scholarly journals Profiling of Cytokines Secreted by Conventional Aqueous Outflow Pathway Endothelial Cells Activated In Vitro and Ex Vivo With Laser Irradiation

2015 ◽  
Vol 56 (12) ◽  
pp. 7100 ◽  
Author(s):  
Jorge A. Alvarado ◽  
Phuonglan Chau ◽  
Jianfeng Wu ◽  
Richard Juster ◽  
Amde Selassie Shifera ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Laser Irradiation ◽  
Ex Vivo ◽  
Outflow Pathway ◽  
Aqueous Outflow

Antiangiogenic effects of phospholipase A2 Lys49 BnSP-7 from Bothrops pauloensis snake venom on endothelial cells: An in vitro and ex vivo approach

2021 ◽  
Vol 72 ◽  
pp. 105099
Author(s):  
Lorena Polloni ◽  
Fernanda Van Petten Vasconcelos Azevedo ◽  
Samuel Cota Teixeira ◽  
Eloá Moura ◽  
Tassia Rafaela Costa ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Phospholipase A2 ◽  
Snake Venom ◽  
Ex Vivo ◽  
Ex Vivo Approach

scholarly journals Neutrophil extracellular traps induce MCP-1 release from endothelial cells at the plaque rupture site in acute myocardial infarction

2020 ◽  
Vol 41 (Supplement_2) ◽  
Author(s):  
A Ondracek ◽  
T.M Hofbauer ◽  
A Mangold ◽  
T Scherz ◽  
V Seidl ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Acute Myocardial Infarction ◽  
Endothelial Cells ◽  
Plaque Rupture ◽  
Ex Vivo ◽  
Coronary Intervention ◽  
Neutrophil Extracellular Traps ◽  
Funding Source ◽  
Extracellular Traps

Abstract Introduction Leukocyte-mediated inflammation is crucial in acute myocardial infarction (AMI). We recently observed that neutrophil extracellular traps (NETs) are increased at the culprit site, promoting activation and differentiation of fibrocytes, cells with mesenchymal and leukocytic properties. Fibrocyte migration is mediated by monocyte chemoattractant protein (MCP)-1 and C-C chemokine receptor type 2 (CCR2). We investigated the interplay between NETs, fibrocyte function, and MCP-1 in AMI. Methods Culprit site and femoral blood of AMI patients was drawn during percutaneous coronary intervention. We characterized CCR2 expression of fibrocytes by flow cytometry. MCP-1 and the NET marker citrullinated histone H3 (citH3) were measured by ELISA. Fibrocytes were treated in vitro with MCP-1. Human coronary arterial endothelial cells (hCAECs) were stimulated with isolated NETs, and MCP-1 was measured by ELISA and qPCR. The influence of MCP-1 on NET formation in vitro was assessed using isolated neutrophils. Results We have included 50 consecutive AMI patients into the study. NETs and concentrations of MCP-1 were increased at the CLS. NET stimulation of hCAECs induced MCP-1 on mRNA and protein level. Increasing MCP-1 gradient was associated with fibrocyte accumulation at the site of occlusion. In the presence of higher MCP-1 these fibrocytes expressed proportionally less CCR2 than peripheral fibrocytes. In vitro, MCP-1 dose-dependently decreased fibrocyte CCR2 and reduced ex vivo NET release of healthy donor neutrophils. Conclusions NETs induce endothelial MCP-1 release, presumably promoting a chemotactic gradient for leukocyte and fibrocyte migration. MCP-1 mediated inhibition of NET formation could point to a negative feedback loop. These data will shed light on vascular healing. Funding Acknowledgement Type of funding source: Public grant(s) – National budget only. Main funding source(s): Austrian Science Fund

scholarly journals Can Be miR-126-3p a Biomarker of Premature Aging? An Ex Vivo and In Vitro Study in Fabry Disease

Cells ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 356 ◽  
Author(s):  
Alessia Lo Curto ◽  
Simona Taverna ◽  
Maria Assunta Costa ◽  
Rosa Passantino ◽  
Giuseppa Augello ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Fabry Disease ◽  
Healthy Subjects ◽  
Aging Process ◽  
Replicative Senescence ◽  
Ex Vivo ◽  
Umbilical Vein ◽  
Quantitative Polymerase Chain Reaction ◽  
Premature Aging

