Improving endothelial health with food-derived H₂S donors: an in vitro study with S-allyl cysteine and with a black-garlic extract enriched in sulfur-containing compounds

Abstract

A healthy vascular endothelium plays an essential role in modulating vascular tone by producing and releasing vasoactive factors such as nitric oxide (NO). Endothelial dysfunction (ED), the loss of the...

Indole-3-Propionic Acid, a Gut Microbiota-Derived Tryptophan Metabolite, Promotes Endothelial Dysfunction Impairing Purinergic-Induced Nitric Oxide Release in Endothelial Cells

“…As previously described [ 49 ], BAE-1 cells, grown in black 96-well microplates with a clear bottom (Greiner Bio-One, Kremsmünster, Austria), were treated with DMEM + 10% FCS, either alone (control condition) or supplemented with IPA (100 nM, 1 µM, 100 µM) for 30 min or 24 h; treatment occurred with 20 µM of menadione (MEN) for 1 h to induce ROS production, which was then used as a positive control, while the addition of IPA and MEN simultaneously allowed for the recognition of IPA’s antioxidant effects. During the last 30 min of the stimulation, cells were loaded in the dark with 5 µM of CellROX ® Green Reagent, a probe that exhibits bright green fluorescence upon oxidation by ROS.…”

“…As previously described [ 49 ], BAE-1 cells, grown on glass-bottom dishes, were treated for 1 h with 20 µM of MEN to induce ROS production (positive control) or with 1 µM of IPA for 30 min or 24 h, alone or in combination with MEN, in DMEM + 10% FCS; cells were loaded with 5 µM of CellROX ® Green probe for the last 30 min in the dark. Cells were then washed twice with PBS, and fluorescence at 488 nm was acquired with a fluorescence inverted microscope (Olympus IX70, Olympus America Inc., Melville, NY, USA) with a 50× Uplan FI oil-immersion objective.…”

“…As previously described [ 49 ], BAE-1 cells, grown on glass-bottom dishes, were loaded with 5 µM of DAR4M-AM probe (Calbiochem ® ) for 30 min in the dark. Next, cells were washed twice with PBS and treated in PBS at 37 °C in the dark for 5 min with 100 µM of ATP as a positive control or 30 min with 1 µM of IPA alone or with ATP for another 5 min.…”

“…As previously described [9,49], BAE-1 cells were seeded on plastic dishes, 20 cm 2 of growth area, at a density of 10,000 cells/cm 2 , in 10% FCS DMEM, and grown in the cell incubator for 48 h. Cells were then treated with 100 µM of ATP for 5 min as a positive control or with 1 µM of IPA for 30 min or 24 h. Cell monolayers were lysed in 200 µL of RIPA lysis buffer (ThermoFisher Scientific) containing a phosphatase inhibitor cocktail (PhosSTOP, Roche, Mannheim, Germany) forced through a 1 mL syringe needle and centrifuged at 10,000 rpm for 5 min at 4 • C. Proteins (20 µg per lane) were resolved on 8% SDS-PAGE, transferred to as a polyvinylidene fluoride membrane (PVDF, ThermoFisher Scientific) in cold transfer buffer (25 mM Tris pH 8.3, 192 mM glycine, 0.1% SDS, 20% methanol) and blocked for 1 h at 37 • C in TBST (10 mM Tris-HCl, 0.1 M NaCl, 0.1% Tween 20, pH 7.5) plus 5% non-fat dry milk. Blots were incubated overnight at 4 • C with primary antibodies (1:500 monoclonal anti-eNOS; 1:250 polyclonal anti-p-eNOS; 1:2000 monoclonal anti-β-actin) in TBST containing 1% non-fat dry milk.…”

Effect of Encapsulated Purple Garlic Oil on Microvascular Function and the Components of Metabolic Syndrome: A Randomized Placebo-Controlled Study—The ENDOTALLIUM Study

“…Nutrients 2024, 16, 1755 2 of 14 Several approaches have been proposed to measure the patho-physiological function of the vascular endothelium, including circulating biomarkers and non-invasive methods [4]. Most studies aimed at studying endothelial dysfunction have focused on large arteries, mainly assessing the flow-mediated dilatation of the brachial artery.…”