Burkholderia cepacia Complex Species Differ in the Frequency of Variation of the Lipopolysaccharide O-Antigen Expression During Cystic Fibrosis Chronic Respiratory Infection.

Front Cell Infect Microbiol. 2019;9:273

Authors: Hassan AA, Coutinho CP, Sá-Correia I

Abstract
Burkholderia cepacia complex (Bcc) bacteria can adapt to the lung environment of cystic fibrosis (CF) patients resulting in the emergence of a very difficult to eradicate heterogeneous population leading to chronic infections associated with rapid lung function loss and increased mortality. Among the important phenotypic modifications is the variation of the lipopolysaccharide (LPS) structure at level of the O-antigen (OAg) presence, influencing adherence, colonization and the ability to evade the host defense mechanisms. The present study was performed to understand whether the loss of OAg expression during CF infection can be considered a general phenomenon in different Bcc species favoring its chronicity. In fact, it is still not clear why different Bcc species/strains differ in their ability to persist in the CF lung and pathogenic potential. The systematic two-decade-retrospective-longitudinal-screening conducted covered 357 isolates retrieved from 19 chronically infected patients receiving care at a central hospital in Lisbon. The study involved 21 Bcc strains of six/seven Bcc species/lineages, frequently or rarely isolated from CF patients worldwide. Different strains/clonal variants obtained during infection gave rise to characteristic OAg-banding patterns. The two most prevalent and feared species, B. cenocepacia and B. multivorans, showed a tendency to lose the OAg along chronic infection. B. cenocepacia recA lineage IIIA strains known to lead to particularly destructive infections exhibit the most frequent OAg loss, compared with lineage IIIB. The switch frequency increased with the duration of infection and the level of lung function deterioration. For the first time, it is shown that the rarely found B. cepacia and B. contaminans, whose representation in the cohort of patients examined is abnormally high, keep the OAg even during 10- or 15-year infections. Data from co-infections with different Bcc species reinforced these conclusions. Concerning the two other rarely found species examined, B. stabilis exhibited a stable OAg expression phenotype over the infection period while for the single clone of the more distantly related B. dolosa species, the OAg-chain was absent from the beginning of the 5.5-year infection until the patient dead. This work reinforces the relevance attributed to the OAg-expression switch suggesting marked differences in the various Bcc species.

PMID: 31417878 [PubMed - in process]

The CXCL12/CXCR4 Signaling Axis Retains Neutrophils at Inflammatory Sites in Zebrafish.

Front Immunol. 2019;10:1784

Authors: Isles HM, Herman KD, Robertson AL, Loynes CA, Prince LR, Elks PM, Renshaw SA

Abstract
The inappropriate retention of neutrophils at inflammatory sites is a major driver of the excessive tissue damage characteristic of respiratory inflammatory diseases including COPD, ARDS, and cystic fibrosis. The molecular programmes which orchestrate neutrophil recruitment to inflammatory sites through chemotactic guidance have been well-studied. However, how neutrophil sensitivity to these cues is modulated during inflammation resolution is not understood. The identification of neutrophil reverse migration as a mechanism of inflammation resolution and the ability to modulate this therapeutically has identified a new target to treat inflammatory disease. Here we investigate the role of the CXCL12/CXCR4 signaling axis in modulating neutrophil retention at inflammatory sites. We used an in vivo tissue injury model to study neutrophilic inflammation using transgenic zebrafish larvae. Expression of cxcl12a and cxcr4b during the tissue damage response was assessed using in situ hybridization and analysis of RNA sequencing data. CRISPR/Cas9 was used to knockdown cxcl12a and cxcr4b in zebrafish larvae. The CXCR4 antagonist AMD3100 was used to block the Cxcl12/Cxcr4 signaling axis pharmacologically. We identified that cxcr4b and cxcl12a are expressed at the wound site in zebrafish larvae during the inflammatory response. Following tail-fin transection, removal of neutrophils from inflammatory sites is significantly increased in cxcr4b and cxcl12a CRISPR knockdown larvae. Pharmacological inhibition of the Cxcl12/Cxcr4 signaling axis accelerated resolution of the neutrophil component of inflammation, an effect caused by an increase in neutrophil reverse migration. The findings of this study suggest that CXCR4/CXCL12 signaling may play an important role in neutrophil retention at inflammatory sites, identifying a potential new target for the therapeutic removal of neutrophils from the lung in chronic inflammatory disease.

