Activity Of Human Umbilical Vein Endothelial Cells Research Articles

Nepeta angustifolia attenuates responses to vascular inflammation in high glucose-induced human umbilical vein endothelial cells through heme oxygenase-1 induction

Ethnopharmacological relevanceThe traditional folk medicine Nepeta angustifolia C. Y. Wu (NA) reportedly possesses various biological activities, such as anti-inflammatory, analgesic, antihypoxia, and antifatigue effects. In this study, we evaluated the anti-vascular inflammation effects of N. angustifolia extract in human umbilical vein endothelial cells (HUVECs) induced by high glucose (HG) as well as the underlying mechanisms and verified its activity in diabetic rats. Materials and methodsHUVECs were exposed to 25 mM glucose to induce endothelial dysfunction. Adhesion molecule expression and reactive oxygen species (ROS) were assayed. IκB and IκB phosphorylation, nuclear factor-κB (NF-κB), HO-1 and nuclear factor erythroid 2-related factor 2 (Nrf2) were examined by Western blot. Nuclear localisation of Nrf2 was also examined using immunofluorescence. The in vivo study of NA was tested in diabetic rats in which the thoracic aorta and serum were collected to observe aorta histological change, and evaluate endothelial function and vascular inflammation. ResultsThe results revealed that HG can significantly promote the generation of ROS, the expression of cell adhesion molecules (CAMs), and the phosphorylation and degradation of IκB and NF-κB activation in HUVECs. These HG-induced phenomena were suppressed by NA-induced heme oxygenase (HO)− 1 expression in a dose- and time-dependent manner by activating Nrf2. The HO-1 inhibitor tin protoporphyrin also dramatically reversed the NA-induced inhibition of CAM expression and the reduction in ROS production. Furthermore, NA also elicited anti-vascular dysfunction effects in diabetic rats, where endothelial function was improved and vascular inflammation was alleviated. ConclusionAll these findings indicated that NA attenuated high glucose-induced vascular dysfunction in vitro and in vivo.

Osthole alleviates oxidized low-density lipoprotein-induced vascular endothelial injury through suppression of transforming growth factor-β1/Smad pathway

Osthole, a naturally-derived coumarin, has been shown to exhibit pharmacological activities including anti-inflammatory, anti-oxidative and cardiovascular protective effects. However, the effect of osthole on oxidized low-density lipoprotein (ox-LDL)-induced endothelial injury and its underlying mechanism remain unknown. We found that osthole did not affect viability of human umbilical vein endothelial cells (HUVECs) but alleviated ox-LDL-induced cytotoxicity in HUVECs. Osthole repressed ox-LDL-induced release of tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, and IL-6 in HUVECs. Osthole reversed ox-LDL-induced elevation of reactive oxygen species (ROS) production and malondialdehyde (MDA) level, and reduction of superoxide dismutase (SOD) activity in HUVECs. Meanwhile, osthole attenuated ox-LDL-induced increase of mRNA expression and secretion of intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in HUVECs. Osthole increased nitric oxide (NO) production and endothelial nitric oxide synthase (eNOS) phosphorylation in ox-LDL-treated HUVECs. Furthermore, osthole inhibited ox-LDL-induced activation of the transforming growth factor-β1 (TGF-β1)/Smad pathway and activation of TGF-β1/Smad pathway by TGF-β1 attenuated the protective effects of osthole on HUVECs injury. These results suggested that osthole attenuated ox-LDL-induced HUVECs injury by inhibiting the TGF-β1/Smad pathway, suggesting that osthole might be a promising therapeutic agent for the treatment of atherosclerosis.

Fenofibrate Interferes with the Diapedesis of Lung Adenocarcinoma Cells through the Interference with Cx43/EGF-Dependent Intercellular Signaling.

