Additive Effect of Dornase Alfa and Nacystelyn on Transportability and Viscoelasticity of Cystic Fibrosis Sputum

OBJECTIVE: To investigate the effect of dornase alfa (DA), Nacystelyn (NAL) and their combination on mucociliary transportability and mucus viscoelasticity of cystic fibrosis (CF) sputum, and to assess whether the combination possesses an additive effect.DESIGN: Determination of transportability in frog palate and viscoelasticity in vitro.SETTING: Research laboratory at a medical centre. Patients: Sputa from 15 patients with CF, chronically infected with Pseudomonas aeruginosa, were studied.INTERVENTIONS: Sputum samples were incubated without any drug solution as a control, and with normal saline, DA, NAL and a mixture of DA and NAL in concentrations approximating those achieved in clinical practice.RESULTS: Normal saline (10% volume) by itself had a small effect on CF sputum transportability with a mean increase of 9%, and on viscoelasticity with a mean of decrease of 0.22 log units, respectively, compared with control (incubation without saline). DA (200 nM) further increased the transportability by a mean of 35% versus saline and decreased viscoelasticity by a mean of 0.30 log units. NAL (100 µM) increased the transportability by a mean of 32% and decreased viscoelasticity by a mean of 0.22 log units from the levels achieved with saline. The mixture of DA plus NAL at one-half of the above concentration of each agent produced an additional increase in the transportability, by a mean of 18%, and a further decrease in viscoelasticity, by a mean of 0.25 log units, compared with DA or NAL as a single treatment.CONCLUSIONS: The combination of DA and NAL exhibits an additive effect for both the viscoelasticity and transportability of CF sputum samples. The two agents appear to act well together in breaking down the bonding due to extracellular DNA and mucins. Clinical studies should be undertaken to see whether the additive combination at lower concentration produces the anticipated benefits of improved airway clearance and fewer side effects.

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Preliminary Report of In vitro and In vivo Effectiveness of Dornase Alfa on SARS-CoV-2 Infection (Preprint)

10.2196/preprints.20200 ◽

2020 ◽

Author(s):

Hacer Kuzu Okur ◽

Koray Yalcin ◽

Cihan Tastan ◽

Sevda Demir ◽

Bulut Yurtsever ◽

...

Keyword(s):

Cystic Fibrosis ◽

Respiratory Tract ◽

Mononuclear Cells ◽

Multiple Organ ◽

Peripheral Blood Mononuclear ◽

Dornase Alfa ◽

Tract Infections ◽

Adult Peripheral Blood

UNSTRUCTURED Dornase alfa, the recombinant form of the human DNase I enzyme, breaks down neutrophil extracellular traps (NET) that include a vast amount of DNA fragments, histones, microbicidal proteins and oxidant enzymes released from necrotic neutrophils in the highly viscous mucus of cystic fibrosis patients. Dornase alfa has been used for decades in patients with cystic fibrosis to reduce the viscoelasticity of respiratory tract secretions, to decrease the severity of respiratory tract infections, and to improve lung function. Previous studies have linked abnormal NET formations to lung diseases, especially to acute respiratory distress syndrome (ARDS). Coronavirus disease 2019 (COVID-19) pandemic affected more than two million people over the world, resulting in unprecedented health, social and economic crises. The COVID-19, viral pneumonia that progresses to ARDS and even multiple organ failure, is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). High blood neutrophil levels are an early indicator of SARS-CoV-2 infection and predict severe respiratory diseases. A similar mucus structure is detected in COVID-19 patients due to the accumulation of excessive NET in the lungs. Here, we show our preliminary results with dornase alfa that may have an in-vitro anti-viral effect against SARS-CoV-2 infection in a bovine kidney cell line, MDBK without drug toxicity on healthy adult peripheral blood mononuclear cells. In this preliminary study, we also showed that dornase alfa can promote clearance of NET formation in both an in-vitro and three COVID-19 cases who showed clinical improvement in radiological analysis (2-of-3 cases), oxygen saturation (SpO2), respiratory rate, disappearing of dyspnea and coughing.

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Synergistic Activity of Berberine with Azithromycin against Pseudomonas Aeruginosa Isolated from Patients with Cystic Fibrosis of Lung In Vitro and In Vivo

Cellular Physiology and Biochemistry ◽

10.1159/000479411 ◽

2017 ◽

Vol 42 (4) ◽

pp. 1657-1669 ◽

Cited By ~ 10

Author(s):

YongTao Li ◽

JianRong Huang ◽

LanJuan Li ◽

LinSheng Liu

Keyword(s):

