Cigarette smoke extract induced exosome release is mediated by depletion of exofacial thiols and can be inhibited by thiol-antioxidants

Introduction: Airway epithelial cells have been described to release extracellular vesicles (EVs) with pathological properties when exposed to cigarette smoke extract (CSE). As CSE causes oxidative stress, we investigated whether its oxidative components are responsible for inducing EV release and whether this could be prevented using the thiol antioxidants N-acetyl-L-cysteine (NAC) or glutathione (GSH). Methods: BEAS-2B cells were exposed for 24 h to CSE, H2O2, acrolein, 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB), bacitracin, rutin or the anti-protein disulfide isomerase (PDI) antibody clone RL90; with or without NAC or GSH. EVs in media were measured using CD63(+)CD81(+) bead-coupled flow cytometry or tunable resistive pulse sensing (TRPS). For characterization by Western Blotting, cryo-transmission electron microscopy and TRPS, EVs were isolated using ultracentrifugation. Glutathione disulfide and GSH in cells were assessed by a GSH reductase cycling assay, and exofacial thiols using Flow cytometry. Results: CSE augmented the release of the EV subtype exosomes, which could be prevented by scavenging thiol-reactive components using NAC or GSH. Among thiol-reactive CSE components, H2O2 had no effect on exosome release, whereas acrolein imitated the NAC-reversible exosome induction. The exosome induction by CSE and acrolein was paralleled by depletion of cell surface thiols. Membrane impermeable thiol blocking agents, but not specific inhibitors of the exofacially located thiol-dependent enzyme PDI, stimulated exosome release. Summary/conclusion: Thiol-reactive compounds like acrolein account for CSE-induced exosome release by reacting with cell surface thiols. As acrolein is produced endogenously during inflammation, it may influence exosome release not only in smokers, but also in ex-smokers with chronic obstructive pulmonary disease. NAC and GSH prevent acrolein-and CSE-induced exosome release, which may contribute to the clinical benefits of NAC treatment

An Essential Difference between the Flavonoids MonoHER and Quercetin in Their Interplay with the Endogenous Antioxidant Network

Antioxidants can scavenge highly reactive radicals. As a result the antioxidants are converted into oxidation products that might cause damage to vital cellular components. To prevent this damage, the human body possesses an intricate network of antioxidants that pass over the reactivity from one antioxidant to another in a controlled way. The aim of the present study was to investigate how the semi-synthetic flavonoid 7-mono-O-(β-hydroxyethyl)-rutoside (monoHER), a potential protective agent against doxorubicin-induced cardiotoxicity, fits into this antioxidant network. This position was compared with that of the well-known flavonoid quercetin. The present study shows that the oxidation products of both monoHER and quercetin are reactive towards thiol groups of both GSH and proteins. However, in human blood plasma, oxidized quercetin easily reacts with protein thiols, whereas oxidized monoHER does not react with plasma protein thiols. Our results indicate that this can be explained by the presence of ascorbate in plasma; ascorbate is able to reduce oxidized monoHER to the parent compound monoHER before oxidized monoHER can react with thiols. This is a major difference with oxidized quercetin that preferentially reacts with thiols rather than ascorbate. The difference in selectivity between monoHER and quercetin originates from an intrinsic difference in the chemical nature of their oxidation products, which was corroborated by molecular quantum chemical calculations. These findings point towards an essential difference between structurally closely related flavonoids in their interplay with the endogenous antioxidant network. The advantage of monoHER is that it can safely channel the reactivity of radicals into the antioxidant network where the reactivity is completely neutralized

Pleiotropic Benefit of Monomeric and Oligomeric Flavanols on Vascular Health - A Randomized Controlled Clinical Pilot Study

