scholarly journals Treatment with a Combination of Metformin and 2-Deoxyglucose Upregulates Thrombospondin-1 in Microvascular Endothelial Cells: Implications in Anti-Angiogenic Cancer Therapy

Cancers ◽  
2019 ◽  
Vol 11 (11) ◽  
pp. 1737 ◽  
Author(s):  
Samson Mathews Samuel ◽  
Noothan Jyothi Satheesh ◽  
Suparna Ghosh ◽  
Dietrich Büsselberg ◽  
Yasser Majeed ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
Endothelial Cells ◽  
Growth Factor Receptor ◽  
Cyclin B1 ◽  
Mtor Pathway ◽  
Microvascular Endothelial Cells ◽  
Thrombospondin 1 ◽  
Microvascular Endothelial ◽  
Related Proteins

Metformin, the most widely used anti-diabetic drug, also exhibits anti-cancer properties; however, the true potential of metformin as an anticancer drug remains largely unknown. In this study using mouse microvascular endothelial cells (MMECs), we investigated the effects of metformin alone or in combination with the glycolytic inhibitor, 2-deoxyglucose (2DG), on angiogenesis-a process known to be an integral part of tumor growth, cancer cell survival and metastasis. MMECs were exposed to 2DG (1–10 mM) for 48 h in the absence or presence of metformin (2 mM). The status of angiogenic and anti-angiogenic marker proteins, proteins of the mTOR pathway and cell-cycle-related proteins were quantified by Western blot analysis. Assays for cell proliferation, migration and tubulogenesis were also performed. We observed robust up-regulation of anti-angiogenic thrombospondin-1 (TSP1) and increased TSP1-CD36 co-localization with a marked decrease in the levels of phosphorylated vascular endothelial growth factor receptor-2 (pVEGFR2; Y1175) in 2DG (5 mM) exposed cells treated with metformin (2 mM). Additionally, treatment with metformin and 2DG (5 mM) inhibited the Akt/mTOR pathway and down-regulated the cell-cycle-related proteins such as p-cyclin B1 (S147) and cyclins D1 and D2 when compared to cells that were treated with either 2DG or metformin alone. Treatment with a combination of 2DG (5 mM) and metformin (2 mM) also significantly decreased cell proliferation, migration and tubulogenic capacity when compared to cells that were treated with either 2DG or metformin alone. The up-regulation of TSP1, inhibition of cell proliferation, migration and tubulogenesis provides support to the argument that the combination of metformin and 2DG may prove to be an appropriate anti-proliferative and anti-angiogenic therapeutic strategy for the treatment of some cancers.

scholarly journals MicroRNA-126-3p Inhibits Angiogenic Function of Human Lung Microvascular Endothelial Cells via LAT1 (L-Type Amino Acid Transporter 1)-Mediated mTOR (Mammalian Target of Rapamycin) Signaling

2020 ◽  
Vol 40 (5) ◽  
pp. 1195-1206 ◽  
Author(s):  
Danting Cao ◽  
Andrew M. Mikosz ◽  
Alexandra J. Ringsby ◽  
Kelsey C. Anderson ◽  
Erica L. Beatman ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cells ◽  
Amino Acid ◽  
Cell Apoptosis ◽  
Human Lung ◽  
Mammalian Target Of Rapamycin ◽  
Tube Formation ◽  
Microvascular Endothelial Cells ◽  
Microvascular Endothelial

