Abstract 315: Calcium Dose Dependently Influences Endothelial Cell Angiogenesis

2017 ◽  
Vol 37 (suppl_1) ◽  
Author(s):  
Jocelyne Mulangala ◽  
Emma J Akers ◽  
Peter J Psaltis ◽  
Stephen J Nicholls ◽  
Belinda A Di Bartolo
Keyword(s):  
Endothelial Cell ◽  
Arterial Wall ◽  
Hypoxia Inducible Factor ◽  
Vascular Complication ◽  
Human Coronary Artery ◽  
Tubule Formation ◽  
Alkaline Phosphotase ◽  
High Concentrations ◽  
Artery Disease

Background: Peripheral artery disease (PAD) is a progressive occlusive disease of the arteries and a vascular complication in diabetes. Vascular calcification (VC) is implicated as a potential driver of PAD, and although the exact mechanisms are unclear, the site and location of calcification within the arterial wall contributes greatly. Long considered a passive process, VC is now recognised as a tightly regulated active process balancing the promotion and inhibition of calcification in the arterial wall. There is little evidence however, to demonstrate the effect of calcification on endothelial cell angiogenesis. This study sought to investigate the effects of calcium as a known inducer of calcification on in vitro angiogenesis. Methods: Human Coronary Artery Endothelial Cells were cultured and treated with increasing calcium concentrations (CaCl 2 2.45-3.3 mM) for 24h. Proliferation, migration and tubule formation assays were conducted and real-time PCR assessed angiogenic and osteogenic genes. Alkaline phosphotase (ALP) activity was measured in supernatants following treatment. Results: High concentrations of calcium reduced cell proliferation with a corresponding increase in ALP production suggesting release of osteogenic stimuli adversely affects cell viability. Mid-range concentrations of calcium induced a significant increase in cell migration (1.0 vs 2.4±0.3, p<0.05) while higher concentrations elicited no effect. Calcium treatment demonstrated a dose response where mid-range concentrations increased gene expression of hypoxia-inducible factor-1α (>500 fold), and fibroblast growth factor-2 (>150 fold). This increase corresponded with a decrease (1.0 vs 15.02±4.24; p<0.0001) in osteoprotegerin (OPG) at mid-range calcium with a significant increase at the highest concentration (1.0 vs 342±13.27; p<0.01) illustrating calcium-induced expression of OPG, a known protective gene in VC, may also regulate angiogenesis. Conclusion: This is the first demonstration investigating the effects of calcium on endothelial cell angiogenesis. These findings suggest that calcium can directly affect genes involved in regulating angiogenesis, and could therefore provide an opportunity to develop potential treatments.

Inhibition of Platelet Function by Antiarrhythmic Drugs, Verapamil and Disopyramide

1982 ◽  
Vol 47 (02) ◽  
pp. 150-153 ◽  
Author(s):  
P Han ◽  
C Boatwright ◽  
N G Ardlie
Keyword(s):  
Platelet Aggregation ◽  
Platelet Function ◽  
Antiarrhythmic Drugs ◽  
Adenosine Diphosphate ◽  
Inhibitory Effect ◽  
Second Phase ◽  
Ionophore A23187 ◽  
High Concentrations ◽  
Artery Disease

SummaryVarious cardiovascular drugs such as nitrates and propranolol, used in the treatment of coronary artery disease have been shown to have an antiplatelet effect. We have studied the in vitro effects of two antiarrhythmic drugs, verapamil and disopyramide, and have shown their inhibitory effect on platelet function. Verapamil, a calcium channel blocker, inhibited the second phase of platelet aggregation induced by adenosine diphosphate (ADP) and inhibited aggregation induced by collagen. Disopyramide similarly inhibited the second phase of platelet aggregation caused by ADP and aggregation induced by collagen. Either drug in synergism with propranolol inhibited ADP or collagen-induced platelet aggregation. Disopyramide at high concentrations inhibited arachidonic add whereas verapamil was without effect. Verapamil, but not disopyramide, inhibited aggregation induced by the ionophore A23187.

scholarly journals IL-17B Can Impact on Endothelial Cellular Traits Linked to Tumour Angiogenesis

