scholarly journals 17β-Estradiol Enhances Vascular Endothelial Ets-1/miR-126-3p Expression: The Possible Mechanism for Attenuation of Atherosclerosis

2016 ◽  
Vol 102 (2) ◽  
pp. 594-603 ◽  
Author(s):  
Ping Li ◽  
Jinzhi Wei ◽  
Xiaosa Li ◽  
Yang Cheng ