L. Claut

Identification of insulin secretory defects and insulin resistance during oral glucose tolerance test in a cohort of cystic fibrosis patients.

BACKGROUND Cystic fibrosis (CF)-related diabetes is a leading complication of CF and is associated with pulmonary and nutritional deterioration, years before an evident hyperglycemia, possibly because of insulin deficiency and resistance. AIM To evaluate glucose tolerance, insulin secretion, and insulin sensitivity by a widely applicable method suitable for accurate and prospective measurements in a CF population. METHODS A total of 165 CF subjects (80 females) aged 17±5 years and 18 age- and sex-matched healthy controls (CON) received an oral glucose tolerance test with glucose, insulin and C-peptide determinations. Insulin sensitivity was defined on the basis of glucose and insulin concentrations using the oral glucose insulin sensitivity index, whereas β-cell function was determined on the basis of a model relating insulin secretion (C-peptide profile) to glucose concentration. RESULTS Fifteen percent of CF patients had glucose intolerance and 6% had diabetes without fasting hyperglycemia and 3% had diabetes with fasting hyperglycemia. β-cell function was reduced in CF patients compared with CON (70.0±4.1 vs 117.9±11.6  pmol/min per m(2) per mM, P<0.001) and decreased significantly with age by -2.7  pmol/min per m(2) per mM per year (confidence interval (CI) -4.5 to -0.82), i.e. almost 4% yearly. The early insulin secretion index was also reduced. Insulin sensitivity was similar to CON. CF patients who attained glucose tolerance comparable to CON had lower β-cell function and higher insulin sensitivity. CONCLUSION The major alteration in insulin secretion and insulin sensitivity of CF patients is slowly declining β-cell function, consisting of delayed and reduced responsiveness to hyperglycemia, that in CF patients with normal glucose tolerance may be compensated by an increased insulin sensitivity.

A novel donor splice site characterized by CFTR mRNA analysis induces a new pseudo-exon in CF patients.

BACKGROUND The CFTR gene is tightly regulated and differentially expressed in many mucosal epithelial cell types. There is evidence of an increasing number of genomic variations in the intronic regions influencing mRNA splicing, and also the level of normal CFTR transcript. METHODS In the present study, we investigate the molecular defect by RT-PCR analyzing the mRNA of 25 cystic fibrosis (CF) patients in whom only one or no CF allele had been identified after DNA analysis (of all the exons of the CFTR gene). RESULTS mRNA analysis led to the detection of a cryptic exon in two patients: the new exon is a 104 bp insertion between exons 10 and 11 and is caused by a new point mutation c.1584+18672 bp A>G (http://www.hgvs.org/mutnomen/) discovered in intron 10; moreover, they showed the absence of exon 9 skipping. CONCLUSIONS Our results confirm the utility of RNA analysis in discovering new mutations and in investigating their effect on normal splicing processes.

Efficacy and tolerability of a new nasal spray formulation containing hyaluronate and tobramycin in cystic fibrosis patients with bacterial rhinosinusitis

BACKGROUND Chronic rhinosinusitis is common in cystic fibrosis (CF), as CFTR defects equally affect the airway and sinonasal mucosa. However, therapeutic strategies for CF-associated chronic rhinosinusitis lag behind current approaches for pulmonary disease. OBJECTIVE To assess the tolerability and efficacy of a nasal spray formulation containing 0.2% sodium hyaluronate and 3% tobramycin compared to a control formulation containing 0.2% sodium hyaluronate alone in the treatment of bacterial rhinosinusitis in patients with CF. METHODS In a double-blind controlled study, 27 patients with an established diagnosis of CF and a documented nasal infection with Pseudomonas aeruginosa and/or Staphylococcus aureus [22 males (81%), median age of 15 years (range 5-26 yrs)], were randomized to receive the nasal spray formulation containing hyaluronate and tobramycin (N=14) or hyaluronate alone (N=13) for 14 days. Efficacy and local tolerability of the treatments were assessed by ear, nose and throat (ENT) examination and related symptoms. RESULTS The formulation containing hyaluronate and tobramycin was more effective than hyaluronate alone in improving the status of the nasal mucosa, in reducing the mucopurulent secretion at the level of the osteomeatal complex and in improving ENT symptoms (hyposmia/anosmia and headache/facial pain). The treatment was well tolerated without relevant side effects. CONCLUSIONS The present study suggests that the combination therapy with hyaluronate plus tobramycin was more effective than hyaluronate alone in the treatment of bacterial rhinosinusitis in CF. TRIAL REGISTRATION NUMBER EudraCT 2007-003628-39.

A wide methodological approach to identify a large duplication in CFTR gene in a CF patient uncharacterised by sequencing analysis

BACKGROUND PCR-based diagnostic procedures are not able to characterise 6% of CF alleles. Recently, the application of array-CGH and of CFTR mRNA analysis has allowed the identification of new copy number mutations and splicing defects, that account for 2% and 13% of CF alleles, respectively, in the Italian population. METHODS Here, we report the characterisation of a large duplication in CFTR gene through different methods: MLPA assay, RT-PCR and high-resolution array-CGH. RESULTS We identified a large duplication, involving exons 6b-16, in a patient heterozygous for F508del mutation. This duplication produces an abnormal transcript with an out of frame addition of 2244 nucleotides and leads to the insertion of 8 amino-acid residues in the protein, followed by a stop codon. CONCLUSIONS We propose a wide methodological approach based on MLPA assay, RT-PCR and high-resolution array-CGH to routinely analyse CF patients uncharacterised for one or both CFTR alleles.

Can biliary endoscopy play a role in liver disease associated to cystic fibrosis

a Gastroenterology and Endoscopy Unit, Fondazione IRCCS Ca’ Granda, Ospedale Maggiore Policlinico, Department of Pathophysiology and Transplantation, Universita degli Studi di Milano, Milan, Italy b Cystic Fibrosis Reference Center, Fondazione IRCCS Ca’ Granda, Ospedale Maggiore Policlinico, Universita degli Studi di Milano, Milan, Italy c Department of Pathology, Fondazione IRCCS Ca’ Granda, Ospedale Policlinico Milano, Universita degli Studi, Milan, Italy

138 Genetic fingerprinting of Stenotrophomonas maltophilia strains isolated from cystic fibrosis patients

Persons with CF can acquire Burkholderia cepacia complex (Bcc) infection through patient-to-patient contacts or from the environment. The former way of acquisition was well documented in the Prague CF centre where infection rate with a single B. cenocepacia strain ST32 reached over 30% in 1997–2003. The aim of this study was to compare epidemiological situation in the Centre in 2003 vs. 2010, and to evaluate efficacy of infection control in tackling the spread of ST32. System of surveillance exploiting typing techniques (RAPD and MLST) was set up in 2008. A total of 76 out of 211 patients examined by the end of 2003 (36.0%) were infected with Bcc; majority of them (78.9%) harboured epidemic strain ST32. Seven years later, Bcc positivity was detected in 72 out of 374 patients who attended the Prague CF clinic (19.3%). While 39 patients still suffered from infection with ST32 (54.2% of infected), a substantial portion of patients carried strains other than ST32 (see Table; note that ST number is specified only if more than two patients harboured the strain). Notably, only 2 patients within the ST32 group became positive after 2003, with the last case dated in May 2007. Epidemiological situation characterized by increasing heterogeneity of the Bcc population and no occurrence of new ST32 cases is a likely consequence of both healthcare workers’ and patients’ good compliance with strict infection control rules. Supported by NS10543−3, MSM0021620812 and MZ0FNM2005.