Extravasation of circulating cancer cells is regulated by the intercellular/intracellular signaling pathways that locally impair the endothelial barrier function. Co-cultures of human umbilical vein endothelial cells (HUVECs) with lung adenocarcinoma A549 cells enabled us to identify these pathways and to quantify the effect of fenofibrate (FF) on their activity. A549 cells induced the disruption and local activation of endothelial continuum. These events were accompanied by epidermal growth factor (EGF) up-regulation in endothelial cells. Impaired A549 diapedesis and HUVEC activation were seen upon the chemical inhibition of connexin(Cx)43 functions, EGF/ERK1/2-dependent signaling, and RhoA/Rac1 activity. A total of 25 μM FF exerted corresponding effects on Cx43-mediated gap junctional coupling, EGF production, and ERK1/2 activation in HUVEC/A549 co-cultures. It also directly augmented endothelial barrier function via the interference with focal adhesion kinase (FAK)/RhoA/Rac1-regulated endothelial cell adhesion/contractility/motility and prompted the selective transmigration of epithelioid A549 cells. N-acetyl-L-cysteine abrogated FF effects on HUVEC activation, suggesting the involvement of PPARα-independent mechanism(s) in its action. Our data identify a novel Cx43/EGF/ERK1/2/FAK/RhoA/Rac1-dependent signaling axis, which determines the efficiency of lung cancer cell diapedesis. FF interferes with its activity and reduces the susceptibility of endothelial cells to A549 stimuli. These findings provide the rationale for the implementation of FF in the therapy of malignant lung cancers.

Dysregulated Txnip-ROS-Wnt axis contributes to the impaired ischemic heart repair in diabetic mice

Hyperglycemia-induced impairment of angiogenesis contributes to the unfavorable prognosis of myocardial ischemia in long-standing diabetes mellitus. The underlying mechanism remains largely unknown and therapeutic strategies thereby limited. In the present study, we investigated the possible involvement of thioredoxin-interacting protein (TXNIP) and Wnt/β-catenin signaling in the context, and their possible relation was also explored. STZ induced diabetic mice were subjected to myocardial infarction (MI). Adenovirus expressing shTXNIP, shCtnnb1 (β-catenin) driven by VE-Cadherin promoter was administered intramyocardially immediately after MI. Cardiac function, histology, and molecular analyses were performed at predetermined time points. Increased endothelial expression of TXNIP was found in diabetic hearts, which correlated well with reduced nuclear β-catenin expression, insufficient angiogenesis, aggravated cardiac remodeling, and poor survival. Endothelial-specific knockdown of TXNIP significantly rescued β-catenin activity, together with increased angiogenesis, preserved cardiac function, and improved survival rate. Moreover, additional knockdown of β-catenin essentially reversed the beneficial effects of TXNIP downregulation. In vitro, high glucose treatment of human umbilical vein endothelial cells (HUVECs) increased TXNIP levels and ROS concentration, while it reduced β-catenin activity. Silencing TXNIP or ROS scavenger restored the high glucose induced reduction of Wnt/β-catenin activity in HUVECs. In addition, either reduction of TXNIP expression or supplementation of exogenous Wnt3a improved the HUVECs quantity and migration under high glucose conditions. Diabetes-induced increase of TXNIP expression in the endothelium contributes to impaired angiogenesis after MI, especially via the elevation of ROS and the impaired Wnt/β-catenin signaling. Targeting TXNIP-ROS-Wnt is a promising strategy in improving the prognosis.

Gemigliptin Inhibits IL-1ß–Induced Endothelial-Mesenchymal Transition via BMP2/Smad/MAPK/Runx2 Pathway

Objective: Endothelial-to-mesenchymal transition (EndMT) contributes to inflammatory conditions inducing conversion of endothelial cells (ECs) into activated fibroblasts, promoting fibrotic diseases. Pro-inflammatory cytokine interleukin (IL)-1ß is the most potent inducer of EndMT. We investigated inhibition of IL-1ß-induced EndMT by gemigliptin, a DPP-4 inhibitor. Methods: We exposed human umbilical vein ECs (HUVECs) to 10 ng/mL IL-1ß/20 uM gemigliptin and analyzed the expression of endothelial, smooth muscle, mesenchymal, and osteoblastic markers, bone morphogenetic protein (BMP), Smad, and non-Smad signaling pathway proteins. Results: Morphological changes showed gemigliptin blocked IL-1ß-induced EndMT, upregulated EC markers, and downregulated smooth muscle and mesenchymal markers. IL-1ß activation of HUVECs is initiated by the BMP/Smad and canonical BMP signaling pathways. Gemigliptin inhibited IL-1ß induction of BMP2 and 7, activin receptor type IA, BMP receptor type IA, and BMP receptor II. Reversal of IL-1ß-mediated inhibition of BMP-induced Smad 1/5/8, Smad2, and Smad3 phosphorylation by gemigliptin suggests involvement of the Smad pathway in gemigliptin action. In the canonical BMP pathway, gemigliptin treatment significantly increased the deactivation of ERK, p38, and JNK by IL-1ß. Gemigliptin treatment suppressed BMP-2-induced expression of key osteoblastic markers including osterix and runt-related transcription factor 2 during IL-1ß-induced EndMT. Conclusions: We demonstrated a novel protective mechanism of gemigliptin against fibrosis by suppressing IL-1ß-induced EndMT. Disclosure H. Kwon: None. O. Hong: None. Y. Kang: None. H. Kwon: None. S. Yoo: None.

