Abstract P429: Aldehyde Dehydrogenase 2 Regulates Angiogenesis Of Coronary Microvascular Endothelial Cells Via A Novel Signaling Pathway

2021 ◽  
Vol 129 (Suppl_1) ◽  
Author(s):  
Bipradas Roy ◽  
Guodong Pan ◽  
Suresh Palaniyandi
Keyword(s):  
Aldehyde Dehydrogenase ◽  
Cardiac Tissue ◽  
Nuclear Translocation ◽  
Growth Factor Receptor ◽  
Tube Formation ◽  
Microvascular Endothelial Cell ◽  
Tissue Samples ◽  
Angiogenesis Assay ◽  
Microvascular Endothelial ◽  
Coronary Angiogenesis

Coronary microvascular endothelial cell (CMECs) damage is implicated in diabetes-mediated heart failure with preserved ejection fraction (HFpEF). 4-hydroxy-2-nonenal (4HNE), a reactive aldehyde that is increased in diabetic heart, decreases angiogenesis in cultured mouse CMECs by decreasing the mRNA and protein levels of vascular endothelial growth factor receptor (VEGFR)2. Nuclear factor-kappa B (NF-kB), a transcription factor, was shown to transcribe VEGFR2. Thus, we presume 4HNE modulates NF-kB-mediated VEGFR2 transcription and regulates angiogenesis in CMECs. Aldehyde dehydrogenase (ALDH) 2, a mitochondrial enzyme that detoxifies 4HNE and confers cryoprotection. However, ALDH2 activity was reduced in the diabetic hearts which results in the augmentation of 4HNE-induced cardiotoxicity. Thus, we hypothesize that ALDH2 in CMECs reduces 4HNE-mediated cell signaling aberrations, and thereby, preserves coronary angiogenesis. We treated the cultured mouse CMECs with disulfiram (DSF) (2.5 μM), an ALDH2 inhibitor, alda1 (10 μM), an ALDH2 activator and prostratin (1 μM), an NF-κB activator prior to challenging the CMECs with 4HNE (75 μM). Our tube-formation angiogenesis assay revealed that pretreatment with DSF exacerbated a 4HNE-induced decrease in CMECs angiogenesis (P<0.0005 vs con and P<0.05 vs both 4HNE & DSF alone) while pretreatments with alda1 and prostratin attenuated a 4HNE-induced decrease in CMEC angiogenesis (P<0.05 vs 4HNE alone). DSF pretreatment exacerbated 4HNE mediated decrease in ALDH2 (P<0.005 vs con), phospho-IKBα (P<0.0005 vs con and P<0.05 vs both 4HNE and DSF alone), NF-κB levels, and nuclear translocation (P<0.0005 vs con and P<0.05 vs both 4HNE & DSF alone) and VEGFR2 (P<0.0005 vs con and P<0.05 vs both 4HNE and DSF alone) levels in cultured CMECs. Pretreatment with both prostratin and alda1 increased ALDH2 (P<0.0005 vs con), VEGFR2 (P<0.05 vs con) and NF-κB (P<0.005 vs con) levels in CMECs. The cardiac tissue samples of db/db mice when they manifest HFpEF showed increased 4HNE adducts, decreased NF-kB and VEGFR2 levels in CD31+ CMECs besides exhibiting low CMEC density. In conclusion, ALDH2 attenuates 4HNE-mediated decrease in coronary angiogenesis by decreasing VEGFR2 levels via low NF-κB mediated transcription.

scholarly journals Differential effects of cyclic and static stretch on coronary microvascular endothelial cell receptors and vasculogenic/angiogenic responses

2008 ◽  
Vol 295 (2) ◽  
pp. H794-H800 ◽  
Author(s):  
Wei Zheng ◽  
Lance P. Christensen ◽  
Robert J. Tomanek
Keyword(s):  
Growth Factor ◽  
Growth Factor Receptor ◽  
Tube Formation ◽  
Receptor Expression ◽  
Cyclic Stretch ◽  
Tube Length ◽  
Microvascular Endothelial Cell ◽  
Vegf Receptor ◽  
Static Stretch ◽  
Microvascular Endothelial

