlli, e in 45 casi si è ottenuta una coltura positiva. Utilizzando cellule in coltura: i) abbiamo dimostrato l’espressione ampiamente eterogenea del CFTR nei pazienti e nei controlli; ii) abbiamo definito l’effetto di splicing di una mutazione sul gene CFTR; iii) abbiamo valutato l’attività di gating di CFTR in pazienti portatori di differenti mutazioni; iv) abbiamo dimostrato che il butirrato migliora in modo significativo l’espressione di CFTR. I dati provenienti dal nostro studio sperimentale dimostrano che l’uso del modello ex-vivo di cellule epiteliali nasali è un importante e valido strumento di ricerca e di diagnosi nella studio della FC e può anche essere mirato alla sperimentazione ed alla verifica di nuovi farmaci. In definitiva, in base ai nostri dati è possibile esprimere le seguenti conclusioni: 1) il prelievo delle cellule epiteliali nasali mediante brushing è applicabile senza alcuna anestesia ed è ben tollerato da tutti i pazienti affetti da FC (bambini e adulti), è scarsamente invasivo e facilmente ripetibile, è anche in grado di ottenere una sufficiente quantità di HNECs rappresentative, ben conservate, idonee allo studio della funzionalità di CFTR; 2) la conservazione delle cellule prelevate è possibile fino a 48 ore prima che si provveda all’allestimento della coltura e ciò permette di avviare studi multicentrici con prelievi in ogni sede e quindi di ottenere una ampia numerosità campionaria; 3) la coltura di cellule epiteliali nasali può essere considerata un modello adatto a studiare l’effetto molecolare di nuove mutazioni del gene CFTR e/o mutazioni specifiche di pazienti “carriers” dal significato incerto; 4) il modello ex-vivo delle HNECs consente inoltre di valutare, prima dell’impiego nell’uomo, l’effetto di farmaci (potenziatori e/o correttori) sulle cellule di pazienti portatori di mutazioni specifiche di CFTR; tali farmaci possono modulare l’espressione genica del canale CFTR aprendo così nuove frontiere terapeutiche e migliori prospettive di vita per pazienti affetti da una patologia cronica come la Fibrosi Cistica; 5) la metodologia da noi istituita risulta essere idonea alla misura quantitativa, mediante fluorescenza, dell’attività di gating del canale CFTR presente nelle membrane delle cellule epiteliali nasali prelevate da pazienti portatori di differenti genotipi; in tal modo è possibile individuare: a) pazienti FC portatori di 2 mutazioni gravi con un’attività10% (in rapporto ai controlli -100%), b) soggetti FC portatori contemporaneamente di una mutazione grave e di una lieve con un’attività tra 10-30%, c) i cosiddetti portatori “carriers”- eterozigoti - con un’attività tra 40-70%. In conclusione la possibilità di misurare l’attività del canale CFTR in HNECs fornisce un importante contributo alla diagnosi di FC, mediante individuazione di un “cut-off diagnostico”, ed anche alla previsione della gravità fenotipica della malattia; quindi quanto rilevabile dalla misura del suddetto canale permette di prospettare per il futuro la possibilità di valutare meglio i pazienti per i quali il test del sudore ha dato risultati ambigui (borderline o negativi). La metodica da noi sperimentata consente anche di monitorare i pazienti durante il trattamento farmacologico, valutando in tal modo i reali effetti delle nuove terapie.

International audience; Cystic fibrosis (CF) is a lethal monogenic disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene that entails the (diagnostic) increase in sweat electrolyte concentrations, progressive lung disease with chronic inflammation and recurrent bacterial infections, pancreatic insufficiency, and male infertility. Therapies aimed at restoring the CFTR defect have emerged. Thus, a small molecule which facilitates chloride channel opening, the potentiator Ivacaftor, has been approved for the treatment of CF patients bearing a particular class of rare CFTR mutations. However, small molecules that directly target the most common misfolded CFTR mutant, F508del, and improve its intracellular trafficking in vitro, have been less effective than expected when tested in CF patients, even in combination with Ivacaftor. Thus, new strategies are required to circumvent the F508del-CFTR defect. Airway and intestinal epithelial cells from CF patients bearing the F508del-CFTR mutation exhibit an impressive derangement of cellular proteostasis, with oxidative stress, overactivation of the tissue transglutaminase (TG2), and disabled autophagy. Proteostasis regulators such as cysteamine can rescue and stabilize a functional F508del-CFTR protein through suppressing TG2 activation and restoring autophagy in vivo in F508del-CFTR homozygous mice, in vitro in CF patient-derived cell lines, ex vivo in freshly collected primary patient's nasal cells, as well as in a pilot clinical trial involving homozygous F508del-CFTR patients. Here, we discuss how the therapeutic normalization of defective proteostasis can be harnessed for the treatment of CF patients with the F508del-CFTR mutation.