Cystic Fibrosis ◽

Pseudomonas Aeruginosa ◽

Lung Infection ◽

Extracellular Dna ◽

Pcr Analysis ◽

Infection Model ◽

Synergistic Activity ◽

Time Kill

Background/Aims: Pseudomonas aeruginosa (PA) is one of the major opportunistic pathogens which can cause chronic lung infection of cystic fibrosis (CF). The formation of PA biofilm promotes CF development and restricts the antimicrobial efficacies of current antibiotics. Methods: The antimicrobial effects of azithromycin (AZM) and berberine (BER) alone and in combination were evaluated using microdilution method, checkerboard assay, time-kill test, qRT-PCR analysis and absorption method. The treatments of AZM and/or BER were further evaluated in an animal lung infection model via observing survival rate, bacterial burden and histopathology of lung, the levels of pro-/anti-inflammatory cytokines. Results: AZM-BER were demonstrated to be synergistic against ten clinical PA isolates as well as the standard reference PA ATCC27853, in which PA03 was the most susceptible isolate to AZM-BER with FICI of 0.13 and chosen for subsequent experiments. The synergism of AZM-BER was further confirmed against PA03 in time-kill test and scanning electron microscope (SEM) at their concentrations showing synergism. In PA03, we found that AZM-BER could significantly attenuate productions of a series of virulence factors including alginate, LasA protease, LasB protease, pyoverdin, pyocyanin, chitinase as well as extracellular DNA, and remarkably inhibit the levels of quorum sensing (QS) molecules and the expressions of lasI, lasR, rhlI, rhlR at 1/2×MIC, 1×MIC and 2×MIC. In the infection model, the mice survival were increased markedly, the inflammations of infected lungs were improved greatly along with reduced IL-6, IL-8 and ascended IL-10 at 0.8 mg/kg of AZM combined with 3.2 mg/kg of BER. Conclusion: BER might be a promising synergist to enhance the antimicrobial activity of AZM in vitro and in vivo.

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Unfractionated heparin reduces the elasticity of sputum from patients with cystic fibrosis

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00206.2007 ◽

2007 ◽

Vol 293 (5) ◽

pp. L1240-L1249 ◽

Cited By ~ 35

Author(s):

Victoria J. Broughton-Head ◽

Jagdeep Shur ◽

Mary P. Carroll ◽

James R. Smith ◽

Janis K. Shute

Keyword(s):

Cystic Fibrosis ◽

Unfractionated Heparin ◽

Laser Scanning ◽

Laser Scanning Microscopy ◽

Confocal Laser ◽

Boyden Chamber ◽

Target Cells ◽

Maximal Effect ◽

Dornase Alfa

Mucus obstruction of the airway in patients with cystic fibrosis (CF) reduces lung function, invites infection, and limits delivery of inhaled drugs including gene therapy vectors to target cells. Not all patients respond to presently available mucolytics, and new approaches are needed. Our objectives were to investigate the in vitro effects of unfractionated heparin (UFH) on the morphology and rheology of sputum and the effect of UFH on diffusion of 200-nm nanospheres through sputum from adult CF patients. Confocal laser scanning microscopy was used to image fluorescently stained actin and DNA components of CF sputum, and atomic force microscopy was used to image isolated DNA networks. The viscoelasticity of CF sputum was measured using dynamic oscillatory rheometry. Nanosphere diffusion was measured through CF sputum using a Boyden chamber-based assay. Actin-DNA bundles in CF sputum were disaggregated by UFH at concentrations of 0.1–10 mg/ml, and UFH enhanced the endonuclease activity in sputum from patients on dornase alfa therapy. UFH significantly reduced the elasticity and yield stress, but not the viscosity, of CF sputum from patients not receiving dornase alfa therapy. Heparin dose-dependently significantly increased the diffusion of nanospheres through CF sputum from patients not on dornase alfa therapy from 10.5 ± 2.5% at baseline to 36.9 ± 4.4% at 10 mg/ml but was more potent, with maximal effect at 0.1 mg/ml, in patients who were on dornase alfa therapy. Thus the mucoactive properties of UFH indicate its potential as a new therapeutic approach in patients with cystic fibrosis.

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Mixed Communities of Mucoid and NonmucoidPseudomonas aeruginosaExhibit Enhanced Resistance to Host Antimicrobials

mBio ◽

10.1128/mbio.00275-18 ◽

2018 ◽

Vol 9 (2) ◽

Cited By ~ 30

Author(s):

Sankalp Malhotra ◽

Dominique H. Limoli ◽

Anthony E. English ◽

Matthew R. Parsek ◽

Daniel J. Wozniak

Keyword(s):