PMID: 31417560 [PubMed - in process]

Pseudomonas aeruginosa Induced Host Epithelial Cell Mitochondrial Dysfunction.

Sci Rep. 2019 Aug 15;9(1):11929

Authors: Maurice NM, Bedi B, Yuan Z, Goldberg JB, Koval M, Hart CM, Sadikot RT

Abstract
The pathogenicity of P. aeruginosa is dependent on quorum sensing (QS), an inter-bacterial communication system that can also modulate host biology. The innate immune function of the lung mucosal barrier is dependent on proper mitochondrial function. The purpose of this study was to define the mechanism by which bacterial factors modulate host lung epithelial cell mitochondrial function and to investigate novel therapies that ameliorate this effect. 3-oxo-C12-HSL disrupts mitochondrial morphology, attenuates mitochondrial bioenergetics, and induces mitochondrial DNA oxidative injury. Mechanistically, we show that 3-oxo-C12-HSL attenuates expression of peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α), a master regulator of mitochondrial biogenesis, antioxidant defense, and cellular respiration, and its downstream effectors in both BEAS-2B and primary lung epithelial cells. Overexpression of PGC-1α attenuates the inhibition in cellular respiration caused by 3-oxo-C12-HSL. Pharmacologic activation of PGC-1α restores barrier integrity in cells treated with 3-oxo-C12-HSL. These data demonstrate that the P. aeruginosa QS molecule, 3-oxo-C12-HSL, alters mitochondrial pathways critical for lung mucosal immunity. Genetic and pharmacologic strategies that activate the PGC-1α pathway enhance host epithelial cell mitochondrial function and improve the epithelial innate response to P. aeruginosa. Therapies that rescue PGC-1α function may provide a complementary approach in the treatment of P. aeruginosa infection.

PMID: 31417101 [PubMed - in process]

The impact of CFTR modulator therapies on CF airway microbiology.

J Cyst Fibros. 2019 Aug 12;:

Authors: Rogers GB, Taylor SL, Hoffman LR, Burr LD

Abstract
Major historical advances in cystic fibrosis (CF) respiratory clinical care, including mechanical airway clearance and inhaled medications, have aimed to address the consequences of cystic fibrosis transmembrane conductance regulator (CFTR) dysfunction. In contrast, CFTR modulator therapies instead target the underlying protein defect that leads to CF lung disease. The extent to which these therapies might reduce susceptibility to chronic lung infections remains to be seen. However, by improving airway clearance, reducing the requirement for antibiotics, and in some cases, through direct antimicrobial effects, CFTR modulators are likely to result in substantial changes in CF airway microbiology. These changes could contribute substantially to the clinical benefit associated with modulator therapies, as well as providing an important indicator of treatment efficacy and residual pathophysiology. Indeed, the widespread introduction of modulator therapies might require us to re-consider our models of CF airway microbiology.

PMID: 31416774 [PubMed - as supplied by publisher]

Post-Translational Modifications in NETosis and NETs-Mediated Diseases.

Biomolecules. 2019 Aug 14;9(8):

Authors: Hamam HJ, Palaniyar N

Abstract
: Neutrophils undergo a unique form of cell death that generates neutrophil extracellular traps (NETs) that may help to neutralize invading pathogens and restore homeostasis. However, uncontrolled NET formation (NETosis) can result in numerous diseases that adversely affect health. Recent studies further elucidate the mechanistic details of the different forms of NETosis and their common end structure, as NETs were constantly found to contain DNA, modified histones and cytotoxic enzymes. In fact, emerging evidence reveal that the post translational modifications (PTMs) of histones in neutrophils have a critical role in regulating neutrophil death. Histone citrullination is shown to promote a rapid form of NET formation independent of NADPH oxidase (NOX), which relies on calcium influx. Interestingly, few studies suggest an association between histone citrullination and other types of PTMs to control cell survival and death, such as histone methylation. Even more exciting is the finding that histone acetylation has a biphasic effect upon NETosis, where histone deacetylase (HDAC) inhibitors promote baseline, NOX-dependent and -independent NETosis. However, increasing levels of histone acetylation suppresses NETosis, and to switch neutrophil death to apoptosis. Interestingly, in the presence of NETosis-promoting stimuli, high levels of HDACis limit both NETosis and apoptosis, and promote neutrophil survival. Recent studies also reveal the importance of the PTMs of neutrophils in influencing numerous pathologies. Histone modifications in NETs can act as a double-edged sword, as they are capable of altering multiple types of neutrophil death, and influencing numerous NET-mediated diseases, such as acute lung injury (ALI), thrombosis, sepsis, systemic lupus erythematosus, and cancer progression. A clear understanding of the role of different PTMs in neutrophils would be important for an understanding of the molecular mechanisms of NETosis, and to appropriately treat NETs-mediated diseases.