2010 ◽  
Vol 2010 ◽  
pp. 1-5 ◽  
Author(s):  
Andrew J. Sanders ◽  
Xiaoxia Guo ◽  
Malcolm D. Mason ◽  
Wen G. Jiang
Keyword(s):  
Rheumatoid Arthritis ◽  
Endothelial Cell ◽  
Inflammatory Response ◽  
Cellular Migration ◽  
Tumour Angiogenesis ◽  
Tubule Formation ◽  
Founding Member ◽  
Cell Matrix ◽  
Cell Matrix Adhesion

IL-17B is a member of the IL-17 cytokine family which have been implicated in inflammatory response and autoimmune diseases such as rheumatoid arthritis. The founding member of this family, IL-17 (or IL-17A), has also been implicated in promoting tumour angiogenesis through the induction of other proangiogenic factors. Here we examine the potential of recombinant human IL-17B to contribute to the angiogenic process. In vitro rhIL-17B was able to inhibit HECV endothelial cell-matrix adhesion and cellular migration and also, at higher concentrations, could substantially reduce tubule formation compared to untreated HECV cells in a Matrigel tubule formation assay. This data suggests that IL-17B may act in an antiangiogenic manner.

In Vitro Analyses of Umbilical Cord Blood (UCB) Derived Monocytes Support Adjunct Topical Application to Augment Platelet Rich Plasma (PRP) in Wound Healing.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1263-1263 ◽  
Author(s):  
Graham C. Chapman ◽  
Nicholas Greco ◽  
Richard Patrick Weitzel ◽  
Phil Paul ◽  
Peter Haviernik ◽  
...  
Keyword(s):  
Wound Healing ◽  
Endothelial Cell ◽  
Topical Application ◽  
Time Course ◽  
Basal Medium ◽  
Fold Increase ◽  
Conditioned Media ◽  
Tubule Formation ◽  
Cell Networks

Abstract Currently, PRP is used clinically as a topical application to augment healing of both surgical and chronic, non-healing wounds. PRP exerts its efficacy by releasing growth factors that enhance clot formation and vasculogenesis. We conducted in vitro functional analyses comparing PRP and/or UCB-derived monocytes including cytokine production, cell migration, and HUVEC tubule formation in standard matrigel assays to test the hypothesis whether topical concurrent application of PRP and UCB-derived monocytes may serve to augment wound healing beyond the ability of topical PRP alone. UCB was obtained according to institutional guidelines and collected into bags with citrate dextrose (Allegiance). MNC were separated on a Histopaque-1077 (Sigma) density gradient. UCB CD14+ monocytes were isolated using AutoMACS magnetic cell sorter (Miltenyi), and cultured in RPMI with 1% HSA. PRP was isolated from adult peripheral blood by centrifugation. To determine if the addition of UCB monocytes may improve the wound healing effects of PRP alone, VEGF, bFGF, and PDGF secreted by monocytes alone, PRP alone, and monocytes supplemented with 3% PRP, were measured by ELISA (RayBiotech) daily over 4 days. PRP alone elicited no measurable secretion of VEGF. UCB-derived monocytes alone showed a low, constant production of VEGF over the four days of 0.868ng/ml. PRP supplemented with UCB-derived monocytes secreted VEGF at a 7.6-fold increase over either PRP or UCB monocytes alone, with a peak production at day three of 6.638ng/ml. PRP alone produced no measurable secretion of bFGF over the four day time course. UCB monocytes alone secreted bFGF in an increasing manner during the same time course. During days one to four, bFGF secreted by UCB monocytes was 33.8, 27.9, 115.4, and 452.1pg/ml, respectively. The presence of PRP suppressed this secretion, as PRP combined with UCB monocytes constantly secreted bFGF at an average of 39.9pg/ml throughout days one to four. Finally, secretion of PDGF was highest in conditions including PRP combined with UCB monocytes. PRP alone constantly produced PDGF at an average of 3,144pg/ml over a 4 day time course. Monocytes alone secreted PDGF constantly at a lower average of 597pg/ml. PRP supplemented with UCB monocytes secreted PDGF at a concentration 5.9-fold higher than PRP alone, producing an average of 18,534pg/ml over four days. To determine whether UCB-derived monocytes respond to cytokines elicited by injured vascular endothelial cells, we measured UCB-derived monocyte chemotaxis to HUVEC conditioned media in hypoxic conditions (5% O2). Migration experiments were conducted using Transwell plates with 8.0 μm pores. Monocytes were cultured in RPMI with 5% FBS at a concentration of 5×106/ml and were allowed to migrate for four hours to either: media alone, PRP, HUVEC-conditioned media, or HUVEC-conditioned media supplemented with PRP. We observed a 3.3 fold increase in the migration of the monocytes to HUVECconditioned media over that of basal media. Experiments with PRP alone showed no significant difference in monocyte migration compared to basal medium. To determine whether UCB-derived monocytes may serve to augment endothelial cell function beyond that elicited by PRP alone, matrigel experiments were conducted by adding HUVEC in endothelial cell basal medium. HUVEC tubule formation in matrigel in basal media was compared in three conditions including media conditioned with: 1) PRP alone, 2) UCB monocytes alone, or 3) a combination of PRP + UCB monocytes. We compared the kinetics and stability of enclosed endothelial cell networks formed by HUVEC. No significant benefit was seen with addition of PRP conditioned media. The number of enclosed endothelial cell networks reached a higher maximum with the addition of monocyte conditioned media (137 networks) as well as PRP + monocyte conditioned media (142 networks), compared to non-conditioned media (80 networks). UCB monocyte and PRP + UCB monocyte conditioned media also improved the stability of the enclosed cell networks in culture as structures persisted beyond 24h, while none were present in the PRP-conditioned or non-conditioned media matrigel cultures. Figure Figure In summary, these in vitro analyses support the hypothesis that UCB-derived monocytes significantly improve efficacy of PRP alone in augmentation of vasculogenesis and cell migration to vascular endothelial injury, thereby supporting potential concurrent topical application of UCB-derived monocytes to PRP in wound healing.

