DNase Research Articles

Physiological strength of lipoic acid in copper intoxication

IntroductionThe average concentration of copper in the air, e.g. in the USA it ranges from 5–20 ng/m3, in soil from 5–70 mg/kg, and the intake of copper from food is 1.0–1.3 mg/days for adults (0.014–0.019 mg/kg/day) (Barceloux, 1999). The effect of lipoic acid is reflected in the intensification of ATP synthesis, participates in the assimilation of lactic acid, activates the enzyme cycle of tricarboxylic acid, stimulates the growth of lactic acid bacteria by replacing acetate (acetate transfer factor), stimulates CoA synthesis (fatty acid utilization), prevents liver damage by various toxins, normalizes aldolase and transferase levels.ObjectivesThe aim of this study is to show the useful role of a supplement, lipoic acid, as an antioxidant in the prevention of oxidative stress.MethodsAll procedures were performed after anesthesia of albino rats with ketal in accordance with the principles of sacrifice in laboratories. After medial laparotomy albino rates Wistar soy, a 10% homogenate of brain tissue was made in an appropriate medium and an analysis of acid and alkaline DNase activity was performed (Kocić i sar., 2004).ResultsDNases are thought to be the main executors of apoptosis, responsible for internucleosomal DNA fragmentation, which is the breakdown of chromosomal DNA into oligonucleosome-sized fragments. Administration of lipoic acid has been shown to protect against oxidative stress caused by copper.ConclusionsBased on the results of this research, it can be concluded that lipoic acid is a powerful and powerful antioxidant.DisclosureNo significant relationships.

Effects of nucleases on cell-free extrachromosomal circular DNA.

Cell-free extrachromosomal circular DNA (eccDNA) as a distinct topological form from linear DNA has recently gained increasing research interest, with possible clinical applications as a class of biomarkers. In this study, we aimed to explore the relationship between nucleases and eccDNA characteristics in plasma. By using knockout mouse models with deficiencies in deoxyribonuclease 1 (DNASE1) or deoxyribonuclease 1 like 3 (DNASE1L3), we found that cell-free eccDNA in Dnase1l3−/− mice exhibited larger size distributions than that in wild-type mice. Such size alterations were not found in tissue eccDNA of either Dnase1−/− or Dnase1l3−/− mice, suggesting that DNASE1L3 could digest eccDNA extracellularly but did not seem to affect intracellular eccDNA. Using a mouse pregnancy model, we observed that in Dnase1l3−/− mice pregnant with Dnase1l3+/− fetuses, the eccDNA in the maternal plasma was shorter compared with that of Dnase1l3−/− mice carrying Dnase1l3−/− fetuses, highlighting the systemic effects of circulating fetal DNASE1L3 degrading the maternal eccDNA extracellularly. Furthermore, plasma eccDNA in patients with DNASE1L3 mutations also exhibited longer size distributions than that in healthy controls. Taken together, this study provided a hitherto missing link between nuclease activity and the biological manifestations of eccDNA in plasma, paving the way for future biomarker development of this special form of DNA molecules.

Identification and characterization of Deoxyribonuclease II in planarian Dugesia japonica

Deoxyribonuclease II (DNase II) has been found to regulate inflammation, autoimmunity and apoptosis in vertebrates and invertebrates. The strong capacity of degrading DNA makes DNase II play an important role in the immune process. Planarian has become one of the model references due to its strong immune system, the environment they live makes planarians face the threat of microorganisms and injury, the strong immune system can protect planarians from the threat of bacterial and infection. In this study, we found that there was DNase in the lysis buffer of planarians, then we acquired the sequence of DjDN2s (Dugesia japonica DNase2s) and confirmed the DjDN2s were conserved DNase IIs. The predicted structure showed the active sites and binding patterns of DjDN2s. Whole-mount in situ hybridization results showed DjDN2s mainly expressed in immune organs. Quantitative real-time PCR revealed that the expression of DjDN2s upregulated in varying degrees when got hurt and challenged with bacteria, and the knockdown of DjDN2s led to the slower repair of wound. The recombinant phages which take DjDN2 also had the ability to degrade DNA and clear young biofilm of Gram-negative bacteria. Collectively, DNase II of planarian might play a role in the antimicrobial response and wound-induced response.

Contribution of Impaired DNASE1L3 Activity to Anti-DNA Autoantibody Production in Systemic Lupus Erythematosus.

