scholarly journals MALAT1 modulates miR-146’s protection of microvascular endothelial cells against LPS-induced NF-κB activation and inflammatory injury

2019 ◽  
Vol 25 (7) ◽  
pp. 433-443
Author(s):  
Lin-Lin Feng ◽  
Wei-Na Xin ◽  
Xiu-Li Tian
Keyword(s):  
Endothelial Cells ◽  
Cell Viability ◽  
Combined Treatment ◽  
Luciferase Assay ◽  
Luciferase Reporter ◽  
Microvascular Endothelial Cells ◽  
Dependent Manner ◽  
Inflammatory Injury ◽  
Tnf Α ◽  
Microvascular Endothelial

To investigate the role of miR-146 and its possible relationship with MALAT1 in LPS-induced inflammation in human microvascular endothelial cells (HMECs), HMEC-1 cells were treated with LPS to construct an inflammatory injury cell model, and the cell viability, TNF-α and IL-6 secretion and the expression levels of VCAM-1, SELE and ICAM-1 were analysed as markers of inflammatory injury. The regulation mechanisms of miR-146 interacted with MALAT1 and the downstream NF-κB signalling were also verified by dual-luciferase assay and knockdown technology. LPS significantly decreased the cell viability, increased levels of VCAM-1, SELE and ICAM-1 and also up-regulated miR-146a/b, TNF-α and IL-6 in a dose-dependent manner. Over-expression of miR-146a resulted in down-regulation of TNF-α and IL-6, as well as VCAM-1, SELE and ICAM-1, while inhibition of miR-146a led to opposite results. The dual-luciferase reporter assay showed both miR-146a and miR-146b directly targeted and negatively regulated the expression of MALAT1. Silencing of MALAT1 suppressed LPS-induced NF-κB activation and TNF-α and IL-6 secretion, reducing the cell inflammatory injury, but these changes were reversed after combined treatment with miR-146a inhibitor. Taken together, we demonstrate that miR-146 protects HMECs against inflammatory injury by inhibiting NF-κB activation. This process is modulated by MALAT1.

scholarly journals Cigarette Smoke Extract and Lipopolysaccharides Induce Pyroptosis in Rats Pulmonary Microvascular Endothelial Cells

2022 ◽  
Author(s):  
Qizhi Wang ◽  
Min Liu ◽  
Yu Liu ◽  
Zhen Zhang ◽  
Zhengping Bai
Keyword(s):  
Flow Cytometry ◽  
Endothelial Cells ◽  
Cell Viability ◽  
Cigarette Smoke ◽  
Cigarette Smoke Extract ◽  
Microvascular Endothelial Cells ◽  
Dependent Manner ◽  
Pulmonary Microvascular Endothelial Cells ◽  
Microvascular Endothelial ◽  
Dose Dependent

Abstract Objective: To investigate the effects of cigarette smoke extract (CSE) and lipopolysaccharide (LPS) on the activity and pyroptosis of pulmonary microvascular endothelial cells (PMVECs). Methods: PMVECs were cultured without treatment or with CSE (1%-25%), LPS, or CSE+LPS. Cell viability was detected using the CCK8 method. Apoptosis was evaluated by flow cytometry. Cell morphology was evaluated using optical microscopy. The content of IL-1β and IL-18 was measured by ELISA. Results: CSE decreased cell viability in a dose-dependent manner. The cells in the CSE+LPS group showed the most obvious cytomorphological changes and the highest pyroptosis rate under the microscope. Flow cytometry showed that the CSE and LPS groups showed higher apoptosis rates than the blank group; the apoptotic rate in the CSE+LPS group was even higher (P<0.01). Compared with the bkank group, the levels of IL-18 and IL-1β in the cell supernatant of the CSE, LPS, and CSE+LPS groups increased significantly, with significant differences (P<0.01). There were no differences between the CSE and LPS groups (P>0.05). Compared with the CSE and LPS groups, the CSE+LPS group had higher IL-18 and IL-1β (P<0.01). Conclusion: The effect of CSE on cell viability is dose-dependent. CSE+LPS can induce cell pyroptosis and increase the levels of inflammatory cytokines in PMVECs. These observations demonstrated that pyroptosis caused by CSE and LPS might play an important role in pulmonary vascular remodeling.

scholarly journals Anti-Inflammatory Effect and Cellular Uptake Mechanism of Carbon Nanodots in in Human Microvascular Endothelial Cells