An in vitro study of the angiogenic effects of concentrate growth factor on human umbilical vein endothelial cells

This study aimed to explore the effects of concentrate growth factor extracts (CGFe) on human umbilical vein endothelial cells (HUVECs) in vitro. Concentrate growth factor (CGF) were prepared from the peripheral blood of healthy donors, followed by CGFe. Four groups were designed based on cell culture medium, as follows: 2%CGFe, 5%CGFe, 10%CGFe, and control. The proliferation activity of HUVECs was detected by cell cycle and CCK-8 assays. The migration of HUVECs was detected by scratch assay. The mRNA expression levels of vascular endothelial growth factor (VEGF), chemokine receptor 4 (CXCR4), and platelet derived growth factor (PDGF) were examined by quantitative real time polymerase chain reaction (qRT-PCR). Results of CCK-8 and cell cycle assays showed that CGFe promoted the proliferation capability of HUVECs in a dose-dependent manner, and the data had statistical significance among four groups (P< 0.05). The cell migration assay indicated that CGF accelerated wound closure in a dose-dependent manner after 12 h of culture (P<0.05). The results of qRT-PCR showed that CGF upregulated the expression levels of VEGF, CXCR4, and PDGF in HUVECs. CGFe can promote the proliferation, migration, and angiogenic differentiation of HUVECs. Thus, CGF might be an appropriate cure for dental pulp revascularization.

Ticagrelor, a P2Y12 antagonist, attenuates vascular dysfunction and inhibits atherogenesis in apolipoprotein-E-deficient mice

Background and aimsTicagrelor reduces cardiovascular events in patients with acute coronary syndrome (ACS). Recent studies demonstrated the expression of P2Y12 on vascular cells including endothelial cells, as well as platelets, and suggested its contribution to atherogenesis. We investigated whether ticagrelor attenuates vascular dysfunction and inhibits atherogenesis in apolipoprotein E-deficient (apoe−/−) mice. MethodsEight-week-old male apoe−/− mice were fed a western-type diet (WTD) supplemented with 0.1% ticagrelor (approximately 120 mg/kg/day). Non-treated animals on WTD served as control. Atherosclerotic lesions were examined by en-face Sudan IV staining, histological analyses, quantitative RT-PCR analysis, and western blotting. Endothelial function was analyzed by acetylcholine-dependent vasodilation using aortic rings. Human umbilical vein endothelial cells (HUVEC) were used for in vitro experiments. ResultsTicagrelor treatment for 20 weeks attenuated atherosclerotic lesion progression in the aortic arch compared with control (p < 0.05). Ticagrelor administration for 8 weeks attenuated endothelial dysfunction (p < 0.01). Ticagrelor reduced the expression of inflammatory molecules such as vascular cell adhesion molecule-1, macrophage accumulation, and lipid deposition. Ticagrelor decreased the phosphorylation of JNK in the aorta compared with control (p < 0.05). Ticagrelor and a JNK inhibitor ameliorated impairment of endothelium-dependent vasodilation by adenosine diphosphate (ADP) in wild-type mouse aortic segments. Furthermore, ticagrelor inhibited the expression of inflammatory molecules which were promoted by ADP in HUVEC (p < 0.001). Ticagrelor also inhibited ADP-induced JNK activation in HUVEC (p < 0.05). ConclusionsTicagrelor attenuated vascular dysfunction and atherogenesis through the inhibition of inflammatory activation of endothelial cells. These effects might be a potential mechanism by which ticagrelor decreases cardiovascular events in patients with ACS.

