Succinate links mitochondria to deadly bacteria in cystic fibrosis.

Ann Transl Med. 2019 Dec;7(Suppl 8):S263

Authors: Esposito S, Villella VR, Rossin F, Tosco A, Raia V, Luciani A

PMID: 32015982 [PubMed]

Pseudomonas aeruginosa Alginate Benefits Staphylococcus aureus?

J Bacteriol. 2020 Feb 03;:

Authors: Schurr MJ

Abstract
In this month's Journal of Bacteriology, Price et al. show that the Pseudomonas aeruginosa produced exopolysaccharide alginate, protects Staphylococcus aureus by dampening the expression of P. aeruginosa virulence products that usually inhibit S. aureus respiration and cell membrane integrity when the two organisms compete in other environments. This is the first report that exogenously added alginate affects P. aeruginosa competition and provides a partial explanation for S. aureus and P. aeruginosa co-infections in Cystic Fibrosis.

PMID: 32015142 [PubMed - as supplied by publisher]

Activity of antibiotics against Pseudomonas aeruginosa in an in vitro model of biofilms in the context of cystic fibrosis : influence of the culture medium.

Antimicrob Agents Chemother. 2020 Feb 03;:

Authors: Diaz Iglesias Y, Van Bambeke F

Abstract
Pseudomonas aeruginosa is a major cause of respiratory biofilm-related infections in patients with cystic fibrosis. We developed an in-vitro pharmacodynamic model to study the activity of antipseudomonal antibiotics against PAO1 biofilms grown in artificial sputum medium with agar (ASM(+)) vs. trypticase soy broth supplemented with glucose and NaCl (TGN). We measured bacterial counts, metabolic activity (fluorescein diacetate [FDA] hydrolysis) and biomass (crystal violet absorbance). Biofilms grew slower in ASM(+) vs. TGN but reached the same cfu counts and metabolic activity, and slightly higher biomass after 48h. Concentration-response curves of antibiotics after 24h incubation with mature biofilms showed maximal effects ranging between -3 (ciprofloxacin) and -1.5 (ceftazidime, meropenem) log10 cfu decrease, tobramycin and colistin showing intermediate values. These maximal reductions in cfus were similar in both media for ciprofloxacin and β-lactams but lower in ASM(+) than in TGN for tobramycin and colistin; they were reached at concentrations lower than human Cmax for ciprofloxacin and β-lactams only. Reduction in metabolic activity and in biomass were low in both media. Small colony variants were selected by tobramycin in ASM(+) and by ciprofloxacin in both media. The model was then successfully applied to 4 isolates from patients with cystic fibrosis. These biofilms showed similar cfus counts but higher biomass than PAO1 in ASM(+) and moderate differences in their susceptibility to antibiotics as compared to PAO1 biofilms grown in this medium. This model proved useful to establish the pharmacodynamic profile of drugs against P. aeruginosa biofilms in the context of cystic fibrosis.

PMID: 32015047 [PubMed - as supplied by publisher]

In-vivo crystals reveal critical features of the interaction between CFTR and the PDZ2 domain of Na+/H+ exchange cofactor NHERF1.

J Biol Chem. 2020 Feb 02;:

Authors: Martin ER, Barbieri A, Ford RC, Robinson RC

Abstract
Crystallization of recombinant proteins has been fundamental to our understanding of protein function, dysfunction and molecular recognition. However, this information has often been gleaned under extremely non-physiological protein, salt, and H+ concentrations. Here, we describe the development of a robust Inka1-Box (iBox)-PAK4cat system that spontaneously crystallizes in several mammalian cell types. The semi-quantitative assay described here allows the measurement of in-vivo protein--protein interactions using a novel GFP-linked reporter system which produces fluorescent readouts from protein crystals. We combined this assay with in-vitro X-ray crystallography and molecular dynamics studies to characterize the molecular determinants of the interaction between PDZ2 domain of Na+/H+ exchange regulatory cofactor NHE-RF1 (NHERF1) and cystic fibrosis transmembrane conductance regulator (CFTR), a protein complex pertinent to the genetic disease cystic fibrosis. These experiments revealed the crystal structure of the extended PDZ domain of NHERF1 and indicated, contrary to what has been previously reported, that residue selection at positions -1 and -3 of the PDZ-binding motif influences the affinity and specificity of the NHERF1 PDZ2-CFTR interaction. Our results suggest that this system could be utilized to screen additional protein-protein interactions, provided they can be accommodated within the spacious iBox-PAK4cat lattice.