Objective: MicroRNA-126-3p (miR-126) is required for angiogenesis during organismal development or the repair of injured arterial vasculature. The role of miR-126 in lung microvascular endothelial cells, which are essential for gas exchange and for lung injury repair and regeneration, remains poorly understood. Considering the significant heterogeneity of endothelial cells from different vascular beds, we aimed to determine the role of miR-126 in regulating lung microvascular endothelial cell function and to elucidate its downstream signaling pathways. Approach and Results: Overexpression and knockdown of miR-126 in primary human lung microvascular endothelial cells (HLMVEC) were achieved via transfections of miR-126 mimics and antisense inhibitors. Increasing miR-126 levels in HLMVEC reduced cell proliferation, weakened tube formation, and increased cell apoptosis, whereas decreased miR-126 levels stimulated cell proliferation and tube formation. Whole-genome RNA sequencing revealed that miR-126 was associated with an antiangiogenic and proapoptotic transcriptomic profile. Using validation assays and knockdown approaches, we identified that the effect of miR-126 on HLMVEC angiogenesis was mediated by the LAT1 (L-type amino acid transporter 1), via regulation of mTOR (mammalian target of rapamycin) signaling. Furthermore, downregulation of miR-126 in HLMVEC inhibited cell apoptosis and improved endothelial tube formation during exposure to environmental insults such as cigarette smoke. Conclusions: miR-126 inhibits HLMVEC angiogenic function by targeting the LAT1-mTOR signaling axis, suggesting that miR-126 inhibition may be useful for conditions associated with microvascular loss, whereas miR-126 augmentation may help control unwanted microvascular angiogenesis.

scholarly journals Thrombin Modulates the Expression of a Set of Genes Including Thrombospondin-1 in Human Microvascular Endothelial Cells

2005 ◽  
Vol 280 (23) ◽  
pp. 22172-22180 ◽  
Author(s):  
Joseph N. McLaughlin ◽  
Maria R. Mazzoni ◽  
John H. Cleator ◽  
Laurie Earls ◽  
Ana Luisa Perdigoto ◽  
...  
Keyword(s):  
Gene Expression ◽  
Extracellular Matrix ◽  
Endothelial Cells ◽  
Umbilical Vein ◽  
Microvascular Endothelial Cells ◽  
Microvascular Endothelium ◽  
Disease States ◽  
Similar Expression Profile ◽  
Thrombospondin 1 ◽  
Microvascular Endothelial

Thrombospondin-1 (THBS1) is a large extracellular matrix glycoprotein that affects vasculature systems such as platelet activation, angiogenesis, and wound healing. Increases in THBS1 expression have been liked to disease states including tumor progression, atherosclerosis, and arthritis. The present study focuses on the effects of thrombin activation of the G-protein-coupled, protease-activated receptor-1 (PAR-1) on THBS1 gene expression in the microvascular endothelium. Thrombin-induced changes in gene expression were characterized by microarray analysis of ∼11,000 different human genes in human microvascular endothelial cells (HMEC-1). Thrombin induced the expression of a set of at least 65 genes including THBS1. Changes in THBS1 mRNA correlated with an increase in the extracellular THBS1 protein concentration. The PAR-1-specific agonist peptide (TFLLRNK-PDK) mimicked thrombin stimulation of THBS1 expression, suggesting that thrombin signaling is through PAR-1. Further studies showed THBS1 expression was sensitive to pertussis toxin and protein kinase C inhibition indicating Gi/o- and Gq-mediated pathways. THBS1 up-regulation was also confirmed in human umbilical vein endothelial cells stimulated with thrombin. Analysis of the promoter region of THBS1 and other genes of similar expression profile identified from the microarray predicted an EBOX/EGRF transcription model. Expression of members of each family, MYC and EGR1, respectively, correlated with THBS1 expression. These results suggest thrombin formed at sites of vascular injury increases THBS1 expression into the extracellular matrix via activation of a PAR-1, Gi/o, Gq, EBOX/EGRF-signaling cascade, elucidating regulatory points that may play a role in increased THBS1 expression in disease states.

scholarly journals Protective Effects of Scutellarin on Human Cardiac Microvascular Endothelial Cells against Hypoxia-Reoxygenation Injury and Its Possible Target-Related Proteins

2015 ◽  
Vol 2015 ◽  
pp. 1-13 ◽  
Author(s):  
Meina Shi ◽  
Yingting Liu ◽  
Lixing Feng ◽  
Yingbo Cui ◽  
Yajuan Chen ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Expression Profiles ◽  
Interaction Network ◽  
Treated Group ◽  
Microvascular Endothelial Cells ◽  
Protective Effects ◽  
Microvascular Endothelial ◽  
Related Proteins ◽  
Cardiac Microvascular Endothelial Cells ◽  
Hypoxia Reoxygenation