SB202190 inhibits endothelial cell apoptosis via induction of autophagy and heme oxygenase-1.

Activation of the p38 mitogen-activated protein kinase (MAPK) pathway has been implicated in various detrimental events finally leading to endothelial dysfunction. The present study therefore investigates the impact of the p38 MAPK inhibitor SB202190 on the expression of the cytoprotective enzyme heme oxygenase-1 (HO-1) as well as metabolic activity, apoptosis and autophagy of endothelial cells. Using human umbilical vein endothelial cells (HUVEC) SB202190 was found to cause a time- and concentration-dependent induction of HO-1 protein. Induction of HO-1 protein expression was mimicked by SB203580, another p38 MAPK inhibitor, but not by SB202474, an inactive structural analogue of p38 MAPK inhibitors. HO-1 induction by both SB202190 and SB203580 was also demonstrated by analysis of mRNA expression. On the functional level, SB202190 was shown to increase metabolic activity and autophagy of HUVEC along with diminishing basal apoptosis. Treatment of cells with tin protoporphyrin IX (SnPPIX), a well-characterised HO-1 enzymatic inhibitor, or HO-1 siRNA left SB202190-modulated metabolic activity and autophagy virtually unaltered but caused a significant reversal of the anti-apoptotic action of SB202190. Conversely, however, HO-1 expression by SB202190 became completely suppressed by the autophagy inhibitor bafilomycin A1. Bafilomycin A1 likewise fully reversed effects of SB202190 on metabolic activity and apoptosis, albeit significantly inducing apoptosis per se. Collectively, this work demonstrates SB202190 to confer upstream induction of autophagy followed by HO-1 induction resulting in potential protective effects against apoptosis. On the other hand, our data oppose HO-1 to contribute to SB202190-mediated increases in metabolic activity and autophagy, respectively.

Expression of microRNA Genes MIR221, MIR222, and MIR181B1 Increases during Induction of Inflammation in the Endothelial Barrier Model.

We studied expression profile of microRNA and their target genes in human umbilical vein endothelial cells (HUVEC) during proinflammatory activation with TNFα. TNFα-induced activation of HUVEC was accompanied by a decrease in the expression of CDKN1B, HIST1H3D, and OIP5 genes that are the common target genes for mature microRNA encoded by MIR221, MIR222, and MIR181B1 genes, whose expression increases in activated cells. Proteins encoded by HIST1H3D and OIP5 genes are associated with chromatin compaction and cell cycle. Our results suggest that fetal endothelial microRNA can appear in the maternal blood during various pathological states, e.g., under conditions of preeclampsia.

Over expressing miR-19b-1 suppress breast cancer growth by inhibiting tumor microenvironment induced angiogenesis.

Metastastic breast cancer, especially triple-negative (TN), is one of the most common causes of cancer related deaths in women. Recent years, tumor-associate vessel formation is considered as a more generally and effective target to patient with malignant cancer. Our previous studies indicated that miR-19b-1 controls the intrinsic angiogenic activity of human umbilical vein endothelial cells (HUVECs) in vitro. In this study, in silico analysis indicated VEZF1 that implicates in angiogenesis and triple-negative breast cancer progression might be the target of miR-19b-1. Further investigation showed that overexpressing miR-19b-1 in human triple-negative breast cancer cell line(MDA-MB-231) led to impaired angiogenic activity of HUVECs in vitro and in vivo. Additionally, miR-19b-1 upregulation inhibited the vessel imitation mediated by MDA-MB-231. Furthermore, enforced expression of miR-19b-1 in MDA-MB-231 has impact on HUVECs apoptosis and altered the mitochondrial membrane potential, which suggested the possible mechanism involved in interaction between breast cancer and endothelial cells. What's more, we found that miR-19b-1 stable overexpression in MDA-MB-231 caused tumor growth arresting entirely. Besides, our results suggested that miR-19b-1 could inhibit angiogenesis by possibly targeting VEGF receptor endocytosis signaling pathway. Taking together, our findings suggest an important role of miR-19b-1 in crosstalk between metastatic breast cancer and endothelial cells and provide us new insights for exploring miR-19b-1 and its multiple targets as promising therapeutic candidates to interfere breast cancer progression.

