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Journal of Translational Medicine Mar 2024Thrombo-inflammation and neutrophil extracellular traps (NETs) are exacerbated in severe cases of COVID-19, potentially contributing to disease exacerbation. However,...
BACKGROUND
Thrombo-inflammation and neutrophil extracellular traps (NETs) are exacerbated in severe cases of COVID-19, potentially contributing to disease exacerbation. However, the mechanisms underpinning this dysregulation remain elusive. We hypothesised that lower DNase activity may be associated with higher NETosis and clinical worsening in patients with COVID-19.
METHODS
Biological samples were obtained from hospitalized patients (15 severe, 37 critical at sampling) and 93 non-severe ambulatory cases. Our aims were to compare NET biomarkers, functional DNase levels, and explore mechanisms driving any imbalance concerning disease severity.
RESULTS
Functional DNase levels were diminished in the most severe patients, paralleling an imbalance between NET markers and DNase activity. DNase1 antigen levels were higher in ambulatory cases but lower in severe patients. DNase1L3 antigen levels remained consistent across subgroups, not rising alongside NET markers. DNASE1 polymorphisms correlated with reduced DNase1 antigen levels. Moreover, a quantitative deficiency in plasmacytoid dendritic cells (pDCs), which primarily express DNase1L3, was observed in critical patients. Analysis of public single-cell RNAseq data revealed reduced DNase1L3 expression in pDCs from severe COVID-19 patient.
CONCLUSION
Severe and critical COVID-19 cases exhibited an imbalance between NET and DNase functional activity and quantity. Early identification of NETosis imbalance could guide targeted therapies against thrombo-inflammation in COVID-19-related sepsis, such as DNase administration, to avert clinical deterioration.
TRIAL REGISTRATION
COVERAGE trial (NCT04356495) and COLCOV19-BX study (NCT04332016).
Topics: Humans; Extracellular Traps; Neutrophils; Deoxyribonucleases; COVID-19; Deoxyribonuclease I; Inflammation; Nervous System Diseases
PubMed: 38454482
DOI: 10.1186/s12967-024-05044-7 -
Frontiers in Cellular and Infection... 2022Anti-infection strategies against pleural empyema include the use of antibiotics and drainage treatments, but bacterial eradication rates remain low. A major challenge...
Anti-infection strategies against pleural empyema include the use of antibiotics and drainage treatments, but bacterial eradication rates remain low. A major challenge is the formation of biofilms in the pleural cavity. DNase has antibiofilm efficacy , and intrapleural therapy with DNase is recommended to treat pleural empyema, but the relevant mechanisms remain limited. Our aim was to investigate whether DNase I inhibit the early biofilm formation in - or -induced empyema models. We used various assays, such as crystal violet staining, confocal laser scanning microscopy (CLSM) analysis, peptide nucleic acid-fluorescence hybridization (PNA-FISH), and scanning electron microscopy (SEM) analysis. Our results suggested that DNase I significantly inhibited early biofilm formation in a dose-dependent manner, without affecting the growth of or . CLSM analysis confirmed that DNase I decreased the biomass and thickness of both bacterial biofilms. The PNA-FISH and SEM analyses also revealed that DNase I inhibited early (24h) biofilm formation in two empyema models. Thus, the results indicated that DNase inhibited early (24h) biofilm formation in - or -induced rabbit empyema models and showed its therapeutic potential against empyema biofilms.
Topics: Animals; Rabbits; Pseudomonas aeruginosa; Staphylococcus aureus; Deoxyribonucleases; In Situ Hybridization, Fluorescence; Staphylococcal Infections; Biofilms; Anti-Bacterial Agents; Deoxyribonuclease I; Empyema, Pleural
PubMed: 36310876
DOI: 10.3389/fcimb.2022.917038 -
Journal of Biochemistry Oct 1980A simple purification method for pancreatic deoxyribonuclease I (DNase I) [EC 3.1.4.3] was developed by utilizing the technique of isoelectric focusing. The active...
