ECFS standards of care on CFTR-related disorders: Towards a comprehensive program for affected individuals.

After three publications defining an updated guidance on the diagnostic criteria for people with cystic fibrosis transmembrane conductance regulator (CFTR)-related disorders (pwCFTR-RDs), establishing its relationship to CFTR-dysfunction and describing the individual disorders, this fourth and last paper in the series addresses some critical challenges facing health care providers and pwCFTR-RD. Topics included are: 1) benefits and obstacles to collect data from pwCFTR-RD are discussed, together with the opportunity to integrate them into established CF-registries; 2) the potential of infants designated CRMS/CFSPID to develop a CFTR-RD and how to communicate this information; 3) a description of the challenges in genetic counseling, with particular regard to phenotypic variability, unknown long-term evolution, CFTR testing and pregnancy termination 4) a proposal for the assessment of potential barriers to the implementation and dissemination of the produced documents to health care professionals involved in the care of pwCFTR-RD and a process to monitor the implementation of the CFTR-RD recommendations; 5) clinical trials investigating the efficacy of CFTR modulators in CFTR-RD and how endpoints and outcomes might be adapted to the heterogeneity of these disorders.

A new highly discriminatory multiplex capillary-based MLVA assay as a tool for the epidemiological survey of Pseudomonas aeruginosa in cystic fibrosis patients.

Multiple locus variable number of tandem repeats (VNTR) analysis (MLVA) has been shown to provide a high level of information for epidemiological investigations and the follow-up of Pseudomonas aeruginosa chronic infection. In the present study, an automatized MLVA assay has been developed for the analysis of 16 VNTRs in two multiplex polymerase chain reactions (PCRs), followed by capillary electrophoresis. The result in the form of a code is directly usable for clustering analyses. This MLVA-16(Orsay) scheme was applied to the genotyping of 83 isolates from eight cystic fibrosis patients, demonstrating that the same genotype persisted during eight years of chronic infection in the majority of cases. Comparison with pulsed-field gel electrophoresis (PFGE) analysis showed that both methods were congruent, MLVA providing, in some cases, additional informativity. The evolution of strains during long-term infection was revealed by the presence of VNTR variants.

Socio-economic status influences blood pressure control despite equal access to care.

OBJECTIVE: Denmark has a health care system with free and equal access to care irrespective of age and socio-economic status (SES). We conducted a cross-sectional study to investigate a possible association between SES and blood pressure (BP) control of hypertensive patients treated in general practice. METHODS: We enrolled 184 general practices and 5260 hypertensive patients. The general practitioners reported information about BP and diagnosis of diabetes. Information about education, income, antihypertensive drug treatment and other co-morbidity was retrieved from relevant registers from Statistics Denmark. The outcome measure was BP control defined as BP <140/90 mmHg in general and <130/80 mmHg in diabetics. RESULTS: Patients <65 years and with an educational level of 10-12 years had increased odds ratio (OR) of BP control compared to patients with an educational level <10 years. Patients ≥65 years had increased OR of BP control if they were married/cohabiting as compared to being single, whereas education and income had no impact in this age group. Diabetics had significantly reduced odds of BP control irrespective of age, educational or income level. CONCLUSIONS: Despite equal access to care for all patients, SES had significant impact on BP control in this survey. Diabetes and cardiovascular disease also had a substantial influence irrespective of age, educational and income level.

Audit report from Greenland on nurses' tasks and perceived competency.

INTRODUCTION: Despite all efforts, recruitment of healthcare personnel has become increasingly difficult in Greenland as in other remote areas. The aim of this observational study was to describe the extent of health care delivered by nurses in Greenland's healthcare system. Reasons for encounter, diagnostic procedures, treatments and need for a physician's assistance, as well as the nurses' self-perceived competency, were also analysed. METHODS: A total of 42 nurses registered all patient encounters for 10 days in late autumn 2006 in 14 out of 16 healthcare districts in Greenland. RESULTS: Nurses treated 1117 encounters (60%) singlehandedly. The nurses felt competent in what they were doing in 1415 encounters (76%). In 525 encounters (31%), a physician's advice was sought. Either the physician was asked to come or the physician's advice was obtained by telephone. In four cases the nurses did not feel completely competent, but did not seek advice from the physician on call. Feeling competent did not depend on length of experience in Greenland. CONCLUSION: In Greenland, nurses independently receive, diagnose and treat a substantial number of primary healthcare patients. The nurses take care of the patients and perform a number of clinical and laboratory procedures with great confidence. There has been speculation that part of the difficulty in recruiting doctors and healthcare personnel in remote areas may be due to uneasiness about professional responsibilities and, to some extent, lack of confidence. At least among the registering nurses in this study, this did not seem to be a problem.