PMID: 32014995 [PubMed - as supplied by publisher]

Applying whole-genome sequencing in relation to phenotype and outcomes in siblings with cystic fibrosis.

Cold Spring Harb Mol Case Stud. 2020 Feb;6(1):

Authors: Wilk MA, Braun AT, Farrell PM, Laxova A, Brown DM, Holt JM, Birch CL, Sosonkina N, Wilk BM, Worthey EA

Abstract
Variations in disease onset and/or severity have often been observed in siblings with cystic fibrosis (CF), despite the same CFTR genotype and environment. We postulated that genomic variation (modifier and/or pharmacogenomic variants) might explain these clinical discordances. From a cohort of patients included in the Wisconsin randomized clinical trial (RCT) of newborn screening (NBS) for CF, we identified two brothers who showed discordant lung disease courses as children, with one milder and the other more severe than average, and a third, eldest brother, who also has severe lung disease. Leukocytes were harvested as the source of DNA, and whole-genome sequencing (WGS) was performed. Variants were identified and analyzed using in-house-developed informatics tools. Lung disease onset and severity were quantitatively different between brothers during childhood. The youngest, less severely affected brother is homozygous for HFE p.H63D. He also has a very rare PLG p.D238N variant that may influence host-pathogen interaction during chronic lung infection. Other variants of interest were found differentially between the siblings. Pharmacogenomics findings were consistent with the middle, most severely affected brother having poor outcomes to common CF treatments. We conclude that genomic variation between siblings with CF is expected. Variable lung disease severity may be associated with differences acting as genetic modifiers and/or pharmacogenomic factors, but large cohort studies are needed to assess this hypothesis.

PMID: 32014855 [PubMed - in process]

What Spanish consensus meant for the treatment of Pseudomonas aeruginosa infection in patients with cystic fibrosis in the initial lung infection?

Med Clin (Barc). 2020 Jan 31;:

Authors: Prados C, Zamarrón E, Girón R, en representación del Grupo de trabajo de Jornadas Ayre-Consenso español, Grupo de trabajo Jornadas Ayre-Consenso español

PMID: 32014243 [PubMed - as supplied by publisher]

Neutrophil Adaptations upon Recruitment to the Lung: New Concepts and Implications for Homeostasis and Disease.

Int J Mol Sci. 2020 Jan 28;21(3):

Authors: Giacalone VD, Margaroli C, Mall MA, Tirouvanziam R

Abstract
Neutrophils have a prominent role in all human immune responses against any type of pathogen or stimulus. The lungs are a major neutrophil reservoir and neutrophilic inflammation is a primary response to both infectious and non-infectious challenges. While neutrophils are well known for their essential role in clearance of bacteria, they are also equipped with specific mechanisms to counter viruses and fungi. When these defense mechanisms become aberrantly activated in the absence of infection, this commonly results in debilitating chronic lung inflammation. Clearance of bacteria by phagocytosis is the hallmark role of neutrophils and has been studied extensively. New studies on neutrophil biology have revealed that this leukocyte subset is highly adaptable and fulfills diverse roles. Of special interest is how these adaptations can impact the outcome of an immune response in the lungs due to their potent capacity for clearing infection and causing damage to host tissue. The adaptability of neutrophils and their propensity to influence the outcome of immune responses implicates them as a much-needed target of future immunomodulatory therapies. This review highlights the recent advances elucidating the mechanisms of neutrophilic inflammation, with a focus on the lung environment due to the immense and growing public health burden of chronic lung diseases such as cystic fibrosis (CF) and chronic obstructive pulmonary disease (COPD), and acute lung inflammatory diseases such as transfusion-related acute lung injury (TRALI).

PMID: 32013006 [PubMed - in process]

Alpha-1 Antitrypsin-A Target for MicroRNA-Based Therapeutic Development for Cystic Fibrosis.