Mechanical stretch, an important growth stimulus, results not only from pulsatile blood flow and diastolic stretch of the ventricles [cyclic stretch (CS)] but also from tissue expansion during growth [constant static stretch (SS)]. We compared growth factor receptor expression and vasculogenic/angiogenic responses of rat coronary microvascular endothelial cells (ECs) by exposing cells to CS (10% elongation at 30 cycles/min) and SS (constant 10% elongation). Both CS and SS increased VEGF receptor (VEGF-R)2 protein levels and the extent of tube formation and branching. Moreover, both CS and SS enhanced VEGF-induced cell proliferation and tube formation, indicating that both types of stretch increase the sensitivity of ECs to VEGF. Blockade of VEGF-R2 prevented the increases in EC proliferation and aggregate tube length. However, CS but not SS enhanced EC Tie-2 protein and migration. CS affected a greater increase in tube length and branch formation than did SS. A unique finding was that SS but not CS increased VEGFR-1 in ECs. Our study is the first to distinguish between the effects of CS and SS on growth factor receptor expression and rat coronary microvascular EC proliferation, migration, and tube formation. In conclusion, EC angiogenic responses to these two types of stretch display both differences and similarities, but both CS and SS are dependent on VEGF-R2 signaling for their vasculogenic/angiogenic effects.

scholarly journals MiR-375-3p regulates rat pulmonary microvascular endothelial cell activity by targeting Notch1 during hypoxia

2020 ◽  
Vol 48 (7) ◽  
pp. 030006052092685
Author(s):  
Yuan An ◽  
Ziquan Liu ◽  
Hui Ding ◽  
Qi Lv ◽  
Haojun Fan ◽  
...  
Keyword(s):  
Inflammatory Response ◽  
Cell Activity ◽  
Tube Formation ◽  
Cell Counting ◽  
Luciferase Reporter ◽  
Microvascular Endothelial Cell ◽  
Adaptation To Hypoxia ◽  
Pulmonary Microvascular Endothelial Cell ◽  
Reporter Assays ◽  
Microvascular Endothelial

Objective Pulmonary microvascular endothelial cells (PMECs) exhibit specific responses in adaptation to hypoxia. However, the mechanisms regulating PMEC activities during hypoxia remain unclear. This study investigated the potential involvement of a microRNA, miR-375-3p, in the regulation of PMEC activities. Methods Primary PMECs were isolated from rats. The expression levels of miR-375-3p and Notch1 in the PMECs were detected by quantitative PCR and western blotting. Luciferase reporter assays were performed to explore the transcriptional regulation of Notch1 by miR-375-3p. The proliferation and chemotaxis of the PMECs were measured with the Cell Counting Kit-8 and Transwell invasion assays, respectively. Additionally, the capacity of hypoxia-treated PMECs for angiogenesis and inflammatory response was determined with tube formation assays and ELISA, respectively. Results The expression of miR-375-3p and Notch1 in the PMECs was significantly down-regulated and up-regulated during hypoxia, respectively. The results demonstrated that miR-375-3p directly targets Notch1 in PMECs, thereby suppressing the transcriptional expression of Notch1. It was further revealed that miR-375-3p regulates the proliferation, chemotaxis, angiogenesis, and inflammatory response of PMECs. Conclusions Our findings revealed the important role of miR-375-3p in the regulation of PMEC function and suggest the potential involvement of miR-375-3p in the development of lung diseases.

Cerebral microvascular endothelial cell tube formation: role of astrocytic epoxyeicosatrienoic acid release

2000 ◽  
Vol 278 (4) ◽  
pp. H1163-H1167 ◽  
Author(s):  
Diane H. Munzenmaier ◽  
David R. Harder
Keyword(s):  
Thymidine Incorporation ◽  
Tube Formation ◽  
Microvascular Endothelial Cell ◽  
Epoxyeicosatrienoic Acids ◽  
Epoxyeicosatrienoic Acid ◽  
Acid Release ◽  
Microvascular Endothelial ◽  
Cytochrome P 450 ◽  
The Brain