scholarly journals Crocus Sativus and Its Active Compound, Crocin Inhibits the Endothelial Activation and Monocyte-Endothelial Cells Interaction in Stimulated Human Coronary Artery Endothelial Cells

2020 ◽  
Author(s):  
Noor Alicezah Mohd Kasim ◽  
Nurul Ain Abu Bakar ◽  
Radzi Ahmad ◽  
Iman Nabilah Abd Rahim ◽  
Thuhairah Hasrah Abdul Rahman ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Coronary Artery ◽  
Endothelial Activation ◽  
Crocus Sativus ◽  
Cellular Interactions ◽  
Human Coronary Artery ◽  
Gene Expressions ◽  
Protein Expressions

AbstractCrocus sativus L. or saffron has been shown to have anti-atherogenic effects. However, its effects on key events in atherogenesis such as endothelial activation and monocyte-endothelial cell binding in lipolysaccharides (LPS)-stimulated in vitro model have not been extensively studied.ObjectivesTo investigate the effects of saffron and its bioactive derivative crocin on the gene and protein expressions of biomarkers of endothelial activation in LPS stimulated human coronary artery endothelial cells (HCAECs).MethodologyHCAECs were incubated with different concentrations of aqueous ethanolic extracts of saffron and crocin together with LPS. Protein and gene expressions of endothelial activation biomarkers were measured using ELISA and qRT-PCR, respectively. Adhesion of monocytes to HCAECs was detected by Rose Bengal staining. Methyl-thiazol-tetrazolium assay was carried out to assess cytotoxicity effects of saffron and crocin.ResultsSaffron and crocin up to 25.0 and 1.6 μg/ml respectively exhibited >85% cell viability. Saffron treatment reduced sICAM-1, sVCAM-1 and E-selectin proteins (concentrations: 3.13, 6.25, 12.5 and 25.0 μg/ml; 3.13, 12.5 and 25.0 μg/ml; 12.5 and 25.0, respectively) and gene expressions (concentration: 12.5 and 25.0μg/ml; 3.13, 6.25 and 25.0 μg/ml; 6.25, 12.5 25.0; respectively). Similarly, treatment with crocin reduced protein expressions of sICAM-1, sVCAM-1 and E-selectin (concentration: 0.2, 0.4, 0.8 and 1.6 μg/ml; 0.4, 0.8 and 1.6 μg/ml; 0.8 and 1.6 μg/ml; respectively] and gene expression (concentration: 0.8 and 1.6 μg/ml; 0.4, 0.8 and 1.6 μg/ml; and 1.6 μg/ml, respectively). Monocyte-endothelial cell interactions were reduced following saffron treatment at concentrations 6.3, 12.5 and 25.00 μg/ml. Similarly, crocin also suppressed cellular interactions at concentrations 0.04, 0.08, 1.60 μg/ml.ConclusionSaffron and crocin exhibits potent inhibitory action for endothelial activation and monocyte-endothelial cells interaction suggesting its potential anti-atherogenic properties.

