Arlene A. Stecenko

Introduction People with cystic fibrosis (PwCF) are at high risk for developing cystic fibrosis (CF)-related diabetes (CFRD), which worsens morbidity and mortality. Although the pathological events leading to the development of CFRD are complex and not completely understood, dietary factors may play a role. For example, habitual intake of dietary added sugar (i.e., sugar not naturally occurring in foods) has been shown to be increased in PwCF and this excess intake of added sugar could increase the risk of CFRD. Methods and analysis/design The goal of this ongoing double-blind, randomised, parallel-group clinical trial is to recruit approximately 60 clinically stable adults with CF to determine if a low-added sugar intervention improves beta-cell responsiveness and insulin sensitivity (Aim 1), reduces visceral adipose tissue (VAT) and other ectopic fat deposition (Aim 2) and improves plasma redox status (Aim 3) over 8 weeks compared with a typical CF diet. All foods will be provided. Participant selection criteria include confirmed CF diagnosis without CFRD, ≥18 years of age, and baseline estimated daily total added sugar intake >16 tsp. Eligible participants will be randomised to one of two arms: a low-added sugar diet (Ethics and dissemination The National Institutes of Health (NIH) funds this study (R01 DK133523). The study protocol was approved by the Emory Institutional Review Board (IRB approval number: 000004517). Any protocol modifications will be reviewed and approved by the IRB prior to implementation and communicated with the study team and participants, as relevant. We will provide reports of the findings to the NIH and Emory IRB in regular progress reports and post the findings on www.Clinicaltrials.gov. We will inform the findings of the study to the scientific community through presentations and peer-reviewed publications. Authorship for any resulting publications will follow the guidelines established by the International Committee of Medical Journal Editors. Trial registration number This research study is registered at www.Clinicaltrials.gov (NCT05766774).

Non-cystic fibrosis bronchiectasis (NCFB) is a disease characterized by abnormal dilatation of the airways, airflow obstruction, persistent cough, excessive sputum production and recurrent lung infections. NCFB exhibits clinical and pathological manifestations similar to key features of cystic fibrosis (CF) lung disease. In CF, pathogenesis results from dysfunction of the cystic fibrosis transmembrane conductance regulator (CFTR), and diagnosis is made by demonstrating elevated sweat chloride concentrations (typically ≥60 mEq/L), two CFTR mutations known to be causal, multi-organ tissue injury, or combination(s) of these findings. Based on a considerable body of evidence, we believe many patients with NCFB have disease likely to benefit from drugs such as elexacaftor/tezacaftor/ivacaftor (ETI) that activate CFTR-dependent ion transport. ETI is currently prescribed solely for treatment of CF, and has not been adequately tested or proposed for patients with NCFB, many of whom exhibit decreased CFTR function. Accordingly, we are conducting a clinical trial of ETI in subjects carrying a diagnosis of NCFB. Participants will exhibit one disease-causing CFTR mutation and/or sweat chloride measurements of 30-59 mEq/L. Cutaneous punch biopsy or blood samples will be obtained for iPS cell differentiation into airway epithelial monolayers – which will then be tested for response to ETI. Each patient will be given CFTR modulator treatment for approximately four weeks, with monitoring of clinical endpoints that include FEV ₁ , sweat chloride, quality of life questionnaire, and weight. The study will evaluate response of patients with NCFB to ETI, and test usefulness of iPSC-derived airway epithelial monolayers as a novel in vitro technology for predicting clinical benefit. FUNDING SOURCE: Marcus Foundation, Inc “Main Study” Summary: Objectives: ‘Lead in’ study: Incidence of diminished CFTR activity among patients with NCFB: In order to gain information regarding numbers of individuals with NCFB who may be eligible for our ‘main’ study, patients at Emory followed with bronchiectasis (who do not have clinical criteria sufficient for a diagnosis of cystic fibrosis) will be asked to consider participating in a ‘lead in’ study that will determine the subject’s CFTR genotype and sweat chloride level. A separate consent form will be utilized for the ‘lead in’ study. Subjects who exhibit a single CF-causing mutation in CFTR and/or sweat chloride 30-59 mEq/L will be approached about their interest in reviewing the consent form for the ‘main’ study (“iPSC derivation and in vivo ETI treatment for 4 weeks”; see following section). Up to 200 subjects will be included in the ‘lead-in’ study. ‘Main’ study: iPSC derivation and in vivo ETI treatment for 4 weeks: We propose a clinical trial of subjects with a diagnosis of NCFB. Participants will exhibit one disease-causing CFTR mutation and/or sweat chloride measurements of 30-59 mEq/L. Each patient will be given ETI for approximately four weeks. We will monitor clinical endpoints that include FEV ₁ , sweat chloride, quality of life questionnaire, and weight. We will also collect cutaneous punch biopsy material or blood samples from subjects so iPS cells can be differentiated into airway epithelial monolayers and tested for response to ETI.

