Seminal cell-free DNA levels measured by PicoGreen fluorochrome are associated with sperm fertility criteria

SummaryPrevious investigations suggested that elevated cell-free DNA (cfDNA) can indicate non-healthy states. However, the potential association between cfDNA seminal plasma levels and fertility sperm parameters has not yet been determined. Therefore, the present study evaluated the association between seminal cfDNA levels and sperm fertility criteria to determine the use of seminal cfDNA quantification. An in vivo protocol quantified cfDNA levels of semen samples obtained from 163 male patients using fluorescent PicoGreen dye staining. To confirm if semen cfDNA quantification is realistic, an in vitro complementary test was performed using three or four semen samples. The fresh sperm samples were exposed to paraquat that generates high levels of superoxide anion causing oxidative stress and cell mortality. The results showed significant association between dsDNA levels and several sperm fertility parameters, such as low viability and alterations of motility and morphology. The in vitro analysis confirmed the association between dsDNA levels and sperm viability. Together, these results suggest that dsDNA levels could be an important biomarker to test sperm fertility.

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Anti-viral properties of interferon beta treatment in patients with multiple sclerosis

Multiple Sclerosis Journal ◽

10.1191/1352458502ms794oa ◽

2002 ◽

Vol 8 (3) ◽

pp. 237-242 ◽

Cited By ~ 29

Author(s):

J Hong ◽

M V Tejada-Simon ◽

V M Rivera ◽

Y CQ Zang ◽

J Z Zhang

Keyword(s):

Multiple Sclerosis ◽

Treatment Efficacy ◽

Interferon Beta ◽

Viral Infections ◽

Human Herpesvirus ◽

Virion Protein ◽

Cell Free Dna ◽

Free Dna

Viral infections are potentially associated with the etiology and pathogenesis of multiple sclerosis (MS). It has been speculated that the treatment efficacy of interferon beta (IFN beta) in MS may relate to its anti-viral properties. The study was undertaken to evaluate the in vivo anti-viral effects of IFN beta-1a in patients with MS. Human herpesvirus-6 (HHV-6) was studied as an example for being a latent neurotropic virus. IFN beta used at concentrations of approximately 0.5 mg/ml was shown to significantly reduce in vitro HHV-6 replication in a susceptible T-cell line. Sera derived from 23 MS patients treated with IFN beta-1a were examined for serum cell-free DNA of HHV-6 as an indicator for viral replication and the reactivity of IgM antibodies to a recombinant HHV-6 virion protein containing a known immunoreactive region. The results were compared with those of control sera obtained from untreated MS (n=29) and healthy individuals (n=21). The findings indicated that IFN beta treatment significantly reduced HHV-6 replication as evident by decreased cell-free DNA in treated MS specimens. The results correlated with decreased IgM reactivity to the HHV-6 antigen in treated MS patients compared to untreated controls, suggesting reduced exposure to HHV-6. The findings were confirmed in paired sera obtained from seven MS patients before and after the treatment. The study provides new evidence indicating that IFN beta has potent in vivo anti-viral effects that may contribute to the treatment efficacy in MS.

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Methylated Cell-Free DNA In Vitro and In Vivo

Circulating Nucleic Acids in Plasma and Serum ◽

10.1007/978-90-481-9382-0_25 ◽

2010 ◽

pp. 185-194 ◽

Cited By ~ 2

Author(s):

Tatyana E. Skvortsova ◽

Olga E. Bryzgunova ◽

Alena O. Lebedeva ◽

Viktoria V. Mak ◽

Valentin V. Vlassov ◽

...

Keyword(s):

Cell Free Dna ◽

Free Dna

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Increased Cell-Free DNA Plasma Concentration Following Liver Transplantation Is Linked to Portal Hepatitis and Inferior Survival

Journal of Clinical Medicine ◽

10.3390/jcm9051543 ◽

2020 ◽

Vol 9 (5) ◽

pp. 1543 ◽

Cited By ~ 2

Author(s):

Felix Krenzien ◽

Shadi Katou ◽

Alba Papa ◽

Bruno Sinn ◽

Christian Benzing ◽

...