Anti-DNA autoantibodies are pathogenic in systemic lupus erythematosus (SLE). Cell-free chromatin associated long DNA fragments are antigens for anti-DNA antibodies. In health state, released by cell death and actively secreted by live cells, these cell-free DNA are cleared by deoxyribonucleases (DNASES). In SLE, cell-free DNA are accumulated. The defective clearance of long fragments of cell-free DNA in SLE is largely attributed to impaired deoxyribonuclease 1 like 3 (DNASE1L3). DNASE1L3 null mutation results in monogenic SLE. The SLE risk single-nucleotide polymorphism (rs35677470) encodes R260C variant DNASE1L3, which is defective in secretion, leading to reduced levels of DNASE1L3. In addition, neutralizing autoantibodies to DNASE1L3 are produced in SLE to inhibit its enzymatic activity.

Deoxyribonuclease (DNAse) and Its Inhibitor

Deoxyribonuclease (DNAse) is an enzyme that plays an important role in cell apoptosis and has the function of hydrolyzing DNA. There are two types of DNAse, namely i) DNAse I which works well at neutral pH (6.5-8.0) and requires bivalent ions such as magnesium (Mg2+) and calcium (Ca2+) for activation, and ii) DNAse II which works by good at acidic pH and does not require bivalent ions. DNAse inhibitors are required to control the activity of DNAse. Both DNAse and its inhibitors have a role in the pathogenesis of various diseases, such as in the autoimmune disease systemic lupus erythematosus. DNase inhibitors can be obtained from natural, conventional, or chemical sources.

Occurrence of Multidrug-Resistant Staphylococcus aureus among Humans, Rodents, Chickens, and Household Soils in Karatu, Northern Tanzania.

We conducted this study to investigate the isolation frequency and phenotypic antibiotic resistance pattern of Staphylococcus aureus isolated from rodents, chickens, humans, and household soils. Specimens were plated onto mannitol salt agar (Oxoid, Basingstoke, UK) and incubated aerobically at 37 °C for 24 h. Presumptive colonies of S. aureus were subjected to Gram staining, as well as catalase, deoxyribonuclease (DNAse), and coagulase tests for identification. Antibiotic susceptibility testing was performed by using the Kirby–Bauer disc diffusion method on Mueller–Hinton agar (Oxoid, Basingstoke, UK). The antibiotics tested were tetracycline (30 μg), erythromycin (15 μg), gentamicin (10 μg), ciprofloxacin (5 μg), clindamycin (2 μg), and amoxicillin-clavulanate (20 μg/10 μg). The S. aureus strain American Type Culture Collection (ATCC) 25,923 was used as the standard organism. We found that 483 out of 956 (50.2%) samples were positive for S. aureus. The isolation frequencies varied significantly between samples sources, being 52.1%, 66.5%, 74.3%, and 24.5%, respectively, in chickens, humans, rodents, and soil samples (p < 0.001). S. aureus isolates had high resistance against clindamycin (51.0%), erythromycin (50.9%), and tetracycline (62.5%). The overall prevalence of multidrug-resistant (MDR) S. aureus isolates was 30.2%, with 8.7% resistant to at least four different classes of antibiotics.

Levels of marker lysosomal hydrolases in women with coronary heart disease depending on age and sex hormone level

Aim of the study was to evaluate the serum concentrations of three marker lysosomal hydrolases (cathepsin D, acid phosphatase (AP) and acid DNase (aDNAase)) in women with coronary heart disease (CHD) depending on the level of follicle-­stimulating hormone (FSH), testosterone (T), age and find if those parameters associated with anthropometric parameters, glycemia, insulinemia and HOMA-IR index, biomarkers of atherosclerosis. The study included 285 women aged 35–65 years (median age was 54.4 years (25% and 75% percentiles — 43.2 and 61.3 years, respectively) who had had myocardial infarction no earlier than 30 days before the examination. Patients were divided into the following age groups: 35–55 and 56–65 years (first and second age groups, respectively), and into groups according to the levels of sex hormones: FSH ≥ and &lt;30 mIU/mL and testosterone ≥ and &lt;3 nmol/L. Results of comparative and correlation analyzes demonstrates that in women 35–65 years old with FSH ≥30 mIU/mL, the levels of cathepsin D are higher (p&lt;0.05) than in patients with FSH &lt;30 mIU/mL, and in women 35–55 years old, the content of AP was also higher (p=0.025). Associations of a high level of androgen with lysosomal hyperenzymemia were demonstrated only in the second age group, where at a level of T ≥3 nmol/L, higher values of all three lysosomal enzymes were recorded. Multivariate analysis in both age groups is confirmed direct impact of periand postmenopausal periods on the levels of lysosomal enzymemia and, accordingly, a negative effect on the state of lysosomal membranes. Thus, FSH levels directly determined the concentrations of AP and cardiotropic cathepsin D. The levels of aDNAase in women with CHD of 56–65 years of age were positively correlated with indicators that determine insulin-­glucose homeostasis: glycemia (p&lt;0.001), HOMA-IR index (p&lt;0.001). Such associations of three marker lysosomal enzymes demonstrate the primary contribution of FSH ≥30 mIU/mL to an increase in the concentration of lysosomal hydrolases in women with CHD35–65 years old and the correlation of aDNAase with the processes triggered by insulin resistance.