Cerebral microvascular endothelial cells (CMVEC) form tubes when cocultured with astrocytes (AS). Therefore, it appears that AS may be important in mediating angiogenesis in the brain. We hypothesized that AS modulate CMVEC tube formation by releasing a soluble factor. Thymidine incorporation in cultured CMVEC increased 305% when incubated with 50% conditioned AS medium for 24 h [control: 52,755 ± 4,838 counts per minute (cpm) per well, conditioned 161,082 ± 12,099 cpm/well, n = 8]. Because our laboratory has previously shown that AS can produce epoxyeicosatrienoic acids (EETs), which are known mitogens, we investigated whether release of EETs by AS is responsible for tube formation in the CMVEC-AS coculture. AS were seeded on Lab-Tek slides, CMVEC were seeded on the AS the next day, and cultures were allowed to progress for another 5 days with and without cytochrome P-450 epoxygenase blockade by 17-octadecynoic acid (17-ODYA). Tube formation in cocultures receiving 17-ODYA was significantly inhibited compared with control (93.8%). These data suggest that tube formation requires the release of EETs by AS.

scholarly journals Aminopeptidase inhibitor bestatin stimulates microvascular endothelial cell invasion in a fibrin matrix

2003 ◽  
Vol 90 (11) ◽  
pp. 921-929 ◽  
Author(s):  
Yvette Hensbergen ◽  
Erna Peters ◽  
Sareena Rana ◽  
Yvonne Elderkamp ◽  
Victor van Hinsbergh ◽  
...  
Keyword(s):  
Statistical Significance ◽  
Tube Formation ◽  
Model Systems ◽  
Microvascular Endothelial Cell ◽  
Fibrin Matrix ◽  
Angiogenic Effect ◽  
Microvascular Endothelial ◽  
High Concentrations ◽  
Blocking Antibodies ◽  
First Time

SummaryThe aminopeptidase inhibitor bestatin has been shown to have anti-angiogenic effects in a number of model systems. These effects are thought to result from inhibition of CD13 activity. Because tumor angiogenesis can evolve in a fibrin-rich stroma matrix we have studied for the first time the effects of bestatin on microvascular endothelial capillary-like tube formation in a fibrin matrix. Bestatin enhanced the formation of capillary-like tubes dose-dependently. Its effects were apparent at 8 µM; the increase was 3.7-fold at 125 µM; while high concentrations (>250 µM), that were shown to have anti-angiogenic effects in other systems, caused extensive matrix degradation. Specific CD13-blocking antibodies WM15 and MY-7, and the aminopeptidase inhibitors amastatin and actinonin also enhanced capillary-like tube formation (maximally 1.5-fold), but these effects did not reach statistical significance. The effect of bestatin was not due to a change in uPAR availability because the relative involvement of the u-PA/u-PAR activity was not altered by bestatin. In view of the present findings we hypothesize that aminopeptidases other than CD13 predominantly contribute to the observed pro-angiogenic effect of bestatin in a fibrin matrix. The identification of this novel effect of bestatin is important in the light of the proposed use of bestatin as anti-angiogenic and/or anti-tumor agent.

scholarly journals Circular RNA circPHKA2 Relieves OGD-Induced Human Brain Microvascular Endothelial Cell Injuries through Competitively Binding miR-574-5p to Modulate SOD2

2021 ◽  
Vol 2021 ◽  
pp. 1-17
Author(s):  
Xiaobo Yang ◽  
Xiuying Li ◽  
Chuanhong Zhong ◽  
Jianhua Peng ◽  
Jinwei Pang ◽  
...  
Keyword(s):  
Human Brain ◽  
Western Blotting ◽  
B Cell Lymphoma ◽  
Circular Rna ◽  
Tube Formation ◽  
Regulatory Subunit ◽  
Luciferase Reporter ◽  
Microvascular Endothelial Cell ◽  
Cerebral Stroke ◽  
Microvascular Endothelial