Keyword(s):

Liver Transplantation ◽

Immune Responses ◽

Viral Infections ◽

Inflammatory Responses ◽

Term Survival ◽

Cell Free Dna ◽

Reactive Protein ◽

Free Dna

Donor organ quality is crucial for transplant survival and long-term survival of patients after liver transplantation. Besides bacterial and viral infections, endogenous damage-associated molecular patterns (DAMPs) can stimulate immune responses. Cell-free DNA (cfDNA) is one such DAMP that exhibits highly proinflammatory effects via DNA sensors. Herein, we measured cfDNA after liver transplantation and found elevated levels when organs from resuscitated donors were transplanted. High levels of cfDNA were associated with high C-reactive protein, leukocytosis as well as granulocytosis in the recipient. In addition to increased systemic immune responses, portal hepatitis was observed, which was associated with increased interface activity and a higher numbers of infiltrating neutrophils and eosinophils in the graft. In fact, the cfDNA was an independent significant factor in multivariate analysis and increased concentration of cfDNA was associated with inferior 1-year survival. Moreover, cfDNA levels were found to be decreased significantly during the postoperative course when patients underwent continuous veno-venous haemofiltration. In conclusion, patients receiving livers from resuscitated donors were characterised by high postoperative cfDNA levels. Those patients showed pronounced portal hepatitis and systemic inflammatory responses in the short term leading to a high mortality. Further studies are needed to evaluate the clinical relevance of cfDNA clearance by haemoadsorption and haemofiltration in vitro and in vivo.

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Short-Term Treadmill Running as a Model for Studying Cell-Free DNA Kinetics In Vivo

Clinical Chemistry ◽

10.1373/clinchem.2010.158030 ◽

2011 ◽

Vol 57 (4) ◽

pp. 633-636 ◽

Cited By ~ 58

Author(s):

Thomas Beiter ◽

Annunziata Fragasso ◽

Jens Hudemann ◽

Andreas M Nieß ◽

Perikles Simon

Keyword(s):

Time Course ◽

Treadmill Exercise ◽

Plasma Concentrations ◽

High Mobility ◽

Treadmill Running ◽

Short Term ◽

Cell Free Dna ◽

Free Dna ◽

Cross Country

BACKGROUND Increased plasma concentrations of cell-free DNA (cf-DNA) are considered a hallmark of various clinical conditions. Despite intensive research in this field, limited data are available concerning the time course of release and clearance of cf-DNA in vivo. METHODS We extracted cf-DNA from plasma samples taken before and immediately after a 10-km cross-country run, and from samples taken before, immediately after, and 30 min after exhaustive short-term treadmill exercise. The contribution of nuclear (nDNA) and mitochondrial DNA (mtDNA) was measured by quantitative real-time PCR. The incremental treadmill exercise setup was exploited to delineate the precise sequencing and timing of cf-nDNA, lactate, and high-mobility group box 1 protein (HMGB1) release during the exercise and recovery phases. RESULTS Postexercise plasma cf-nDNA concentrations in cross-country and treadmill runners were significantly increased, by 7.6-fold and 9.9-fold, respectively (P < 0.001). cf-nDNA concentrations were not correlated with age, sex, or body mass index. Plasma concentrations of cf-nDNA and HMGB1 in postexercise samples of treadmill runners were significantly correlated (r = 0.84; P = 0.004). cf-mtDNA concentrations were not affected by treadmill exercise. Time-course analyses demonstrated that cf-nDNA is released within minutes after the onset of exercise and is rapidly cleared from the circulation after the cessation of exercise. Nearly congruent kinetics for cf-nDNA, lactate, and HMGB1 were observed during the exercise phase. CONCLUSIONS A single bout of exhaustive short-term treadmill exercise constitutes a versatile model system suitable for addressing basic questions about cf-DNA biology.

