Isolation and activation of human neutrophils in vitro. The importance of the anticoagulant used during blood collection

Freitas, Marisa; Porto, Graça; Lima, José L.F.C.; Fernandes, Eduarda

doi:10.1016/j.clinbiochem.2007.12.021

Cited by 107 publications

(75 citation statements)

References 26 publications

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“…Human neutrophils in culture Blood samples (10 mL) were drawn from the forearm cubital vein of volunteers using EDTA--containing Vacutainer ® kits. Heparinized tubes were avoided, since eventual iron contamination could accelerate oxidative degradation of samples (Freitas et al 2008). Blood samples were diluted in the same volume of phosphate buffered saline solution (0.137 M NaCl, 2.7 mM KCl, 8.0 mM Na2HPO4, pH 7.4) and neutrophils were separated using Histopaque 1077 according to the manufacturer's instructions (Sigma--Aldrich, MO, USA).…”

Section: Materials and Methods Chemicalsmentioning

confidence: 99%

Supra-physiological doses of testosterone affect membrane oxidation of human neutrophils monitored by the fluorescent probe C11-BODIPY581/591

et al. 2012

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Section: Materials and Methods Chemicalsmentioning

confidence: 99%

Supra-physiological doses of testosterone affect membrane oxidation of human neutrophils monitored by the fluorescent probe C11-BODIPY581/591

et al. 2012

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“…The isolation of the human neutrophils was performed by the density gradient centrifugation method as previously described [19]. Cell viability and cell yield were evaluated by the trypan blue exclusion method, using a Neubauer chamber and an optic microscope with ×40 magnification.…”

Section: Isolation and Treatment Of Human Neutrophilsmentioning

confidence: 99%

2,3-Diarylxanthones as Potential Inhibitors of Arachidonic Acid Metabolic Pathways

et al. 2017

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Abstract-In response to an inflammatory stimulus, arachidonic acid (AA), the main polyunsaturated fatty acid present in the phospholipid layer of cell membranes, is released and metabolized to a series of eicosanoids. These bioactive lipid mediators of inflammation arise physiologically through the action of the enzymes 5-lipoxygenase (5-LOX) and cyclooxygenases (constitutive COX-1 and inducible COX-2). It is believed that dual inhibition of 5-LOX and COXs may have a higher beneficial impact in the treatment of inflammatory disorders rather than the inhibition of each enzyme. With this demand for new dual-acting antiinflammatory agents, a range of 2,3-diarylxanthones were tested through their ability to interact in the AA metabolism. In vitro anti-inflammatory activity was evaluated through the inhibition of 5-LOX-catalyzed leukotriene B 4 (LTB 4 ) formation in human neutrophils and inhibition of COX-1-and COX-2-catalyzed prostaglandin E 2 (PGE 2 ) formation in human whole blood. The results showed that some of the studied arylxanthones were able to prevent LTB 4 production in human neutrophils, in a concentration-dependent manner. The xanthone with a 2-catechol was the most active one (IC 50 ∼ 9 μM). The more effective arylxanthones in preventing COX-1-catalyzed PGE 2 production presented IC 50 values from 1 to 7 μM, exhibiting a structural feature with at least one non-substituted aryl group. All the studied arylxanthones were ineffective to prevent the formation of PGE 2 catalyzed by COX-2, up to the maximum concentration of 100 μM. The ability of the tested 2,3-diarylxanthones to interact with both 5-LOX and COX-1 pathways constitutes an important step in the research of novel dual-acting anti-inflammatory drugs.

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“…The isolation of human neutrophils was performed by the density gradient centrifugation method, and the cell viability and cell yield were evaluated by the trypan blue exclusion method as previously reported (Freitas et al, 2008).…”

Section: Isolation Of Human Neutrophilsmentioning

confidence: 99%

“…The neutrophils' oxidative burst was measured by monitoring the oxidation of luminol by neutrophil-generated reactive species by chemiluminescence (Freitas et al, 2008). The reaction mixtures contained neutrophils (at final concentration of 2 Â 10 6 cells/mL) and the following reagents at the indicated final concentrations (in a final volume of 250 mL): stem bark and flower extracts of V. cauliflora (0.3-10 mg/mL), luminol (500 mM), and PMA (160 nM).…”

Section: Luminol-amplified Chemiluminescence Assaymentioning

confidence: 99%

Stem bark and flower extracts ofVismia caulifloraare highly effective antioxidants to human blood cells by preventing oxidative burst in neutrophils and oxidative damage in erythrocytes

Ribeiro

Berto

Ribeiro

et al. 2015

Pharmaceutical Biology

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(2015) Stem bark and flower extracts of Vismiacauliflora are highly effective antioxidants to human blood cells by preventing oxidative burst in neutrophils and oxidative damage in erythrocytes, Pharmaceutical Biology, 53:11, 1691-1698, DOI: 10.3109/13880209.2014 Previous research on V. cauliflora extracts suggests its potential to neutralize cellular oxidative damages related to the production of reactive oxygen and nitrogen species.Objective: To determine the activity of stem bark and flower extracts of V. cauliflora on the modulation of oxidative burst in human neutrophils, as well as its potential to inhibit oxidative damage in human erythrocytes. Materials and methods:The modulation of neutrophil's oxidative burst by the ethanolic extracts (0.3-1000 mg/mL) was determined by the oxidation of specific probes by reactive species. Additionally, the potential of these extracts to inhibit oxidative damage in human erythrocytes was evaluated by monitoring its biomarkers of oxidative stress. Results: Vismia cauliflora extracts presented remarkable capacity to prevent the oxidative burst in activated human neutrophils (IC 50 515 mg/mL). However, the maximum percentage of inhibition achieved against hydrogen peroxide was 45%. Concerning the oxidative damage in human erythrocytes, the extracts were able to minimize the tert-butyl hydroperoxide-induced hemoglobin oxidation and lipid peroxidation in a very low concentration range (2.7-18 mg/mL). Furthermore, only stem bark extract (100 mg/mL) was able to inhibit the depletion of glutathione (13%). Discussion and conclusion: These results reinforce the therapeutic potential of stem bark and flower extracts of V. cauliflora to heal topical skin disease, namely in the treatment of neutrophil-related dermatosis and skin conditions related to oxidative stress, including skin aging.

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Isolation and activation of human neutrophils in vitro. The importance of the anticoagulant used during blood collection

Cited by 107 publications

References 26 publications

Supra-physiological doses of testosterone affect membrane oxidation of human neutrophils monitored by the fluorescent probe C11-BODIPY581/591

Supra-physiological doses of testosterone affect membrane oxidation of human neutrophils monitored by the fluorescent probe C11-BODIPY581/591

2,3-Diarylxanthones as Potential Inhibitors of Arachidonic Acid Metabolic Pathways

Stem bark and flower extracts ofVismia caulifloraare highly effective antioxidants to human blood cells by preventing oxidative burst in neutrophils and oxidative damage in erythrocytes

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