Cystic Fibrosis ◽

Extracellular Dna ◽

Cationic Antimicrobial Peptide ◽

Alginate Production ◽

Enhanced Resistance ◽

Extracellular Release ◽

Mucoid Conversion ◽

Mixed Populations

ABSTRACTPseudomonas aeruginosacauses chronic pulmonary infections in patients with cystic fibrosis (CF).P. aeruginosamucoid conversion, defined by overproduction of the exopolysaccharide alginate, correlates with accelerated decline in CF patient lung function. Recalcitrance of the mucoid phenotype to clearance by antibiotics and the immune response is well documented. However, despite advantages conferred by mucoidy, mucoid variants often revert to a nonmucoid phenotype bothin vitroandin vivo. Mixed populations of mucoid isolates and nonmucoid revertants are recovered from CF lungs, suggesting a selective benefit for coexistence of these variants. In this study, cocultures of mucoid and nonmucoid variants exhibited enhanced resistance to two host antimicrobials: LL-37, a cationic antimicrobial peptide, and hydrogen peroxide (H2O2). Alginate production by mucoid isolates protected nonmucoid variants in consortia from LL-37, as addition of alginate exogenously to nonmucoid variants abrogated LL-37 killing. Conversely, nonmucoid revertants shielded mucoid variants from H2O2stress via catalase (KatA) production, which was transcriptionally repressed by AlgT and AlgR, central regulators of alginate biosynthesis. Furthermore, extracellular release of KatA by nonmucoid revertants was dependent onlys, encoding an endolysin implicated in autolysis and extracellular DNA (eDNA) release. Overall, these data provide a rationale to study interactions ofP. aeruginosamucoid and nonmucoid variants as contributors to evasion of innate immunity and persistence within the CF lung.IMPORTANCEP. aeruginosamucoid conversion within lungs of cystic fibrosis (CF) patients is a hallmark of chronic infection and predictive of poor prognosis. The selective benefit of mixed populations of mucoid and nonmucoid variants, often isolated from chronically infected CF patients, has not been explored. Here, we show that mixed-variant communities ofP. aeruginosademonstrate advantages in evasion of innate antimicrobials via production of shared goods: alginate and catalase. These data argue for therapeutically targeting multiple constituents (both mucoid and nonmucoid variants) within diversifiedP. aeruginosacommunitiesin vivo, as these variants can differentially shield one another from components of the host response.

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Preliminary Report of In vitro and In vivo Effectiveness of Dornase Alfa on SARS-CoV-2 Infection (Preprint)

10.2196/preprints.20121 ◽

2020 ◽

Author(s):

Hacer Kuzu Okur ◽

Koray Yalcin ◽

Cihan Tastan ◽

Sevda Demir ◽

Bulut Yurtsever ◽

...

Keyword(s):

Cystic Fibrosis ◽

Respiratory Tract ◽

Mononuclear Cells ◽

Multiple Organ ◽

Peripheral Blood Mononuclear ◽

Dornase Alfa ◽

Tract Infections ◽

Adult Peripheral Blood

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Clinical and in vitro Effect of Dornase Alfa in Mechanically Ventilated Pediatric Non-Cystic Fibrosis Patients with Atelectases

Cellular Physiology and Biochemistry ◽

10.1159/000204109 ◽

2009 ◽

Vol 23 (1-3) ◽

pp. 205-210 ◽

Cited By ~ 16

Author(s):

Joachim Riethmueller ◽

Matthias Kumpf ◽

Thomas Borth-Bruhns ◽

Wolfgang Brehm ◽

Jakub Wiskirchen ◽

...

Keyword(s):

Cystic Fibrosis ◽

Mechanically Ventilated ◽

Dornase Alfa

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Comparative In Vitro Activities of Meropenem, Imipenem, Temocillin, Piperacillin, and Ceftazidime in Combination with Tobramycin, Rifampin, or Ciprofloxacin against Burkholderia cepacia Isolates from Patients with Cystic Fibrosis

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.43.2.213 ◽

1999 ◽

Vol 43 (2) ◽

pp. 213-217 ◽

Cited By ~ 47

Author(s):

Stephane Bonacorsi ◽

Frederic Fitoussi ◽

Sylvie Lhopital ◽

Edouard Bingen

Keyword(s):

Cystic Fibrosis ◽

Synergistic Effect ◽

Bactericidal Activity ◽

Burkholderia Cepacia ◽

Drug Combinations ◽

Killing Activity ◽

Time Kill

We evaluated the activities of meropenem, imipenem, temocillin, piperacillin, and ceftazidime by determination of the MICs for 66 genotypically characterized Burkholderia cepacia isolates obtained from the sputum of cystic fibrosis patients. In vitro synergy assays, as performed by the time-kill methodology, of two- and three-drug combinations of the β-lactams with tobramycin, rifampin, and/or ciprofloxacin were also performed with 10 strains susceptible, intermediate, or resistant to fluoroquinolones. On the basis of the MICs, meropenem and temocillin were the most active β-lactam agents, with MICs at which 90% of isolates are inhibited of 8 and 32 μg/ml, respectively. The addition of ciprofloxacin significantly enhanced the killing activities of piperacillin, imipenem, and meropenem against the 10 strains tested (P < 0.05). The best killing activity was obtained with the combination of meropenem and ciprofloxacin, with bactericidal activity of 3.31 ± 0.36 log10 CFU/ml (P < 0.05). Compared to the activity of the two-drug β-lactam–ciprofloxacin combination, the addition of rifampin or tobramycin did not significantly increase the killing activity (P > 0.05). The three-drug combinations (with or without ciprofloxacin) significantly enhanced the killing activities of piperacillin, imipenem, and meropenem relative to the activities of the β-lactams used alone (P < 0.05). The combination β-lactam–ciprofloxacin–tobramycin was the combination with the most consistently synergistic effect.

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