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Type-4 Phosphodiesterase (PDE4) Blockade Reduces NETosis in Cystic Fibrosis

Frontiers in Pharmacology ◽

10.3389/fphar.2021.702677 ◽

2021 ◽

Vol 12 ◽

Author(s):

Licia Totani ◽

Concetta Amore ◽

Antonio Piccoli ◽

Giuseppe Dell’Elba ◽

Angelo Di Santo ◽

...

Keyword(s):

Cystic Fibrosis ◽

Pseudomonas Aeruginosa ◽

Lung Disease ◽

Pde4 Inhibitor ◽

Free Dna ◽

Impaired Lung Function ◽

Cellular Integrity ◽

Net Release

Neutrophilic inflammation is a key determinant of cystic fibrosis (CF) lung disease. Neutrophil-derived free DNA, released in the form of extracellular traps (NETs), significantly correlates with impaired lung function in patients with CF, underlying their pathogenetic role in CF lung disease. Thus, specific approaches to control NETosis of neutrophils migrated into the lungs may be clinically relevant in CF. We investigated the efficacy of phosphodiesterase (PDE) type-4 inhibitors, in vitro, on NET release by neutrophils from healthy volunteers and individuals with CF, and in vivo, on NET accumulation and lung inflammation in mice infected with Pseudomonas aeruginosa. PDE4 blockade curbed endotoxin-induced NET production and preserved cellular integrity and apoptosis in neutrophils, from healthy subjects and patients with CF, challenged with endotoxin, in vitro. The pharmacological effects of PDE4 inhibitors were significantly more evident on CF neutrophils. In a mouse model of Pseudomonas aeruginosa chronic infection, aerosol treatment with roflumilast, a selective PDE4 inhibitor, gave a significant reduction in free DNA in the BALF. This was accompanied by reduced citrullination of histone H3 in neutrophils migrated into the airways. Roflumilast-treated mice showed a significant improvement in weight recovery. Our study provides the first evidence that PDE4 blockade controls NETosis in vitro and in vivo, in CF-relevant models. Since selective PDE4 inhibitors have been recently approved for the treatment of COPD and psoriasis, our present results encourage clinical trials to test the efficacy of this class of drugs in CF.

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Exposure of surfactant protein A to ozone in vitro and in vivo impairs its interactions with alveolar cells

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.1992.262.1.l63 ◽

1992 ◽

Vol 262 (1) ◽

pp. L63-L68 ◽

Cited By ~ 12

Author(s):

R. S. Oosting ◽

J. F. Van Iwaarden ◽

L. Van Bree ◽

J. Verhoef ◽

L. M. Van Golde ◽

...

Keyword(s):

Superoxide Anion ◽

Protein A ◽

Surfactant Protein ◽

Surfactant Protein A ◽

Alveolar Type ◽

Type Ii ◽

Superoxide Anion Production ◽

Anion Production

This study focused on the question of whether exposure of surfactant protein A (SP-A) to ozone affected properties of this protein that may be involved in regulating alveolar type II cell and alveolar macrophage functions. In vitro exposure of human or canine SP-A to ozone reduced the ability of this protein to inhibit phorbol-ester induced secretion of [3H]phosphatidylcholine by alveolar type II cells in culture. Ozone-exposed human SP-A showed a decreased ability to enhance phagocytosis of herpes simplex virus and to stimulate superoxide anion production by alveolar macrophages. Experiments with elastase showed that ozone-exposed canine SP-A was more susceptible to proteolysis. A conformational change of the protein could underlie this phenomenon. Surfactant isolated from ozone-exposed rats (0.4 ppm ozone for 12 h) was also less able to stimulate superoxide anion production by alveolar macrophages than surfactant from control rats, which suggested that SP-A in vivo was also susceptible to ozone. The results of this study suggest that SP-A-alveolar cell interactions can be inhibited by ozone exposure, which may contribute to the toxicity of ozone in the lungs.