Background. Circular RNA phosphorylase kinase regulatory subunit alpha 2 (circPHKA2; hsa_circ_0090002) has a significantly, specifically different expression in acute ischemic stroke (AIS) patients’ blood. Here, we intended to investigate the role and mechanism of circPHKA2 in oxygen-glucose deprivation- (OGD-) induced stoke model in human brain microvascular endothelial cells (HBMEC). Methods. Expression of circPHKA2, microRNA- (miR-) 574-5p, and superoxide dismutase-2 (SOD2) was detected by quantitative PCR and western blotting. Cell injury was measured by detecting cell proliferation (EdU assay and CCK-8 assay), migration (transwell assay), neovascularization (tube formation assay), apoptosis (flow cytometry and western blotting), endoplasmic reticulum stress (western blotting), and oxidative stress (assay kits). Direct intermolecular interaction was determined by bioinformatics algorithms, dual-luciferase reporter assay, biotin-labelled miRNA capture, and argonaute 2 RNA immunoprecipitation. Results. circPHKA2 was downregulated in AIS patients’ blood in SOD2-correlated manner. Reexpressing circPHKA2 rescued EdU incorporation, cell viability and migration, tube formation, B cell lymphoma-2 (Bcl-2) expression, and SOD activity of OGD-induced HBMEC and alleviate apoptotic rate and levels of Bcl-2-associated protein (Bax), glucose-regulated protein 78 kD (GRP78), C/EBP-homologous protein (CHOP), caspase-12, reactive oxygen species (ROS), and malondialdehyde (MDA). Additionally, blocking SOD2 partially attenuated these roles of circPHKA2 overexpression. Molecularly, circPHKA2 upregulated SOD2 expression via interacting with miR-574-5p, and miR-574-5p could target SOD2. Similarly, allied to neurovascular protection of circPHKA2 was the downregulation of miR-574-5p. Conclusion. circPHKA2 could protect HBMEC against OGD-induced cerebral stroke model via the miR-574-5p/SOD2 axis, suggesting circPHKA2 as a novel and promising candidate in ischemic brain injury.

Abstract 24: Hydrogen Sulfide Potentiates Bone Marrow-derived Angiogenic Progenitor Cell-mediated Ischemic Limb Angiogenesis in Diabetic Animals

2016 ◽  
Vol 119 (suppl_1) ◽  
Author(s):  
Zhongjian Cheng ◽  
Venkata NS Garikipati ◽  
Emily Nickoloff ◽  
Chunlin Wang ◽  
David Polhemus ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Bone Marrow ◽  
Hydrogen Sulfide ◽  
Critical Role ◽  
Tube Formation ◽  
Microvascular Endothelial Cell ◽  
Diabetic Patients ◽  
No Production ◽  
Therapeutic Effects ◽  
Microvascular Endothelial

Rationale: Critical hindlimb ischemia (CLI) represents an outcome of peripheral artery disease with a high incidence in diabetic patients. Recently, insufficient hydrogen sulfide (H 2 S) production has been implicated in cardiovascular disease. Objective: Explore the role of H 2 S in diabetes-induced bone marrow-derived angiogenic progenitor cells (BMAPC) dysfunction and its therapeutic effects on ischemic hindlimb injury of diabetes. Methods and Results: Diabetic BMAPCs from db/db mice showed decreased intracellular H 2 S production and blunted cystathionine γ-lyase (CSE) expression-an enzyme required for H 2 S generation. High glucose (HG) inhibited H 2 S production, migration and increased apoptosis of BMPAC that was rescued by H 2 S donor diallyl trisulfide (DATS) or overexpression of CSE. Administration DATS or local injection of diabetic BMAPCs overexpressing CSE significantly potentiated BMAPC-mediated blood flow recovery, capillary and arteriole formation, skeletal muscle architecture preservation, cell survival and decrease of perivascular infiltration of monocytes (CD68 + ) cells in ischemic skeletal muscle. Moreover, overexpression of CSE increased diabetic BMAPCs homing and engraftment in ischemic hindlimb. HG-impaired human cardiac microvascular endothelial cells (HCMVECs) tube formation and migration were rescued by DATS or overexpression of CSE. Mechanistically, HG increased threonine-495 phosphorylation of eNOS (eNOS-pT495) and inhibited nitric oxide (NO) production in HCMVECs which were rescued by DATS or overexpression of CSE. Silencing CSE by siRNA impaired tube formation and increased eNOS-pT495 expression in HCMVECs. Finally, HG-treated BMAPCs impaired mouse microvascular endothelial cell tube formation that was rescued by DATS. Conclusions: Our data suggests that CSE downregulation-induced H 2 S insufficient plays a critical role in diabetes-mediated BMAPC dysfunction. Administration of DATS or overexpression of CSE improves diabetic BMAPC-mediated angiogenesis/neovascularization via, at least partially, eNOS inactivation/NO reduction pathways. Our data indicate that H 2 S and overexpression of CSE in diabetic BMAPCs may open novel avenues for cell-based therapeutics of CLI in diabetic patients.

scholarly journals PLD1-dependent PKCγ activation downstream to Src is essential for the development of pathologic retinal neovascularization

