J Bronchology Interv Pulmonol. 2022 May 27. doi: 10.1097/LBR.0000000000000874. Online ahead of print.

ABSTRACT

BACKGROUND: Currently, no consensus guidelines recommend routine bronchoscopy procedure in cystic fibrosis (CF), as no evidence is available concerning its use as either a diagnostic or therapeutic tool. Its efficacy is controversial, and no randomized controlled prospective trials are available to check its effectiveness. The aims of the present study were to evaluate the effectiveness of bronchoscopy as a diagnostic/therapeutic tool in CF children and adolescents; and to verify the effect of serial bronchoscopy on lung disease progression in subjects with CF not responding to a single procedure.

METHODS: Data of patients who received bronchoscopy at 2 Italian CF centers were collected. Bronchoalveolar lavage was performed during the procedure including airway clearance with mucolytics, inhaled antibiotics, and/or surfactant instillation.

RESULTS: A total of 16 patients in center 1 and 17 in center 2 underwent, respectively, 28 and 23 bronchoscopic procedure in the study period. Five patients in each center underwent >1 procedure. All procedures were generally well tolerated. No patient required admission to the pediatric intensive therapy unit. In 19.6% of bronchoalveolar lavages, growth of Aspergillus fumigatus was evident, although not detected by sputum analyses. After the procedure, an increase in mean percent predicted forced expiratory volume in the 1 second >10% was observed, and a significant decrease in pulmonary exacerbations yearly was evident.

CONCLUSION: Based on the results, we suggest bronchoscopy is not to be considered an obsolete tool, and it remains useful in CF management, although in selected cases. We encourage to support longitudinal observational studies to standardize the procedure, focusing on the choice of drugs to be instilled, modalities and timing of serial bronchoscopy and subsequent follow-up in selected severe clinical conditions.

PMID:35698279 | DOI:10.1097/LBR.0000000000000874

Commun Biol. 2022 Jun 13;5(1):580. doi: 10.1038/s42003-022-03448-z.

ABSTRACT

Reduced glomerular filtration rate (GFR) can progress to kidney failure. Risk factors include genetics and diabetes mellitus (DM), but little is known about their interaction. We conducted genome-wide association meta-analyses for estimated GFR based on serum creatinine (eGFR), separately for individuals with or without DM (nDM = 178,691, nnoDM = 1,296,113). Our genome-wide searches identified (i) seven eGFR loci with significant DM/noDM-difference, (ii) four additional novel loci with suggestive difference and (iii) 28 further novel loci (including CUBN) by allowing for potential difference. GWAS on eGFR among DM individuals identified 2 known and 27 potentially responsible loci for diabetic kidney disease. Gene prioritization highlighted 18 genes that may inform reno-protective drug development. We highlight the existence of DM-only and noDM-only effects, which can inform about the target group, if respective genes are advanced as drug targets. Largely shared effects suggest that most drug interventions to alter eGFR should be effective in DM and noDM.

PMID:35697829 | PMC:PMC9192715 | DOI:10.1038/s42003-022-03448-z

Orphanet J Rare Dis. 2022 Jun 13;17(1):222. doi: 10.1186/s13023-022-02373-y.

ABSTRACT

BACKGROUND: Cystic fibrosis (CF) has a vast and heterogeneous mutational spectrum in Europe. This variability has also been described in Spain, and there are numerous studies linking CFTR variants with the symptoms of the disease. Most of the studies analysed determinate clinical manifestations or specific sequence variants in patients from clinical units. Others used registry data without addressing the genotype-phenotype relationship. Therefore, the objective of this study is to describe the genetic and clinical characteristics of people with CF and to analyse the relationship between both using data from the rare disease registry of a region in southeastern Spain.

METHODS: A cross-sectional study was carried out in people with a confirmed diagnosis of CF registered in the Rare Diseases Information System (SIER) of the Region of Murcia (Spain). The patients were classified into two genotypes according to the functional consequence that the genetic variants had on the CFTR protein.