PMID: 31416265 [PubMed - in process]

Neutrophil Extracellular Trap Formation: Physiology, Pathology, and Pharmacology.

Biomolecules. 2019 Aug 14;9(8):

Authors: Ravindran M, Khan MA, Palaniyar N

Abstract
Neutrophil extracellular traps (NETs), a unique DNA framework decorated with antimicrobial peptides, have been in the scientific limelight for their role in a variety of pathologies ranging from cystic fibrosis to cancer. The formation of NETs, as well as relevant regulatory mechanisms, physiological factors, and pharmacological agents have not been systematically discussed in the context of their beneficial and pathological aspects. Novel forms of NET formation including vital NET formation continue to be uncovered, however, there remain fundamental questions around established mechanisms such as NADPH-oxidase (Nox)-dependent and Nox-independent NET formation. Whether NET formation takes place in the tissue versus the bloodstream, internal factors (e.g. reactive oxygen species (ROS) production and transcription factor activation), and external factors (e.g. alkaline pH and hypertonic conditions), have all been demonstrated to influence specific NET pathways. Elements of neutrophil biology such as transcription and mitochondria, which were previously of unknown significance, have been identified as critical mediators of NET formation through facilitating chromatin decondensation and generating ROS, respectively. While promising therapeutics inhibiting ROS, transcription, and gasdermin D are being investigated, neutrophil phagocytosis plays a critical role in host defense and any therapies targeting NET formation must avoid impairing the physiological functions of these cells. This review summarizes what is known in the many domains of NET research, highlights the most relevant challenges in the field, and inspires new questions that can bring us closer to a unified model of NET formation.

PMID: 31416173 [PubMed - in process]

Anti-inflammatory role of CD11b+Ly6G+ neutrophilic cells in allergic airway inflammation in mice.

Immunol Lett. 2018 12;204:67-74

Authors: Nowroozilarki N, Öz HH, Schroth C, Hector A, Nürnberg B, Hartl D, Kolahian S

Abstract
Asthma is a chronic inflammatory disease driven by overactivation of T helper cell type 2 (Th2) responses. In the present study, we investigated the functional relevance of CD11b+Ly6G+ neutrophilic cells in allergic airway inflammation in vivo. Allergic airway inflammation in mice was induced by house dust mite (HDM) or ovalbumin (OVA) sensitization and challenge. CD11b+Ly6G+ neutrophilic cells and T cell phenotypes were quantified by flow cytometry. To assess the functional in vivo relevance, CD11b+Ly6G+ neutrophilic cells were adoptively transferred intravenously or intratracheally and consequences on airway inflammation were studied. Adoptively transferred CD11b+Ly6G+ neutrophilic cells attenuated Th2 and Th17 responses and airway inflammation in vivo. Collectively, our results demonstrate that CD11b+Ly6G+ neutrophilic cells suppress airway inflammation in allergic mice in vivo. Adoptive cellular transfer of suppressive neutrophilic cells may represent an attractive therapeutic strategy for allergic airway inflammation.

PMID: 30392943 [PubMed - indexed for MEDLINE]

Robot-assisted radical prostatectomy due to a primary carcinoid prostatic tumour in a three-year-old child: a case report.