scholarly journals Association of carbamylated high-density lipoprotein with coronary artery disease in type 2 diabetes mellitus: carbamylated high-density lipoprotein of patients promotes monocyte adhesion

2020 ◽  
Vol 18 (1) ◽  
Author(s):  
Zhongli Chen ◽  
Song Ding ◽  
Yan Ping Wang ◽  
Liang Chen ◽  
Jing Yan Mao ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Endothelial Cell ◽  
High Density Lipoprotein ◽  
Density Lipoprotein ◽  
High Density ◽  
Control Group ◽  
Diabetic Patients ◽  
Artery Disease ◽  
Endothelial Cell Adhesion

Abstract Background Increasing evidence showed that carbamylated lipoprotein accelerated atherosclerosis. However, whether such modification of high-density lipoprotein (HDL) particles alters in type 2 diabetes mellitus (T2DM) patients and facilitates vascular complications remains unclear. We aimed to investigate the alteration of the carbamylation in HDL among T2DM patients and clarify its potential role in atherogenesis. Methods A total of 148 consecutive T2DM patients undergoning angiography and 40 age- and gender-matched control subjects were included. HDL was isolated from plasma samples, and the concentration of HDL carbamyl-lysine (HDL-CBL) was measured. Furthermore, the HDL from subjects and in-vitro carbamylated HDL (C-HDL) was incubated with endothelial cells and monocyte to endothelial cell adhesion. Adhesion molecule expression and signaling pathway were detected. Results Compared with the control group, the HDL-CBL level was remarkably increased in T2DM patients (6.13 ± 1.94 vs 12.00 ± 4.06 (ng/mg), P < 0.001). Of note, HDL-CBL demonstrated a more significant increase in T2DM patients with coronary artery disease (CAD) (n = 102) than those without CAD (n = 46) (12.75 ± 3.82 vs. 10.35 ± 4.11(ng/mg), P = 0.001). Multivariate logistic regression analysis demonstrated that higher HDL-CBL level was independently associated with a higher prevalence of CAD in diabetic patients after adjusting for established cofounders (adjusted odds ratio 1.174, 95% confidence Interval 1.045–1.319, p = 0.017). HDL from diabetic patients with CAD enhanced greater monocyte adhesion than that from the non-CAD or the control group (P < 0.001). Such pro-atherogenic capacity of diabetic HDL positively correlated with HDL-CBL level. Furthermore, in-vitro incubation of carbamylated HDL (C-HDL) with endothelial promoted monocyte to endothelial cell adhesion, induced upregulation of cell adhesion molecules expression, and activated NF-κB/p65 signaling in endothelial cells. Inhibiting carbamylation of HDL or NF-κB activation attenuated the monocyte to endothelial cell adhesion and cell surface adhesion molecules expression. Conclusions Our study identified elevated carbamylation modification of HDL from T2DM patients, especially in those with concomitant CAD. We also evidenced that C-HDL enhanced monocyte to endothelial cell adhesion, indicating a potential pro-atherogenic role of C-HDL in atherosclerosis among T2DM patients. Trial registrationhttps://register.clinicaltrials.gov, NCT04390711 Registered on 14 May 2020; Retrospectively registered

Influence of Umbilical Cord Blood-Derived Endothelial Precursor Cells on a Human Umbilical Vein Endothelial Cell Co-Culture In Vitro Angiogenesis Assay.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4218-4218
Author(s):  
Nicholas J. Greco ◽  
Brandon Eilertson ◽  
Jason J. Banks ◽  
Paul Scheid ◽  
Marcie Finney ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Growth Factor ◽  
Cord Blood ◽  
Umbilical Cord ◽  
Umbilical Cord Blood ◽  
Umbilical Vein ◽  
Tubule Formation ◽  
Human Umbilical Vein ◽  
In Vitro Angiogenesis

