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The NF-κB/miR-425-5p/MCT4 axis: A novel insight into diabetes-induced endothelial dysfunction.

Endothelial cells (ECs) primarily rely on glycolysis for their energy metabolism, and the final product of glycolysis-lactate-is transferred out of cells via monocarboxylate transporter 4 (MCT4). We previously showed that MCT4 downregulation is involved in diabetic endothelial injury. However, the underlying regulatory mechanisms of MCT4 in diabetes remain unclear. This study showed that miR-425-5p was significantly upregulated in diabetic patients and human umbilical vein endothelial cells (HUVECs) treated with high glucose (HG) and interleukin-1β (IL-1β). MCT4 was shown to be a direct target gene of miR-425-5p, and miR-425-5p expression led to MCT4 downregulation, lactate accumulation and increased apoptosis in HUVECs. Furthermore, the results indicated that NF-κB signaling activation increased miR-425-5p levels and induced MCT4 downregulation, lactate accumulation and apoptosis in HUVECs. In conclusion, NF-κB/miR-425-5p/MCT4 axis activation plays a crucial role in the EC injury induced by HG and IL-1β.

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Co-culture of human induced pluripotent stem cell-derived retinal pigment epithelial cells and endothelial cells on double collagen-coated honeycomb films.

In vitro cell culture models representing the physiological and pathological features of the outer retina are urgently needed. Artificial tissue replacements for patients suffering from degenerative retinal diseases are similarly in great demand. Here, we developed a co-culture system based solely on the use of human induced pluripotent stem cell (hiPSC)-derived cells. For the first time, hiPSC-derived retinal pigment epithelium (RPE) and endothelial cells (EC) were cultured on opposite sides of porous polylactide substrates prepared by breath figures (BF), where both surfaces had been collagen-coated by Langmuir-Schaefer (LS) technology. Small modifications of casting conditions during material preparation allowed the production of free-standing materials with distinct porosity, wettability and ion diffusion capacity. Complete pore coverage was achieved by the collagen coating procedure, resulting in a detectable nanoscale topography. Primary retinal endothelial cells (ACBRI181) and umbilical cord vein endothelial cells (hUVEC) were utilised as EC references. Mono-cultures of all ECs were prepared for comparison. All tested materials supported cell attachment and growth. In mono-culture, properties of the materials had a major effect on the growth of all ECs. In co-culture, the presence of hiPSC-RPE affected the primary ECs more significantly than hiPSC-EC. In consistency, hiPSC-RPE were also less affected by hiPSC-EC than by the primary ECs. Finally, our results show that the modulation of the porosity of the materials can promote or prevent EC migration. In short, we showed that the behaviour of the cells is highly dependent on the three main variables of the study: the presence of a second cell type in co-culture, the source of endothelial cells and the biomaterial properties. The combination of BF and LS methodologies is a powerful strategy to develop thin but stable materials enabling cell growth and modulation of cell-cell contact. Statement of Significance Artificial blood-retinal barriers (BRB), mimicking the interface at the back of the eye, are urgently needed as physiological and disease models, and for tissue transplantation targeting patients suffering from degenerative retinal diseases. Here, we developed a new co-culture model based on thin, biodegradable porous films, coated on both sides with collagen, one of the main components of the natural BRB, and cultivated endothelial and retinal pigment epithelial cells on opposite sides of the films, forming a three-layer structure. Importantly, our hiPSC-EC and hiPSC-RPE co-culture model is the first to exclusively use human induced pluripotent stem cells as cell source, which have been widely regarded as an practical candidate for therapeutic applications in regenerative medicine.

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Endothelial pyroptosis underlies systemic inflammatory response following radiofrequency ablation of hepatic hemangiomas.

This study investigated the relationship between endothelial pyroptosis and the occurrence of systemic inflammatory response (SIR) after radiofrequency (RF) ablation of hepatic hemangiomas. Thirty-two patients with hepatic hemangiomas were treated with RF ablation and blood samples of the patients were collected at different time points. Immunohistochemistry staining was performed to evaluate the expression of caspase-1, gasdermin D (GSDMD), IL-1β and IL-18 in hepatic hemangioma and subablated hemangioma tissue. In vitro experiments, human umbilical vein endothelial cells (HUVECs) were treated with sub-ablative hyperthermia with or without the addition of caspase-1 inhibitor, Ac-YVAD-CMK in the medium. Lactate dehydrogenase (LDH), IL-18, IL-1β, caspase-1 and GSDMD were measured by enzyme-linked immunosorbent assay, real-time PCR and Western blot methods. An elevation of general SIR parameters (CRP and WBC), pyroptosis-related inflammatory cytokines (IL-1β and IL-18) and LDH were observed 1-day post-RF ablation and their peak values were significantly correlated with ablated volume ( < .001) and ablation time ( < .001). Moreover, levels of pyroptosis-related inflammatory cytokines correlated well with general SIR parameters, respectively ( < .001). Immunohistochemical analysis showed the increased expression of caspase-1, GSDMD, IL-18 and IL-1β in the endothelial cells of subablated hemangioma. In vitro experiments showed that subablative hyperthermia induced the caspase-1-associated endothelial pyroptosis and Ac-YVAD-CMK attenuated pyroptosis. In conclusion, SIR in patients treated by RF ablation for hepatic hemangiomas was significantly associated with the ablated volume and ablation time and endothelial pyroptosis may involve in the occurrence of SIR following RF ablation of hepatic hemangiomas.