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Neutrophil Extracellular Traps May Contribute to the Pathogenesis in Adult-onset Still Disease

The Journal of Rheumatology ◽

10.3899/jrheum.181058 ◽

2019 ◽

Vol 46 (12) ◽

pp. 1560-1569 ◽

Cited By ~ 4

Author(s):

Mi-Hyun Ahn ◽

Jae Ho Han ◽

Young-Jun Chwae ◽

Ju-Yang Jung ◽

Chang-Hee Suh ◽

...

Keyword(s):

Immunohistochemical Analysis ◽

Serum Levels ◽

Neutrophil Extracellular Traps ◽

Polymorphonuclear Neutrophils ◽

Adult Onset ◽

Cell Free Dna ◽

Extracellular Traps ◽

Free Dna ◽

Still Disease

Objective.Release of neutrophil extracellular traps (NET) has been described as an effector mechanism of polymorphonuclear neutrophils in several inflammatory diseases. Thus, this study was performed to evaluate the role of NET in the pathogenesis of adult-onset Still disease (AOSD).Methods.We determined the serum levels of NET molecules and investigated their associations with clinical disease activities in patients with AOSD. Further, we analyzed the differences in the NETosis response in AOSD patients compared to healthy controls (HC). To explore the in vivo involvement of NET in AOSD, we performed immunohistochemical analysis of skin and lymph node (LN) biopsies for proteins related to NET in patients with active AOSD.Results.Serum levels of cell-free DNA, myeloperoxidase (MPO)-DNA complex, and α-defensin were significantly increased in patients with AOSD compared to HC. Serum levels of the NET molecules, cell-free DNA, MPO-DNA, and α-defensin were correlated with several disease activity markers for AOSD. In followup of patients with AOSD after treatment with corticosteroid, the levels of cell-free DNA and α-defensin decreased significantly. On immunohistochemistry, neutrophil elastase–positive and MPO-positive inflammatory cells were detected in skin and LN of patients with AOSD, and were expressed in fiber form in the lesions. The serum from patients with active AOSD induced NETosis in neutrophils from HC. NET molecules induced interleukin 1β production in monocytes, representing a novel mechanism in the pathogenesis of AOSD.Conclusion.The findings presented here suggest that NET may contribute to the inflammatory response and pathogenesis in AOSD.

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Influence of SIN-1 on Platelet Ca2+ Handling in Patients with Suspected Coronary Artery Disease: Ex Vivo and In Vitro Studies

Thrombosis and Haemostasis ◽

10.1055/s-0037-1613904 ◽

2000 ◽

Vol 83 (05) ◽

pp. 752-758 ◽

Cited By ~ 3

Author(s):

Claude Le Feuvre ◽

Annie Brunet ◽

Thuc Do Pham ◽

Jean-Philippe Metzger ◽

André Vacheron ◽

...

Keyword(s):

Superoxide Anion ◽

Vascular Tone ◽

Ex Vivo ◽

Suspected Coronary Artery Disease ◽

In Vitro Studies ◽

H2o2 Formation ◽

Artery Disease ◽

Dose Dependent

SummaryThe 3-morpholinosydnonimine (SIN-1) generates both nitric oxide (NO) and superoxide anion (O2−). It elicits dose-dependent vasodilation in vivo, in spite of the opposite effects of its breakdown products on vascular tone and platelet aggregation.This study was designed to investigate the influence of intravenous SIN-1 injection on platelet Ca2+ handling in patients undergoing coronary angiography. SIN-1 administration reduced cytosolic [Ca2+] in unstimulated platelets by decreasing Ca2+ influx. It attenuated Ca2+ mobilization from internal stores evoked by thrombin or thapsigargin. In vitro studies were used as an approach to investigate how simultaneous productions of NO and O2− from SIN-1 modify thrombin- or thapsigargin-induced platelet Ca2+ mobilization. Superoxide dismutase, the O2− scavenger, enhanced the capacity of SIN-1 to inhibit Ca2+ mobilization but catalase had no effect.This suggests that the effects of SIN-1 on platelet Ca2+ handling resemble those of NO, but are modulated by simultaneous O2− release, independently of H2O2 formation.

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