Int J Mol Sci. 2020 Jan 28;21(3):

Authors: Hunt AMD, Glasgow AMA, Humphreys H, Greene CM

Abstract
Cystic fibrosis (CF) is an autosomal recessive genetic disorder arising from mutations to the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Disruption to normal ion homeostasis in the airway results in impaired mucociliary clearance, leaving the lung more vulnerable to recurrent and chronic bacterial infections. The CF lung endures an excess of neutrophilic inflammation, and whilst neutrophil serine proteases are a crucial part of the innate host defence to infection, a surplus of neutrophil elastase (NE) is understood to create a net destructive effect. Alpha-1 antitrypsin (A1AT) is a key antiprotease in the control of NE protease activity but is ineffective in the CF lung due to the huge imbalance of NE levels. Therapeutic strategies to boost levels of protective antiproteases such as A1AT in the lung remain an attractive research strategy to limit the damage from excess protease activity. microRNAs are small non-coding RNA molecules that bind specific cognate sequences to inhibit expression of target mRNAs. The inhibition of miRNAs which target the SERPINA1 (A1AT-encoding gene) mRNA represents a novel therapeutic approach for CF inflammation. This could involve the delivery of antagomirs that bind and sequester the target miRNA, or target site blockers that bind miRNA recognition elements within the target mRNA to prevent miRNA interaction. Therefore, miRNA targeted therapies offer an alternative strategy to drive endogenous A1AT production and thus supplement the antiprotease shield of the CF lung.

PMID: 32012925 [PubMed - in process]

Is Phosphatidylinositol 3- kinase (PI3K) a Villain in Cystic Fibrosis?

Am J Respir Cell Mol Biol. 2020 Feb 03;:

Authors: Natarajan V

PMID: 32011906 [PubMed - as supplied by publisher]

Challenges facing microRNA therapeutics for cystic fibrosis lung disease.

Epigenomics. 2020 Feb 03;:

Authors: Santi C, Greene CM

PMID: 32011168 [PubMed - as supplied by publisher]

Maternal Genetic Disorders and Fetal Development.

Prenat Diagn. 2020 Feb 03;:

Authors: Mardy AH, Chetty SP, Norton ME

Abstract
With improvements in early diagnosis and management of genetic diseases, more women with genetic disorders are reaching reproductive age and becoming pregnant. While pregnancy can have a significant impact on a woman's health when there is an underlying genetic disorder, there can also be fetal effects, including embryopathy, fetal growth restriction and brain injury. Some maternal genetic disorders are associated with adverse perinatal outcomes, including a high risk of perinatal loss and preterm birth. In this article, we review several maternal genetic disorders associated with fetal risk that are important for clinicians and patients to understand and manage appropriately. These include phenylalanine hydroxylase (PAH) deficiency and other inborn errors of metabolism, tuberous sclerosis complex, myotonic dystrophy, cystic fibrosis, Turner syndrome, sickle cell disease and connective tissue disorders. This article is protected by copyright. All rights reserved.

PMID: 32010984 [PubMed - as supplied by publisher]

Sodium antiporters of Pseudomonas aeruginosa in challenging conditions: effects on growth, biofilm formation, and swarming motility.

J Genet Eng Biotechnol. 2020 Feb 03;18(1):4

Authors: Schubiger CB, Hoang KHT, Häse CC

Abstract
BACKGROUND: Pseudomonas aeruginosa is a bacterial pathogen that can cause grave and sometimes chronic infections in patients with weakened immune systems and cystic fibrosis. It is expected that sodium/proton transporters in the cellular membrane are crucial for the organism's survival and growth under certain conditions, since many cellular processes rely on the maintenance of Na+ and H+ transmembrane gradients.
RESULTS: This study focused on the role of the primary and secondary proton and/or sodium pumps Mrp, Nuo, NhaB, NhaP, and NQR for growth, biofilm formation, and swarming motility in P. aeruginosa. Using mutants with gene deletions, we investigated the impact of each sodium pump's absence on the overall growth, biofilm formation, motility, and weak acid tolerance of the organism. We found that the absence of some, but not all, of the sodium pumps have a deleterious effect on the different phenotypes of P. aeruginosa.
CONCLUSION: The absence of the Mrp sodium/proton antiporter was clearly important in the organism's ability to survive and function in environments of higher pH and sodium concentrations, while the absence of Complex I, which is encoded by the nuo genes, had some consistent impact on the organism's growth regardless of the pH and sodium concentration of the environment.