Standards of care guidance for sweat testing; phase two of the ECFS quality improvement programme.

More than five decades after the introduction of the quantitative pilocarpine iontophoresis technique, surveys still highlight inconsistencies in the performance and reporting of sweat tests in Europe. The sweat test remains key for the Cystic Fibrosis (CF) diagnostic pathway for all age groups, as it reflects the basic pathophysiological defect in the sweat gland. It is also critical following newborn screening as a confirmatory diagnostic step. Despite its importance, sweat test quality is variable whether performed in the laboratory or as a point of care test. The ECFS DNWG aims to improve sweat test performance, taking into account the barriers and issues identified in the European survey; the previous step in the ECFS sweat test project. This manuscript proposes a grading of sweat test guidance from "acceptable" to "optimal", aiming to pragmatically improve quality while taking into account local situations, especially in resource-limited settings.

Treatment of 5413 hypertensive patients: a cross-sectional study.

BACKGROUND: Most hypertensive patients are managed in primary care in Denmark, but previous studies have shown that only 21-43% of hypertensive patients achieve optimal blood pressure (BP) control. Antihypertensive drug treatment, risk factors and cardiovascular disease (CVD) are some of the important factors to consider when optimizing the individual treatment strategy in hypertensive patients. OBJECTIVE: To examine treatment of BP according to Danish guidelines (BP < 140/90 mmHg generally and <130/80 mmHg for diabetics) in a population from general practice in relation to risk factors, CVD and diagnosis of diabetes. METHODS: A cross-sectional study comprising 184 practices and 5413 hypertensive patients was carried out in Denmark. The general practitioners filled in information on each patient's risk factors, CVD and antihypertensive drug treatment. Patients filled in a questionnaire on risk factors. The outcome measures were optimal BP control according to Danish guidelines and antihypertensive drug treatment. RESULTS: Mean patient age was 65.9 years [95% confidence interval (CI): 65.6-66.1]. Optimal BP control was achieved in 29.1% (95% CI: 27.9-30.3) of the study population. Among 842 diabetics with or without CVD, optimal BP control was achieved in 10.9% (95% CI: 8.8-10.3), while 38.7% (35.5-41.9) of patients with CVD achieved optimal BP control. The majority of all patients were treated with 1 (32.5%, 95% CI: 32.5 (31.3-33.8)) or two antihypertensive drugs (39.0%, 95% CI: 38.2-40.8). In hypertensive diabetics, 17.7% were not treated with an angiotensin-converting enzyme inhibitor or angiotensin-receptor blocker. CONCLUSION: In general practice, the proportion of hypertensive patients achieving optimal BP control is inadequate. The majority of hypertensive patients are treated with only one or two antihypertensive drugs.

T cell growth factor receptors. Quantitation, specificity, and biological relevance.

To examine directly the hypothesis that T cell growth factor (TCGF) interacts with target cells in a fashion similar to polypeptide hormones, the binding of radiolabeled TCGF to various cell populations was investigated. The results indicate that TCGF interacts with activated T cells via a receptor through which it initiates the T cell proliferative response. Internally radiolabeled TCGF, prepared from a human T leukemia cell line and purified by gel filtration and isoelectric focusing, retained biological activity and was uniform with respect to size and charge. Binding of radiolabeled TCGF to TCGF-dependent cytolytic T cells occurred rapidly (within 15 rain at 37 degrees C) and was both saturable and largely reversible. In addition, at 37 degrees C, a receptor- and lysosome-dependent degradation of TCGF occurred. Radiolabeled TCGF binding was specific for activated, TCGF-responsive T cells. Whereas unstimulated lymphocytes of human or murine origin and lipopolysaccharide-activated B cell blasts expressed few if any detectable binding sites, lectin- or alloantigen-activated cells had easily detectable binding sites. Moreover, compared with lectin- or alloantigen-activated T cells, long-term TCGF-dependent cytolytic and helper T cell lines and TCGF-dependent neo-plastic T cell lines bound TCGF with a similar affinity (dissociation constant of 5-25 pM) and expressed a similar number of receptor sites per cell (5,000-15,000). In contrast, a number of TCGF-independent cell lines of T cell, B cell, or myeloid origin did not bind detectable quantities of radiolabeled TCGF. Binding of radiolabeled TCGF to TCGF-responsive cells was specific, in that among several growth factors and polypeptide hormones tested, only TCGF competed for binding. Finally, the relative magnitude of T cell proliferation induced by a given concentration of TCGF closely paralleled the fraction of occupied receptor sites. As the extent of T cell clonal expansion depends on TCGF and on the TCGF receptor, the dissection of the molecular events surrounding the interaction of TCGF and its receptor that these studies permit, should provide new insight into the hormonelike regulation of the immune response by this lymphokine.