Intracellular acidification reduces l-arginine transport via system y+L but not via system y+/CATs and nitric oxide synthase activity in human umbilical vein endothelial cells

l-Arginine is taken up via the cationic amino acid transporters (system y+/CATs) and system y+L in human umbilical vein endothelial cells (HUVECs). l-Arginine is the substrate for endothelial NO synthase (eNOS) which is activated by intracellular alkalization, but nothing is known regarding modulation of system y+/CATs and system y+L activity, and eNOS activity by the pHi in HUVECs. We studied whether an acidic pHi modulates l-arginine transport and eNOS activity in HUVECs. Cells loaded with a pH-sensitive probe were subjected to 0.1–20 mmol/L NH4Cl pulse assay to generate pHi 7.13–6.55. Before pHi started to recover, l-arginine transport (0–20 or 0–1000 μmol/L, 10 s, 37 °C) in the absence or presence of 200 μmol/L N-ethylmaleimide (NEM) (system y+/CATs inhibitor) or 2 mmol/L l-leucine (systemy+L substrate) was measured. Protein abundance for eNOS and serine1177 or threonine495 phosphorylated eNOS was determined. The results show that intracellular acidification reduced system y+L but not system y+/CATs mediated l-arginine maximal transport capacity due to reduced maximal velocity. Acidic pHi reduced NO synthesis and eNOS serine1177 phosphorylation. Thus, system y+L activity is downregulated by an acidic pHi, a phenomenon that may result in reduced NO synthesis in HUVECs.

Inhibition of angiogenesis by leflunomide via targeting the soluble ephrin-A1/EphA2 system in bladder cancer

Angiogenesis plays an important role in bladder cancer (BCa). The immunosuppressive drug leflunomide has attracted worldwide attention. However, the effects of leflunomide on angiogenesis in cancer remain unclear. Here, we report the increased expression of soluble ephrin-A1 (sEphrin-A1) in supernatants of BCa cell lines (RT4, T24, and TCCSUP) co-cultured with human umbilical vein endothelial cells (HUVECs) compared with that in immortalized uroepithelial cells (SV-HUC-1) co-cultured with HUVECs. sEphrin-A1 is released from BCa cells as a monomeric protein that is a functional form of the ligand. The co-culture supernatants containing sEphrin-A1 caused the internalization and down-regulation of EphA2 on endothelial cells and dramatic functional activation of HUVECs. This sEphrin-A1/EphA2 system is mainly functional in regulating angiogenesis in BCa tissue. We showed that leflunomide (LEF) inhibited angiogenesis in a N-butyl-N-(4-hydroxybutyl)-nitrosamine (BBN)-induced bladder carcinogenesis model and a tumor xenograft model, as well as in BCa cell and HUVEC co-culture systems, via significant inhibition of the sEphrin-A1/EphA2 system. Ephrin-A1 overexpression could partially reverse LEF-induced suppression of angiogenesis and subsequent tumor growth inhibition. Thus, LEF has a significant anti-angiogenesis effect on BCa cells and BCa tissue via its inhibition of the functional angiogenic sEphrin-A1/EphA2 system and may have potential for treating BCa beyond immunosuppressive therapy.

Biomimetic Composite Scaffold Containing Small Intestinal Submucosa and Mesoporous Bioactive Glass Exhibits High Osteogenic and Angiogenic Capacity.

Biomaterials with excellent osteogenic and angiogenic activities are desirable to repair massive bone defects. Decellularized matrix from porcine small intestinal submucosa (SIS) has attracted particular attention for tissue regeneration because it has strong angiogenic effects and retains plentiful bioactive components. However, it has inferior osteoinductivity and osteoconductivity. In this study, we developed porous composite of SIS combined with mesoporous bioactive glass (SIS/MBG) with the goal of improving the mechanical and biological properties. SIS/MBG scaffolds showed uniform interconnected macropores (∼150 μm), high porosity (∼76%), and enhanced compressive strength (∼0.87 MPa). The proliferation and osteogenic gene expression (Runx2, ALP, Ocn, and Col-Iα) of rat bone marrow stromal cells (rBMSCs) as well as the proliferation, angiogenic gene expression (VEGF, bFGF, and KDR), and tube formation capacity of human umbilical vein endothelial cells (HUVECs) in SIS/MBG scaffolds were significantly upregulated compared with nonmesoporous bioactive glass (BG)-modified SIS (SIS/BG) and SIS-only scaffolds. Western blot analysis revealed that SIS/MBG induced rBMSCs to osteogenic differentiation through the activation of Wnt/β-Catenin signaling pathway, and SIS/MBG enhanced angiogenic activity of HUVEC through the activation of PI3k/Akt pathways. The in vivo results demonstrated that SIS/MBG scaffolds significantly enhanced new bone formation and neovascularization simultaneously in critical-sized rat calvarial defects as compared with SIS/BG and SIS. Collectively, the osteostimulative and angiostimulative biomimetic composite scaffold SIS/MBG represents an exciting biomaterial option for bone regeneration.