Abstract To assess in vitro angiogenesis, cellular co-culture assays have been utilized to study adherence, spreading, differentiation and proliferation, and migration of endothelial cells. Formation of tubule or capillary-like networks is influenced by the presence of vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF) but other factors provided by cell sources and/or direct contact with multiple cell types may facilitate this formation. The hypothesis of this study is that umbilical cord blood (UCB)-derived endothelial precursor cells (EPCs) may influence the formation of human umbilical vein endothelial cell (HUVEC) tubule structures during angiogenesis. Methods: UCB-derived EPCs were isolated from CD133negative cells after a 7-day culture on human fibronectin in EGM-2 media. Tubule formation was evaluated (passage 1–2, 20 x 103 or 2 x 103 cells) by adding HUVECs without or with EPCs to cultures of human bone marrow-derived mesenchymal stromal cells (MSCs) under normoxic (20%) conditions (37°C, 5% CO2, containing VEGF, epidermal growth factor, FGF, insulin-like growth factor, heparin, hydrocortisone, and ascorbic acid in EGM-2 medium) for a 2-week period. HUVECs were added to cultures without or with labeling with Vybrant® CM-DiI which allows the temporal observation of tubule formation progress and cellular incorporation. Final tubule formation was confirmed using a primary anti-CD31 (PECAM) antibody followed by a FITC-conjugated secondary antibody for signal amplification. Results: After 2–4 days, linear aggregates of labeled HUVECs (2-D arrangement) were observed. After 14 days, there was remodeling of HUVECs into the development of a 3D network of linear and branched tubule structures. EPCs facilitated the formation of tubules affecting both the extent of tubule formation and also enhanced proliferation of HUVEC cells. A minority (&lt; 5%) of EPCs were incorporated into developing tubules (estimated using CM-Dil-labeled EPCs). To quantify tubule formation, digital pictures of representative areas of culture wells (2–4/well) were acquired. Using Image Pro Plus software, tubules were quantified using multi-parameter analysis with respect to length, area, and perimeter. The presence of EPCs (equal to the number of added HUVECs) significantly enhanced all parameters. In comparison to control samples, the presence of EPCs increased the area, perimeter and size by 15.2-fold, 3.4-fold, and 3.2-fold, respectively. Confocal microscopy revealed that the co-cultures formed anatamoses, indicating the formation of a connected network. Conclusions: Taken together, these results suggest that the presence of cord blood-derived EPCs facilitate tubule formation and development via a heterotypic cell-cell interaction without integrating into the angiogenic structures. Further studies will evaluate the secretion of cytokines and growth factors.

scholarly journals Modified LDLs induce proliferation-mediated death of human vascular endothelial cells through MAPK pathway

2007 ◽  
Vol 292 (4) ◽  
pp. H1836-H1846 ◽  
Author(s):  
Eugene O. Apostolov ◽  
Alexei G. Basnakian ◽  
Xiaoyan Yin ◽  
Ercan Ok ◽  
Sudhir V. Shah
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cells ◽  
Cell Death ◽  
Endothelial Cell ◽  
Mapk Pathway ◽  
Oxidized Ldl ◽  
Mitogen Activated Protein Kinase ◽  
Cell Multiplication ◽  
Human Coronary Artery