Scutellarin (SCU) is one of the main components of traditional Chinese medicine plantErigeron breviscapus (Vant.)Hand.-Mazz. In this paper, we studied the protective effects of SCU on human cardiac microvascular endothelial cells (HCMECs) against hypoxia-reoxygenation (HR) injury and its possible target-related proteins. Results of MTT assay showed that pretreatment of SCU at doses of 1, 5, and 10 μM for 2 h could significantly inhibit the decrease in cell viability of HCMECs induced by HR injury. Subcellular fractions of cells treated with vehicle control, 1 μM SCU, HR injury, or 1 μM SCU + HR injury were separated by ultracentrifugation. The protein expression profiles of cytoplasm and membrane/nuclei fractions were checked using protein two-dimensional electrophoresis (2-DE). Proteins differentially expressed between control and SCU-treated group, control and HR group, or HR and SCU + HR group were identified using mass spectrometry (MS/MS). Possible interaction network of these target-related proteins was predicted using bioinformatic analysis. The influence of SCU on the expression levels of these proteins was confirmed using Western blotting assay. The results indicated that proteins such as p27BBP protein (EIF6), heat shock 60 kDa protein 1 (HSPD1), and chaperonin containing TCP1 subunit 6A isoform (CCT6A) might play important roles in the effects of SCU.

scholarly journals CD36 Mediates the In Vitro Inhibitory Effects of Thrombospondin-1 on Endothelial Cells

1997 ◽  
Vol 138 (3) ◽  
pp. 707-717 ◽  
Author(s):  
David W. Dawson ◽  
S. Frieda A. Pearce ◽  
Ruiqin Zhong ◽  
Roy L. Silverstein ◽  
William A. Frazier ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Fusion Proteins ◽  
Solid Phase ◽  
Umbilical Vein ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Microvascular Endothelial Cells ◽  
Thrombospondin 1 ◽  
Microvascular Endothelial

Thrombospondin-1 (TSP-1) is a naturally occurring inhibitor of angiogenesis that is able to make normal endothelial cells unresponsive to a wide variety of inducers. Here we use both native TSP-1 and small antiangiogenic peptides derived from it to show that this inhibition is mediated by CD36, a transmembrane glycoprotein found on microvascular endothelial cells. Both IgG antibodies against CD36 and glutathione-S-transferase–CD36 fusion proteins that contain the TSP-1 binding site blocked the ability of intact TSP-1 and its active peptides to inhibit the migration of cultured microvascular endothelial cells. In addition, antiangiogenic TSP-1 peptides inhibited the binding of native TSP-1 to solid phase CD36 and its fusion proteins, as well as to CD36-expressing cells. Additional molecules known to bind CD36, including the IgM anti-CD36 antibody SM∅, oxidized (but not unoxidized) low density lipoprotein, and human collagen 1, mimicked TSP-1 by inhibiting the migration of human microvascular endothelial cells. Transfection of CD36-deficient human umbilical vein endothelial cells with a CD36 expression plasmid caused them to become sensitive to TSP-1 inhibition of their migration and tube formation. This work demonstrates that endothelial CD36, previously thought to be involved only in adhesion and scavenging activities, may be essential for the inhibition of angiogenesis by thrombospondin-1.

scholarly journals Hemozoin impairs cell cycle progression and promotes chemokine release in human microvascular endothelial cells

2012 ◽  
Vol 11 (S1) ◽  
Author(s):  
Nicoletta Basilico ◽  
Daniela Girelli ◽  
Sarah D’Alessandro ◽  
Yolanda Corbett ◽  
Mauro Prato ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Endothelial Cells ◽  
Cell Cycle Progression ◽  
Microvascular Endothelial Cells ◽  
Cycle Progression ◽  
Microvascular Endothelial

scholarly journals Hypoxic proliferation requires EGFR-mediated ERK activation in human pulmonary microvascular endothelial cells