Exosomal Metastasis‑Associated Lung Adenocarcinoma Transcript 1 Promotes Angiogenesis and Predicts Poor Prognosis in Epithelial Ovarian Cancer.

Exosomes mediate cell-cell crosstalk in cancer progression by transferring their molecular cargos, including long noncoding RNAs (lncRNAs). Metastasis‑associated lung adenocarcinoma transcript 1 (MALAT1) is a well-known lncRNA associated with cancer angiogenesis and metastasis. However, the presence of MALAT1 in exosomes and the roles and clinical values of exosomal MALAT1 in epithelial ovarian cancer (EOC) remain unknown. The present study focused on the crosstalk between EOC cells and endothelial cells mediated by exosomal MALAT1 and aimed to explore the roles of exosomes and exosomal MALAT1 in EOC angiogenesis and to reveal the clinical relevance and prognostic predictive value of serum exosomal MALAT1 in EOC. We observed that MALAT1 was increased in both metastatic EOC cells and their secreted exosomes. Exosomal MALAT1 derived from EOC cells was transferred to recipient human umbilical vein endothelial cells (HUVECs) via exosomes. In vitro and in vivo experiments demonstrated that MALAT1 knockdown impaired the exosome-mediated proangiogenic activity of HUVECs through certain key angiogenesis-related genes. Clinically, elevated serum exosomal MALAT1 was highly correlated with an advanced and metastatic phenotype of EOC and was an independent predictive factor for EOC overall survival (OS). Moreover, a prognostic nomogram model we constructed showed a good prediction of the probability of 3-year OS of EOC patients according to the c-index (0.751, 95% confidence interval [CI]=0.691-0.811) and calibration curve. Collectively, our data provide a novel mechanism by which EOC cells transfer MALAT1 via exosomes to recipient HUVECs and influence HUVECs by stimulating angiogenesis-related gene expression, eventually promoting angiogenesis. Additionally, circulating exosomal MALAT1 can serve as a promising serum-based, noninvasive predictive biomarker for EOC prognosis.

Ultrasound-Stimulated Microbubbles Enhance Radiosensitization of Nasopharyngeal Carcinoma

Background/Aims: Recent studies indicate that therapies targeting the vasculature can significantly sensitize tumors to radiation. Ultrasound-stimulated microbubbles (USMBs) are regarded as a promising radiosensitizer. In this study, we investigated the effect of USMBs on the sensitivity of nasopharyngeal carcinoma (NPC) to radiation. Methods: Human NPC (CNE-2) cells and human umbilical vein endothelial cells (HUVECs) were exposed to radiation (0, 2, and 8 Gy) alone or in combination with USMBs. Cell viability and apoptosis were measured with the MTT assay and flow cytometry, respectively. The angiogenic activity of HUVECs was detected using matrigel tubule formation. The in vitro effects induced by these treatments were confirmed in vivo with xenograft models of CNE-2 cells in nude mice by examining vascular integrity using color Doppler flow imaging and cell survival using immunohistochemistry. Additionally, the in vivo and in vitro expressions of angiotensin II (ANG II) and its receptor (AT1R) were detected by immunohistochemistry and western blotting, respectively. With CNE-2 cells and HUVECs transfected with control, ANG II, or AT1R, perindopril (an inhibitor of angiotensin-converting enzyme) and candesartan (an inhibitor of AT1R) were used to verify the role of ANG II and AT1R in the radiosensitivity of tumor and endothelial cells by USMBs, by determining cell viability and apoptosis and angiogenic activity. Results: In the NPC xenografts, USMBs slightly reduced blood flow and CD34 expression, increased tumor cell death and ANG II and AT1R expression, and significantly enhanced the effects of radiation. With CNE-2 cells and HUVECs, the USMBs further enhanced the inhibition of tumor cell viability and endothelial tubule formation and further enhanced the increase in ANG II and AT1R due to radiation. Furthermore, perindopril and candesartan significantly enhanced the inhibitory effect of radiation and USMBs on tumor cell growth and angiogenesis in vitro. Conclusions: We have demonstrated for the first time that USMB exposure can significantly enhance the destructive effect on NPC of radiation, and this effect might be further increased by ANG II and AT1R inhibition. Our findings suggest that USMBs can be used as a promising sensitizer of radiotherapy to treat NPC, and the clinical effect might be increased by ANG II and AT1R inhibition.