Fabry disease (FD) is a lysosomal storage disorder (LSD) characterized by lysosomal accumulation of glycosphingolipids in a wide variety of cytotypes, including endothelial cells (ECs). FD patients experience a significantly reduced life expectancy compared to the general population; therefore, the association with a premature aging process would be plausible. To assess this hypothesis, miR-126-3p, a senescence-associated microRNA (SA-miRNAs), was considered as an aging biomarker. The levels of miR-126-3p contained in small extracellular vesicles (sEVs), with about 130 nm of diameter, were measured in FD patients and healthy subjects divided into age classes, in vitro, in human umbilical vein endothelial cells (HUVECs) “young” and undergoing replicative senescence, through a quantitative polymerase chain reaction (qPCR) approach. We confirmed that, in vivo, circulating miR-126 levels physiologically increase with age. In vitro, miR-126 augments in HUVECs underwent replicative senescence. We observed that FD patients are characterized by higher miR-126-3p levels in sEVs, compared to age-matched healthy subjects. We also explored, in vitro, the effect on ECs of glycosphingolipids that are typically accumulated in FD patients. We observed that FD storage substances induced in HUVECs premature senescence and increased of miR-126-3p levels. This study reinforces the hypothesis that FD may aggravate the normal aging process.

scholarly journals A tumor-promoting role for soluble TβRIII in glioblastoma

2021 ◽  
Author(s):  
Isabel Burghardt ◽  
Judith Johanna Schroeder ◽  
Tobias Weiss ◽  
Dorothee Gramatzki ◽  
Michael Weller
Keyword(s):  
Endothelial Cells ◽  
Ex Vivo ◽  
Microvascular Endothelial Cells ◽  
Smad2 Phosphorylation ◽  
Cell Gene Expression ◽  
Microvascular Endothelial ◽  
Cell Gene ◽  
Mouse Glioma

Abstract Purpose Members of the transforming growth factor (TGF)-β superfamily play a key role in the regulation of the malignant phenotype of glioblastoma by promoting invasiveness, angiogenesis, immunosuppression, and maintaining stem cell-like properties. Betaglycan, a TGF-β coreceptor also known as TGF-β receptor III (TβRIII), interacts with members of the TGF-β superfamily and acts as membrane-associated or shed molecule. Shed, soluble TβRIII (sTβRIII) is produced upon ectodomain cleavage of the membrane-bound form. Elucidating the role of TβRIII may improve our understanding of TGF-β pathway activity in glioblastoma Methods Protein levels of TβRIII were determined by immunohistochemical analyses and ex vivo single-cell gene expression profiling of glioblastoma tissue respectively. In vitro, TβRIII levels were assessed investigating long-term glioma cell lines (LTCs), cultured human brain-derived microvascular endothelial cells (hCMECs), glioblastoma-derived microvascular endothelial cells, and glioma-initiating cell lines (GICs). The impact of TβRIII on TGF-β signaling was investigated, and results were validated in a xenograft mouse glioma model Results Immunohistochemistry and ex vivo single-cell gene expression profiling of glioblastoma tissue showed that TβRIII was expressed in the tumor tissue, predominantly in the vascular compartment. We confirmed this pattern of TβRIII expression in vitro. Specifically, we detected sTβRIII in glioblastoma-derived microvascular endothelial cells. STβRIII facilitated TGF-β-induced Smad2 phosphorylation in vitro and overexpression of sTβRIII in a xenograft mouse glioma model led to increased levels of Smad2 phosphorylation, increased tumor volume, and decreased survival Conclusions These data shed light on the potential tumor-promoting role of extracellular shed TβRIII which may be released by glioblastoma endothelium with high sTβRIII levels.

scholarly journals C5a and C5aR1 are key drivers of microvascular platelet aggregation in clinical entities spanning from aHUS to COVID-19

2021 ◽  
Author(s):  
Sistiana Aiello ◽  
Sara Gastoldi ◽  
Miriam Galbusera ◽  
Piero Luigi Ruggenenti ◽  
Valentina Portalupi ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Bacterial Infections ◽  
Viral Infections ◽  
Ex Vivo ◽  
Inflammatory Diseases ◽  
Thrombus Formation ◽  
Alternative Pathway ◽  
Atypical Hemolytic Uremic Syndrome ◽  
Uremic Syndrome