Urology. 2019 Aug 12;:

Authors: Llorens de Knecht E, Guadarrama Vega S, Donate G, Palou Redorta J, Bujons Tur A

Abstract
We present an extremely rare case of a 3-year-old child with a primary carcinoid tumour of the prostate. A 3-year-old boy presented with failure to thrive, constipation, recurrent respiratory tract infections and pain in the genital area. His karyotype was normal and cystic fibrosis and coeliac disease were excluded prior to further investigation. An abdominopelvic CT scan revealed a prostatic mass. Transrectal ultrasound-guided prostate biopsy was therefore performed and pathological examination revealed a carcinoid tumour. A robotic radical prostatectomy was performed. As this is an innovative surgical approach, we describe the surgical technique used.

PMID: 31415779 [PubMed - as supplied by publisher]

Measures of Cystic Fibrosis Airway Microbiota during Periods of Clinical Stability.

Ann Am Thorac Soc. 2019 Aug 15;:

Authors: Caverly LJ, Lu J, Carmody LA, Kalikin LM, Shedden K, Opron K, Azar M, Cahalan S, Foster B, VanDevanter DR, Simon RH, LiPuma JJ

Abstract
RATIONALE: Differences in cystic fibrosis (CF) airway microbiota between periods of clinical stability and exacerbation of respiratory symptoms have been investigated in efforts to better understand microbial triggers of CF exacerbations. Prior studies have often relied on a single or a limited number of samples to represent airway microbiota. However, the variability in airway microbiota during periods of clinical stability is not well known.
OBJECTIVES: We sought to determine the temporal variability of measures of airway microbiota during periods of clinical stability, and to identify factors associated with this variability.
METHODS: Sputum samples (n=527), obtained daily from six adults with CF during ten periods of clinical stability, underwent sequencing of the V4 region of the bacterial 16S rRNA gene. The variability in airway microbiota among samples within each period of clinical stability was calculated as the average of the Bray-Curtis similarity measures of each sample to every other sample within the same period. Outlier samples were defined as samples outside of 1.5 times the interquartile range within a baseline period with respect to the average Bray-Curtis similarity. Total bacterial load was measured with droplet digital PCR.
RESULTS: The variation in Bray-Curtis similarity and total bacterial load among samples within the same baseline period was greater than the variation observed in technical replicate control samples. Overall, 6% of samples were identified as outliers. Within baseline periods, changes in bacterial community structure occurred coincident with changes in maintenance antibiotics (p<0.05, AMOVA). Within subjects, bacterial community structure changed between baseline periods (p<0.01, AMOVA). Sample-to-sample similarity within baseline periods was greater with fewer interval days between sampling.
CONCLUSIONS: During periods of clinical stability, airway bacterial community structure and bacterial load vary among daily sputum samples from adults with CF. This day-to-day variation has bearing on study design and interpretation of results, particularly in analyses that rely on single samples to represent periods of interest (e.g., clinical stability versus pulmonary exacerbation). These data also emphasize the importance of accounting for maintenance antibiotic use and granularity of sample collection in studies designed to assess the dynamics of CF airway microbiota relative to changes in clinical state.

PMID: 31415187 [PubMed - as supplied by publisher]

Iterative Optimization of the Cyclic Peptide SFTI-1 Yields Potent Inhibitors of Neutrophil Proteinase 3.

ACS Med Chem Lett. 2019 Aug 08;10(8):1234-1239

Authors: Tian S, Swedberg JE, Li CY, Craik DJ, de Veer SJ

Abstract
Neutrophils produce at least four serine proteases that are packaged within azurophilic granules. These enzymes contribute to antimicrobial defense and inflammation but can be destructive if their activities are not properly regulated. Accordingly, they represent therapeutic targets for several diseases, including chronic obstructive pulmonary disease, cystic fibrosis, and rheumatoid arthritis. In this study, we focused on proteinase 3 (PR3), a neutrophil protease with elastase-like specificity, and engineered potent PR3 inhibitors based on the cyclic peptide sunflower trypsin inhibitor-1 (SFTI-1). We used an iterative optimization approach to screen targeted substitutions at the P1, P2, P2', and P4 positions of SFTI-1, and generated several new inhibitors with K i values in the low nanomolar range. These SFTI-variants show high stability in human serum and are attractive leads for further optimization.

PMID: 31413811 [PubMed]

A BAC Transgene Expressing Human CFTR under Control of Its Regulatory Elements Rescues Cftr Knockout Mice.