PMID: 32009221 [PubMed]

Azithromycin inhibits constitutive airway epithelial sodium channel activation in vitro and modulates downstream pathogenesis in vivo.

Biol Pharm Bull. 2020 Jan 31;:

Authors: Fujikawa H, Kawakami T, Nakashima R, Nasu A, Kamei S, Nohara H, Eto Y, Ueno-Shuto K, Takeo T, Nakagata N, Suico MA, Kai H, Shuto T

Abstract
Epithelial sodium channel (ENaC) is an amiloride-sensitive sodium ion channel that is expressed in epithelial tissues. ENaC overexpression and/or hyperactivation in airway epithelial cells cause sodium over-absorption and dysregulated ciliary movement for mucus clearance; however, the agents that suppress constitutive airway ENaC activation are yet to be clinically available. Here, we focused on macrolides, which are widely used antibiotics that have many potential immunomodulatory effects. We examined whether macrolides could modulate constitutive ENaC activity and downstream events that typify cystic fibrosis (CF) and chronic obstructive pulmonary diseases (COPD) in in vitro and in vivo models of ENaC overexpression. Treatment of ENaC-overexpressing human bronchial epithelial cells (β/γENaC-16HBE14o- cells) with three macrolides (erythromycin, clarithromycin, azithromycin) confirmed dose-dependent suppression of ENaC function. For in vivo studies, mice harboring airway specific βENaC overexpression (C57BL/6J-βENaC-Tg mice) were treated orally with azithromycin, a well-established antimicrobial agent that has been widely prescribed. Azithromycin treatment modulated pulmonary mechanics, emphysematous phenotype and pulmonary dysfunction. Notably, a lower dose (3 mg kg-1) of azithromycin significantly increased forced expiratory volume in 0.1 second (FEV0.1), an inverse indicator of bronchoconstriction. Although not statistically significant, improvement of pulmonary obstructive parameters such as emphysema and lung dysfunction (FEV0.1%) was observed. Our results demonstrate that macrolides directly attenuate constitutive ENaC function in vitro and may be promising for the treatment of obstructive lung diseases with defective mucociliary clearance, possibly by targeting ENaC hyperactivation.

PMID: 32009028 [PubMed - as supplied by publisher]

Emerging Themes in PDZ Domain Signaling: Structure, Function, and Inhibition.

Int Rev Cell Mol Biol. 2019;343:129-218

Authors: Liu X, Fuentes EJ

Abstract
Post-synaptic density-95, disks-large and zonula occludens-1 (PDZ) domains are small globular protein-protein interaction domains widely conserved from yeast to humans. They are composed of ∼90 amino acids and form a classical two α-helical/six β-strand structure. The prototypical ligand is the C-terminus of partner proteins; however, they also bind internal peptide sequences. Recent findings indicate that PDZ domains also bind phosphatidylinositides and cholesterol. Through their ligand interactions, PDZ domain proteins are critical for cellular trafficking and the surface retention of various ion channels. In addition, PDZ proteins are essential for neuronal signaling, memory, and learning. PDZ proteins also contribute to cytoskeletal dynamics by mediating interactions critical for maintaining cell-cell junctions, cell polarity, and cell migration. Given their important biological roles, it is not surprising that their dysfunction can lead to multiple disease states. As such, PDZ domain-containing proteins have emerged as potential targets for the development of small molecular inhibitors as therapeutic agents. Recent data suggest that the critical binding function of PDZ domains in cell signaling is more than just glue, and their binding function can be regulated by phosphorylation or allosterically by other binding partners. These studies also provide a wealth of structural and biophysical data that are beginning to reveal the physical features that endow this small modular domain with a central role in cell signaling.

PMID: 30712672 [PubMed - indexed for MEDLINE]

Bronchiectasis in rheumatoid arthritis. A clinical appraisial.