Unrestrained activation of the complement system till the terminal products, C5a and C5b-9, plays a pathogenetic role in acute and chronic inflammatory diseases. In endothelial cells, complement hyperactivation may translate into cell dysfunction, favoring thrombus formation. The aim of this study was to investigate the role of the C5a/C5aR1 axis as opposite to C5b-9 in inducing endothelial dysfunction and loss of anti-thrombogenic properties. In vitro and ex vivo assays with serum from patients with atypical hemolytic uremic syndrome (aHUS) -a prototype rare disease of complement-mediated microvascular thrombosis due to genetically determined alternative pathway dysregulation- and cultured microvascular endothelial cells, demonstrated that the C5a/C5aR1 axis is a key player of endothelial thromboresistance loss. C5a added to normal human serum, fully recapitulated the pro-thrombotic effects of aHUS serum. Mechanistic studies showed that C5a caused RalA-mediated exocytosis of vWF and P-selectin from Weibel-Palade bodies, which favored further vWF binding on the endothelium and platelet adhesion and aggregation. In patients with severe COVID-19 -who suffered from acute activation of complement triggered by SARS-CoV-2 infection- we found the same C5a-dependent pathogenic mechanisms. These results highlight C5a/C5aR1 as a common pro-thrombogenic effector spanning from genetic rare diseases to viral infections, and may have clinical implications. Selective C5a/C5aR1 blockade could have advantages over C5 inhibition, since the former preserves the formation of C5b-9 that is critical to control bacterial infections that often develop as comorbidities in severely ill patients. (Clinicaltrials.gov identifier NCT02464891)

scholarly journals Metformin-stimulated AMPK-α1 promotes microvascular repair in acute lung injury

2013 ◽  
Vol 305 (11) ◽  
pp. L844-L855 ◽  
Author(s):  
Ming-Yuan Jian ◽  
Mikhail F. Alexeyev ◽  
Paul E. Wolkowicz ◽  
Jaroslaw W. Zmijewski ◽  
Judy R. Creighton
Keyword(s):  
Endothelial Cells ◽  
Acute Lung Injury ◽  
Lung Injury ◽  
Ex Vivo ◽  
Endothelial Permeability ◽  
Beneficial Effects ◽  
Isolated Lung

Acute lung injury secondary to sepsis is a leading cause of mortality in sepsis-related death. Present therapies are not effective in reversing endothelial cell dysfunction, which plays a key role in increased vascular permeability and compromised lung function. AMP-activated protein kinase (AMPK) is a molecular sensor important for detection and mediation of cellular adaptations to vascular disruptive stimuli. In this study, we sought to determine the role of AMPK in resolving increased endothelial permeability in the sepsis-injured lung. AMPK function was determined in vivo using a rat model of endotoxin-induced lung injury, ex vivo using the isolated lung, and in vitro using cultured rat pulmonary microvascular endothelial cells (PMVECs). AMPK stimulation using N1-(α-d-ribofuranosyl)-5-aminoimidizole-4-carboxamide or metformin decreased the LPS-induced increase in permeability, as determined by filtration coefficient ( Kf) measurements, and resolved edema as indicated by decreased wet-to-dry ratios. The role of AMPK in the endothelial response to LPS was determined by shRNA designed to decrease expression of the AMPK-α1 isoform in capillary endothelial cells. Permeability, wounding, and barrier resistance assays using PMVECs identified AMPK-α1 as the molecule responsible for the beneficial effects of AMPK in the lung. Our findings provide novel evidence for AMPK-α1 as a vascular repair mechanism important in the pulmonary response to sepsis and identify a role for metformin treatment in the management of capillary injury.

scholarly journals Kisspeptin-10 Inhibits Angiogenesis in Human Placental Vessels ex Vivo and Endothelial Cells in Vitro

Endocrinology ◽  
2010 ◽  
Vol 151 (12) ◽  
pp. 5927-5934 ◽  
Author(s):  
Thayalini Ramaesh ◽  
James J. Logie ◽  
Antonia K. Roseweir ◽  
Robert P. Millar ◽  
Brian R. Walker ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Blood Vessels ◽  
Ex Vivo ◽  
Umbilical Vein ◽  
Biologically Active ◽  
Trophoblast Invasion ◽  
In Vitro Assays ◽  
Dependent Manner ◽  
Inhibition Of Proliferation