A simple purification method for pancreatic deoxyribonuclease I (DNase I) [EC 3.1.4.3] was developed by utilizing the technique of isoelectric focusing. The active protein was resolved in to at least four forms with different isoelectric points; the major components a, b, and c had isoelectric points at pH 5.2, 4.9, and 4.8, respectively, and that of the minor component d was at 4.7. The four components (a, b, c, and d) exhibited peaks similar to those observed by Salnikow et al. after phosphocellulose chromatography (A, B, C, and D). The four components were all free from RNase and protease activities and were very stable at 0-2 degrees C for at least four weeks. Further, each of the four peaks exhibited a single protein band after polyacrylamide electrophoresis. DNase I-a antibody was prepared; it was very specific for DNase I and precipitated with the other components (b, c, and d). The mode of endonucleolytic action of pancreatic DNase I-a purified from Worthington DP grade DNase I was investigated. The sedimentation patterns in neutral sucrose gradients of digest of circular duplex DNA in an early stage of hydrolysis suggested that DNase I produces single strand scissions in the initial attack in the presence of divalent metal ions.
Topics: Animals; Cattle; Deoxyribonuclease I; Deoxyribonucleases; Endonucleases; Immune Sera; Immunoassay; Immunodiffusion; Pancreas; Ribonucleases
PubMed: 6256339
DOI: 10.1093/oxfordjournals.jbchem.a133064 -
Journal of Biochemistry Dec 1977Human pancreatic DNase I was purified extensively from duodenal juice of healthy subjects by a procedure including ammonium sulfate fractionation, ethanol fractionation,...
Human pancreatic DNase I was purified extensively from duodenal juice of healthy subjects by a procedure including ammonium sulfate fractionation, ethanol fractionation, phosphocellulose fractionation, isoelectric focusing, and gel filtration. The final preparation was free of DNase II, pancreatic RNase, alkaline phosphatase, and protease. The enzyme had a molecular weight of approximately 30,000, as determined by gel filtration on Sephadex G-100, and showed maximum activity at pH 7.2-7.6. It required divalent cations for activity, and caused single-strand breaks by endonucleolytic attack on double- as well as single-stranded DNA molecules. The enzyme was inhibited by actin and bovine pancreatic DNase I antibody.
Topics: Deoxyribonucleases; Humans; Immunodiffusion; Molecular Weight; Pancreas
PubMed: 413831
DOI: 10.1093/oxfordjournals.jbchem.a131875 -
International Journal of Molecular... Feb 2023Deoxyribonucleases (DNases) cleave extracellular DNA (ecDNA) and are under intense research as interventions for diseases associated with high ecDNA, such as acute live...
Deoxyribonucleases (DNases) cleave extracellular DNA (ecDNA) and are under intense research as interventions for diseases associated with high ecDNA, such as acute live injury. DNase I treatment decreases morbidity and mortality in this animal model. Endogenous DNase activity has high interindividual variability. In this study, we tested the hypothesis that high endogenous DNase activity is beneficial in an animal model of acute liver failure. DNase activity was measured in the plasma of adult male mice taken before i.p. injection of thioacetamide to induce acute liver failure. The survival of mice was monitored for 48 h. Mice were retrospectively divided into two groups based on the median DNase activity assessed using the gel-based single-radial enzyme diffusion assay. In acute liver failure, mice with a higher baseline DNase activity had lower mortality after 48 h (by 25%). Different protection of ecDNA against nucleases by vesicles or DNA-binding proteins could play a role and should be further evaluated. Similarly, the role of endogenous DNase activity should be analyzed in other disease models associated with high ecDNA.
Topics: Male; Mice; Animals; Deoxyribonucleases; Retrospective Studies; DNA; Deoxyribonuclease I; Models, Animal; Liver Failure, Acute
PubMed: 36769306
DOI: 10.3390/ijms24032984 -
Proceedings of the National Academy of... Mar 1974A system for genetic exchange in Rhodopseudomonas capsulata has been discovered. Each genetic marker thus far examined can be transferred, and many strains of Rps....