Nanomaterials ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 1247
Author(s):  
Sarah Belperain ◽  
Zi Yae Kang ◽  
Andrew Dunphy ◽  
Brandon Priebe ◽  
Norman H. L. Chiu ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Interleukin 1 ◽  
Antioxidant Properties ◽  
Synthesis Method ◽  
Microvascular Endothelial Cells ◽  
Carbon Nanodots ◽  
Uptake Mechanism ◽  
Tnf Α ◽  
Anti Inflammatory ◽  
Microvascular Endothelial

Cardiovascular disease (CVD) has become an increasingly important topic in the field of medical research due to the steadily increasing rates of mortality caused by this disease. With recent advancements in nanotechnology, a push for new, novel treatments for CVD utilizing these new materials has begun. Carbon Nanodots (CNDs), are a new form of nanoparticles that have been coveted due to the green synthesis method, biocompatibility, fluorescent capabilities and potential anti-antioxidant properties. With much research pouring into CNDs being used as bioimaging and drug delivery tools, few studies have been completed on their anti-inflammatory potential, especially in the cardiovascular system. CVD begins initially by endothelial cell inflammation. The cause of this inflammation can come from many sources; one being tumor necrosis factor (TNF-α), which can not only trigger inflammation but prolong its existence by causing a storm of pro-inflammatory cytokines. This study investigated the ability of CNDs to attenuate TNF-α induced inflammation in human microvascular endothelial cells (HMEC-1). Results show that CNDs at non-cytotoxic concentrations reduce the expression of pro-inflammatory genes, mainly Interleukin-8 (IL-8), and interleukin 1 beta (IL-1β). The uptake of CNDs by HMEC-1s was examined. Results from the studies involving channel blockers and endocytosis disruptors suggest that uptake takes place by endocytosis. These findings provide insights on the interaction CNDs and endothelial cells undergoing TNF-α induced cellular inflammation.

scholarly journals A Feedback Loop Involving MicroRNA-150 and MYB Regulates VEGF Expression in Brain Microvascular Endothelial Cells After Oxygen Glucose Deprivation

2021 ◽  
Vol 12 ◽  
Author(s):  
Song Zhang ◽  
Anqi Chen ◽  
Xiaolu Chen
Keyword(s):  
Endothelial Cells ◽  
Feedback Loop ◽  
Glucose Deprivation ◽  
Luciferase Reporter ◽  
Microvascular Endothelial Cells ◽  
Oxygen Glucose Deprivation ◽  
Brain Microvascular Endothelial Cells ◽  
Vegf Expression ◽  
Direct Target ◽  
Microvascular Endothelial

Vascular endothelial growth factor (VEGF) plays a pivotal role in regulating cerebral angiogenesis after stroke. Meanwhile, excessive VEGF expression induces increased microvascular permeability in brain, probably leading to neurological deterioration. Therefore, the appropriate level of VEGF expression is significant to the recovery of brain exposed to stroke. In this work, we demonstrate that microRNA-150 (miR-150) and its predicted target MYB form a negative feedback loop to control the level of post-stroke VEGF expression. Repression of MYB leads to decreased expression of miR-150 in brain microvascular endothelial cells (BMVECs) exposed to oxygen glucose deprivation (OGD), thus miR-150 was predicted to be down-regulated by MYB. Moreover, MYB was confirmed to be a direct target of miR-150 by using dual luciferase reporter assay. In our previous work, we have validated VEGF as another direct target of miR-150. Therefore, MYB participates in regulation of VEGF via miR-150 under OGD, forming a feedback loop with miR-150. We also find that high levels of miR-150 inhibitors combined with MYB silence contribute to further enhancement of VEGF expression in BMVECs in response to OGD. These observations suggest that the feedback loop comprised of miR-150 and MYB, which is a pivotal endogenous epigenetic regulation to control the expression levels of VEGF in BMVECs subjected to OGD.