RESULTS: There were 192 people diagnosed with CF reported in the Region of Murcia as of 31 December 2018. Seventy-six genotypes and 49 different variants were described, with c.1521_1523delCTT (p. Phe508del) being the most common in 58.3% of the CF patients and 37.0% of the alleles. In addition, 67% of the patients were classified as a high-risk genotype, which was associated with a lower percentage of FEV1 (OR: 5.3; 95% CI: 1.2, 24.4), an increased risk of colonization by Pseudomonas aeruginosa (OR: 7.5; 95% CI: 1.7, 33.0) and the presence of pancreatic insufficiency (OR: 28.1; 95% CI: 9.3, 84.4) compared to those with a low-risk genotype.

CONCLUSIONS: This is the first study in Spain that describes the mutational spectrum and its association with clinical manifestations in patients with CF using data from a rare disease registry. The results obtained allow planning for the health resources needed by people with this disease, thus contributing to the development of personalized medicine that helps to optimize health care in CF patients.

PMID:35698092 | DOI:10.1186/s13023-022-02373-y

Eur J Intern Med. 2022 Jun 8:S0953-6205(22)00217-5. doi: 10.1016/j.ejim.2022.06.005. Online ahead of print.

ABSTRACT

BACKGROUND: The coronavirus disease 2019 (COVID-19) presents an urgent threat to global health. Prediction models that accurately estimate mortality risk in hospitalized patients could assist medical staff in treatment and allocating limited resources.

AIMS: To externally validate two promising previously published risk scores that predict in-hospital mortality among hospitalized COVID-19 patients.

METHODS: Two prospective cohorts were available; a cohort of 1028 patients admitted to one of nine hospitals in Lombardy, Italy (the Lombardy cohort) and a cohort of 432 patients admitted to a hospital in Leiden, the Netherlands (the Leiden cohort). The endpoint was in-hospital mortality. All patients were adult and tested COVID-19 PCR-positive. Model discrimination and calibration were assessed.

RESULTS: The C-statistic of the 4C mortality score was good in the Lombardy cohort (0.85, 95CI: 0.82-0.89) and in the Leiden cohort (0.87, 95CI: 0.80-0.94). Model calibration was acceptable in the Lombardy cohort but poor in the Leiden cohort due to the model systematically overpredicting the mortality risk for all patients. The C-statistic of the CURB-65 score was good in the Lombardy cohort (0.80, 95CI: 0.75-0.85) and in the Leiden cohort (0.82, 95CI: 0.76-0.88). The mortality rate in the CURB-65 development cohort was much lower than the mortality rate in the Lombardy cohort. A similar but less pronounced trend was found for patients in the Leiden cohort.

CONCLUSION: Although performances did not differ greatly, the 4C mortality score showed the best performance. However, because of quickly changing circumstances, model recalibration may be necessary before using the 4C mortality score.

PMID:35697562 | DOI:10.1016/j.ejim.2022.06.005

Crit Rev Microbiol. 2022 Jun 13:1-16. doi: 10.1080/1040841X.2022.2082268. Online ahead of print.

ABSTRACT

Mycobacterium abscessus is a nontuberculous mycobacterium, associated with broncho-pulmonary infections in individuals suffering from cystic fibrosis, bronchiectasis, and pulmonary diseases. The risk factors for transmission include biofilms, contaminated water resources, fomites, and infected individuals. M. abscessus is extensively resistant to antibiotics. To date, there is no vaccine and combination antibiotic therapy is followed. However, drug toxicities, low cure rates, and high cost of treatment make it imperfect. Over the last 20 years, bioinformatic studies on M. abscessus have advanced our understanding of the pathogen. This review integrates knowledge from the analysis of genomes, microbiomes, genomic variations, phylogeny, proteome, transcriptome, secretome, antibiotic resistance, and vaccine design to further our understanding. The utility of genome-based studies in comprehending disease progression, surveillance, tracing transmission routes, and epidemiological outbreaks on a global scale has been highlighted. Furthermore, this review underlined the importance of using computational methodologies for pinpointing factors responsible for pathogen survival and resistance. We reiterate the significance of interdisciplinary research to fight M. abscessus. In a nutshell, the outcome of computational studies can go a long way in creating novel therapeutic avenues to control M. abscessus mediated pulmonary infections.