Joint Bone Spine. 2020 Jan 30;:

Authors: Duarte AC, Porter J, Leandro MJ

Abstract
Bronchiectasis is defined as irreversibly damaged and dilated bronchi and is one of the most common pulmonary manifestations in patients with rheumatoid arthritis (RA). The model of RA-associated autoimmunity induced in some individuals by chronic bacterial infection in bronchiectasis is becoming increasingly acceptable, although a genetic predisposition to RA-associated bronchiectasis has also been demonstrated. Bronchiectasis should be suspected in RA patients with chronic cough and sputum production or frequent respiratory infections and the diagnosis must be confirmed by thoracic high-resolution computed tomography. Management of patients with RA-associated bronchiectasis includes a multimodal treatment approach. Similar to all patients with non-cystic fibrosis bronchiectasis, patients with RA-associated bronchiectasis benefit from a pulmonary rehabilitation program, including an exercise/muscle strengthening program and an education program with a specific session on airway clearance techniques. Prophylactic antibiotics are recommended for patients with frequent (3 or more infective exacerbations per year) or severe infections requiring hospitalization/intravenous antibiotics and inhaled corticosteroids and long-acting β2-agonists should be used in patients with non-cystic fibrosis bronchiectasis and associated airway hyper-responsiveness. In patients with RA-associated bronchiectasis the use of immunomodulatory drugs has to be carefully considered, as they are essential to control disease activity, despite being associated with an increased infectious risk. Pneumococcal and influenza vaccines are advised to all patients with RA-associated bronchiectasis in order to reduce the risk of infection. Patients with RA-associated bronchiectasis have a poorer prognosis than those with either RA or bronchiectasis alone and require regular follow-up, under the joint care of a rheumatologist and a pulmonologist.

PMID: 32007647 [PubMed - as supplied by publisher]

ppGpp ribosome dimerization model for bacterial persister formation and resuscitation.

Biochem Biophys Res Commun. 2020 Jan 29;:

Authors: Song S, Wood TK

Abstract
Stress is ubiquitous for bacteria and can convert a subpopulation of cells into a dormant state known as persistence, in which cells are tolerant to antimicrobials. These cells revive rapidly when the stress is removed and are likely the cause of many recurring infections such as those associated with tuberculosis, cystic fibrosis, and Lyme disease. However, how persister cells are formed is not understood well. Here we propose the ppGpp ribosome dimerization persister (PRDP) model in which the alarmone guanosine pentaphosphate/tetraphosphate (henceforth ppGpp) generates persister cells directly by inactivating ribosomes via the ribosome modulation factor (RMF), the hibernation promoting factor (Hpf), and the ribosome-associated inhibitor (RaiA). We demonstrate that persister cells contain a large fraction of 100S ribosomes, that inactivation of RMF, HpF, and RaiA reduces persistence and increases single-cell persister resuscitation and that ppGpp has no effect on single-cell persister resuscitation. Hence, a direct connection between ppGpp and persistence is shown along with evidence of the importance of ribosome dimerization in persistence and for active ribosomes during resuscitation.

PMID: 32007277 [PubMed - as supplied by publisher]

Pulmonary Cystic Disease and Its Mimics.

Surg Pathol Clin. 2020 Mar;13(1):141-163

Authors: Jones KD

Abstract
Cystic diseases of the lung encompass a fairly broad variety of different diseases with causes including genetic abnormalities, smoking-related problems, developmental disorders, malignant neoplasms, and inflammatory processes. In addition, there are several diagnoses that closely resemble cystic lung disease, including cavitary diseases, cystic bronchiectasis, emphysema, and cystic changes in fibrosing interstitial lung disease. This article provides a review of cystic lung disease and its gross and histologic mimics.

PMID: 32005429 [PubMed - in process]

Evaluating the Clinical Implications of an Innovative Anatomy Lab Suturing Curriculum for Physician Assistant Students.