The ability of modified low-density lipoptoteins (LDLs) to induce both proliferation and death of endothelial cells is considered to be a mechanism of early atherosclerosis development. We previously showed that carbamylated LDL (cLDL) induces human coronary artery endothelial cell (HCAEC) death in vitro. This effect is similar to the atherogenic action of oxidized LDL (oxLDL) that induces the proliferation and death of endothelial cells. The present study was designed to analyze a potential proliferative effect of cLDL and whether proliferation caused by modified LDLs is related to cell death. Cultured HCAECs were exposed to different concentrations of modified LDL or native LDL for varying periods of time. Cell proliferation measured by bromodeoxyuridine incorporation and S-phase analysis was dose-dependently increased in the presence of cLDL (6.25–200 μg/ml). The proliferation induced by cLDL or oxLDL was associated with cell death and increased phosphorylation of extracellular signal-regulated kinase (ERK) and c-Jun NH2-terminal kinase (JNK). Inhibition of cLDL- or oxLDL-induced proliferation by aphidicolin (1 μg/ml) was protective against both short-term cell death measured by lactate dehydrogenase release into the medium and long-term cell viability visualized by cell multiplication. Inhibition of ERK phosphorylation led to a significant decrease of DNA synthesis and cell rescue from injury by modified LDLs, while inhibition of JNK phosphorylation had an only partial rescue effect without involvement in cell proliferation. These data are the first evidence that endothelial cell death induced by cLDL or oxLDL is mediated by cell proliferation through the mitogen-activated protein kinase pathway.

Endothelial Cell Attachment and Proliferation Studies on Modified Metal Stent Surfaces

2009 ◽  
Vol 1236 ◽  
Author(s):  
Vipul Davè ◽  
Charito Buensuceso ◽  
David Colter ◽  
Jonathon Zhao ◽  
Robert Falotico
Keyword(s):  
Endothelial Cell ◽  
Cell Attachment ◽  
Screening Method ◽  
Cell Number ◽  
Chromium Alloy ◽  
Cobalt Chromium ◽  
Metal Stent ◽  
Human Coronary Artery ◽  
Endothelial Cell Attachment

AbstractFlat coupons prepared from cobalt chromium alloy (CoCr) were modified using different methods (low energy excimer laser processing, electron beam irradiation, and immobilized covalently-bound heparin coating). Human coronary artery endothelial cell (HCAEC) attachment and growth kinetics were investigated on unmodified and modified metal surfaces. Results showed that HCAEC attach to unmodified CoCr coupons and surface-modified CoCr coupons. No change in cell number was observed when cells were grown on CoCr coupons and heparin coated coupons throughout the 72h study period. A decrease in cell number was observed for excimer treated coupons. HCAEC seeding on CoCr stents indicated that cells attached and proliferated on the stents over a ten day study period. This research showed that physical modifications did not improve cell proliferation. Very few non-viable cells were observed for unmodified and surface bound heparin coupons, and cells attached to the surface up to 72h. This shows that heparin can be coated on a stent surface to provide anti-thrombotic properties without any negative effect on cell attachment and proliferation. In vitro screening method of testing endothelial cell attachment and proliferation on modified metal stent surfaces can be used to gain insight for developing next generation drug eluting stents with improved endothelialization behavior.

Mechanisms of endothelial cell activation by endocannabinoid 2-arachidonoylglycerol

2020 ◽  
Vol 41 (Supplement_2) ◽  
Author(s):  
J Jehle ◽  
L Eich ◽  
E Avraamidou ◽  
V Tiyerili ◽  
L Bindila ◽  
...  
Keyword(s):  
Coronary Artery Disease ◽  
Endothelial Cell ◽  
Coronary Artery ◽  
Carotid Artery ◽  
Adhesion Molecules ◽  
Common Carotid Artery ◽  
Endothelial Repair ◽  
The Common ◽  
Artery Disease