2017 ◽  
Vol 312 (5) ◽  
pp. L649-L656 ◽  
Author(s):  
Hilary A. White ◽  
Yi Jin ◽  
Louis G. Chicoine ◽  
Bernadette Chen ◽  
Yusen Liu ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Cellular Proliferation ◽  
Growth Factor Receptor ◽  
Viable Cell ◽  
Microvascular Endothelial Cells ◽  
Cell Numbers ◽  
Pulmonary Microvascular Endothelial Cells ◽  
Protein Levels ◽  
Microvascular Endothelial ◽  
Arginase Ii

We have previously shown that hypoxic proliferation of human pulmonary microvascular endothelial cells (hPMVECs) depends on epidermal growth factor receptor (EGFR) activation. To determine downstream signaling leading to proliferation, we tested the hypothesis that hypoxia-induced proliferation in hPMVECs would require EGFR-mediated activation of extracellular signal-regulated kinase (ERK) leading to arginase II induction. To test this hypothesis, hPMVECs were incubated in either normoxia (21% O2, 5% CO2) or hypoxia (1% O2, 5% CO2) and Western blotting was performed for EGFR, arginase II, phosphorylated-ERK (pERK), and total ERK (ERK). Hypoxia led to greater EGFR, pERK, and arginase II protein levels than did normoxia in hPMVECs. To examine the role of EGFR in these hypoxia-induced changes, hPMVECs were transfected with siRNA against EGFR or a scrambled siRNA and placed in hypoxia. Inhibition of EGFR using siRNA attenuated hypoxia-induced pERK and arginase II expression as well as the hypoxia-induced increase in viable cell numbers. hPMVECs were then treated with vehicle, an EGFR inhibitor (AG1478), or an ERK pathway inhibitor (U0126) and placed in hypoxia. Pharmacologic inhibition of EGFR significantly attenuated the hypoxia-induced increase in pERK level. Both AG1478 and U0126 also significantly attenuated the hypoxia-induced increase in viable hPMVECs numbers. hPMVECs were transfected with an adenoviral vector containing arginase II (AdArg2) and overexpression of arginase II rescued the U0126-mediated decrease in viable cell numbers in hypoxic hPMVECs. Our findings suggest that hypoxic activation of EGFR results in phosphorylation of ERK, which is required for hypoxic induction of arginase II and cellular proliferation.

scholarly journals Comparative proteome analysis of rat brain and coronary microvascular endothelial cells

2007 ◽  
pp. 159-168
Author(s):  
L Lu ◽  
PY Yang ◽  
Y-Ch Rui ◽  
H Kang ◽  
J Zhang ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Proteome Analysis ◽  
Microvascular Endothelial Cells ◽  
Ionization Time ◽  
Proteomic Approach ◽  
Microvascular Endothelial ◽  
Signal Transduction Proteins ◽  
Ph Range ◽  
Related Proteins ◽  
Dimensional Electrophoresis

The endothelium of different organs displays a remarkable heterogeneity, although it presents many common functional and morphological features. However, despite our knowledge of heterogeneity among endothelial cells from different sites, the differences between brain microvascular endothelial cells (BMEC) and coronary microvascular endothelial cells (CMEC) are poorly defined. The aim of this study was to investigate whether BMEC are distinct from CMEC at the protein level. Using the proteomic approach, we comparatively analyzed the proteome of cultured BMEC and CMEC. We reproducibly separated over 2000 polypeptides by using two-dimensional electrophoresis (2-DE) at pH range of 3-10. Using PDQuest software to process the 2-DE gel images, forty-seven protein spots were differentially expressed in the two-endothelial cells. Of these, thirty-five proteins are highly expressed in BMEC, whereas twelve proteins are highly expressed in CMEC. Fifteen proteins in BMEC and seven proteins in CMEC were identified with high confidence by matrix-associated laser desorption ionization time-of-flight mass spectrometer (MALDI-TOF-MS). Our data suggested that BMEC and CMEC were different in several aspects including cytokine and growth-related molecules, stress-related proteins, metabolic enzymes, signal transduction proteins and others. The identification of a set of proteins preferentially expressed in BMEC and CMEC provided new data on the heterogeneity of the endothelium.

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