PMID:35696783 | DOI:10.1080/1040841X.2022.2082268

Pancreas. 2022 Jun 11. doi: 10.1097/MPA.0000000000002016. Online ahead of print.

ABSTRACT

OBJECTIVES: We sought data on the validity, reliability, responsiveness, and feasibility of the coefficient of fat absorption (CFA) as a measure of pancreatic enzyme replacement therapy (PERT) efficacy in people with cystic fibrosis (pwCF) and reviewed the literature for alternative measures.

METHODS: We searched PubMed for the Medical Subject Heading cystic fibrosis and the key words cystic fibrosis, fat absorption, CFA, and fecal fat imbalance; historical articles; and citations in bibliographies.

RESULTS: The lower the CFA, the greater its variability; thus, it is less variable in healthy individuals who have higher CFA than pwCF. In addition, the test-retest values for CFA are more variable in pwCF than the general population. There is no correlation between CFA and body mass index or PERT dose but CFA is related to gastrointestinal signs and symptoms. Research-quality CFA studies are expensive, time consuming, and odious to pwCF and research staff. Sparse stool tests, breath tests, and blood tests of fat absorption have been studied as potential alternatives to CFA to measure PERT efficacy.

CONCLUSIONS: Based on the evidence, we conclude that CFA as a measure of the efficacy of PERT is more of a "coal standard" than a gold standard; developing suitable alternatives should be a priority.

PMID:35695742 | DOI:10.1097/MPA.0000000000002016

mBio. 2022 Jun 13:e0019122. doi: 10.1128/mbio.00191-22. Online ahead of print.

ABSTRACT

Bacteria change phenotypically in response to their environment. Free swimming cells transition to biofilm communities that promote cellular cooperativity and resistance to stressors and antibiotics. We uncovered three subpopulations of cells with diverse phenotypes from a single-species Pseudomonas aeruginosa PA14 biofilm, and used a series of steps to isolate, characterize, and map these cell subpopulations in a biofilm. The subpopulations were distinguishable by size and morphology using dynamic light scattering (DLS) and scanning electron microscopy (SEM). Additionally, growth and dispersal of biofilms originating from each cell subpopulation exhibited contrasting responses to antibiotic challenge. Cell subpopulation surface charges were distinctly different, which led us to examine the ionizable surface molecules associated with each subpopulation using mass spectrometry. Matrix assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry analysis of cell subpopulations revealed ions unique to each subpopulation of cells that significantly co-localized with ions associated with quorum sensing. Transcript levels of algR, lasR, and rhlI in subpopulations isolated from biofilms differed from levels in planktonic stationary and mid-log cell subpopulations. These studies provide insight into diverse phenotypes, morphologies, and biochemistries of PA14 cell subpopulations for potential applications in combating bacterial pathogenesis, with medical, industrial, and environmental complications. IMPORTANCE Pseudomonas aeruginosa biofilms can cause chronic infections in burn wounds, grow on medical equipment, and proliferate in the lungs of people with cystic fibrosis. These inherently antibiotic tolerant biofilms are difficult to eradicate largely due to the complexity of the biofilm environment. Developing more effective biofilm treatments is reliant upon understanding biofilm heterogeneity. We identified and characterized three separate cell subpopulations found in P. aeruginosa PA14 biofilms. The distinct morphologies, phenotypes, and biochemistries of each of these cell subpopulations indicate that they contribute differently to the overall biofilm environment. These findings demonstrate that bacterial cells of the same species exhibit diversity that implies distinct roles in biofilm initiation, maturation, and maintenance.