Recent studies suggest that kisspeptin (a neuropeptide central to the regulation of gonadotrophin secretion) has diverse roles in human physiology, including a putative role in implantation and placental function. Kisspeptin and its receptor are present in human blood vessels, where they mediate vasoconstriction, and kisspeptin is known to inhibit tumor metastasis and trophoblast invasion, both processes involving angiogenesis. We hypothesized that kisspeptin contributes to the regulation of angiogenesis in the reproductive system. The presence of the kisspeptin receptor was confirmed in human placental blood vessels and human umbilical vein endothelial cells (HUVEC) using immunochemistry. The ability of kisspeptin-10 (KP-10) (a shorter biologically active processed peptide) to inhibit angiogenesis was tested in explanted human placental arteries and HUVEC using complementary ex vivo and in vitro assays. KP-10 inhibited new vessel sprouting from placental arteries embedded in Matrigel and tube-like structure formation by HUVEC, in a concentration-dependent manner. KP-10 had no effect on HUVEC viability or apoptosis but induced concentration-dependent inhibition of proliferation and migration. In conclusion, KP-10 has antiangiogenic effects and, given its high expression in the placenta, may contribute to the regulation of angiogenesis in this tissue.

Abstract 23: Microrna302-367 Sphingosine 1 Phosphate Receptor 1 Pathway Prevents Tumor Growth via Restricting Angiogenesis and Enhancing Vascular Stability

2016 ◽  
Vol 36 (suppl_1) ◽  
Author(s):  
Jinjiang Pi ◽  
Ting Tao ◽  
Tao Zhuang ◽  
Huimin Sun ◽  
Xiaoli Chen ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Tumor Growth ◽  
Ex Vivo ◽  
Downstream Target ◽  
Sprouting Angiogenesis ◽  
Pathological Angiogenesis ◽  
Vascular Stability ◽  
Sphingosine 1 Phosphate

Angiogenic hypersprouting and leaky immature vessels of pathological angiogenesis are essential for tumor growth. MicroRNAs have unique therapeutic advantages by targeting multiple pathways of tumor-associated angiogenesis, but the function of individual miRNAs in angiogenesis and tumors has not yet been fully evaluated. Here, we show that miR302-367 elevation in endothelial cells reduces retina sprouting angiogenesis and promotes vascular stability in vivo, ex vivo and in vitro. Erk1/2 are identified as direct targets of miR302-367, and down-regulation of Erk1/2 upon miR302-367 elevation in endothelial cells increases the expression of Klf2 and in turn S1pr1 and its downstream target VE-cadherin, suppressing angiogenesis and improving vascular stability. Conversely, both pharmacological blockade and genetic deletion of S1pr1 in endothelial cells reverse the anti-angiogenic and vascular stabilizing effect of miR302-367 in mice. Pathological angiogenesis in tumors shares features of developmental angiogenesis, and endothelial specific elevation of miR302-367 reduces tumor growth by restricting sprout angiogenesis and decreasing vascular permeability via the same Erk1/2-Klf2-S1pr1 pathways. In conclusion, miR302-367 regulation of an Erk1/2-Klf2-S1pr1 pathway in the endothelium advances our understanding of angiogenesis, meanwhile also provides opportunities for therapeutic intervention of tumor growth.

scholarly journals N-Acetylcysteine and Sulodexide Reduce the Prothrombotic Effect of Uremic Serum on the Venous Endothelial Cells

2019 ◽  
Vol 44 (2) ◽  
pp. 277-285
Author(s):  
Patrycja Sosinska-Zawierucha ◽  
Beata Mackowiak ◽  
Andrzej Breborowicz
Keyword(s):  
Gene Expression ◽  
Renal Failure ◽  
Endothelial Cells ◽  
Ex Vivo ◽  
Secretory Activity ◽  
Serum Samples ◽  
Uremic Serum ◽  
Uremic Patients ◽  
End Stage