Interaction of glucocorticoids with macrophages. Identification of glucocorticoid receptors in monocytes and macrophages.

Glucocorticoid binding was measured in resident and thioglycollate-elicited mouse peritoneal macrophages, rabbit alveolar macrophages, and human monocytes. Two assays of binding were used--an assay with intact cells in suspension or monolayers, and an assay of cytosol and nuclear forms of glucocorticoid receptors. The mononuclear phagocytes contained approximately equal to 4--10 X 10(3) high affinity receptor sites per cell, with dissociation constants of approximately equal to 2--8 nM dexamethasone. The binding to the saturable sites was specific for steroids with glucocorticoid or antiglucocorticoid activity. Cortisol, corticosterone, and progesterone competed with dexamethasone for binding, whereas estradiol, dihydrotestosterone, and 11-epicortisol competed very little. Binding of dexamethasone to cytosol and nuclear forms of the receptor complex and temperature-sensitive translocation of cytosol forms to nuclear forms were shown. At 37 degrees C the predominant form of the hormone-receptor complex was nuclear. These results demonstrate that corticosteroids interact with macrophages at physiological concentrations.

The very low penetrance of cystic fibrosis for the R117H mutation: a reappraisal for genetic counselling and newborn screening.

BACKGROUND: Cystic fibrosis (CF) is caused by compound heterozygosity or homozygosity of CF transmembrane conductance regulator gene (CFTR) mutations. Phenotypic variability associated with certain mutations makes genetic counselling difficult, notably for R117H, whose disease phenotype varies from asymptomatic to classical CF. The high frequency of R117H observed in CF newborn screening has also introduced diagnostic dilemmas. The aim of this study was to evaluate the disease penetrance for R117H in order to improve clinical practice. METHODS: The phenotypes in all individuals identified in France as compound heterozygous for R117H and F508del, the most frequent CF mutation, were described. The allelic prevalences of R117H (p(R117H)), on either intron 8 T5 or T7 background, and F508del (p(F508del)) were determined in the French population, to permit an evaluation of the penetrance of CF for the [R117H]+[F508del] genotype. RESULTS: Clinical details were documented for 184 [R117H]+[F508del] individuals, including 72 newborns. The disease phenotype was predominantly mild; one child had classical CF, and three adults' severe pulmonary symptoms. In 5245 healthy adults, p(F508del) was 1.06%, p(R117H;T7) 0.27% and p(R117H;T5)<0.01%. The theoretical number of [R117H;T7]+[F508del] individuals in the French population was estimated at 3650, whereas only 112 were known with CF related symptoms (3.1%). The penetrance of classical CF for [R117H;T7]+[F508del] was estimated at 0.03% and that of severe CF in adulthood at 0.06%. CONCLUSIONS: These results suggest that R117H should be withdrawn from CF mutation panels used for screening programmes. The real impact of so-called disease mutations should be assessed before including them in newborn or preconceptional carrier screening programmes.

Newborn screening for CF in France: An exemplary national experience.

Newborn screening (NBS) for cystic fibrosis (CF) was implemented throughout France since 2002, with a 3-tiered strategy consisting in an immunoreactive trypsinogen (IRT) measurement at day-3, a search for the most common mutations responsible for CF when the IRT value is above the cut-off level, and, if necessary, a safetynet retesting of IRT at day-21. Coordination and follow-up are ensured at the national level and NBS is carried out through a regional organization involving NBS centers, biochemical and molecular genetics laboratories. Sweat testing and comprehensive mutation gene analysis are then performed according to a defined algorithm. Between 2002 and 2014, screening for the 30 most common mutations identified 87% of the alleles and comprehensive mutation gene analysis performed when applicable identified more than 300 additional mutations and resulted in a detection rate of 99.8% of the mutated alleles. Program surveillance ensured at a national level allowed to carry out adaptation of cut-off levels and removal of the p.Arg117His mutation. Thanks to these modifications, the performance of the French NBS program for CF meets the European guideline standards regarding positive predictive values, sensitivity and time to initial visit at the CF center, thus making the strategy effective. © 2020 French Society of Pediatrics. Published by Elsevier Masson SAS. All rights reserved.

Penetrance is a critical parameter for assessing the disease liability of CFTR variants.