PMID:35695457 | DOI:10.1128/mbio.00191-22

Oncoimmunology. 2022 May 31;11(1):2081415. doi: 10.1080/2162402X.2022.2081415. eCollection 2022.

ABSTRACT

Natural Killer (NK) cells are known for their high intrinsic cytotoxic capacity, and the possibility to be applied as 'off-the-shelf' product makes them highly attractive for cell-based immunotherapies. In patients with multiple myeloma (MM), an elevated number of NK cells has been correlated with higher overall-survival rate. However, NK cell function can be impaired by upregulation of inhibitory receptors, such as the immune checkpoint NKG2A. Here, we developed a CRISPR-Cas9-based gene editing protocol that allowed us to knockout about 80% of the NKG2A-encoding killer cell lectin like receptor C1 (KLRC1) locus in primary NK cells. In-depth phenotypic analysis confirmed significant reduction in NKG2A protein expression. Importantly, the KLRC1-edited NK cells showed significantly increased cytotoxicity against primary MM cells isolated from a small cohort of patients, and maintained the NK cell-specific cytokine production. In conclusion, KLRC1-editing in primary NK cells has the prospect of overcoming immune checkpoint inhibition in clinical applications.

PMID:35694192 | PMC:PMC9176243 | DOI:10.1080/2162402X.2022.2081415

Curr Protein Pept Sci. 2022 Jun 10. doi: 10.2174/1389203723666220610124303. Online ahead of print.

ABSTRACT

The pathogenesis of SARS-CoV-2 infection is related to the direct cytopathic effect and associated hyper-inflammatory due to exaggerated immune response. Different experimental and clinical studies revealed that other biomarkers could be used to determine the Covid-19 severity, such as D-dimer, procalcitonin, C-reaction protein (CRP), IL-6, and ferritin. Calprotectin (CP) is associated with intestinal inflammation, intestinal injury, and different respiratory diseases such as cystic fibrosis. Thus, CP might be a possible biomarker linking intestinal injury and acute lung injury (ALI) in Covid-19. Therefore, this study aimed to find a potential role of CP regarding GITI and ALI in Covid-19. CP is a complex protein consisting of S100A8 and S100A9, belongs to the Ca+2-binding proteins S100 family abundant in the cytosol of neutrophils and expressed on the monocyte membranes, macrophages, and intestinal epithelial cells. CP is a proinflammatory protein that acts through activation of the receptor for the advanced glycation end product (RAGE) and toll-like receptor 4 (TLR4). CP is a biomarker of neutrophil activation and is released following the turnover of neutrophils. CP could be controversial; it increases airway inflammation or protects lung and airway epithelium from an exaggerated immune response. Therefore, a high level of CP in different respiratory disorders might be protective and compensate against abnormal immune responses. CP level is high in Covid-19 and correlated with Covid-19 severity and oxygen demand due to activation release of proinflammatory cytokines and inflammatory signaling pathways. Therefore, CP level is elevated in both ALI and intestinal inflammation so that it could be a potential biomarker link the respiratory and intestinal injury in Covid-19.

PMID:35692161 | DOI:10.2174/1389203723666220610124303

J Cyst Fibros. 2022 Jun 9:S1569-1993(22)00151-5. doi: 10.1016/j.jcf.2022.05.013. Online ahead of print.

ABSTRACT

BACKGROUND: In cystic fibrosis (CF), genotype-phenotype correlation is complicated by the large number of CFTR variants, the influence of modifier genes, environmental effects, and the existence of complex alleles. We document the importance of complex alleles, in particular the F508C variant present in cis with the S1251N disease-causing variant, by detailed analysis of a patient with CF, with the [S1251N;F508]/G542X genotype and a very mild phenotype, contrasting it to that of four subjects with the [S1251N;F508C]/F508del genotype and classical CF presentation.