REST upregulates gremlin to modulate diffuse intrinsic pontine glioma vasculature.

Diffuse intrinsic pontine glioma (DIPG) is a highly aggressive glial tumor that occurs in children. The extremely poor median and 5-year survival in children afflicted with DIPG highlights the need for novel biology-driven therapeutics. Here, we have implicated the chromatin remodeler and regulator of brain development called RE1 Silencing Transcription Factor (REST), in DIPG pathology. We show that REST protein is aberrantly elevated in at least 21% of DIPG tumors compared to normal controls. Its knockdown in DIPG cell lines diminished cell growth and decreased their tumorigenicity in mouse intracranial models. DIPGs are vascularized tumors and interestingly, REST loss in DIPG cells also caused a substantial decline in tumor vasculature as measured by a decrease in CD31 and VEGFR2 staining. These observations were validated in vitro, where a significant decline in tube formation by human umbilical vein endothelial cells (HUVEC) was seen following REST-loss in DIPG cells. Mechanistically, REST controlled the secretion of a pro-angiogenic molecule and ligand for VEGFR2 called Gremlin-1 (GREM-1), and was associated with enhanced AKT activation. Importantly, the decline in tube formation caused by REST loss could be rescued by addition of recombinant GREM-1, which also caused AKT activation in HUVECs and human brain microvascular endothelial cells (HBMECs). In summary, our study is the first to demonstrate autocrine and paracrine functions for REST in DIPG development. It also provides the foundation for future investigations on anti-angiogenic therapies targeting GREM-1 in combination with drugs that target REST-associated chromatin remodeling activities.

Teneligliptin enhances the beneficial effects of GLP-1 in endothelial cells exposed to hyperglycemic conditions.

High-glucose-induced oxidative stress contributes to cardiovascular endothelial damage in diabetes. Glucagon-like peptide 1 (GLP-1) is beneficial to endothelial cells, but its effects are diminished when cells are continuously exposed to high glucose. Teneligliptin is a dipeptidyl peptidase-4 (DPP-4) inhibitor that prevents oxidative stress, apoptosis and the metabolic memory effect. We explored the potential additive effects of Teneligliptin and GLP-1 in hyperglycemia-damaged endothelial cells. Human umbilical vein endothelial cells (HUVECs) were exposed to normal-glucose (5 mmol/L) or high-glucose (HG, 25 mmol/L) for 21 days, or to HG for 14 days followed by normal-glucose for 7 days (HM). These cells were continually treated with Teneligliptin 3.0 μmol/L, alone or in combination with an acute dose of GLP-1 50 nmol/L. DPP-4 was upregulated under hyperglycemic conditions, but Teneligliptin reduced DPP-4 expression and activity. Simultaneous Teneligliptin and GLP-1 synergistically increased the antioxidant response and reduced ROS levels in HG- and HM-exposed HUVECs. Concurrent treatment also enhanced cell proliferation, reduced apoptotic gene expression and ameliorated endoplasmic reticulum stress in HG- and HM-exposed HUVECs. Thus, long-term Teneligliptin treatment reduced DPP-4 levels and activity in HUVECs exposed to chronic hyperglycemia. Moreover, Teneligliptin enhanced the beneficial effects of GLP-1 on oxidative stress, proliferation, apoptosis and endoplasmic reticulum homeostasis.

