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Exosome-like nanoparticles (ELNs) are attracting interest as important vehicles of intercellular communication, both in prokaryotes and eukaryotes. Recently, dietary nanoparticles similar to mammalian exosomes have attracted attention for these features. In particular they appear to be relevant in the modulation of several cellular processes as well as candidate carriers of bioactive molecules (proteins, lipids, and nucleic acids, including miRNAs) with therapeutic value. Herein, we investigated the cellular uptake of blueberry-derived ELNs (B-ELNs) by a human stabilized endothelial cell line (EA.hy926) and the ability of B-ELNs to modulate the expression of inflammatory genes as the response of tumor necrosis factor-α (TNF-α). Our results indicate that 1) EA.hy926 cells internalize B-ELNs in a dose-dependent manner; 2) pretreatment with B-ELNs counters TNF-α-induced reactive oxygen species (ROS) generation and loss of cell viability and modulates the differential expression of 29 genes (fold change > 1.5) induced by TNF-α compared to control; 3) pathway analysis reveals their involvement in a total of 340 canonical pathways, 121 KEGG pathways, and 121 GO Biological processes; and 4) the intersection between differentially expressed (DE) genes and miRNAs contained in B-ELNs unveils a set of candidate target genes, such as prostaglandin I2 synthase (PTGIS), mitogen-activated protein kinase 14 (MAPK14), and phosphodiesterase 7A (PDE7A), for ELNs-contained cargo. In conclusion, our study indicates that B-ELNs can be considered candidate therapeutic carriers of bioactive compounds potentially able to protect vascular system against various stressors.

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The term “normobaric oxygen paradox” (NOP), describes the response to the return to normoxia after a hyperoxic event, sensed by tissues as oxygen shortage, and resulting in up-regulation of the Hypoxia-inducible factor 1α (HIF-1α) transcription factor activity. The molecular characteristics of this response have not been yet fully characterized. Herein, we report the activation time trend of oxygen-sensitive transcription factors in human peripheral blood mononuclear cells (PBMCs) obtained from healthy subjects after one hour of exposure to mild (MH), high (HH) and very high (VHH) hyperoxia, corresponding to 30%, 100%, 140% O2, respectively. Our observations confirm that MH is perceived as a hypoxic stress, characterized by the activation of HIF-1α and Nuclear factor (erythroid-derived 2)-like 2 (NRF2), but not Nuclear Factor kappa-light-chain-enhancer of activated B cells (NF-κB). Conversely, HH is associated to a progressive loss of NOP response and to an increase in oxidative stress leading to NRF2 and NF-kB activation, accompanied by the synthesis of glutathione (GSH). After VHH, HIF-1α activation is totally absent and oxidative stress response, accompanied by NF-κB activation, is prevalent. Intracellular GSH and Matrix metallopeptidase 9 (MMP-9) plasma levels parallel the transcription factors activation pattern and remain elevated throughout the observation time. In conclusion, our study confirms that, in vivo, the return to normoxia after MH is sensed as a hypoxic trigger characterized by HIF-1α activation. On the contrary, HH and VHH induce a shift toward an oxidative stress response, characterized by NRF2 and NF-κB activation in the first 24 h post exposure.

Intestinal epithelium represents a protective physical barrier and actively contributes to the mucosal immune system. Polarized basolateral intestinal secretion of inflammatory mediators, followed by activation of NF-κB signaling and inflammatory pathways in endothelial cells, efficiently triggers extravasation of neutrophils from the vasculature, therefore contributing to the development and maintenance of intestinal inflammation. Proper regulation of NF-κB activation at the epithelial interface is crucial for the maintenance of physiological tissue homeostasis. Many papers reported that anthocyanins, a group of compounds belonging to flavonoids, possess anti-inflammatory effects and modulate NF-κB activity. In this study, by using a coculture in vitro system, we aimed to evaluate the effects of TNF-α-stimulated intestinal cells on endothelial cells activation, as well as the protective effects of cyanidin-3-glucoside (C3G). In this model, TNF-α induced nuclear translocation of NF-κB and TNF-α and IL-8 gene expression in Caco-2 cells, whereas C3G pretreatment dose-dependently reduced these effects. Furthermore, TNF-α-stimulated Caco-2 cells induced endothelial cells activation with increased E-selectin and VCAM-1 mRNA, leukocyte adhesion, and NF-κB levels in HUVECs, which were inhibited by C3G. We demonstrated that selective inhibition of the NF-κB pathway in epithelial cells represents the main mechanism by which C3G exerts these protective effects. Thus, anthocyanins could contribute to the management of chronic gut inflammatory diseases.

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Hyperglycemia contributes to dysregulate endothelial function associated with diabetes, leading to initiation and propagation of vascular complications and dysfunction. Caffeic acid (CA), a dietary hydroxycinnamic acid abundant in coffee, has been reported to exert antidiabetic effects in rat models. Herein, we investigated the molecular effects of physiological concentrations of CA (10 nM) against endothelial dysfunction induced by high glucose (HG) in human endothelial cells (HUVECs). HUVECs were exposed to HG 25 mM, to mimic diabetic condition, in presence of CA. Intracellular redox status (reduced glutathione, superoxide dismutase (SOD) and total antioxidant activity levels), and NF-κB pathway were examined. We also evaluated the involvement of NF-E2-related factor 2 (Nrf2)/electrophile responsive element (EpRE) pathway. Our data show that CA inhibits HG-induced nuclear translocation of NF-κB and the downstream expression of endothelial adhesion molecule 1 and restores antioxidant levels by upregulating Nrf2/EpRE pathway. Our data suggest that CA can suppress several aspects of HG-induced endothelial dysfunction through the modulation of intracellular redox status controlled by the transcription factor Nrf2. These findings highlight that low physiological concentration of CA achievable specifically upon food consumption are able to prevent endothelial dysfunction associated with inflammation and oxidative stress induced by high concentration of glucose. © 2016 BioFactors, 43(1):54-62, 2017.