METHODS: Genetic differences were identified by Sanger sequencing and CFTR function was quantified using rectal organoids in rectal organoid morphology analysis (ROMA) and forskolin-induced swelling (FIS) assays. CFTR variants were further characterised in CF bronchial epithelial (CFBE) cell lines. The impact of involved amino acid changes in the CFTR 3D protein structure was evaluated.

RESULTS: Organoids of the patient [S1251N;F508] with mild CF phenotype confirmed the CF diagnosis but showed higher residual CFTR function compared to the four others [S1251N;F508C]. CFBE cell lines showed a decrease in [S1251N;F508C]-CFTR function but not in processing when compared to [S1251N;F508]-CFTR. Analysis of the 3D CFTR structure suggested an additive deleterious effect of the combined presence of S1251N and F508C with respect to NBD1-2 dimerisation.

CONCLUSIONS: In vitro and in silico data show that the presence of F508C in cis with S1251N decreases CFTR function without affecting processing. Complex CFTR alleles play a role in clinical phenotype and their identification is relevant in the context of personalised medicine for each patient with CF.

PMID:35690578 | DOI:10.1016/j.jcf.2022.05.013

J Heart Lung Transplant. 2022 May 20:S1053-2498(22)01947-7. doi: 10.1016/j.healun.2022.05.008. Online ahead of print.

ABSTRACT

BACKGROUND: Improved predictive models are needed in lung transplantation in the setting of a proposed allocation system that incorporates longer-term post-transplant survival in the United States. Allocation systems require accurate mortality predictions to justly allocate organs.

METHODS: Utilizing the United Network for Organ Sharing database (2005-2017), we fit models to predict 1-year mortality based on the Lung Allocation Score (LAS), the Chan, et al, 2019 model, a novel "clinician" model (a priori clinician selection of pre-transplant covariates), and two machine learning models (Least Absolute Shrinkage and Selection Operator; LASSO and Random Forests) for predicting 1-year and 3-year post-transplant mortality. We compared predictive accuracy among models. We evaluated the calibration of models by comparing average predicted probability vs observed outcome per decile. We repeated analyses fit for 3-year mortality, disease category, including donor covariates, and LAS era.

RESULTS: The area under the cure for all models was low, ranging from 0.55 to 0.62. All exhibited reasonable negative predictive values (0.87-0.90), but the positive predictive value for was poor (all <0.25). Evaluating LAS calibration found 1-year post-transplant estimates consistently overestimated risk of mortality, with greater differences in higher deciles. LASSO, Random Forests, and clinician models showed no improvement when evaluated by disease category or with the addition of donor covariates and performed worse for 3-year outcomes.

CONCLUSIONS: The LAS overestimated patients' risk of post-transplant death, thus underestimating transplant benefit in the sickest candidates. Novel models based on pre-transplant recipient covariates failed to improve prediction. There should be wariness in post-transplant survival predictions from available models.

PMID:35690561 | DOI:10.1016/j.healun.2022.05.008

Curr Opin Pharmacol. 2022 Jun 8;65:102248. doi: 10.1016/j.coph.2022.102248. Online ahead of print.

ABSTRACT

In the lungs, defective CFTR associated with cystic fibrosis (CF) represents the nidus for abnormal mucus clearance in the airways and consequently a progressive lung disease. Defective CFTR-mediated Cl- secretion results in altered mucus properties, including concentration, viscoelasticity, and the ratio of the two mucins, MUC5B and MUC5AC. In the past decades, therapies targeting the CF mucus defect, directly or indirectly, have been developed; nevertheless, better treatments to prevent the disease progression are still needed. This review summarizes the existing knowledge on the defective mucus in CF disease and highlights it as a barrier to the development of future inhaled genetic therapies. The use of new mucus-targeting treatments is also discussed, focusing on their potential role to halt the progress of CF lung disease.

PMID:35689870 | DOI:10.1016/j.coph.2022.102248

Pharmacotherapy. 2022 Jun 11. doi: 10.1002/phar.2710. Online ahead of print.

ABSTRACT

Elexacaftor/tezacaftor/ivacaftor (ELX/TEZ/IVA) is a highly effective therapy for patients with cystic fibrosis (CF) with potential benefits in lung transplant recipients (LTRs) for extrapulmonary CF manifestations, however tolerability and efficacy in this population is largely unknown. We report our experience with ELX/TEZ/IVA in LTRs for extrapulmonary complications of CF including tolerability, drug-drug interactions, and therapeutic benefit. All LTRs at a single center initiated on ELX/TEZ/IVA were reviewed. Adverse events and patient-reported outcomes attributed to ELX/TEZ/IVA were documented. Pulmonary function, tacrolimus requirements in mg/kg/dL, body mass index (BMI), and reason for initiation were assessed at the initiation of ELX/TEZ/IVA, and at 12 months post-initiation or at the time of discontinuation for those in whom therapy was discontinued. Thirteen LTRs were initiated on ELX/TEZ/IVA at a mean of 115 ± 92 months post-transplant. All were initiated on ELX/TEZ/IVA for sinus or sinus and gastrointestinal CF manifestations. Five (38.4%) patients discontinued therapy due to declining pulmonary function (2/5, 40%), mood disturbances (2/5, 40%), or lack of benefit (1/5, 20%). Of the eight patients who remain on ELX/TEZ/IVA, four reported adverse effects and three LTRs temporarily held therapy. Six (46.2%) LTRs reported improvement in sinus symptoms, while four (30.7%) reported improved gastrointestinal symptoms. Weight declined in the cohort overall. Tacrolimus dose requirements decreased following initiation of ELX/TEZ/IVA therapy, with a 50% decline in dose requirements observed. In our experience, ELX/TEZ/IVA in LTRs is poorly tolerated with modest perceived extrapulmonary benefit and a significant effect on tacrolimus dose requirements. More data is needed to determine the benefits of ELX/TEZ/IVA therapy in LTRs.

PMID:35689451 | DOI:10.1002/phar.2710

Bioorg Med Chem Lett. 2022 Jun 7:128843. doi: 10.1016/j.bmcl.2022.128843. Online ahead of print.

ABSTRACT

Cystic fibrosis (CF) is an autosomal recessive disease resulting from mutations on both copies of the CFTR gene. Phenylalanine deletion at position 508 of the CFTR protein (F508del-CFTR) is the most frequent mutation in CF patients. Currently, the most effective treatments of CF use a dual or triple combination of CFTR correctors and potentiators. In triple therapy, two correctors (C1 and C2) and a potentiator are employed. Herein, we describe the identification and exploration of the SAR of a series of 4-aminopyrrolidine-2-carboxylic acid C2 correctors of CFTR to be used in conjunction with our existing C1 corrector series for the treatment of CF.

PMID:35688367 | DOI:10.1016/j.bmcl.2022.128843

Lett Appl Microbiol. 2022 Jun 10. doi: 10.1111/lam.13759. Online ahead of print.

ABSTRACT

Pseudomonas aeruginosa is one of the most worrisome infectious bacteria due to its intrinsic and acquired resistance against several antibiotics and the recalcitrance of its infections; hence, the development of novel antimicrobials able to inhibit it is mandatory. In this work, silver nanoparticles obtained by green synthesis using a leaf extract and fungi were tested against a battery of clinical strains from cystic fibrosis, pneumonia, and burnt patients, some of them with multidrug resistance. Both nanoparticles showed a potent antibacterial effect, causing severe damage to the cell wall, membrane, and DNA, and inducing the production of reactive oxygen species. Moreover, the nanoparticles derived from fungi showed synergistic antibacterial effects with the antibiotics meropenem and levofloxacin for some clinical strains and both kinds of nanoparticles were nontoxic for larvae of the moth Galleria mellonella, encouraging further research for their implementation in the treatment of P. aeruginosa infections.