Abstract 235: Hypoxia Enhanced Delivery of Mitochondria-Targeted Catalase Protects Choroid Retinal Microvascular Endothelial Cells from Oxidative Stress

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Christopher J Dougherty ◽  
Howard Prentice ◽  
Kathleen Dorey ◽  
Keith A Webster ◽  
Janet C Blanks
Keyword(s):  
Oxidative Stress ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
High Glucose ◽  
Endothelial Permeability ◽  
Luciferase Assay ◽  
Expression Vectors ◽  
Microvascular Endothelial Cells ◽  
Tissue Specific ◽  
Microvascular Endothelial

Loss of pericytes is a critical event early in the progression of microvascular dysfunction in diabetic retinopathy. Pericyte loss may be linked to high glucose mediated reactive oxygen species generation, blocking N-cadherin trafficking to the endothelial cell surface preventing pericyte recruitment and vessel stabilization. Hydrogen peroxide has been identified as a major free radical produced during high glucose exposure in endothelial cells. The goal of this research is to determine if tissue-specific hypoxia-regulated expression of a mitochondria-targeted catalase can prevent or limit RF/6A microvascular endothelial cell apoptosis and decrease vascular permeability by limiting cellular oxidative stress. For the development of tissue-specific and hypoxia-enhanced expression vectors, promoters were constructed with nine tandem combinations of HREs. This 9x HRE oligomer enhancer was inserted together into a pGL3 firefly luciferase plasmid with the Tie2( short ) promoter for endothelial-specific expression. The 9xHRE-Tie2( sh ) promoter construct was highly selective for RF/6A cells producing a basal amount of mitochondria-targeted catalase equivalent to the Tie2( short ) promoter alone. In response to hypoxia ( pO 2 = 1% ), the 9xHRE-Tie2( short ) promoter showed a 21-fold hypoxia-inducible activation similar in strength to the CMV promoter , measured by dual luciferase assay. The hybrid promoters were incorporated into a replication deficient AAV delivery system for apoptosis and cell culture based endothelial permeability assays. In preliminary assays using RF/6A microvascular endothelial cells, apoptosis was reduced by 58% and permeability was reduced by 46%. The results suggest that mitochondria-targeted catalase protects RF/6A microvascular endothelial cells from apoptosis and reduces endothelial permeability in a high-glucose, low-oxygen environment.

scholarly journals MicroRNA-429/Cxcl1 Axis Protective Against Oxygen Glucose Deprivation/Reoxygenation-Induced Injury in Brain Microvascular Endothelial Cells

Dose-Response ◽  
2020 ◽  
Vol 18 (2) ◽  
pp. 155932582091378
Author(s):  
Jun Leng ◽  
Wei Liu ◽  
Li Li ◽  
Fang Yue Wei ◽  
Meng Tian ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Ischemia Reperfusion ◽  
Glucose Deprivation ◽  
Luciferase Reporter ◽  
Microvascular Endothelial Cells ◽  
Oxygen Glucose Deprivation ◽  
Brain Microvascular Endothelial Cells ◽  
Public Data ◽  
Microvascular Endothelial

Objective: The objective of the present work was to study the role of Cxcl1 in cerebral ischemia–reperfusion (I/R) injury and to in-depth explore its pathogenesis. Methods: The expression of Cxcl1 based on the public data was analyzed. Then, we constructed an oxygen glucose deprivation/reoxygenation (OGD/R) model in vitro using mice brain microvascular endothelial cells (BMECs) to simulate cerebral I/R in vivo. Results: The results of quantitative real-time polymerase chain reaction assay uncovered that Cxcl1 showed higher expression while miR-429 showed lower expression in BMECs damaged by OGD/R, whereas overexpression of Cxcl1 or inhibition of miR-429 expression can strengthen this effect. Hereafter, through dual luciferase reporter assay, we verified that miR-429 directly targets Cxcl1 and negatively regulates Cxcl1 expression. Furthermore, the results also revealed that overexpression of Cxcl1 can reverse the miR-429-mediated effects. Conclusion: We concluded that miR-429 exerts protective effects against OGD/R-induce injury in vitro through modulation of Cxcl1 and nuclear factor kinase B pathway, hoping provide a new view on the pathogenesis of cerebral I/R injury and a feasible potential therapeutic target.

scholarly journals LncRNA MALAT1 up-regulates VEGF-A and ANGPT2 to promote angiogenesis in brain microvascular endothelial cells against oxygen–glucose deprivation via targetting miR-145

2019 ◽  
Vol 39 (3) ◽  
Author(s):  
Lanfen Ren ◽  
Chunxia Wei ◽  
Kui Li ◽  
Zuneng Lu
Keyword(s):  
Endothelial Cells ◽  
Glucose Deprivation ◽  
Luciferase Assay ◽  
Microvascular Endothelial Cells ◽  
Oxygen Glucose Deprivation ◽  
Brain Microvascular Endothelial Cells ◽  
Qrt Pcr ◽  
Lncrna Malat1 ◽  
Microvascular Endothelial