A system for genetic exchange in Rhodopseudomonas capsulata has been discovered. Each genetic marker thus far examined can be transferred, and many strains of Rps. capsulata can participate in genetic exchange. The mechanism of gene transfer seems unlike that of any previously described bacterial system, since genes can be transferred by cell-free filtrates, but the vector is resistant to deoxyribonuclease and has a sedimentation constant of about 70 S.
Topics: Cell-Free System; Deoxyribonucleases; Drug Resistance, Microbial; Genetics, Microbial; Phenotype; Recombination, Genetic; Rhodopseudomonas; Rifampin; Streptomycin
PubMed: 4522805
DOI: 10.1073/pnas.71.3.971 -
The Journal of Antibiotics Feb 1982
Topics: Agar; Deoxyribonucleases; Methods
PubMed: 7076568
DOI: 10.7164/antibiotics.35.245 -
FEBS Letters Feb 1999A RecA/Rad51 homologue from Pyrococcus kodakaraensis KOD1 (Pk-REC) is the smallest protein among various RecA/Rad51 homologues. Nevertheless, Pk-Rec is a super...
A RecA/Rad51 homologue from Pyrococcus kodakaraensis KOD1 (Pk-REC) is the smallest protein among various RecA/Rad51 homologues. Nevertheless, Pk-Rec is a super multifunctional protein and shows a deoxyribonuclease activity. This deoxyribonuclease activity was inhibited by 3 mM or more ATP, suggesting that the catalytic centers of the ATPase and deoxyribonuclease activities are overlapped. To examine whether these two enzymatic activities share the same active site, a number of site-directed mutations were introduced into Pk-REC and the ATPase and deoxyribonuclease activities of the mutant proteins were determined. The mutant enzyme in which double mutations Lys-33 to Ala and Thr-34 to Ala were introduced, fully lost both of these activities, indicating that Lys-33 and/or Thr-34 are important for both ATPase and deoxyribonuclease activities. The mutation of Asp-112 to Ala slightly and almost equally reduced both ATPase and deoxyribonuclease activities. In addition, the mutation of Glu-54 to Gln did not seriously affect the ATPase, deoxyribonuclease, and UV tolerant activities. These results strongly suggest that the active sites of the ATPase and deoxyribonuclease activities of Pk-REC are common. It is noted that unlike Glu-96 in Escherichia coli RecA, which has been proposed to be a catalytic residue for the ATPase activity, the corresponding residual Glu-54 in Pk-REC is not involved in the catalytic function of the protein.
Topics: Adenosine Triphosphatases; DNA-Binding Proteins; Deoxyribonucleases; Mutagenesis, Site-Directed; Pyrococcus; Rad51 Recombinase; Rec A Recombinases
PubMed: 10069383
DOI: 10.1016/s0014-5793(99)00107-6 -
European Journal of Biochemistry Aug 2000An extracellular nuclease from Basidiobolus haptosporus (designated as nuclease Bh1) was purified to homogeneity by ammonium sulfate precipitation, heat treatment,...
Purification and characterization of the single-strand-specific and guanylic-acid-preferential deoxyribonuclease activity of the extracellular nuclease from Basidiobolus haptosporus.