PMID:35687297 | DOI:10.1111/lam.13759

Hum Gene Ther. 2022 Jun 10. doi: 10.1089/hum.2022.036. Online ahead of print.

ABSTRACT

Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene cause cystic fibrosis (CF), a chronic disease that affects multiple organs including the lung. We developed a CF ferret model of a scarless G551→D substitution in CFTR (CFTRG551D-KI), enabling approaches to correct this gating mutation in CF airways via gene editing. Homology-directed repair (HDR) was tested in Cas9-expressing CF airway basal cells (Cas9-GKI) from this model, as well as reporter basal cells (Y66S-Cas9-GKI) that express an integrated non-fluorescent Y66S-EGFP mutant gene to facilitate rapid assessment of HDR by the restoration of fluorescence. rAAV vectors were used to deliver two DNA templates and sgRNAs for dual gene editing at the EGFP and CFTR genes, followed by fluorescence activated cell sorting (FACS) of EGFPY66S-corrected cells. When gene-edited airway basal cells were polarized at an air-liquid interface, unsorted and EGFPY66S-corrected sorted populations gave rise to 26.0% and 70.4% CFTR-mediated Cl- transport of that observed in non-CF cultures, respectively. The consequences of gene editing at the CFTRG551D locus by HDR and non-homology end joining (NHEJ) were assessed by targeted gene next generation sequencing (NGS) against a specific amplicon. NGS revealed HDR corrections of 3.1% of G551 sequences in the unsorted population of rAAV-infected cells, and 18.4% in the EGFPY66S-corrected cells. However, the largest proportion of sequences had indels surrounding the CRISPR cut site, demonstrating that NHEJ was the dominant repair pathway. This approach to simultaneously co-edit at two genomic loci using rAAV may have utility as a model system for optimizing gene editing efficiencies in proliferating airway basal cells through the modulation of DNA repair pathways in favor of HDR.

PMID:35686451 | DOI:10.1089/hum.2022.036

Comput Struct Biotechnol J. 2022 May 23;20:2587-2599. doi: 10.1016/j.csbj.2022.05.036. eCollection 2022.

ABSTRACT

Cystic fibrosis (CF) is a frequent genetic disease in Caucasians that is caused by the deletion of F508 (ΔF508) in the nucleotide binding domain 1 (NBD1) of the CF transmembrane conductance regulator (CFTR). The ΔF508 compromises the folding energetics of the NBD1, as well as the folding of three other CFTR domains. Combination of FDA approved corrector molecules can efficiently but incompletely rescue the ΔF508-CFTR folding and stability defect. Thus, new pharmacophores that would reinstate the wild-type-like conformational stability of the ΔF508-NBD1 would be highly beneficial. The most prominent molecule, 5-bromoindole-3-acetic acid (BIA) that can thermally stabilize the NBD1 has low potency and efficacy. To gain insights into the NBD1 (un)folding dynamics and BIA binding site localization, we combined molecular dynamics (MD) simulations, atomic force spectroscopy (AFM) and hydrogen-deuterium exchange (HDX) experiments. We found that the NBD1 α-subdomain with three adjacent strands from the β-subdomain plays an important role in early folding steps, when crucial non-native interactions are formed via residue F508. Our AFM and HDX experiments showed that BIA associates with this α-core region and increases the resistance of the ΔF508-NBD1 against mechanical unfolding, a phenomenon that could be exploited in future developments of folding correctors.

PMID:35685375 | PMC:PMC9160490 | DOI:10.1016/j.csbj.2022.05.036

Front Physiol. 2022 May 24;13:882525. doi: 10.3389/fphys.2022.882525. eCollection 2022.

