Neutrophil Elastase Enhances Sputum Solubilization in Cystic Fibrosis Patients Receiving DNase Therapy

Cystic fibrosis patients suffer from chronic lung infection and inflammation due to the secretion of viscous sputum. Sputum viscosity is caused by extracellular DNA, some of which originates from the release of neutrophil extracellular traps (NETs). During NET formation neutrophil elastase (NE) partially processes histones to decondense chromatin. NE is abundant in CF sputum and is thought to contribute to tissue damage. Exogenous nucleases are a palliative treatment in CF as they promote sputum solubilization. We show that in a process reminiscent of NET formation, NE enhances sputum solubilization by cleaving histones to enhance the access of exogenous nucleases to DNA. In addition, we find that in Cf sputum NE is predominantly bound to DNA, which is known to downregulate its proteolytic activity and may restrict host tissue damage. The beneficial role of NE in CF sputum solubilization may have important implications for the development of CF therapies targeting NE

Activation of the Aryl Hydrocarbon Receptor Interferes with Early Embryonic Development.

The transcriptional program of early embryonic development is tightly regulated by a set of well-defined transcription factors that suppress premature expression of differentiation genes and sustain the pluripotent identity. It is generally accepted that this program can be perturbed by environmental factors such as chemical pollutants; however, the precise molecular mechanisms remain unknown. The aryl hydrocarbon receptor (AHR) is a widely expressed nuclear receptor that senses environmental stimuli and modulates target gene expression. Here, we have investigated the AHR interactome in embryonic stem cells by mass spectrometry and show that ectopic activation of AHR during early differentiation disrupts the differentiation program via the chromatin remodeling complex NuRD (nucleosome remodeling and deacetylation). The activated AHR/NuRD complex altered the expression of differentiation-specific genes that control the first two developmental decisions without affecting the pluripotency program. These findings identify a mechanism that allows environmental stimuli to disrupt embryonic development through AHR signaling

Liposome Co-sedimentation and Co-flotation Assays to Study Lipid-Protein Interactions

A large proportion of proteins are expected to interact with cellular membranes to carry out their physiological functions in processes such as membrane transport, morphogenesis, cytoskeletal organization, and signal transduction. The recruitment of proteins at the membrane-cytoplasm interface and their activities are precisely regulated by phosphoinositides, which are negatively charged phospholipids found on the cytoplasmic leaflet of cellular membranes and play critical roles in membrane homeostasis and cellular signaling. Thus, it is important to reveal which proteins interact with phosphoinositides and to elucidate the underlying mechanisms. Here, we present two standard in vitro methods, liposome co-sedimentation and co-flotation assays, to study lipid-protein interactions. Liposomes can mimic various biological membranes in these assays because their lipid compositions and concentrations can be varied. Thus, in addition to mechanisms of lipid-protein interactions, these methods provide information on the possible specificities of proteins toward certain lipids such as specific phosphoinositide species and can hence shed light on the roles of membrane interactions on the functions of membrane-associated proteins.Peer reviewe

Human Erythroid Progenitors Are Directly Infected by SARS-CoV-2: Implications for Emerging Erythropoiesis in Severe COVID-19 Patients

We document here that intensive care COVID-19 patients suffer a profound decline in hemoglobin levels but show an increase of circulating nucleated red cells, suggesting that SARS-CoV-2 infection either directly or indirectly induces stress erythropoiesis. We show that ACE2 expression peaks during erythropoiesis and renders erythroid progenitors vulnerable to infection by SARS-CoV-2. Early erythroid progenitors, defined as CD34-CD117+CD71+CD235a-, show the highest levels of ACE2 and constitute the primary target cell to be infected during erythropoiesis. SARS-CoV-2 causes the expansion of colony formation by erythroid progenitors and can be detected in these cells after 2 weeks of the initial infection. Our findings constitute the first report of SARS-CoV-2 infectivity in erythroid progenitor cells and can contribute to understanding both the clinical symptoms of severe COVID-19 patients and how the virus can spread through the circulation to produce local inflammation in tissues, including the bone marrow

Anti-inflammatory therapy with nebulised dornase alfa in patients with severe COVID-19 pneumonia A Randomised Clinical Trial