An extracellular nuclease from Basidiobolus haptosporus (designated as nuclease Bh1) was purified to homogeneity by ammonium sulfate precipitation, heat treatment, negative adsorption on DEAE-cellulose, and chromatography on phenyl-Sepharose followed by FPLC on phenyl-Superose. The overall yield was 26%. The Mr of the purified enzyme, determined by gel filtration, was 41 000 whereas by SDS/PAGE (after deglycosylation) it was 30 000. It is a glycoprotein with a pI of 6.8. The optimum pH and temperature for DNA hydrolysis were 8. 5 and 60 degrees C, respectively. Nuclease Bh1 is a metalloprotein but has no obligate requirement for metal ions to be active, nor is its activity stimulated in the presence of metal ions. The enzyme was inhibited by Zn2+, Ag2+, Hg2+, Fe3+ and Al3+, inorganic phosphate, pyrophosphate, dithiothreitol, 2-mercaptoethanol, NaCl and KCl. It was stable to high concentrations of organic solvents and urea but susceptible to low concentrations of SDS and guanidine hydrochloride. Nuclease Bh1 is a multifunctional enzyme and its substrate specificity is in the order of ssDNA approximately 3'AMP >> RNA > dsDNA. Studies on its mode of action showed that it cleaved supercoiled pUC 18 DNA and phage M13 DNA, endonucleolytically, generating single base nicks. The enzyme hydrolyzed DNA with preferential liberation of 5'dGMP, suggesting it to be a guanylic acid preferential endoexonuclease. 5'dGMP, the end product of hydrolysis, was a competitive inhibitor of the enzyme. The absence of 5'dCMP as a hydrolytic product, coupled with the resistance of (dC)10 and deoxyribodinucleoside monophosphates having cytosine either at the 3' or the 5' end, indicates that C-linkages are resistant to cleavage by nuclease Bh1.
Topics: Amino Acid Sequence; Cations; Chromatography, DEAE-Cellulose; Chromatography, Gel; Chromatography, Ion Exchange; Deoxyribonucleases; Endonucleases; Entomophthorales; Enzyme Stability; Exonucleases; Fungal Proteins; Hot Temperature; Kinetics; Molecular Sequence Data; Molecular Weight; Sequence Alignment; Sequence Homology, Amino Acid; Thermodynamics
PubMed: 10931196
DOI: 10.1046/j.1432-1327.2000.01580.x -
Oncogene Sep 2010Chemotherapy and radiotherapy commonly damage DNA and trigger p53-dependent apoptosis through intrinsic apoptotic pathways. Two unfortunate consequences of this...
Chemotherapy and radiotherapy commonly damage DNA and trigger p53-dependent apoptosis through intrinsic apoptotic pathways. Two unfortunate consequences of this mechanism are resistance due to blockade of p53 or intrinsic apoptosis pathways, and mutagenesis of non-malignant surviving cells which can impair cellular function or provoke second malignancies. Death ligand-based drugs, such as tumor necrosis factor-related apoptosis inducing ligand (TRAIL), stimulate extrinsic apoptotic signaling, and may overcome resistance to treatments that induce intrinsic apoptosis. As death receptor ligation does not damage DNA as a primary mechanism of pro-apoptotic action, we hypothesized that surviving cells would remain genetically unscathed, suggesting that death ligand-based therapies may avoid some of the adverse effects associated with traditional cancer treatments. Surprisingly, however, treatment with sub-lethal concentrations of TRAIL or FasL was mutagenic. Mutations arose in viable cells that contained active caspases, and overexpression of the caspase-8 inhibitor crmA or silencing of caspase-8 abolished TRAIL-mediated mutagenesis. Downregulation of the apoptotic nuclease caspase-activated DNAse (CAD)/DNA fragmentation factor 40 (DFF40) prevented the DNA damage associated with TRAIL treatment. Although death ligands do not need to damage DNA in order to induce apoptosis, surviving cells nevertheless incur DNA damage after treatment with these agents.
Topics: Animals; Antineoplastic Agents; Cell Line, Tumor; Cell Survival; Cells, Cultured; Cisplatin; Deoxyribonucleases; Drug Resistance, Neoplasm; Fibroblasts; Mice; Models, Biological; Mutation; Neoplasms; RNA, Small Interfering; TNF-Related Apoptosis-Inducing Ligand
PubMed: 20639907
DOI: 10.1038/onc.2010.242