Background:Low cholesterol is typically observed in the plasma of patients with cystic fibrosis (CF) contrasting with the subcellular accumulation of cholesterol demonstrated in CF cells and in mice models. However, the homeostasis of cholesterol has not been well investigated in patients with CF.Methods:We studied the plasma of 26 patients with CF and 33 unaffected controls campesterol and β-sitosterol as markers of intestinal absorption and lathosterol as a marker of de novo cholesterol biosynthesis by gas chromatography (GC-FID and GC-MS).Results:Plasma campesterol and β-sitosterol results were significantly (p=0.01) lower while plasma lathosterol was significantly higher (p=0.001) in patients with CF as compared to control subjects. Plasma cholesterol results were significantly lower (p=0.01) in CF patients.Conclusions:Our data suggest that the impaired intestinal absorption of exogenous sterols in patients with CF stimulates the endogenous synthesis of cholesterol, but the levels of total cholesterol in plasma remain lower. This may be due to the CFTR dysfunction that reduces cholesterol blood excretion causing the accumulation of cholesterol in liver cells and in other tissues contributing to trigger CF chronic inflammation.

Objectives Sweat chloride test is the gold standard test for cystic fibrosis (CF) diagnosis. In 2014 the Istituto Superiore di Sanita established the Italian pilot external quality assessment program for CF sweat test (IEQA-ST). Design and methods Ten laboratories, included among the 33 Italian CF Referral Centers, were selected and enrolled on the basis of their attitude to perform sweat test (ST) analysis by using methods recommended by the Italian Guidelines. They received three different sweat-like samples (normal, borderline and pathologic chloride concentration), with mock clinical indications, for analysis according to routine procedures. Assessment, performed by a panel of experts, covered analytical performance, interpretation and reporting of results; categories of “poor” and “satisfactory” performance were not defined. All data were managed through a web utility. Results The program identified important areas of interest and, in some case, of concern. It is important to underline that results are referred to a small proportion, i.e. about 30%, of Italian laboratories performing CF ST in the context of the Referral Centers. Conclusions Data collected highlight the importance of participation in EQA programs as it may improve laboratory/clinical performance; our study represents a model for the setting up of a large-scale EQA scheme for ST.

Channel activators (potentiators) of cystic fibrosis (CF) transmembrane conductance regulator (CFTR), can be used for the treatment of the small subset of CF patients that carry plasma membrane-resident CFTR mutants. However, approximately 90% of CF patients carry the misfolded Delta F508-CFTR and are poorly responsive to potentiators, because Delta F508-CFTR is intrinsically unstable at the plasma membrane (PM) even if rescued by pharmacological correctors. We have demonstrated that human and mouse CF airways are autophagy deficient due to functional sequestration of BECN1 and that the tissue transglutaminase-2 inhibitor, cystamine, or antioxidants restore BECN1-dependent autophagy and reduce SQSTM1/p62 levels, thus favoring Delta F508-CFTR trafficking to the epithelial surface. Here, we investigated whether these treatments could facilitate the beneficial action of potentiators on Delta F508-CFTR homozygous airways. Cystamine or the superoxide dismutase (SO D)/catalase-mimetic EUK-134 stabilized Delta F508-CFTR at the plasma membrane of airway epithelial cells and sustained the expression of CFTR at the epithelial surface well beyond drug withdrawal, overexpressing BECN1 and depleting SQSTM1. This facilitates the beneficial action of potentiators in controlling inflammation in ex vivo Delta F508-CFTR homozygous human nasal biopsies and in vivo in mouse Delta F508-CFTR lungs. Direct depletion of Sqstm1 by shRNAs in vivo in Delta F508-CFTR mice synergized with potentiators in sustaining surface CFTR expression and suppressing inflammation. Cystamine pre-treatment restored Delta F508-CFTR response to the CFTR potentiators genistein, Vrx-532 or Vrx-770 in freshly isolated brushed nasal epithelial cells from Delta F508-CFTR homozygous patients. These findings delineate a novel therapeutic strategy for the treatment of CF patients with the Delta F508-CFTR mutation in which patients are first treated with cystamine and subsequently pulsed with CFTR potentiators.