Glutathione as a powerful antioxidant in oxidative stress in the brain tissue of rats caused by the pathophysiological action of copper

IntroductionDue to increased human activity, the amount of copper in air, soil and water has increased. Copper, at minimum concentrations, is essential for the normal functioning of the organism (cellular respiration, hemoglobin formation, growth and reproduction). At higher concentrations, copper is deposited in the liver, brain tissue, and bone marrow.ObjectivesTo investigate the protective role of the supplement, glutathione (GSH), the S-donor ligand, in conditions of chronic copper intoxication via the parameters of oxidative stress, ie. Alkaline and acidic DNase values in brain tissue in albino rats of Wistar strain.MethodsThe model system for testing the effects of copper exposure and the protective effect of GSH was a study on female albino rats of Wistar strain, stored in the vivarium of the Scientific Research Center for Biomedicine, Faculty of Medicine, Niš, Serbia. Endonucleases, alkaline and acidic DNase activities were determined spectrophotometrically from homogenates of brain tissue.ResultsCopper is believed to be a likely cause of oxidative damage to the DNA molecule, as manifested by increased alkaline and acidic DNase activity. The results of this study show that GSH is a potent chelator that binds copper and enables its elimination from the body.ConclusionsIn this experiment, the beneficial role of GSH supplements, which has an antioxidant character, in the prevention and reduction of the adverse effects of chronic copper intoxication was demonstrated. In this way, GSH acts as a powerful protector and antioxidant.DisclosureNo significant relationships.

The Role of Nucleases and Nucleic Acid Editing Enzymes in the Regulation of Self-Nucleic Acid Sensing.

Detection of microbial nucleic acids by the innate immune system is mediated by numerous intracellular nucleic acids sensors. Upon the detection of nucleic acids these sensors induce the production of inflammatory cytokines, and thus play a crucial role in the activation of anti-microbial immunity. In addition to microbial genetic material, nucleic acid sensors can also recognize self-nucleic acids exposed extracellularly during turn-over of cells, inefficient efferocytosis, or intracellularly upon mislocalization. Safeguard mechanisms have evolved to dispose of such self-nucleic acids to impede the development of autoinflammatory and autoimmune responses. These safeguard mechanisms involve nucleases that are either specific to DNA (DNases) or RNA (RNases) as well as nucleic acid editing enzymes, whose biochemical properties, expression profiles, functions and mechanisms of action will be detailed in this review. Fully elucidating the role of these enzymes in degrading and/or processing of self-nucleic acids to thwart their immunostimulatory potential is of utmost importance to develop novel therapeutic strategies for patients affected by inflammatory and autoimmune diseases.

High-mobility group box protein-1 induces acute pancreatitis through activation of neutrophil extracellular trap and subsequent production of IL-1β