Abstract Background Endothelial dysfunction promotes atherogenesis, vascular inflammation, and thrombus formation. Reendothelialization after angioplasty is required in order to restore vascular function and to prevent stent thrombosis. The endocannabinoid (eCB) 2-arachidonoylglycerol (2-AG) is a known modulator of inflammation. Earlier studies have demonstrated the relevance of this endocannabinoid in human pathophysiology during coronary artery disease and in murine experimental atherogenesis. However, evidence on the impact of 2-AG on endothelial cell function remains scarce. Methods Endothelial repair was studied in two treatment groups of wildtype mice following electrical denudation of the common carotid artery. One group received the monoacylglycerol lipase (MAGL)-inhibitor JZL184, which impairs 2-AG degradation and thus causes elevated 2-AG levels, the other group received DMSO. The residual endothelial gap at five days was visualized by Evan's blue staining in either group. In vitro, the effect of 2-AG on human coronary artery endothelial cell (HCAEC) viability was assessed by an XTT-based assay. Endothelial activation was studied by an adhesion assay of THP-1 monocytes to 2-AG-preconditioned HCAEC. Activation of HCAEC adhesion molecules was characterized by flow cytometry. Results Elevated 2-AG levels significantly impaired reendothelialization in wildtype mice following electrical injury of the common carotid artery, resulting in a residual denudation at 5 days of 2291±286 μm vs. 1505±223 μm (n=18–19; p&lt;0.05). In vitro, 2-AG significantly reduced viability of HCAEC at 24 hours (0.31±0.10 vs. 1.00±0.08; n=3; p&lt;0.01). Finally, 2-AG promoted HCAEC activation resulting in a significant increase in THP-1 monocyte adhesion to HCAEC following pre-treatment of HCAEC with 2-AG (0.17±0.03 THP-1 cells per HCAEC vs. 0.07±0.01 THP-1 cells per HCAEC; n=3; p&lt;0.05). Adhesion molecules E-selectin, ICAM-1 and VCAM-1, that are known to be regulated by 2-AG in the venous endothelium, remained unchanged in arterial endothelial cells. Besides, HCAEC migration, ROS-production, expression of NADPH oxidases and secretion of inflammatory cytokines were unaffected by 2-AG. Conclusion Elevated 2-AG levels hamper endothelial repair and impair HCAEC proliferation while facilitating adhesion of monocytes. Intriguingly, the underlying mechanisms in the arterial vascular bed appear distinct from venous endothelium. Given that 2-AG is elevated during coronary artery disease in humans, 2-AG might impair reendothelialization after angioplasty and thus impact on clinical outcomes. Funding Acknowledgement Type of funding source: None

MicroRNA-mediated vascular intercellular communication is altered in chronic kidney disease

2020 ◽  
Author(s):  
Andreas Zietzer ◽  
Eva Steffen ◽  
Sven Niepmann ◽  
Philip Düsing ◽  
Mohammed Rabiul Hosen ◽  
...  
Keyword(s):  
Chronic Kidney Disease ◽  
Endothelial Cell ◽  
Coronary Artery ◽  
Kidney Disease ◽  
Intercellular Communication ◽  
Recipient Cell ◽  
Flow Cytometric Analysis ◽  
Human Coronary Artery ◽  
Artery Disease ◽  
Nuclear Ribonucleoprotein

Abstract Aims Chronic kidney disease (CKD) is an independent risk factor for the development of coronary artery disease (CAD). For both, CKD and CAD, the intercellular transfer of microRNAs (miRs) through extracellular vesicles (EVs) is an important factor of disease development. Whether the combination of CAD and CKD affects endothelial function through cellular crosstalk of EV-incorporated miRs is still unknown. Methods and results Out of 172 screened CAD patients, 31 patients with CAD + CKD were identified and matched with 31 CAD patients without CKD. Additionally, 13 controls without CAD and CKD were included. Large EVs from CAD + CKD patients contained significantly lower levels of the vasculo-protective miR-130a-3p and miR-126-3p compared to CAD patients and controls. Flow cytometric analysis of plasma-derived EVs revealed significantly higher numbers of endothelial cell-derived EVs in CAD and CAD + CKD patients compared to controls. EVs from CAD + CKD patients impaired target human coronary artery endothelial cell (HCAEC) proliferation upon incubationin vitro. Consistent with the clinical data, treatment with the uraemia toxin indoxyl sulfate (IS)-reduced miR-130a-3p levels in HCAEC-derived EVs. EVs from IS-treated donor HCAECs-reduced proliferation and re-endothelialization in EV-recipient cells and induced an anti-angiogenic gene expression profile. In a mouse-experiment, intravenous treatment with EVs from IS-treated endothelial cells significantly impaired endothelial regeneration. On the molecular level, we found that IS leads to an up-regulation of the heterogenous nuclear ribonucleoprotein U (hnRNPU), which retains miR-130a-3p in the cell leading to reduced vesicular miR-130a-3p export and impaired EV-recipient cell proliferation. Conclusion Our findings suggest that EV-miR-mediated vascular intercellular communication is altered in patients with CAD and CKD, promoting CKD-induced endothelial dysfunction.

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