BACKGROUND: Major issues of newborn screening (NBS) for CF are the assessment of disease liability of variants and of the penetrance of clinical CF, notably in inconclusive diagnosis. The penetrance of CF is defined as the risk of a particular genotype to lead to a CF phenotype. METHODS: We aimed to get insight into the penetrance of CF for fifteen CFTR variants: 5 frequent CF-causing and 10 classified as of varying clinical consequence (VCC) or associated with a CFTR-related disorder (CFTR-RD) in CFTR2 or CFTR-France databases. The penetrance was approached by: (1) comparison of variant allelic frequencies in CF patients (CFTR2) and in the general population; (2) estimation of the likelihood of a positive NBS test for the 14 compound heterozygous with F508del and the F508del homozygous genotypes, defined as the ratio of detected/expected number of neonates with a given genotype in the 2002-2017 period. RESULTS: A full penetrance was observed for severe CF-causing variants. Five variants were more frequently found in the general population than in CF patients: TG11T5, TG12T5, TG13T5, L997F and R117H;T7. The likelihood of a positive NBS test was 0.03% for TG11T5, 0.3% for TG12T5, 1.9% for TG13T5, 0.6% for L997F, 11.7% for D1152H, and 17.8% for R117H;T7. Penetrance varied greatly for variants with discrepant classification between CFTR2 and CFTR-France: 5.1% for R117C, 12.3% for T338I, 43.5% for D110H and 52.6% for L206W. CONCLUSION: These results illustrate the contribution of genetics population data to assess the disease liability of variants for diagnosis and genetic counselling purposes.

Phenotypic characterisation of patients with intermediate sweat chloride values: towards validation of the European diagnostic algorithm for cystic fibrosis.

BACKGROUND: In patients with symptoms suggestive of cystic fibrosis (CF) and intermediate sweat chloride values (30-60 mmol/l), extensive CFTR gene mutation analysis and nasal potential difference (NPD) measurement are used as additional diagnostic tests and a positive result in either test provides evidence of CFTR dysfunction. To define the phenotype of such patients and confirm the validity of grouping them, patients with intermediate sweat chloride values in whom either additional CF diagnostic test was abnormal were compared with subjects in whom this was not the case and patients with classic CF. METHODS: The phenotypic features of four groups were compared: 59 patients with CFTR dysfunction, 46 with an intermediate sweat chloride concentration but no evidence of CFTR dysfunction (CF unlikely), 103 patients with CF and pancreatic sufficiency (CF-PS) and 62 with CF and pancreatic insufficiency (CF-PI). RESULTS: The CFTR dysfunction group had more lower respiratory tract infections (p = 0.01), more isolation of CF pathogens (p<0.001) and clubbing (p = 0.001) than the CF unlikely group, but less frequent respiratory tract infections with CF pathogens than the CF-PS group (p = 0.05). Patients in the CF-PS group had a milder phenotype than those with PI. Many features showed stepwise changes through the patient groups. CONCLUSION: Patients with intermediate sweat chloride values and two CFTR mutations or an abnormal NPD measurement have a CF-like phenotype compatible with CFTR dysfunction and, as a group, differ phenotypically from patients with intermediate sweat chloride values in whom further CF diagnostic tests are normal as well as from CF-PS and CF-PI patients.

Absence of microsomal triglyceride transfer protein in individuals with abetalipoproteinemia.

Abetalipoproteinemia is a human genetic disease that is characterized by a defect in the assembly or secretion of plasma very low density lipoproteins and chylomicrons. The microsomal triglyceride transfer protein (MTP), which is located in the lumen of microsomes isolated from the liver and intestine, has been proposed to function in lipoprotein assembly. MTP activity and the 88-kilodalton component of MTP were present in intestinal biopsy samples from eight control individuals but were absent in four abetalipoproteinemic subjects. This finding suggests that a defect in MTP is the basis for abetalipoproteinemia and that MTP is indeed required for lipoprotein assembly.

A European consensus for the evaluation and management of infants with an equivocal diagnosis following newborn screening for cystic fibrosis.

Screening newborns for cystic fibrosis (CF) is considered to be an ethical undertaking in regions with a significant incidence of the condition. Current screening protocols result in recognition of infants with an equivocal diagnosis. A survey of European practice suggested inconsistencies in the evaluation and management of these infants. We have undertaken a consensus process using a modified Delphi method. This has enabled input of CF specialists from a wide geographical area to a rigorous process that has provided a clear pathway to a consensus statement. A core group produced 21 statements, which were modified over a series of three rounds (including a meeting arranged at the European CF Conference). A final document of 19 statements was produced, all of which achieved a satisfactory level of consensus. The statements cover four themes; sweat testing, further assessments and investigations, review arrangements and database. This consensus document will provide guidance to CF specialists with established screening programmes and those who are in the process of implementing newborn screening in their region.