Blood ◽  
2010 ◽  
Vol 116 (8) ◽  
pp. 1377-1385 ◽  
Author(s):  
Qiuhua Zhang ◽  
Dong Wang ◽  
Venkatesh Kundumani-Sridharan ◽  
Laxmisilpa Gadiparthi ◽  
Dianna A. Johnson ◽  
...  
Keyword(s):  
Tube Formation ◽  
Dominant Negative ◽  
Retinal Neovascularization ◽  
Endothelial Cell Migration ◽  
Microvascular Endothelial Cell ◽  
Dominant Negative Mutant ◽  
Vascular Endothelial ◽  
Retinal Angiogenesis ◽  
Microvascular Endothelial ◽  
Endothelial Growth Factor

Abstract Vascular endothelial growth factor (VEGF) appears to be an important mediator of pathologic retinal angiogenesis. In understanding the mechanisms of pathologic retinal neovascularization, we found that VEGF activates PLD1 in human retinal microvascular endothelial cells, and this event is dependent on Src. In addition, VEGF activates protein kinase C-γ (PKCγ) via Src-dependent PLD1 stimulation. Inhibition of Src, PLD1, or PKCγ via pharmacologic, dominant negative mutant, or siRNA approaches significantly attenuated VEGF-induced human retinal microvascular endothelial cell migration, proliferation, and tube formation. Hypoxia also induced Src-PLD1-PKCγ signaling in retina, leading to retinal neovascularization. Furthermore, siRNA-mediated down-regulation of VEGF inhibited hypoxia-induced Src-PLD1-PKCγ activation and neovascularization. Blockade of Src-PLD1-PKCγ signaling via the siRNA approach also suppressed hypoxia-induced retinal neovascularization. Thus, these observations demonstrate, for the first time, that Src-dependent PLD1-PKCγ activation plays an important role in pathologic retinal angiogenesis.

Abstract 387: Role of Aldehyde Dehydrogenase (aldh) 2 in Angiotensin II-induced Decrease in Angiogenesis of Coronary Endothelial Cells

2020 ◽  
Vol 127 (Suppl_1) ◽  
Author(s):  
Bipradas Roy ◽  
Suresh S Palaniyandi
Keyword(s):  
Angiotensin Ii ◽  
Aldehyde Dehydrogenase ◽  
Molecular Mechanisms ◽  
Growth Factor Receptor ◽  
Tube Formation ◽  
Vascular Endothelial ◽  
Ang Ii ◽  
Endothelial Growth Factor

Diabetes-induced coronary endothelial cell (CEC) dysfunction following defective angiogenesis is reported in cardiovascular diseases (CVD). Angiotensin II (Ang II), a vasoactive molecule, is upregulated in diabetes. However, the underlying molecular mechanisms of Ang II-induced CEC dysfunction are not fully understood. Aldehyde dehydrogenase (ALDH) 2 is cytoprotective in diabetic CVD. Thus, we hypothesize that ALDH2 improves Ang II-mediated defective CEC angiogenesis. To test our hypothesis, we treated the cultured mouse CECs with Ang II (0.1, 1 and 10 μM) for 2 and 4 hours. Next, we treated CEC with Alda-1 (10 μM), an ALDH2 activator or disulfiram (2.5 μM), an ALDH2 inhibitor, before challenging MCECs with Ang II. We found that Ang II attenuated tube formation (P<0.05 vs control) which indicates in vitro angiogenesis. Next, we found that Ang II have downregulated the mRNA expressions of vascular endothelial growth factor receptor VEGFR1 (p<0.05) and upregulated angiotensin II type-2 receptor (AT2R) (P<0.05) in cultured CECs compared to controls. ALDH2 inhibition with disulfiram potentiated Ang II-induced decrease in angiogenesis (P<0.005) by decreasing the expressions of VEGFR1 (P<0.0005) and increasing the expression of AT2R (p<0.05) relative to Ang II alone. Additionally, activation of ALDH2 activity with Alda-1 rescued Ang II-induced decrease in angiogenesis (P<0.05) by increasing the expression of VEGFR1 (P<0.05) and decreasing the expression of AT2R (P<0.05) relative to Ang II alone. Finally, we conclude that ALDH2 can be an important therapeutic target to improve coronary angiogenesis in diabetic CVD.

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