Background/Aims: Thromboembolic episodes are a frequent problem in end stage renal failure patients. The pathomechanism of the disorder is complex, including bioincompatibility of renal replacement therapy, endothelial dysfunction, increased blood level of procoagulant factors and uremic toxins. We studied changes in the functional properties of venous endothelial cells (VEC) in the presence of uremic serum and evaluated their possible modulation by N-acetylcysteine (NAC) or sulodexide (SUL). Methods: Serum samples from 12 uremic patients treated with hemodialysis were studied ex vivo on in vitro cultured VEC. In separate experiments, NAC 1 mmol/L or SUL 0.5 LRU/mL were added to uremic serum samples. Both changes in the gene expression and secretory activity of VEC were studied. Results: Uremic serum increased the expression of the following genes: IL6 +97%, p < 0.002; VEGF +28%, p < 0.002; vWF +47%, p < 0.002; PECAM +76%, p < 0.002; ICAM-1 +275%, p < 0.002; t-PA +96%, p < 0.002. Changes in gene expression were reflected by the increased secretory activity of VEC treated with the uremic serum. Exposure of VEC to uremic serum supplemented with NAC or SUL resulted in weaker stimulation of the studied genes’ expression. Also, secretion of the studied solutes, with the exception of ICAM-1, was reduced in the presence of NAC: IL6 –34%, p < 0.01; VEGF –40%, p < 0.005; vWF –25%, p < 0.001; t-PA –47%, p < 0.01, and MMP9 –37%, p < 0.001. SUL reduced the uremic serum-induced secretion of all solutes: IL6 –24%, p < 0.05; ICAM-1 –43%, p < 0.01; VEGF –38%, p < 0.01; vWF –23%, p < 0.01; t-PA –49%, p < 0.01, and MMP9 –25%, p < 0.05. Conclusions: Uremic serum induces prothrombotic changes in VEC, which may cause a predisposition to thrombotic disorders in patients with renal failure. NAC and SUL reduce the effects of the uremic serum in VEC, which suggests their potential therapeutic application in uremic patients.

scholarly journals Exosomes derived from induced vascular progenitor cells promote angiogenesis in vitro and in an in vivo rat hindlimb ischemia model

2019 ◽  
Vol 317 (4) ◽  
pp. H765-H776 ◽  
Author(s):  
Takerra K. Johnson ◽  
Lina Zhao ◽  
Dihan Zhu ◽  
Yang Wang ◽  
Yan Xiao ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Progenitor Cells ◽  
Ex Vivo ◽  
Aortic Ring ◽  
Size Exclusion ◽  
Hindlimb Ischemia ◽  
Vascular Progenitor Cells ◽  
Angiogenic Effect

Induced vascular progenitor cells (iVPCs) were created as an ideal cell type for regenerative medicine and have been reported to positively promote collateral blood flow and improve cardiac function in a rat model of myocardial ischemia. Exosomes have emerged as a novel biomedicine that mimics the function of the donor cells. We investigated the angiogenic activity of exosomes from iPVCs (iVPC-Exo) as a cell-free therapeutic approach for ischemia. Exosomes from iVPCs and rat aortic endothelial cells (RAECs) were isolated using a combination of ultrafiltration and size-exclusion chromatography. Nanoparticle tracking analysis revealed that exosome isolates fell within the exosomal diameter (<150 nm). These exosomes contained known markers Alix and TSG101, and their morphology was validated using transmission electron microscopy. When compared with RAECs, iVPCs significantly increased the secretion of exosomes. Cardiac microvascular endothelial cells and aortic ring explants were pretreated with RAEC-Exo or iVPC-Exo, and basal medium was used as a control. iVPC-Exo exerted an in vitro angiogenic effect on the proliferation, tube formation, and migration of endothelial cells and stimulated microvessel sprouting in an ex vivo aortic ring assay. Additionally, iVPC-Exo increased blood perfusion in a hindlimb ischemia model. Proangiogenic proteins (pentraxin-3 and insulin-like growth factor-binding protein-3) and microRNAs (-143-3p, -291b, and -20b-5p) were found to be enriched in iVPC-Exo, which may mediate iVPC-Exo induced vascular growth. Our findings demonstrate that treatment with iVPC-Exo promotes angiogenesis in vitro, ex vivo, and in vivo. Collectively, these findings indicate a novel cell-free approach for therapeutic angiogenesis. NEW & NOTEWORTHY The results of this work demonstrate exosomes as a novel physiological mechanism by which induced vascular progenitor cells exert their angiogenic effect. Moreover, angiogenic cargo of proteins and microRNAs may define the biological contributors in activating endothelial cells to form a new capillary plexus for ischemic vascular diseases. Listen to this article's corresponding podcast at https://ajpheart.podbean.com/e/angiogenic-exosomes-from-vascular-progenitor-cells/ .

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