J Physician Assist Educ. 2020 Jan 30;:

Authors: Fankhanel C, Talwalkar JS, Rienzo R

Abstract
PURPOSE: The purpose of this study was to evaluate the outcomes of a new suturing curriculum incorporated throughout the anatomy lab for Yale Physician Associate (YPA) students.
METHODS: This controlled before-and-after study evaluated the class of 2017 (n = 39) as the control group and the class of 2018 (n = 37) as the intervention group. Suturing competency data were collected on all students from their clinical preceptor evaluations. Students completed surveys to measure perceived confidence in suturing skills.
RESULTS: Preceptor evaluations showed a 14.98% increase in suturing competence between the control and intervention groups (P < .05). Student surveys showed no significant difference in self-perceived confidence in suturing skills between the 2 cohorts.
CONCLUSIONS: After initiation of an innovative anatomy lab suturing curriculum, YPA students demonstrated improvement in preceptor-perceived suturing competency during clinical rotations.

PMID: 32004249 [PubMed - as supplied by publisher]

Expanded carrier screening for preconception reproductive risk assessment: Prevalence of carrier status in a Mexican population.

Prenat Diagn. 2020 Jan 31;:

Authors: Hernandez-Nieto C, Alkon-Meadows T, Lee J, Cacchione T, Iyune-Cojab E, Garza-Galvan M, Luna-Rojas M, Copperman AB, Sandler B

Abstract
OBJECTIVE: Genetic carrier screening has the potential to identify couples at risk of having a child affected with an autosomal recessive or X-linked disorder. However, the current prevalence of carrier status for these conditions in developing countries is not well defined. This study assesses the prevalence of carrier status of selected genetic conditions utilizing an expanded, pan-ethnic genetic carrier screening panel (ECS) in a large population of Mexican patients.
METHODS: Retrospective chart review of all patients tested with a single ECS panel at an international infertility center from 2012-2018 were included, the prevalence of positive carrier status in a Mexican population was evaluated.
RESULTS: 805 individuals were analyzed with ECS testing for 283 genetic conditions. 352 carriers (43.7%) were identified with 503 pathogenic variants in 145 different genes. Seventeen of the 391 participating couples (4.34%) were identified as being at-risk couples. The most prevalent alleles found were associated with alpha-thalassemia, cystic fibrosis, GJB2 non-syndromic hearing loss, biotinidase deficiency, and familial Mediterranean fever.
CONCLUSION: Based on the prevalence and severity of Mendelian disorders, we recommend that couples, who wish to conceive regardless of their ethnicity background, explore carrier screening and genetic counseling prior to reproductive medical treatment. This article is protected by copyright. All rights reserved.

PMID: 32003480 [PubMed - as supplied by publisher]

Clinical characteristics and genetic analysis of cystic fibrosis transmembrane conductance reseptor- related disease.

Pediatr Int. 2020 Jan 30;:

Authors: Kilinc AA, Alishbayli G, Taner HE, Cokugras FC, Cokugras H

Abstract
BACKGROUND: Cystic fibrosis transmembrane conductance receptor (CFTR)-related disease is diagnosed in patients affected by CFTR dysfunction but who do not fully meet the CF diagnostic criteria. Only 2% of all CF patients have CFTR-related disease. We define the demographic characteristics of such patients, perform mutational analyses, and describe the clinical findings.
METHODS: Twenty-four patients were followed-up in terms of CFTR-related disease. Patients with CF symptoms but who did not completely fulfil the CF diagnostic criteria were enrolled. Age, body mass index (BMI) at the times of diagnosis and admission, symptoms, pulmonary function and fecal elastase test results, gene analyses, and clinical findings during follow-up, were evaluated.
RESULTS: Ten patients (42%) were female and 14 (58%) male, of mean age 15.3 years (min-max 6-20 years). The mean age at diagnosis was 8.5 years (min-max 3-14 years) and the most common presenting complaint was a cough (n=19). During follow-up, chronic sinusitis developed in 15 patients, bronchiectasis in 13, nasal polyposis in six, failure to thrive in three, recurrent pancreatitis in two, asthma in one, and congenital bilateral absence of the vas deferens (CBAVD) in one. Fecal elastase levels were low in only one of the three patients who failed to thrive. In terms of CFTR gene mutations, two were found in 10 patients, one in eight patients, and none in six.
CONCLUSION: CFTR-related disease presents with various clinical findings. Serious symptoms may develop later in life. Late diagnosis significantly compromises the quality and duration of life in such patients.

PMID: 32003094 [PubMed - as supplied by publisher]