Cystic fibrosis (CF) airway disease is characterized by chronic polymicrobial infections and an infiltration of neutrophils (PMNs). Staphylococcus aureus has been the most prevalent respiratory pathogen in CF. In particular, methicillin-resistant S. aureus (MRSA) represents a huge clinical burden in CF due to its association with lung disease and increased resistance to antibiotics. In CF, PMNs are unable to kill and clear MRSA. The reason for this remains largely unknown. Our study found that CF PMNs are as equally capable of killing MRSA as healthy PMNs. We show that the CF sputum, however, significantly impairs the ability of human PMNs to kill CF MRSA isolates. In the absence of CF sputum, PMNs kill MRSA via intracellular mechanisms mediated by phagocytosis, rather than extracellular mechanisms via NET formation. CF sputum does not affect the phagocytosis of MRSA via healthy or CF PMNs. Our results demonstrate that CF sputum exposure impairs phagosomal levels of reactive oxygen species (ROS) in MRSA-phagocytosing PMNs. While phagosomal co-localizations of MRSA with primary granule markers, myeloperoxidase and cathepsin D, were significantly reduced upon CF sputum exposure, that of a third azurophilic granule marker, neutrophil elastase, remained unaffected. This suggests that CF sputum does not compromise the fusion of primary granules with phagosomes but diminishes phagosomal ROS levels via another, likely more specific, mechanism. Overall, we identified the airway environment as an important factor that restricts neutrophils’ oxidative microbicidal activities in CF against MRSA. These results deliver new details of the complex host–pathogen interactions present in the CF lung.

Cystic fibrosis (CF) airway disease is characterized by chronic microbial infections and infiltration of inflammatory polymorphonuclear (PMN) granulocytes. Staphylococcus aureus (S. aureus) is a major lung pathogen in CF that persists despite the presence of PMNs and has been associated with CF lung function decline. While PMNs represent the main mechanism of the immune system to kill S. aureus, it remains largely unknown why PMNs fail to eliminate S. aureus in CF. The goal of this study was to observe how the CF airway environment affects S. aureus killing by PMNs. PMNs were isolated from the blood of healthy volunteers and CF patients. Clinical isolates of S. aureus were obtained from the airways of CF patients. The results show that PMNs from healthy volunteers were able to kill all CF isolates and laboratory strains of S. aureus tested in vitro. The extent of killing varied among strains. When PMNs were pretreated with supernatants of CF sputum, S. aureus killing was significantly inhibited suggesting that the CF airway environment compromises PMN antibacterial functions. CF blood PMNs were capable of killing S. aureus. Although bacterial killing was inhibited with CF sputum, PMN binding and phagocytosis of S. aureus was not diminished. The S. aureus-induced respiratory burst and neutrophil extracellular trap release from PMNs also remained uninhibited by CF sputum. In summary, our data demonstrate that the CF airway environment limits killing of S. aureus by PMNs and provides a new in vitro experimental model to study this phenomenon and its mechanism.