PurposeThe aim of this study is to evaluate acute pancreatitis (AP)-associated NET activation mediated by a novel inflammatory mediator (high-mobility group box protein-1 [HMGB1]) and proinflammatory cytokine responses. MethodsIn this study, primary neutrophils, monocytes, and monocytic cell line Thp-1-derived macrophages were isolated and treated with HMGB1, lipopolysaccharide (LPS), adenosine triphosphate (ATP), and ATP + ATP inhibitor. The effects of HMGB1, ATP, and deoxyribonuclease (DNAse) were then examined for their in vivo effects using a newly established AP mouse model. ResultsThe mRNA and protein levels of inflammasome and interleukin IL-1β in cells, blood, and pancreatic tissues were examined. Within-cell nuclear DNA signal, cell-free DNA concentration, and pancreatic tissue damage were investigated. Our study showed that HMGB1 triggers NET formation in neutrophils and promotes the activation of inflammasome complexes (the NLR family, pyrin domain containing 3, and NLRP3; ASC; and caspase-1); therefore, the production of IL-1β is induced in human monocytes/macrophages. HMGB1 and NET cooperatively stimulate IL-1β processing in macrophages. Furthermore, the AP mouse model confirmed these HMGB1-mediated molecular mechanisms in vivo and indicated that HMGB1 is required for NET activation. ConclusionsWe found that NET inhibition reverses HMGB1-stimulated inflammasome activation and IL-1β production. HMGB1 thus leads to pancreatic injury through the activation of NET and subsequently induces IL-1β processing from neutrophils to pancreatic tissues. These findings demonstrate that HMGB1 and NET are new therapeutic targets for inflammation suppression in severe AP.

Accelerated Evolution of the Regulatory Sequences of Brain Development in the Human Genome.

Genetic modifications in noncoding regulatory regions are likely critical to human evolution. Human-accelerated noncoding elements are highly conserved noncoding regions among vertebrates but have large differences across humans, which implies human-specific regulatory potential. In this study, we found that human-accelerated noncoding elements were frequently coupled with DNase I hypersensitive sites (DHSs), together with monomethylated and trimethylated histone H3 lysine 4, which are active regulatory markers. This coupling was particularly pronounced in fetal brains relative to adult brains, non-brain fetal tissues, and embryonic stem cells. However, fetal brain DHSs were also specifically enriched in deeply conserved sequences, implying coexistence of universal maintenance and human-specific fitness in human brain development. We assessed whether this coexisting pattern was a general one by quantitatively measuring evolutionary rates of DHSs. As a result, fetal brain DHSs showed a mixed but distinct signature of regional conservation and outlier point acceleration as compared to other DHSs. This finding suggests that brain developmental sequences are selectively constrained in general, whereas specific nucleotides are under positive selection or constraint relaxation simultaneously. Hence, we hypothesize that human- or primate-specific changes to universally conserved regulatory codes of brain development may drive the accelerated, and most likely adaptive, evolution of the regulatory network of the human brain.

Effects of sample processing modes on high-throughput sequencing of coronavirus whole genome

Objective To study the effects of different pre-sequencing sample processing modes on the results of whole genome sequencing with high-throughput sequencing (HTS) by taking the largest RNA virus (human coronavirus, HCoV) as the representative. Methods Cell-cultured human coronavirus HCoV-OC43 strains were used as the representative samples and divided into different groups based on pre-sequencing processing modes as follows: untreated group, DNase and RNase treatment before nucleic acid extraction group, DNase treatment after nucleic acid extraction group, and DNase and RNase treatment before nucleic acid extraction and DNase treatment after nucleic acid extraction group. Nucleic acid samples of each group were analyzed by direct RNA sequencing (without amplification) and DNA sequencing after sequence independent single primer amplification (SISPA), respectively. Results No significant difference in viral genome coverage rates was observed between different groups. The highest genome coverage and sequencing accuracy were obtained in DNase treatment after nucleic acid extraction group by direct RNA sequencing, and the ratio of viral reads and the sequencing depth of each locus were effectively improved by SISPA amplification. Conclusions This study provided an optimized technical strategy for whole genome sequencing of RNA viruses such as coronavirus. Key words: Human coronavirus; Sequence independent single primer amplification; Whole genome sequencing; High-throughput sequencing

The Biology of Cell-free DNA Fragmentation and the Roles of DNASE1, DNASE1L3, and DFFB

Cell-free DNA (cf.DNA) is a powerful noninvasive biomarker for cancer and prenatal testing, and it circulates in plasma as short fragments. To elucidate the biology of cf.DNA fragmentation, we explored the roles of deoxyribonuclease 1 (DNASE1), deoxyribonuclease 1 like 3 (DNASE1L3), and DNA fragmentation factor subunit beta (DFFB) with mice deficient in each of these nucleases. By analyzing the ends of cf.DNA fragments in each type of nuclease-deficient mice with those in wild-type mice, we show that each nuclease has a specific cutting preference that reveals the stepwise process of cf.DNA fragmentation. Essentially, we demonstrate that cf.DNA is generated first intracellularly with DFFB, intracellular DNASE1L3, and other nucleases. Then, cf.DNA fragmentation continues extracellularly with circulating DNASE1L3 and DNASE1. With the use of heparin to disrupt the nucleosomal structure, we also show that the 10 bp periodicity originates from the cutting of DNA within an intact nucleosomal structure. Altogether, this work establishes a model of cf.DNA fragmentation.

