Standards for the care of people with cystic fibrosis; establishing and maintaining health.

This is the second in a series of four papers updating the European Cystic Fibrosis Society (ECFS) standards for the care of people with CF. This paper focuses on establishing and maintaining health. The guidance is produced using an evidence-based framework and with wide stakeholder engagement, including people from the CF community. Authors provided a narrative description of their topic and statements, which were more directive. These statements were reviewed by a Delphi exercise, achieving good levels of agreement from a wide group for all statements. This guidance reinforces the importance of a multi-disciplinary CF team, but also describes developing models of care including virtual consultations. The framework for health is reinforced, including the need for a physically active lifestyle and the strict avoidance of all recreational inhalations, including e-cigarettes. Progress with cystic fibrosis transmembrane conductance regulator (CFTR) modulator therapy is reviewed, including emerging adverse events and advice for dose reduction and interruption. This paper contains guidance that is pertinent to all people with CF regardless of age and eligibility for and access to modulator therapy.

Imaging the Response to DNA Damage in Heterochromatin Domains.

The eukaryotic genome is assembled in a nucleoprotein complex called chromatin, whose organization markedly influences the repair of DNA lesions. For instance, compact chromatin states, broadly categorized as heterochromatin, present a challenging environment for DNA damage repair. Through transcriptional silencing, heterochromatin also plays a vital role in the maintenance of genomic integrity and cellular homeostasis. It is thus of critical importance to decipher whether and how heterochromatin affects the DNA damage response (DDR) to understand how this chromatin state is preserved after DNA damage. Here, we present two laser micro-irradiation-based methods for imaging the DDR in heterochromatin domains in mammalian cells. These methods allow DNA damage targeting to specific subnuclear compartments, direct visualization of the DDR and image-based quantification of the repair response. We apply them to study DNA double-strand break repair pathways in facultative heterochromatin and the repair of UV photoproducts in constitutive heterochromatin. We discuss the advantages and limitations of these methods compared to other targeted approaches for DNA damage induction.

Corrigendum: Unveiling Interindividual Variability of Human Fibroblast Innate Immune Response Using Robust Cell-Based Protocols.

[This corrects the article DOI: 10.3389/fimmu.2020.569331.].

Imaging the response to DNA damage in heterochromatin domains reveals core principles of heterochromatin maintenance.

Heterochromatin is a critical chromatin compartment, whose integrity governs genome stability and cell fate transitions. How heterochromatin features, including higher-order chromatin folding and histone modifications associated with transcriptional silencing, are maintained following a genotoxic stress challenge is unknown. Here, we establish a system for targeting UV damage to pericentric heterochromatin in mammalian cells and for tracking the heterochromatin response to UV in real time. We uncover profound heterochromatin compaction changes during repair, orchestrated by the UV damage sensor DDB2, which stimulates linker histone displacement from chromatin. Despite massive heterochromatin unfolding, heterochromatin-specific histone modifications and transcriptional silencing are maintained. We unveil a central role for the methyltransferase SETDB1 in the maintenance of heterochromatic histone marks after UV. SETDB1 coordinates histone methylation with new histone deposition in damaged heterochromatin, thus protecting cells from genome instability. Our data shed light on fundamental molecular mechanisms safeguarding higher-order chromatin integrity following DNA damage.

"Il faut continuer à poser des questions" patient reported outcome measures in cystic fibrosis: An anthropological perspective.

BACKGROUND: People with cystic fibrosis (pwCF) are central in the development of patient-led assessment tools. Qualitative analysis of a frequently used CF-specific patient-reported outcome measure (PROM) sought patient recommendations for development of a new quality of life (QoL) tool. METHODS: We performed an inventory of PROMs, symptom-report and QoL tools used in clinical trials within the European Cystic Fibrosis Society Clinical Trial Network (ECFS-CTN) and in routine clinical practice among Cystic Fibrosis Europe and ECFS members. A qualitative study using cognitive interviews with pwCF and their caregivers reviewed the Cystic Fibrosis Questionnaire (CFQ), the French initial form of the Cystic Fibrosis Questionnaire-Revised (CFQ-R). RESULTS: Survey results from 33 countries revealed over 70 tools used in routine clinical practice, utilized by clinical specialists (n=124), pwCF/parents/carers (n=49) and other allied health professionals (n=60). The CFQ-R was the main PROM used in clinical trials. The qualitative study enrolled 99 pwCF, 6 to 11 years (n=31); 12 to 18 years (n=38); >18 years (n=30) and 26 parents. Inductive thematic analysis based on the CFQ, revealed 19 key themes. Themes common across all cohorts included burden of treatment, impact of disease on day-to-day life, relationships/family, stress/mood, and nutrition. Themes unique to individual groups included, treatment when not symptomatic for the paediatric group; education/studies and planning for the future for adolescents, impact of anxiety and depression on day-to-day life for adults, and for parents, questions addressing anxiety and their role as carers. CONCLUSIONS: Patient-centeredness is paramount in development of an up-to-date PROM in the era of novel therapies.

Unveiling Interindividual Variability of Human Fibroblast Innate Immune Response Using Robust Cell-Based Protocols.

The LabEx Milieu Interieur (MI) project is a clinical study centered on the detailed characterization of the baseline and induced immune responses in blood samples from 1,000 healthy donors. Analyses of these samples has lay ground for seminal studies on the genetic and environmental determinants of immunologic variance in a healthy cohort population. In the current study we developed in vitro methods enabling standardized quantification of MI-cohort-derived primary fibroblasts responses. Our results show that in vitro human donor cohort fibroblast responses to stimulation by different MAMPs analogs allows to characterize individual donor immune-phenotype variability. The results provide proof-of-concept foundation to a new experimental framework for such studies. A bio-bank of primary fibroblast lines was generated from 323 out of 1,000 healthy individuals selected from the MI-study cohort. To study inter-donor variability of innate immune response in primary human dermal fibroblasts we chose to measure the TLR3 and TLR4 response pathways, both receptors being expressed and previously studied in fibroblasts. We established high-throughput automation compatible methods for standardized primary fibroblast cell activation, using purified MAMPS analogs, poly I:C and LPS that stimulate TLR3 and TLR4 pathways respectively. These results were in turn compared with a stimulation method using infection by HSV-1 virus. Our "Add-only" protocol minimizes high-throughput automation system variability facilitating whole process automation from cell plating through stimulation to recovery of cell supernatants, and fluorescent labeling. Images were acquired automatically by high-throughput acquisition on an automated high-content imaging microscope. Under these methodological conditions standardized image acquisition provided for quantification of cellular responses allowing biological variability to be measured with low system noise and high biological signal fidelity. Optimal for automated analysis of immuno-phenotype of primary human cell responses our method and experimental framework as reported here is highly compatible to high-throughput screening protocols like those necessary for chemo-genomic screening. In context of primary fibroblasts derived from donors enrolled to the MI-clinical-study our results open the way to assert the utility of studying immune-phenotype characteristics relevant to a human clinical cohort.