Although extracellular host DNA (ecDNA) levels in CF airways were linked to airflow obstruction and recombinant DNAse therapy is beneficial for CF patients, it remains incompletely understood whether ecDNA also leads to an autoimmune response. Here we hypothesized that chronic presence of DNA in CF airways triggers the production of autoantibodies targeting host human DNA. We measured the levels of IgA autoantibodies recognising host double-stranded (ds) DNA in the blood and sputum samples of CF patients and only sera of controls subjects and patients suffering from rheumatoid arthritis and systemic lupus erythematosus (SLE) that served as non-CF, autoimmune disease cohorts. We found that concentrations of anti-dsDNA IgA, but not IgG, autoantibodies in the circulation were significantly elevated in adult CF patients compared to age-matched, control subjects. Systemic levels of anti-dsDNA IgA antibodies negatively correlated with FEV₁% predicted, a measure of lung function, in CF patients. Anti-dsDNA IgA autoantibodies were also detected in CF sputa but sputum levels did not correlate with the degree of airway obstruction or sputum levels of DNA. We also found elevated autoantibody levels in CF children as 76.5% of CF patients younger than 10 years and 87.5% of CF patients 10–21 years had higher blood anti-dsDNA IgA levels than the highest value found in healthy control adults. Overall, our results detect elevated systemic anti-dsDNA IgA autoantibody levels in CF adults, teenagers and young children. We speculate that the appearance of an autoimmune response against host DNA in CF is an early event potentially contributing to disease pathogenesis.

Introduction: The acute respiratory distress syndrome (ARDS), affects up to 150,000 patients per year in the United States. We and other groups have demonstrated that bone marrow derived mesenchymal stromal stem cells prevent ARDS induced by systemic and local administration of endotoxin (lipopolysaccharide (LPS)) in mice. Methods: A study was undertaken to determine the effects of the diverse populations of bone marrow derived cells on the pathophysiology of ARDS, using a unique ex-vivo swine preparation, in which only the ventilated lung and the liver are perfused with autologous blood. Six experimental groups were designated as: 1) endotoxin alone, 2) endotoxin + total fresh whole bone marrow nuclear cells (BMC), 3) endotoxin + non-hematopoietic bone marrow cells (CD45 neg), 4) endotoxin + hematopoietic bone marrow cells (CD45 positive), 5) endotoxin + buffy coat and 6) endotoxin + in vitro expanded swine CD45 negative adherent allogeneic bone marrow cells (cultured CD45neg). We measured at different levels the biological consequences of the infusion of the different subsets of cells. The measured parameters were: pulmonary vascular resistance (PVR), gas exchange (PO₂), lung edema (lung wet/dry weight), gene expression and serum concentrations of the pro-inflammatory cytokines IL-1β, TNF-α and IL-6. Results: Infusion of freshly purified autologous total BMCs, as well as non-hematopoietic CD45(-) bone marrow cells significantly reduced endotoxin-induced pulmonary hypertension and hypoxemia and reduced the lung edema. Also, in the groups that received BMCs and cultured CD45neg we observed a decrease in the levels of IL-1β and TNF-α in plasma. Infusion of hematopoietic CD45(+) bone marrow cells or peripheral blood buffy coat cells did not protect against LPS-induced lung injury. Conclusions: We conclude that infusion of freshly isolated autologous whole bone marrow cells and the subset of non-hematopoietic cells can suppress the acute humoral and physiologic responses induced by endotoxemia by modulating the inflammatory response, mechanisms that do not involve engraftment or trans-differentiation of the cells. These observations may have important implications for the design of future cell therapies for ARDS.

The aim is to provide a detailed review of recent publications on cystic fibrosis-related diabetes (CFRD) with a particular focus on the interplay between cystic fibrosis (CF) lung disease and diabetes. CFRD is a form of diabetes that is distinct from type 1 or type 2 diabetes. CFRD remains very common and increases in prevalence with increasing age so that one in two middle-aged CF persons have CFRD. People with CFRD have lower lung function, worse nutrition, more frequent hospitalization, and worse mortality than CF people without diabetes. The excess mortality previously noted in women with CFRD compared with CF women without diabetes or CF men is much less apparent. CFRD is due to insulin deficiency and peripheral insulin resistance is much less a factor. Genetic susceptibility and oxidant stress are key risk factors for developing CFRD. The lung is the prime end organ target in CFRD and mortality is due to respiratory failure, not vascular complications. Insulin is the mainstay of therapy and early recognition and institution of therapy appear to improve health outcomes. CFRD remains one of the most important co-morbidities in CF. Early recognition of the disease and therapeutic intervention may diminish the negative impact that diabetes has on lung health in CF. Although a clearer understanding of the role of oxidant stress and genetics in the pathogenesis of CFRD is being elucidated, much needs to be learned before more targeted, specific therapies can be developed for this distinct form of diabetes.