Sci Rep. 2019 Aug 14;9(1):11828

Authors: Gawenis LR, Hodges CA, McHugh DR, Valerio DM, Miron A, Cotton CU, Liu J, Walker NM, Strubberg AM, Gillen AE, Mutolo MJ, Kotzamanis G, Bosch J, Harris A, Drumm ML, Clarke LL

Abstract
Small-molecule modulators of cystic fibrosis transmembrane conductance regulator (CFTR) biology show promise in the treatment of cystic fibrosis (CF). A Cftr knockout (Cftr KO) mouse expressing mutants of human CFTR would advance in vivo testing of new modulators. A bacterial artificial chromosome (BAC) carrying the complete hCFTR gene including regulatory elements within 40.1 kb of DNA 5' and 25 kb of DNA 3' to the gene was used to generate founder mice expressing hCFTR. Whole genome sequencing indicated a single integration site on mouse chromosome 8 (8qB2) with ~6 gene copies. hCFTR+ offspring were bred to murine Cftr KO mice, producing hCFTR+/mCftr- (H+/m-) mice, which had normal survival, growth and goblet cell function as compared to wild-type (WT) mice. Expression studies showed hCFTR protein and transcripts in tissues typically expressing mCftr. Functionally, nasal potential difference and large intestinal short-circuit (Isc) responses to cAMP stimulation were similar in magnitude to WT mice, whereas small intestinal cAMP ΔIsc responses were reduced. A BAC transgenic mouse with functional hCFTR under control of its regulatory elements has been developed to enable the generation of mouse models of hCFTR mutations by gene editing for in vivo testing of new CF therapies.

PMID: 31413336 [PubMed - in process]

Role of a patatin-like phospholipase in Plasmodium falciparum gametogenesis and malaria transmission.

Proc Natl Acad Sci U S A. 2019 Aug 14;:

Authors: Singh P, Alaganan A, More KR, Lorthiois A, Thiberge S, Gorgette O, Guillotte Blisnick M, Guglielmini J, Aguilera SS, Touqui L, Singh S, Chitnis CE

Abstract
Transmission of Plasmodium falciparum involves a complex process that starts with the ingestion of gametocytes by female Anopheles mosquitoes during a blood meal. Activation of gametocytes in the mosquito midgut triggers "rounding up" followed by egress of both male and female gametes. Egress requires secretion of a perforin-like protein, PfPLP2, from intracellular vesicles to the periphery, which leads to destabilization of peripheral membranes. Male gametes also develop flagella, which assist in binding female gametes for fertilization. This process of gametogenesis, which is key to malaria transmission, involves extensive membrane remodeling as well as vesicular discharge. Phospholipase A2 enzymes (PLA2) are known to mediate membrane remodeling and vesicle secretion in diverse organisms. Here, we show that a P. falciparum patatin-like phospholipase (PfPATPL1) with PLA2 activity plays a key role in gametogenesis. Conditional deletion of the gene encoding PfPATPL1 does not affect P. falciparum blood stage growth or gametocyte development but reduces efficiency of rounding up, egress, and exflagellation of gametocytes following activation. Interestingly, deletion of the PfPATPL1 gene inhibits secretion of PfPLP2, reducing the efficiency of gamete egress. Deletion of PfPATPL1 also reduces the efficiency of oocyst formation in mosquitoes. These studies demonstrate that PfPATPL1 plays a role in gametogenesis, thereby identifying PLA2 phospholipases such as PfPATPL1 as potential targets for the development of drugs to block malaria transmission.

PMID: 31413195 [PubMed - as supplied by publisher]

Training dogs to differentiate Pseudomonas aeruginosa from other cystic fibrosis bacterial pathogens: not to be sniffed at?

Eur Respir J. 2019 Aug 14;:

Authors: Davies JC, Alton E, Simbo A, Murphy R, Seth I, Williams K, Somerville M, Jolly L, Morant S, Guest C

PMID: 31413160 [PubMed - as supplied by publisher]

Emerging non-genetic therapies for sickle cell disease.