Crystal Structure of the Regulatory Domain of MexT, a Transcriptional Activator of the MexEFOprN Efflux Pump in Pseudomonas aeruginosa.

The Gram-negative opportunistic pathogen, Pseudomonas aeruginosa, has multiple multidrug efflux pumps. MexT, a LysR-type transcriptional regulator, functions as a transcriptional activator of the MexEF-OprN efflux system. MexT consists of an N-terminal DNA-binding domain and a C-terminal regulatory domain (RD). Little is known regarding MexT ligands and its mechanism of activation. We elucidated the crystal structure of the MexT RD at 2.0 Å resolution. The structure comprised two protomer chains in a dimeric arrangement. MexT possessed an arginine-rich region and a hydrophobic patch lined by a variable loop, both of which are putative ligand-binding sites. The three-dimensional structure of MexT provided clues to the interacting ligand structure. A DNase I footprinting assay of full-length MexT identified two MexT-binding sequence in the mexEF-oprN promoter. Our findings enhance the understanding of the regulation of MexT-dependent activation of efflux pumps.

THE ROLE OF ADRENOCORTICAL HORMONES IN THE REGULATION OF ORGANISM RESPONSE TO INTRATRACHEAL INJECTIONS OF ELECTROLYSIS DUST TO WISTAR RATS

Introduction. Adrenocortical hormones play a leading role in the adaptation of human organism to damaging factors. The aim of this study is to compare levels of cortisol and biochemical markers of organism damage (MOD) in blood samples of rats under a model of electrolysis dust (ED) inhalation exposure. ED being component of emissions in aluminum production and has high content of resinous substances (RS), including 3,4-benzpyrene. Material and methods. Male Wistar rats were injected ED intratracheally once per month, in doses corresponding to keeping of animals at ED concentrations of 1.0; 5.2 and 25.1 mg/m3, calculated on RS mass. 2 weeks after the 1st and 2nd ED administration and 6 days after the 3rd one, blood serum samples were used for determination of cortisol levels (ELISA) and the following 6 MOD: the intensity of luminol-enchanced chemiluminescence, activities of catalase, N-acetyl-β-D-glucosaminidase (NAG), acid DNAse, acetylesterase and γ-glutamyl transferase (GGT). Results. 2 weeks after the 1st and 2nd ED administrations (cortisol content 2 times reduced or not changed), similar significant changes were observed in almost all MOD; 2nd experimental point, if comparing with 1st point, had slight increase in oxidative stress and significant rise in GGT activity (a marker of precancer changes) at the maximum dose. On the contrary, 6 days after the 3rd injection of ED, in parallel with the increase of cortisol content in rat blood by 1.5 - 2 times, small significant changes were found for only one marker (NAG). The obtained data fit into existing concepts about protective role of cortisol and biphasic nature of its release into the blood, allow us to consider the absence of MOD, in parallel with rising levels of C, as a transit phase and have common problem aspects with CIRCI syndrome (Critical illness-Related Corticosteroid Insufficiency) in emergency medicine. Conclusion. Determination of cortisol levels by ELISA assay can be easily incorporated into any toxicological protocol as index of organism adaptive response, but further investigations are needed to clarify its characteristics and to build continuous model from separate time-dose points.