Zingerone Suppresses the Shedding of Endothelial Protein C Receptor

Zingerone (ZGR), a phenolic alkanone found in Zingiber officinale, has been reported to have various pharmacological activities including anti-inflammatory and anti-apoptotic activities. The endothelial cell protein C receptor (EPCR) plays an important role in the cytoprotective pathway and activation of protein C EPCR can be shed from the cell surface, which is mediated by tumor necrosis factor-α converting enzyme (TACE). However, little is known about the effects of ZGR on EPCR shedding. We investigated this by monitoring the effects of ZGR on phorbol-12-myristate 13-acetate (PMA)-, tumor necrosis factor (TNF)-a, and interleukin (IL)-1p-induced EPCR shedding in human umbilical vein endothelial cells (HUVECs), and cecal ligation and puncture (CLP)-mediated EPCR shedding in mice, as well as by analyzing the underlying mechanisms. Here, ZGR triggered potent inhibition of PMA-, TNF-α-, IL-1β-and CLP-induced EPCR shedding through the inhibition of phosphorylation of mitogen-activated protein kinases (MAPKs) such as p38, janus kinase (JNK), and extracellular signal-regulated kinase (ERK) 1/2. ZGR also inhibited PMA-induced TACE expression and activity in HUVECs, suggesting that p38, ERK1/2, and JNK could be molecular targets of ZGR. These results demonstrate the potential of ZGR as an agent against PMA- and CLP-mediated EPCR shedding.

Maspin impairs the function of endothelial cells: an implying pathway of preeclampsia

BackgroudWidespread endothelial injury contributes to the occurrence of preeclampsia. Maspin, first identified as a tumor suppressor, plays a critical role in cell invasion and angiogenesis. Our previous studies found that the expression of maspin was increased in preeclampsic placenta. In this research, we studied the function of human umbilical vein endothelial cells (HUVECs) to explore the role and possible mechanism of maspin gene in the pathogenesis of preeclampsia.MethodsHUVECs were treated with different concentration of recombinant human maspin protein (r-maspin) during normoxia and hypoxia, we detected the proliferation, apoptosis, migration and tube formation of HUVECs. We also assessed nitride oxide (NO) synthesis and the expression of matrix metalloproteinase 2 (MMP2) to further explore the underlying molecular mechanism.ResultsThere was only slight maspin expression at mRNA level in HUVECs. Treated HUVECs with r-maspin, the proliferation of HUVECs was significantly promoted both under normoxia and hypoxia. The tubes formed by HUVECs were significantly inhibited and NO synthesis was significantly reduced by r-maspin. Meantime, r-maspin also inhibited MMP2 expression and activity in HUVECs. However, there was no significant change in the migration and apoptosis of HUVECs.ConclusionsMaspin may be an important participant for mediating endothelial function and ultimately leads to the occurence of preeclamsia.

Von Willebrand factor protects against acute CCl4-induced hepatotoxicity through phospho-p38 MAPK signaling pathway inhibition.

The blood glycoprotein von Willebrand factor (vWF) is involved in coagulopathy and inflammation; however, its role in the pathogenesis of acute liver failure, as suggested by its higher expression levels in such patients, remains unknown. In this study, vWF-knockout (KO) mice showed more severe carbon tetrachloride (CCl4)-induced liver injury than wild-type mice. Patients with acute liver injury also showed elevated vWF protein activity and expression in liver tissues, as compared to healthy individuals. Using the mouse model and cultured human umbilical vein endothelial cells (HUVECs), CCl4 was found to directly increase vWF protein expression through interaction with the highly expressed vWF receptor, GPIbα. Microarray analysis revealed that the genes showing the most differential expression in response to CCl4-induced liver injury and vWF deficiency were related to the MAPK signaling pathway. Subsequent inhibition of vWF protein activity in HUVECs led to activation of the MAPK signal pathway and elevated production of FGL2, and treatment with a phospho-p38 inhibitor suppressed the CCl4-induced production of FGL2. Exposure of liver sinusoidal endothelial cells isolated from the vWF-KO acute liver injury model mice to phospho-p38 inhibitor also decreased FGL2 expression. The vWF/GPIbα axis plays a protective role against development of acute liver injury by attenuating FGL2 production through the MAPK signaling pathway. Collectively, these data provide insight into the pathogenesis of acute liver injury and a potential novel strategy for its treatment.