Haematologica. 2019 Aug 14;:

Authors: Carden MA, Little J

Abstract
Sickle cell disease afflicts millions of people worldwide and approximately 100,000 Americans. Complications are myriad and arise as a result of complex pathological pathways 'downstream' to a point mutation in DNA, and include red blood cell membrane damage, inflammation, chronic hemolytic anemia with episodic vaso-occlusion, ischemia and pain, and ultimately risk of cumulative organ damage with reduced patient lifespan. The National Heart, Lung, and Blood Institute's 2014 evidence-based guideline for sickle cell disease -management states additional research is needed before investigational curative therapies will be widely available to most patients with sickle cell disease. To date, sickle cell disease has been cured by hematopoietic stem cell transplantation in approximately 1000 people, most of whom were children, and significantly ameliorated by gene therapy in a handful of subjects who have only limited follow-up thus far. During a timespan in which over 20 agents were approved for Cystic Fibrosis by the Food and Drug Administration, similar approval was granted for only two drugs for sickle cell disease (hydroxyurea and L-glutamine) despite the higher prevalence of sickle cell disease. . This trajectory appears to be changing, as the lack of multimodal agent therapy in sickle cell disease has spurred engagement among many in academia and industry who, in the last decade, have developed new drugs poised to prevent complications and alleviate suffering. Identified therapeutic strategies include fetal hemoglobin induction, inhibition of intracellular HbS polymerization, inhibition of oxidant stress and inflammation, and perturbation of the activation of the endothelium and other blood components (i.e. platelets, white blood cells, coagulation proteins) involved in sickle cell disease pathology. In this review, we present a crash course review of disease modifying approaches (minus hematopoietic stem cell transplant and gene therapy) for patients with sickle cell disease currently, or recently, tested in clinical trials in the post-approval era for hydroxyurea.

PMID: 31413089 [PubMed - as supplied by publisher]

Kinetic assay of starvation sensitivity in yeast autophagy mutants allows for the identification of intermediary phenotypes.

BMC Res Notes. 2019 Aug 14;12(1):505

Authors: Sturgeon CM, Robinson MR, Penton MC, Clemmer DC, Trujillo MA, Khawaja AU, Segarra VA

Abstract
OBJECTIVE: A classical method to quantitatively determine the starvation sensitivity phenotype of autophagy mutant budding yeast strains is to starve them for a period of time and then to assess the proportion of cells that retain the ability to form colonies when the availability of nutrients is restored. The readout of this colony-formation assay is generally evaluated after a fixed period of time following the restoration of nutrients, so that it can be considered an endpoint assay. One drawback we have identified is the inability to characterize subtle intermediary phenotypes that are detectable at the molecular level but fail to reach statistical significance in the colony formation experiment. We set out to determine whether a more dynamic measurement of growth during recovery after starvation would increase the sensitivity with which we are able to detect partial loss-of-function phenotypes.
RESULTS: We describe a 96-well plate-based assay to kinetically assess starvation sensitivity in budding yeast that allows for the quantitative detection of very modest starvation sensitivity phenotypes with statistical significance in autophagy mutant yeast strains lacking the ATG27 gene.

PMID: 31412956 [PubMed - in process]

Night Blindness in Cystic Fibrosis: The Key Role of Vitamin A in the Digestive System.

Nutrients. 2019 Aug 13;11(8):

Authors: Norsa L, Zazzeron L, Cuomo M, Claut L, Bulfamante AMC, Biffi A, Colombo C

Abstract
Vitamin A is a fundamental micronutrient that regulates various cellular patterns. Vitamin A deficiency (VAT) is a worldwide problem and the primary cause of nocturnal blindness especially in low income countries. Cystic fibrosis (CF) is a known risk factor of VAD because of liposoluble vitamin malabsorption due to pancreatic insufficiency. We describe a case of a 9-year-old girl who experienced recurrent episodes of nocturnal blindness due to profound VAD. This little girl is paradigmatic for the explanation of the key role of the gut-liver axis in vitamin A metabolism. She presents with meconium ileus at birth, requiring intestinal resection that led to a transient intestinal failure with parenteral nutrition need. In addition, she suffered from cholestatic liver disease due to CF and intestinal failure-associated liver disease. The interaction of pancreatic function, intestinal absorption and liver storage is fundamental for the correct metabolism of vitamin A.

PMID: 31412557 [PubMed - in process]

Disassembling the complexity of mucus barriers to develop a fast screening tool for early drug discovery.