Comparison of the effect of tenofovir disoproxil fumarate and entecavir in the treatment of chronic hepatitis B with positive E antigen

Objective To compare the efficacy and safety of tenofovir disoproxil fumarate(TDF) and entecavir(ETV) in the treatment of chronic hepatitis B(CHB) with positive hepatitis B E antigen(HBeAg). Methods A total of 104 cases with newly diagnosed HBeAg positive CHB were selected and randomly divided into TDF group and ETV group, with 52 cases in each group.The TDF group was given 300mg/d TDF, and the ETV group was given 0.5mg/d ETV.All the patients were continuously treated for 12 months.The serum HBV DNA, HBeAg and ALT levels before and after treatment were compared between the two groups. Results Before treatment, there were no statistically significant differences in serum HBV DNA, HBeAg and ALT levels between the two groups (t=0.12, 1.51, 1.62, all P>0.05). The serum HBV DNA, HBeAg and ALT levels in the two groups were decreased after treatment, and the decrease of serum HBV DNA level in the TDF group was more significant than that in the ETV group, the difference was statistically significant(t=3.54, P 0.05). The total effective rate of the TDF group was 92.31%(48/52), which was significantly higher than 76.92%(40/52) in the ETV group (χ2=4.73, P 0.05). Conclusion TDF has better clinical effect in treating newly diagnosed HBeAg positive CHB than ETV due to TDF can inhibit HBV DNA replication significantly, but the safety of TDF and ETV is similar. Key words: Hepatitis B, chronic; Hepatitis B e antigens; Alanine transaminase; Deoxyribonucleases; Tenofovir; Entecavir; Drug therapy; Controlled Clinical trial

Optimizing Nanoparticle Design for Gene Therapy: Protection of Oligonucleotides from Degradation Without Impeding Release of Cargo.

Gene therapy delivery systems that rely on synthetic nanocarriers can be optimized by assays of nucleic acid protection and kinetic studies of nucleic acid release. These empirical measurements ensure nanoparticle stability and predict potential in vivo efficacy. Quantitative methods for assessment of the capacity of nanoparticles to protect oligonucleotide cargo and to measure the rate of release of the cargo were developed and tested based on six commercial cationic matrices. in vitro study of drug release kinetics provides predictable release rates under a variety of conditions which can be adapted to appropriate physiological factors that affect release in vivo. In brief, in vitro DNA release and DNase I degradation assays described here will be useful for optimization of nanocarrier-mediated gene therapy administration by various routes.

Reduced deoxyribonuclease enzyme activity in response to high postinjury mitochondrial DNA concentration provides a therapeutic target for Systemic Inflammatory Response Syndrome.

Cell-free mitochondrial DNA (mtDNA) is proinflammatory and has been detected in high concentrations in trauma patients' plasma. Deoxyribonuclease (DNAse) is the free plasma enzyme responsible for the digestion of extracellular DNA. The relationship between mtDNA and DNAse after major trauma is unknown. We hypothesized that DNAse activity would be elevated after injury and trauma surgery and would be associated with high concentrations of extracellular DNA. Two-year prospective study was performed on 103 consecutive trauma patients (male, 81%; age, 38 years [interquartile range, 30-59 years]; injury severity score, 18 [interquartile range, 12-26 years]) who underwent standardized major orthopedic trauma surgical interventions. Blood was collected at five perioperative time points (preoperative, postoperative, 7 hours, 24 hours, and 3 days postoperatively). Healthy control subjects (n = 20) were also sampled. Cell-free mtDNA and nuclear DNA (nDNA) were measured using quantitative polymerase chain reaction. Deoxyribonuclease was also assayed in the same plasma samples. Increased levels of mtDNA (from preoperative 163 ± 86 ng/mL to 3 days 282 ± 201 ng/mL, p < 0.0001) and nDNA (from preoperative 28 ± 20 ng/mL to 3 days 37 ± 27 ng/mL, p < 0.05) were present in trauma patients at all perioperative time points compared with healthy controls (mtDNA: 4 ± 2 ng/mL; nDNA: 10 ± 5 ng/mL). Deoxyribonuclease activity was lower in the trauma cohort (from preoperative 0.06 ± 0.04U/mL to 3 days 0.08 ± 0.04U/mL, p < 0.0001) compared with healthy controls (DNAse: 0.17 ± 0.03U/mL). There was no correlation between DNAse and perioperative DNA concentrations. Elevated mtDNA (but not nDNA) correlated with the development of systemic inflammatory response syndrome (SIRS) (p = 0.026) but not multiple organ failure. The significant perioperative elevation in plasma-free mtDNA concentration is associated with the development of SIRS. The fact that increased cell-free DNA concentrations present with significantly lower than healthy control DNAse activity suggests a potential therapeutic opportunity with DNAse administration to modulate postinjury severe SIRS. Prognostic/Epidemiological, level II.