Rationale Chronic lung inflammation with increased susceptibility to bacterial infections cause much of the morbidity and mortality in patients with cystic fibrosis (CF), the most common severe, autosomal recessively inherited disease in the Caucasian population. Exogenous inhaled hyaluronan (HA) can exert a protective effect against injury and beneficial effects of HA have been shown in experimental models of chronic respiratory diseases. Our objective was to examine whether exogenous administration of nebulized HA might interfere with lung inflammation in CF. Study design/methods F508del homozygous mice (CftrF508del) and transgenic mice overexpressing the ENaC channel β-subunit (Scnn1b-Tg) were treated with nebulized HA (0.5 mg/mouse/day for 7 days). Tumor necrosis factor-alpha (TNFα), macrophage inflammatory protein-2 (MIP-2), myeloperoxidase (MPO) levels, and macrophage infiltration were assessed on lung tissues. IB3-1 and CFBE41o-epithelial cell lines were cultured with HA (24 hr, 100 µg/ml) and Reactive Oxygen Species (ROS), Tissue Transglutaminase (TG2) SUMOylation and Peroxisome Proliferator Activated Receptor gamma (PPARγ) and phospho-p42/p44 levels were measured by dichlorodihydrofluorescein assay, or fluorescence resonance energy transfer (FRET) microscopy or immunoblots. Results Nebulized HA reduced TNFα expression (P

Eur J Clin Invest 2011; 41 (10): 1063–1070 Abstract Background Nasal nitric oxide (nNO) is assessed by nasal aspiration/insufflation via one nostril or by nasal silent exhalation through a facemask and is also measured during humming, a manoeuvre that results in increased nNO in the presence of a patent osteomeatal complex. Humming nNO peak is absent in primary ciliary dyskinesia (PCD) and in cystic fibrosis (CF). Hand-held devices are used successfully for exhaled or nNO analysis. No study compared nNO during silent and humming exhalation using hand-held and stationary analysers. Methods Thirty-eight subjects (14 PCD; 11 CF; 13 healthy individuals) measured nNO with a stationary and a hand-held analyser during silent and humming exhalations. Results No difference between nNO obtained from stationary or hand-held analyser during silent and humming exhalation was found (P > 0·05). Patients with PCD exhibited lower silent and humming nNO than CF or controls (P 90% for discriminating PCD or CF from healthy subjects, and patients with PCD from patients with CF, respectively. Conclusions The hand-held device is as effective as the stationary analyzer for assessing nNO during silent and humming exhalation. Its wider use might result in an increased number of subjects suspected to have PCD.

We describe, to our knowledge, the first case of arterial ischemic stroke after primary herpes simplex virus type 1 (HSV1) infection in a previously healthy child, without signs of encephalitis. A 10-year-old previously healthy girl was admitted to our hospital with acute left-sided hemiparesis which involved the lower half of her face. Submandibular lymphadenitis and oral vesicular lesions were present. MRI confirmed the suspicion of an acute ischemic stroke. Immunoglobulin M antibodies to HSV1 were detected. Cerebrospinal fluid polymerase chain reaction for herpes virus was negative. She was treated with aspirin (3 mg/kg) and intravenous acyclovir (10 mg/kg every 8 hours) for 21 days. Immunoglobulin G antibodies to HSV1 appeared 16 days after admission. Twelve months after her hospitalization the patient’s examination was normal. Stroke should be considered a possible complication of HSV1 primary infection. Guidelines for the management of acute stroke in children are needed.

Background This is the second article related to a review of the literature based on data from national cystic fibrosis (CF) registries up to June 2008 and covering a total of 115 studies. It focuses on two topics: neonatal screening (NS) and nutritional status, with particular reference to growth. Methods Ten papers meeting the inclusion criteria were found on the topic of NS and its impact on the course of the disease, and were analyzed according to a dedicated grid. The issue of nutrition was addressed by 14 studies, analyzed according to similar criteria. Results Most of the studies report benefits of early diagnosis by NS, albeit to variable degrees. The benefits were assessed in terms of better nutritional status and growth, but also in terms of lower overall morbidity rate as compared to subjects diagnosed by symptoms. The main biases of these studies, which partly undermine the validity of their results, are also analyzed. A part of our analysis on nutrition/growth is dedicated to the identification of the most suitable parameters to define malnutrition: in children older than two years the body mass index percentile (BMIp) appears to be the most sensitive and significantly associated with respiratory function. Better nutritional status and satisfactory growth appear to be associated with better lung function and lower risk of death. The relationship between nutritional status and socio-economic status is also of interest. Conclusions CF registry studies support the outcome of cohort observational studies i.e. that pre-symptomatic early diagnosis is beneficial, especially in terms of nutritional status and growth. Studies on nutrition indicate that good nutritional status is associated with better respiratory function and prognosis. Regarding methods, the need emerged to manage potential biases of this kind of non randomized studies, resorting to suitable statistical techniques, such as matching and stratification and, above all, to multivariate methods able to provide estimates adjusted for the main covariates tested.

Objectives: To evaluate growth in Italian patients with cystic fibrosis (CF). Patients and Methods: A multicentre cross-sectional study was carried out on patients with CF attending Italian reference centres. Anthropometric data were evaluated using the Centers for Disease Control and Prevention 2000 reference data. Nutritional failure was defined as height-for-age percentile (HAP)