BACKGROUND: SARS-CoV2 infection causes severe, life-threatening pneumonia. Hyper-inflammation, coagulopathy and lymphopenia are associated with pathology and poor outcomes in these patients. Cell-free (cf) DNA is prominent in COVID-19 patients, amplifies inflammation and promotes coagulopathy and immune dysfunction. We hypothesized that cf-DNA clearance by nebulised dornase alfa may reduce inflammation and improve disease outcomes. Here, we evaluated the efficacy of nebulized dornase alfa in patients hospitalised with severe COVID-19 pneumonia. METHODS: In this randomised controlled single-centre phase 2 proof-of-concept trial, we recruited adult patients admitted to hospital that exhibited stable oxygen saturation (≥94%) on supplementary oxygen and a C-reactive protein (CRP) level ≥30mg/L post dexamethasone treatment. Participants were randomized at a 3:1 ratio to receive twice-daily nebulised dornase alfa in addition to best available care (BAC) or BAC alone for seven days or until hospital discharge. A 2:1 ratio of historical controls to treated individuals (HC, 2:1) were included as the primary endpoint comparators. The primary outcome was a reduction in systemic inflammation measured by blood CRP levels over 7 days post-randomisation, or to discharge if sooner. Secondary and exploratory outcomes included time to discharge, time on oxygen, D-dimer levels, lymphocyte counts and levels of circulating cf-DNA. RESULTS: We screened 75 patients and enrolled 39 participants out of which 30 in dornase alfa arm, and 9 in BAC group. We also matched the recruited patients in the treated group (N=30) to historical controls in the BAC group (N=60). For the the primary outcome, 30 patients in the dornase alfa were compared to 69 patients in the BAC group. Dornase alfa treatment reduced CRP by 33% compared to the BAC group at 7-days (P=0.01). The dornase alfa group least squares mean CRP was 23.23 mg/L (95% CI 17.71 to 30.46) and the BAC group 34.82 mg/L (95% CI 28.55 to 42.47). A significant difference was also observed when only randomised participants were compared. Furthermore, compared to the BAC group, the chance of live discharge was increased by 63% in the dornase alfa group (HR 1.63, 95% CI 1.01 to 2.61, P=0.03), lymphocyte counts were improved (least-square mean: 1.08 vs 0.87, P=0.02) and markers of coagulopathy such as D-dimer were diminished (least-square mean: 570.78 vs 1656.96μg/mL, P=0.004). Moreover, the dornase alfa group exhibited lower circulating cf-DNA levels that correlated with CRP changes over the course of treatment. No differences were recorded in the rates and length of stay in the ICU or the time on oxygen between the groups. Dornase alfa was well-tolerated with no serious adverse events reported. CONCLUSION: In this proof-of-concept study in patients with severe COVID-19 pneumonia, treatment with nebulised dornase alfa resulted in a significant reduction in inflammation, markers of immune pathology and time to discharge. The effectiveness of dornase alfa in patients with acute respiratory infection and inflammation should be investigated further in larger trials

Loss of TET2 in human hematopoietic stem cells alters the development and function of neutrophils

Somatic mutations commonly occur in hematopoietic stem cells (HSCs). Some mutant clones outgrow through clonal hematopoiesis (CH) and produce mutated immune progenies shaping host immunity. Individuals with CH are asymptomatic but have an increased risk of developing leukemia, cardiovascular and pulmonary inflammatory diseases, and severe infections. Using genetic engineering of human HSCs (hHSCs) and transplantation in immunodeficient mice, we describe how a commonly mutated gene in CH, TET2, affects human neutrophil development and function. TET2 loss in hHSCs produce a distinct neutrophil heterogeneity in bone marrow and peripheral tissues by increasing the repopulating capacity of neutrophil progenitors and giving rise to low-granule neutrophils. Human neutrophils that inherited TET2 mutations mount exacerbated inflammatory responses and have more condensed chromatin, which correlates with compact neutrophil extracellular trap (NET) production. We expose here physiological abnormalities that may inform future strategies to detect TET2-CH and prevent NET-mediated pathologies associated with CH

Dynamics of Barred Galaxies

Some 30% of disc galaxies have a pronounced central bar feature in the disc plane and many more have weaker features of a similar kind. Kinematic data indicate that the bar constitutes a major non-axisymmetric component of the mass distribution and that the bar pattern tumbles rapidly about the axis normal to the disc plane. The observed motions are consistent with material within the bar streaming along highly elongated orbits aligned with the rotating major axis. A barred galaxy may also contain a spheroidal bulge at its centre, spirals in the outer disc and, less commonly, other features such as a ring or lens. Mild asymmetries in both the light and kinematics are quite common. We review the main problems presented by these complicated dynamical systems and summarize the effort so far made towards their solution, emphasizing results which appear secure. (Truncated)Comment: This old review appeared in 1993. Plain tex with macro file. 82 pages 18 figures. A pdf version with figures at full resolution (3.24MB) is available at http://www.physics.rutgers.edu/~sellwood/bar_review.pd

Bronchiectasis insanity:Doing the same thing over and over again and expecting different results?

Bronchiectasis is an increasingly common disease with a significant impact on quality of life and morbidity of affected patients. It is also a very heterogeneous disease with numerous different underlying etiologies and presentations. Most treatments for bronchiectasis are based on low-quality evidence; consequently, no treatments have been approved by the US Food and Drug Administration or the European Medicines Agency for the treatment of bronchiectasis. The last several years have seen numerous clinical trials in which the investigational agent, thought to hold great promise, did not demonstrate a clinically or statistically significant benefit. This commentary will review the likely reasons for these disappointing results and a potential approach that may have a greater likelihood of defining evidence-based treatment for bronchiectasis

Biological Activity of CXCL8 Forms Generated by Alternative Cleavage of the Signal Peptide or by Aminopeptidase-Mediated Truncation

Posttranslational modification of chemokines is one of the mechanisms that regulate leukocyte migration during inflammation. Multiple natural NH(2)-terminally truncated forms of the major human neutrophil attractant interleukin-8 or CXCL8 have been identified. Although differential activity was reported for some CXCL8 forms, no biological data are available for others.status: publishe