Features of cellular mechanisms affecting the pathogenesis and course of chronic catarrhal gingivitis

Aim. To investigate the cellular mechanisms of chronic catarrhal gingivitis to develop the appropriate treatment schemes based on the analysis of obtained data. Methods. 57 patients aged 16 to 22 years were observed and divided into the control group (18 subjects with the intact periodontium with no orthopedic constructions) and the study group of patients with chronic catarrhal gingivitis (39 subjects, group 1). The degree of gingival inflammation was evaluated by papillo-marginal alveolar index as well as by gingival bleeding index. In addition, laboratory parameters were measured: spontaneous nitroblue tetrazolium test and zymosan-stimulated nitroblue tetrazolium test. Leukocyte-modulating activity of oral fluid was determined by measurement of luminol-dependent chemiluminescence. The activity of lysosomal enzymes - acid deoxyribonuclease and acid phosphatase - was determined in whole oral fluid, supernatant fraction of oral fluid and in sediment, obtained by centrifugation. Antioxidant activity was determined in whole oral fluid by biochemiluminescent method with hydrogen peroxide. Results. The performed investigations demonstrated that the development of chronic catarrhal gingivitis is accompanied by a sharp increase of the number of neutrophils and macrophages with a significant increase of their functional activity. At the same time, more than two-fold increase of the activity of lysosomal enzymes, lysosomal membrane labilization and release of enzymes outside the organelles and cells were observed, that indicated the increase of destructive potential in chronic catarrhal gingivitis. Use of indometacin in the complex treatment of chronic catarrhal gingivitis contributed to normalization of clinical parameters, increased neutrophil functional activity and enhanced their functional reserve, reduced the activity of phlogogenic factors and normalized the antioxidant activity of oral fluid. Pro- and anti-inflammatory factors secreted into the oral cavity during inflammatory periodontal diseases collectively have leukocyte-modulating activity. Conclusion. The obtained data indicate the important role of both high oxygen-dependent and oxygen-independent biocidal activity of neutrophils and macrophages in the pathogenesis and clinical course of chronic catarrhal gingivitis.

DNase II activated by the mitochondrial apoptotic pathway regulates RIP1-dependent non-apoptotic hepatocyte death via the TLR9/IFN-β signaling pathway.

Cell death, including apoptotic and non-apoptotic cell death, is frequently observed in liver disease. Upon activation of the mitochondrial apoptotic pathway, mitochondria release not only apoptogenic cytochrome c but also mitochondrial DNA (mtDNA) into the cytosol. The impact of DNase II, a lysosomal acid DNase that degrades mtDNA, on hepatocyte death remains unclear. Administration of ABT-737, a Bcl-xL inhibitor, upregulated DNase II activity in murine hepatocyte cell line BNL CL.2 cells and induced apoptosis. In cells treated with DNase II siRNA, ABT-737 led to accumulation of mtDNA in the cytosol and increased expression of interferon (IFN)-β and induction of propidium iodide (PI)-positive cells, in addition to apoptosis. Induced PI-positive cells were suppressed by RIP1 inhibitor, Necrostatin-1, but not by pan-caspase inhibitor, ZVAD-FMK, suggesting non-apoptotic cell death. Both the increase in IFN-β and the induction of non-apoptotic cell death were abolished by administering a TLR9 antagonist, ODN2088, or by the removal of mtDNA from cells with ethidium bromide. Hepatocyte-specific Mcl-1 knockout mice developed hepatocyte apoptosis accompanied by upregulated DNase II activity in their livers. Further knockout of DNase II induced IFN-β expression and RIP1-dependent non-apoptotic hepatocyte death, both of which were suppressed by the administration of ODN2088. Mice fed a high-fat diet (HFD), an obesity-associated fatty liver model, showed increased expression of IFN-β with suppression of DNase II activity in their livers and developed not only hepatocyte apoptosis but also non-apoptotic hepatocyte death. Hepatocyte-specific knockout of DNase II exacerbated HFD-induced non-apoptotic hepatocyte death and liver fibrosis. In conclusion, without DNase II, apoptotic stimulation on hepatocytes induces TLR9-dependent IFN-β production and RIP1-dependent non-apoptotic cell death originating from mtDNA. In fatty livers, DNase II activity is suppressed in contrast to simple inactivation of Bcl-xL or Mcl-1, and both apoptotic and non-apoptotic hepatocyte death can develop, leading to the progression of liver fibrosis.