J Mater Chem B. 2019 Aug 14;7(32):4940-4952

Authors: Pacheco DP, Butnarasu CS, Briatico Vangosa F, Pastorino L, Visai L, Visentin S, Petrini P

Abstract
Mucus is a natural barrier with a protective role that hinders drug diffusion, representing a steric and interactive barrier to overcome for an effective drug delivery to target sites. In diseases like cystic fibrosis (CF), pulmonary mucus exhibits altered features, which hamper clearance mechanisms and drug diffusion, ultimately leading to lung failure. Effectively modelling the passage through mucus still represents an unmet challenge. An airway CF mucus model is herein proposed to disassemble the complexity of the mucus barrier following a modular approach. A hydrogel, mainly composed of mucin in an alginate (Alg) network, is proposed to specifically model the chemical-physical properties of CF mucus. The steric retention of pathological mucus was reproduced by targeting its mesh size (approximately 50 nm) and viscoelastic properties. The interactive barrier was reproduced by a composition inspired from the CF mucus. Optimized mucus models, composed of 3 mg ml-1 Alg and 25 mg ml-1 mucin, exhibited a G' increasing from ∼21.2 to 55.2 Pa and a G'' ranging from ∼5.26 to 28.8 Pa in the frequency range of 0.1 to 20 Hz. Drug diffusion was tested using three model drugs. The proposed mucus model was able to discriminate between the mucin-drug interaction and the steric barrier of a mucus layer with respect to the parallel artificial membrane permeability (PAMPA) that models the phospholipidic cell membrane, the state-of-the-art screening tool for passive drug diffusion. The mucus model can be proposed as an in vitro tool for early drug discovery, representing a step forward to model the mucus layer. Additionally, the proposed methodology allows to easily include other molecules present within mucus, as relevant proteins, lipids and DNA.

PMID: 31411620 [PubMed - in process]

Brazilian consensus on non-cystic fibrosis bronchiectasis.

J Bras Pneumol. 2019 Aug 12;45(4):e20190122

Authors: Pereira MC, Athanazio RA, Dalcin PTR, Figueiredo MRF, Gomes M, Freitas CG, Ludgren F, Paschoal IA, Rached SZ, Maurici R

Abstract
Bronchiectasis is a condition that has been increasingly diagnosed by chest HRCT. In the literature, bronchiectasis is divided into bronchiectasis secondary to cystic fibrosis and bronchiectasis not associated with cystic fibrosis, which is termed non-cystic fibrosis bronchiectasis. Many causes can lead to the development of bronchiectasis, and patients usually have chronic airway symptoms, recurrent infections, and CT abnormalities consistent with the condition. The first international guideline on the diagnosis and treatment of non-cystic fibrosis bronchiectasis was published in 2010. In Brazil, this is the first review document aimed at systematizing the knowledge that has been accumulated on the subject to date. Because there is insufficient evidence on which to base recommendations for various treatment topics, here the decision was made to prepare an expert consensus document. The Brazilian Thoracic Association Committee on Respiratory Infections summoned 10 pulmonologists with expertise in bronchiectasis in Brazil to conduct a critical assessment of the available scientific evidence and international guidelines, as well as to identify aspects that are relevant to the understanding of the heterogeneity of bronchiectasis and to its diagnostic and therapeutic management. Five broad topics were established (pathophysiology, diagnosis, monitoring of stable patients, treatment of stable patients, and management of exacerbations). After this subdivision, the topics were distributed among the authors, who conducted a nonsystematic review of the literature, giving priority to major publications in the specific areas, including original articles, review articles, and systematic reviews. The authors reviewed and commented on all topics, producing a single final document that was approved by consensus.

PMID: 31411280 [PubMed - in process]

Animal models for cystic fibrosis: A systematic search and mapping review of the literature - Part 1: genetic models.

Lab Anim. 2019 Aug 14;:23677219868502

Authors: Leenaars CH, De Vries RB, Heming A, Visser D, Holthaus D, Reijmer J, Elzinga J, Kempkes RW, Punt C, Beumer W, Meijboom FL, Ritskes-Hoitinga M