Bioavailability of oral vitamin E formulations in adult volunteers and children with chronic cholestasis or cystic fibrosis.

PURPOSE: To test bioequivalence of oral vitamin E formulations, water-soluble tocofersolan (test) and water-miscible (reference), in healthy adult volunteers, and their bioavailability in children with chronic cholestasis or cystic fibrosis. METHODS: In a two-way open randomized single dose cross-over design, 1200 IU were administered in 12 healthy volunteers and 100 IU/kg in 12 children with chronic cholestasis or cystic fibrosis. RESULTS: In healthy volunteers, formulations were not bioequivalent with a higher exposure to tocofersolan. In cholestatic children tocofersolan bioavailability was significantly higher than reference formulation (maximum plasma concentration: P = 0.008 and AUC: P = 0.0026). Bioavailability was not statistically different in cystic fibrosis. CONCLUSIONS: Oral tocofersolan was more bioavailable than the reference formulation in children with chronic cholestasis and similarly bioavailable in cystic fibrosis. Tocofersolan may represent an alternative to painful intramuscular vitamin E injections in chronic cholestasis, or to other oral formulations in cystic fibrosis.

Prevention of relapse by mesalazine (Pentasa) in pediatric Crohn's disease: a multicenter, double-blind, randomized, placebo-controlled trial.

AIM: This study aimed to test the efficacy of mesalazine in maintaining remission in pediatric Crohn's disease (CD) following successful flare-up treatment. METHODS: In this double-blind, randomized, placebo-controlled trial, 122 patients received either mesalazine 50mg/kg per day (n=60) or placebo (n=62) for one year. Treatment allocation was stratified according to flare-up treatment (nutrition or medication alone). Recruitment was carried out over two periods, as the first period's results showed a trend favoring mesalazine. Relapse was defined as a Harvey-Bradshaw score more than or equal to 5. Time to relapse was analyzed using the Cox model. RESULTS: The one-year relapse rate was 57% (n=29) and 63% (n=35) in the mesalazine and placebo groups, respectively. We demonstrated a twofold lower relapse risk (P<0.02) in patients taking mesalazine in the medication stratum (first recruitment period), and a twofold higher risk in patients taking mesalazine in the nutrition stratum (second recruitment period), compared with the other groups. None of the children's characteristics, which differed across the two recruitment periods, accounted for the between-period variation in mesalazine efficacy. One serious adverse event was reported in each treatment group. CONCLUSION: Overall, mesalazine does not appear to be an effective maintenance treatment in pediatric CD.

Phosphorylation of steroid hormone receptors.

Many observations with intact cells as well as cell-free systems suggest that receptors of the steroid hormone superfamily, along with other transcription factors, are regulated by phosphorylation. All receptors that have been analyzed carefully so far have turned out to be phosphoproteins. They are basally phosphorylated in the absence of ligand, and in many cases become hyperphosphorylated in the presence of hormone or other agonists, and sometimes of antagonists. Several studies indicate that hyperphosphorylation of receptors follows activation, and may require nuclear binding of the receptor. Serines are the predominant phosphorylated residues detected in receptors, with minor amounts of threonine. Tyrosine phosphorylation of the estrogen receptor is a subject of controversy. With various receptors, evidence has been found for phosphorylation in vivo of the N-terminal, hormone-binding, and DNA-binding domains, as well as of the hinge region. All but one of the phosphorylated sites identified in progesterone and glucocorticoid receptors by phosphopeptide mapping and sequencing are in the N-terminal domain; one is in the hinge region. Even after hormone treatment most of those sites are only partly phosphorylated, which means that several subpopulations of receptors, characterized by different states of phosphorylation and potentially different biological activities, must coexist. The majority of identified phosphorylated sites lie in consensus sequences for the PDPK. Many parallels can be discerned between phosphorylation of receptors and of other transcription factors. For example, several transcription factors become hyperphosphorylated on stimulation, and much indirect evidence points to regulation of both receptors and transcription factors by kinases and phosphatases, with cycling between different phosphorylated states. Functions of receptors that are regulated by phosphorylation are only beginning to be investigated. With transcription factors a substantial body of information is already available, and functions that appear to be thus regulated include dimerization, interactions with other proteins, binding to DNA, nuclear-cytoplasmic localization, and transcriptional activity. These and other functions may be found to be regulated by phosphorylation of receptors.