BACKGROUND: Cardiovascular diseases are expanding to a major social-economic burden in the Western World and undermine man's deep desire for healthy ageing. Epidemiological studies suggest that flavanol-rich foods (e.g. grapes, wine, chocolate) sustain cardiovascular health. For an evidenced-based application, however, sound clinical data on their efficacy are strongly demanded. METHODS: In a double-blind, randomized, placebo-controlled intervention study we supplemented 28 male smokers with 200 mg per day of monomeric and oligomeric flavanols (MOF) from grape seeds. At baseline, after 4 and 8 weeks we measured macro- and microvascular function and a cluster of systemic biomarkers for major pathological processes occurring in the vasculature: disturbances in lipid metabolism and cellular redox balance, and activation of inflammatory cells and platelets. RESULTS: In the MOF group serum total cholesterol and LDL decreased significantly (P ≤ 0.05) by 5% (n = 11) and 7% (n = 9), respectively in volunteers with elevated baseline levels. Additionally, after 8 weeks the ratio of glutathione to glutathione disulphide in erythrocytes rose from baseline by 22% (n = 15, P<0.05) in MOF supplemented subjects. We also observed that MOF supplementation exerts anti-inflammatory effects in blood towards ex vivo added bacterial endotoxin and significantly reduces expression of inflammatory genes in leukocytes. Conversely, alterations in macro- and microvascular function, platelet aggregation, plasma levels of nitric oxide surrogates, endothelin-1, C-reactive protein, fibrinogen, prostaglandin F2alpha, plasma antioxidant capacity and gene expression levels of antioxidant defense enzymes did not reach statistical significance after 8 weeks MOF supplementation. However, integrating all measured effects into a global, so-called vascular health index revealed a significant improvement of overall vascular health by MOF compared to placebo (P ≤ 0.05). CONCLUSION: Our integrative multi-biomarker approach unveiled the pleiotropic vascular health benefit of an 8 weeks supplementation with 200 mg/d MOF in humans. TRIAL REGISTRATION: ClinicalTrials.gov NCT00742287

Swimming physiology of European silver eels (Anguilla anguilla L.): energetic costs and effects on sexual maturation and reproduction

The European eel migrates 5,000–6,000 km to the Sargasso Sea to reproduce. Because they venture into the ocean in a pre-pubertal state and reproduce after swimming for months, a strong interaction between swimming and sexual maturation is expected. Many swimming trials have been performed in 22 swim tunnels to elucidate their performance and the impact on maturation. European eels are able to swim long distances at a cost of 10–12 mg fat/km which is 4–6 times more efficient than salmonids. The total energy costs of reproduction correspond to 67% of the fat stores. During long distance swimming, the body composition stays the same showing that energy consumption calculations cannot be based on fat alone but need to be compensated for protein oxidation. The optimal swimming speed is 0.61–0.67 m s−1, which is ~60% higher than the generally assumed cruise speed of 0.4 m s−1 and implies that female eels may reach the Sargasso Sea within 3.5 months instead of the assumed 6 months. Swimming trials showed lipid deposition and oocyte growth, which are the first steps of sexual maturation. To investigate effects of oceanic migration on maturation, we simulated group-wise migration in a large swim-gutter with seawater. These trials showed suppressed gonadotropin expression and vitellogenesis in females, while in contrast continued sexual maturation was observed in silver males. The induction of lipid deposition in the oocytes and the inhibition of vitellogenesis by swimming in females suggest a natural sequence of events quite different from artificial maturation protocols

Past, Present and Future of the Section “Molecular Toxicology”

‘Forever’ chemicals that unintendedly in the long run pollute the environment, climate change, COVID; life continuously faces all sorts of unforeseen challenges that are an inevitable side product of ‘progress’ [...

Connecting West and East

Despite their similarities, Western medicine and Eastern medicine are very different because they are built on different fundamentals. The general idea has arisen that we will benefit by connecting Western and Eastern medicine. First, both the merits as well as the limitations of both types of medicine are discussed. It was concluded that to create a bridge, we should focus on similarities that inspire the further unravelling of the molecular mechanism of the mode of action and toxicity of Traditional Chinese Medicine. It is suggested that the energy perspective provides a basis to integrate Eastern and Western medicine

The Screening of Anticholinergic Accumulation by Traditional Chinese Medicine

Many Western drugs can give rise to serious side effects due to their ability to bind to acetylcholine receptors in the brain. This aggravates when they are combined, which is known as anticholinergic accumulation (AA). Some bioactives in Traditional Chinese Medicine (TCM) are known to block acetylcholine receptors and thus potentially cause AA. The AA of TCM was screened by quantifying the displacement of [3H] pirenzepine on acetylcholine receptors in a rat brain homogenate. We used a new unit to express AA, namely the Total Atropine Equivalents (TOAT). The TOAT of various herbs used in TCM was very diverse and even negative for some herbs. This is indicative for the broadness of the pallet of ingredients used in TCM. Three TCM formulas were screened for AA: Ma Huang Decotion (MHD), Antiasthma Simplified Herbal Medicine intervention (ASHMI), and Yu Ping Feng San (YPFS). The TOAT of ASHMI was indicative for an additive effect of herbs used in it. Nevertheless, it can be calculated that one dose of ASHMI is probably too low to cause AA. The TOAT of YPFS was practically zero. This points to a protective interaction of AA. Remarkably, MHD gave a negative TOAT, indicating that the binding to the acetylcholine receptors was increased, which also circumvents AA. In conclusion, our results indicate that TCM is not prone to give AA and support that there is an intricate interaction between the various bioactives in TCM to cure diseases with minimal side effects