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Hypoxia Conditioned Mesenchymal Stem Cell-Derived Extracellular Vesicles Induce Increased Vascular Tube Formation .

Mesenchymal stem/stromal cells (MSCs) display a variety of therapeutically relevant effects, such as the induction of angiogenesis, particularly under hypoxic conditions. It is generally recognized that MSCs exert their effects by secretion of paracrine factors and by stimulation of host cells. Furthermore, there is increasing evidence that some therapeutically relevant effects of MSCs are mediated by MSC-derived extracellular vesicles (EVs). Since our current knowledge on MSC-derived EVs released under hypoxic conditions is very limited, we aimed to characterize MSC-derived EVs from normoxic vs. hypoxic conditions (5% O). Adipose-derived MSCs were grown under normoxic and hypoxic conditions, and EVs were analyzed by flow cytometry using lactadherin as a marker for EVs exposing phosphatidylserine, CD63 and CD81 as EV markers, as well as CD73 and CD90 as MSC surface markers. Particle concentration and size distribution were measured by nanoparticle tracking analysis (NTA), and the EV surface antigen signature was characterized using bead-based multiplex flow cytometry. Furthermore, we evaluated the potential of MSC-derived EVs obtained under hypoxic conditions to support angiogenesis using an assay with an hTERT-immortalized human umbilical vein endothelial cell (HUVEC) line. Proliferation and viability of MSCs were increased under hypoxic conditions. EV concentration, size, and surface signature did not differ significantly between normoxic and hypoxic conditions, with the exception of CD44, which was significantly upregulated on normoxic EVs. EVs from hypoxic conditions exhibited increased tube formation as compared to normoxic EVs or to the corresponding supernatants from both groups, indicating that tube formation is facilitated by EVs rather than by soluble factors. In conclusion, hypoxia conditioned MSC-derived EVs appear to be functionally more potent than normoxic MSC-derived EVs regarding the induction of angiogenesis.

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Contribution of different adherent properties of Granulicatella adiacens and Abiotrophia defectiva to their associations with oral colonization and the risk of infective endocarditis.

Granulicatella adiacens (G. adiacens) and Abiotrophia defectiva (A. defectiva) colonize the oral cavity and form part of the normal flora in the intestinal and genitourinary tracts. As reported previously, the frequency of isolation of G. adiacens from the oral cavity was much higher than that of A. defectiva. However, it has been reported that compared with G. adiacens, A. defectiva was isolated at considerably higher frequencies from the blood of patients with infective endocarditis (IE). Hence, in this study, the in vitro interaction of G. adiacens and A. defectiva strains with host surfaces and biofilm formation was examined to assess whether their different adhesive properties contribute to their associations with oral colonization and IE, respectively. G. adiacens exhibited an increased binding ability to saliva-coated hydroxyapatite beads than A. defectiva following the addition of CaCl. Furthermore, biofilm formation was observed only for G. adiacens with the use of a polystyrene tube and scanning electron microscopy analysis. Conversely, A. defectiva displayed significantly greater adherence to human umbilical vein endothelial cells and immobilized fibronectin than G. adiacens. These findings suggest that differences in binding properties to host components imply specific binding mechanisms in G. adiacens and A. defectiva, which might mediate selective colonization in the oral cavity or are associated with the pathogenicity of endocarditis.

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Toxicity of cooking oil fume derived particulate matter: Vitamin D protects tubule formation activation in human umbilical vein endothelial cells.

Cooking oil fumes-derived PM (COFs-derived PM) is the main source of indoor pollution. Exposure to COFs-derived PM can cause oxidative stress and affect angiogenesis. Here we investigated the roles of vitamin D (VD) in protecting tubule formation injury induced by COFs-derived PM, and the roles of ROS/NLRP3/VEGF signaling pathway in the effects. Human umbilical vein endothelial cells (HUVECs) were exposed to 0 (1‰ DMSO), 1000 nmol/l VD, 100 μg/ml PM, and 1000 nmol/l VD + 100 μg/ml PM, respectively. Cell viability and tube formation, as well as protein and mRNA levels were measured. The results showed that exposure of COFs-derived PM dose-and time-dependently reduced the viability of HUVECs, increased the levels of mitochondrial and intracellular ROS, and changed the mitochondrial membrane potential level. While co-incubation with VD rescued these adverse effects. Both Western blot and real-time PCR (RT-PCR) showed that the expressions of NLRP3, caspase-1, Interleukin (IL)-1β, and IL-18 in COFs-derived PM exposure group increased significantly, which could be effectively decreased by co-incubation with VD. COFs-derived PM exposure could also reduce the expression of VEGF, while co-incubating HUVECs with VD evidently up-regulated the protein level of VEGF in HUVECs. In addition, COFs-derived PM could also inhibit the tube formation of HUVECs in vitro, which could be effectively rescued by the co-incubation of VD. Our study proved that COFs-derived PM could damage the tubule formation of HUVECs in vitro, which could be effectively rescue by co-incubation with VD, in which processes the ROS/NLRP3/VEGF signaling pathway played a crucial role. It provides a new theoretical basis for further study on the toxicity of PM to umbilical cord blood vessels.

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Sex-specific metabolic and functional differences in human umbilical vein endothelial cells from twin pairs.

Gonadal hormones are mainly thought to account for sex and gender differences in the incidence, clinical manifestation and therapy of many cardiovascular diseases. However, intrinsic sex differences at the cellular level are mostly overlooked. Here, we assessed sex-specific metabolic and functional differences between male and female human umbilical vein endothelial cells (HUVECs).

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