ABSTRACT

Fluid and anion secretion are important functions of the biliary tract. It has been established that cAMP regulates Na+ absorption through NHE3. However, mechanisms of gallbladder anion transport are less defined. We created organoids and organoid-derived monolayers from human gallbladder tissue to measure organoid swelling and transepithelial electrophysiology. In our in vitro models, forskolin-stimulation caused organoid swelling and increased transepithelial anion transport. Full organoid swelling required Cl-while changes in short-circuit current were HCO3 --dependent. Organoids and monolayers from an individual homozygous for the cystic fibrosis-causing ΔF508 CFTR mutation had no apical expression of CFTR and minimal changes in transepithelial current and conductance with forskolin treatment. However, organoid swelling remained intact. Dilution potential studies revealed that forskolin treatment increased the paracellular permeability to anions relative to cations. These data suggest a novel paracellular contribution to forskolin-stimulated fluid transport across the gallbladder epithelium.

PMID:35685290 | PMC:PMC9171199 | DOI:10.3389/fphys.2022.882525

J Clin Med. 2022 Jun 2;11(11):3189. doi: 10.3390/jcm11113189.

ABSTRACT

BACKGROUND: Assessment of the effect of subgingival instrumentation (SI) on systemic inflammation in periodontitis grades B (BP) and C (CP).

METHODS: In this prospective cohort study, eight BP and 46 CP patients received SI. Data were collected prior to and 12 weeks after SI. Blood was sampled prior to, one day, 6, and 12 weeks after SI. Neutrophil elastase (NE), C-reactive protein (CRP), leukocyte count, lipopolysaccharide binding protein, interleukin 6 (IL-6) and IL-8 were assessed.

RESULTS: Both groups showed significant clinical improvement. NE was lower in BP than CP at baseline and 1 day after SI, while CRP was lower in BP than CP at baseline (p &lt; 0.05). NE and CRP had a peak 1 day after SI (p &lt; 0.05). Between-subjects effects due to CP (p = 0.042) and PISA (p = 0.005) occurred. Within-subjects NE change was confirmed and modulated by grade (p = 0.017), smoking (p = 0.029), number of teeth (p = 0.033), and PISA (p = 0.002). For CRP between-subjects effects due to BMI (p = 0.008) were seen. Within-subjects PISA modulated the change of CRP over time (p = 0.017).

CONCLUSIONS: In untreated CP, NE and CRP were higher than in BP. SI results in better PPD and PISA reduction in BP than CP.

TRIAL REGISTRATION: Deutsches Register Klinischer Studien DRKS00026952 28 October 2021 registered retrospectively.

PMID:35683571 | DOI:10.3390/jcm11113189

J Clin Med. 2022 May 29;11(11):3066. doi: 10.3390/jcm11113066.

ABSTRACT

The acceptable duration of donor warm ischemia time (DWIT) after cardiocirculatory death (DCD) is still debated. We analyzed the biomolecular profile and function during ex vivo lung perfusion (EVLP) of DCD lungs and their correlation with lung transplantation (LuTx) outcomes. Donor data, procurement times, recipient outcomes, and graft function up to 1 year after LuTx were collected. During EVLP, the parameters of graft function and metabolism, perfusate samples to quantify inflammation, glycocalyx breakdown products, coagulation, and endothelial activation markers were obtained. Data were compared to a cohort of extended-criteria donors after brain death (EC-DBD). Eight DBD and seven DCD grafts transplanted after EVLP were analyzed. DCD's DWIT was 201 [188;247] minutes. Donors differed only regarding the duration of mechanical ventilation that was longer in the EC-DBD group. No difference was observed in lung graft function during EVLP. At reperfusion, "wash-out" of inflammatory cells and microthrombi was predominant in DCD grafts. Perfusate biomolecular profile demonstrated marked endothelial activation, characterized by the presence of inflammatory mediators and glycocalyx breakdown products both in DCD and EC-DBD grafts. Early graft function after LuTx was similar between DCD and EC-DBD. DCD lungs exposed to prolonged DWIT represent a potential resource for donation if properly preserved and evaluated.

PMID:35683455 | DOI:10.3390/jcm11113066