Abstract
Animal models for cystic fibrosis (CF) have enhanced our understanding of the pathology and contributed to the development of new treatments. In the field of CF, many animal models have been developed and described. To our knowledge, thus far, none of the reviews of CF animal models has used a systematic methodology. A systematic approach to creating model overviews can lead to an objective, evidence-based choice of an animal model for new research questions. We searched Pubmed and Embase for the currently available animal models for CF. Two independent reviewers screened the results. We included all primary studies describing an animal model for CF. After duplicate removal, 12,304 publications were left. Because of the large number of models, in the current paper, only the genetic models are presented. A total of 636 publications were identified describing genetic animal models for CF in mice, pigs, ferrets, rats and zebrafish. Most of these models have an altered Cftr gene. An overview of basic model characteristics and outcome measures for these genetic models is provided, together with advice on using these data. As far as the authors are aware, this is one of the largest systematic mapping reviews on genetic animal models for CF. It can aid in selecting a suitable model and outcome measures. In general, the reporting quality of the included publications was poor. Further systematic reviews are warranted to determine the quality and translational value of these models further.

PMID: 31411127 [PubMed - as supplied by publisher]

Human protein paucimannosylation: cues from the eukaryotic kingdoms.

Biol Rev Camb Philos Soc. 2019 Aug 14;:

Authors: Tjondro HC, Loke I, Chatterjee S, Thaysen-Andersen M

Abstract
Paucimannosidic proteins (PMPs) are bioactive glycoproteins carrying truncated α- or β-mannosyl-terminating asparagine (N)-linked glycans widely reported across the eukaryotic domain. Our understanding of human PMPs remains limited, despite findings documenting their existence and association with human disease glycobiology. This review comprehensively surveys the structures, biosynthetic routes and functions of PMPs across the eukaryotic kingdoms with the aim of synthesising an improved understanding on the role of protein paucimannosylation in human health and diseases. Convincing biochemical, glycoanalytical and biological data detail a vast structural heterogeneity and fascinating tissue- and subcellular-specific expression of PMPs within invertebrates and plants, often comprising multi-α1,3/6-fucosylation and β1,2-xylosylation amongst other glycan modifications and non-glycan substitutions e.g. O-methylation. Vertebrates and protists express less-heterogeneous PMPs typically only comprising variable core fucosylation of bi- and trimannosylchitobiose core glycans. In particular, the Manα1,6Manβ1,4GlcNAc(α1,6Fuc)β1,4GlcNAcβAsn glycan (M2F) decorates various human neutrophil proteins reportedly displaying bioactivity and structural integrity demonstrating that they are not degradation products. Less-truncated paucimannosidic glycans (e.g. M3F) are characteristic glycosylation features of proteins expressed by human cancer and stem cells. Concertedly, these observations suggest the involvement of human PMPs in processes related to innate immunity, tumorigenesis and cellular differentiation. The absence of human PMPs in diverse bodily fluids studied under many (patho)physiological conditions suggests extravascular residence and points to localised functions of PMPs in peripheral tissues. Absence of PMPs in Fungi indicates that paucimannosylation is common, but not universally conserved, in eukaryotes. Relative to human PMPs, the expression of PMPs in plants, invertebrates and protists is more tissue-wide and constitutive yet, similar to their human counterparts, PMP expression remains regulated by the physiology of the producing organism and PMPs evidently serve essential functions in development, cell-cell communication and host-pathogen/symbiont interactions. In most PMP-producing organisms, including humans, the N-acetyl-β-hexosaminidase isoenzymes and linkage-specific α-mannosidases are glycoside hydrolases critical for generating PMPs via N-acetylglucosaminyltransferase I (GnT-I)-dependent and GnT-I-independent truncation pathways. However, the identity and structure of many species-specific PMPs in eukaryotes, their biosynthetic routes, strong tissue- and development-specific expression, and diverse functions are still elusive. Deep exploration of these PMP features involving, for example, the characterisation of endogenous PMP-recognising lectins across a variety of healthy and N-acetyl-β-hexosaminidase-deficient human tissue types and identification of microbial adhesins reactive to human PMPs, are amongst the many tasks required for enhanced insight into the glycobiology of human PMPs. In conclusion, the literature supports the notion that PMPs are significant, yet still heavily under-studied biomolecules in human glycobiology that serve essential functions and create structural heterogeneity not dissimilar to other human N-glycoprotein types. Human PMPs should therefore be recognised as bioactive glycoproteins that are distinctly different from the canonical N-glycoprotein classes and which warrant a more dedicated focus in glycobiological research.

PMID: 31410980 [PubMed - as supplied by publisher]