Abstract Stroke is one of the leading causes of death and long-term disability around the world. Angiogenesis is supposed to protect brain microvascular endothelial cells (BMECs) from oxidative and ischemic stress. Previous studies indicated that interaction between metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) and miR-145 was involved in myocardial ischemia reperfusion, suggesting MALAT1 and miR-145 were also mediated with the progress of angiogenesis and cell migration in oxygen–glucose deprivation (OGD)-induced BMECs. The present study aimed to investigate the functional roles of MALAT1 in regulating miR-145 and its downstream pro-angiogenesis factors, vascular endothelial growth factor (VEGF)-A and Angiopoietin-2 (ANGPT2) during the progress of angiogenesis in OGD-induced BMECs. An in vitro OGD model was employed in mouse BMECs to mimic brain hypoxic and ischemic conditions; MTT was used to determine cell viability. qRT-PCR was used to determine the expression of long non-coding RNA (lncRNA)-MALAT1 and miR-145 under OGD conditions; in vitro tube formation assay was used to investigate angiogenic effect of MALAT1 and miR-145. The relationship between lncRNA-MALAT1/miR-145 and miR-145/VEGF-A/ANGPT2 was evaluated by qRT-PCR and Western blot, and direct binding was assessed using dual luciferase assay. Results showed that the levels of lncRNA-MALAT1 and miR-145 were up-regulated in OGD-induced BMECs. miR-145 functioned as an anti-angiogenic and pro-apoptotic factor in OGD treated BMECs via down-regulating VEGF-A and ANGPT2 directly. While lncRNA-MALAT1 enhanced the expressions of VEGF-A and ANGPT2 by targetting miR-145 to promote angiogenesis and proliferation of BMECs under OGD conditions. Our present study revealed the inhibitory functions of miR-145 on angiogenesis through direct targetting on VEGF-A and ANGPT2 for the first time and proved the protective role of lncRNA-MALAT1 for BMECs under OGD conditions through the direct regulation of miR-145.

scholarly journals α1G T-type calcium channel determines the angiogenic potential of pulmonary microvascular endothelial cells

2019 ◽  
Vol 316 (3) ◽  
pp. C353-C364 ◽  
Author(s):  
Zhen Zheng ◽  
Hairu Chen ◽  
Peilin Xie ◽  
Carol A. Dickerson ◽  
Judy A. C. King ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Functional Role ◽  
Transient Receptor Potential ◽  
Network Formation ◽  
Microvascular Endothelial Cells ◽  
Dependent Manner ◽  
Channel Blockade ◽  
Angiogenic Potential ◽  
Pulmonary Microvascular Endothelial Cells ◽  
Microvascular Endothelial

Pulmonary microvascular endothelial cells (PMVECs) display a rapid angioproliferative phenotype, essential for maintaining homeostasis in steady-state and promoting vascular repair after injury. Although it has long been established that endothelial cytosolic Ca2+ ([Ca2+]i) transients are required for proliferation and angiogenesis, mechanisms underlying such regulation and the transmembrane channels mediating the relevant [Ca2+]i transients remain incompletely understood. In the present study, the functional role of the microvascular endothelial site-specific α1G T-type Ca2+ channel in angiogenesis was examined. PMVECs intrinsically possess an in vitro angiogenic “network formation” capacity. Depleting extracellular Ca2+ abolishes network formation, whereas blockade of vascular endothelial growth factor receptor or nitric oxide synthase has little or no effect, suggesting that the network formation is a [Ca2+]i-dependent process. Blockade of the T-type Ca2+ channel or silencing of α1G, the only voltage-gated Ca2+ channel subtype expressed in PMVECs, disrupts network formation. In contrast, blockade of canonical transient receptor potential (TRP) isoform 4 or TRP vanilloid 4, two other Ca2+ permeable channels expressed in PMVECs, has no effect on network formation. T-type Ca2+ channel blockade also reduces proliferation, cell-matrix adhesion, and migration, three major components of angiogenesis in PMVECs. An in vivo study demonstrated that the mice lacking α1G exhibited a profoundly impaired postinjury cell proliferation in the lungs following lipopolysaccharide challenge. Mechanistically, T-type Ca2+ channel blockade reduces Akt phosphorylation in a dose-dependent manner. Blockade of Akt or its upstream activator, phosphatidylinositol-3-kinase (PI3K), also impairs network formation. Altogether, these findings suggest a novel functional role for the α1G T-type Ca2+ channel to promote the cell’s angiogenic potential via a PI3K-Akt signaling pathway.

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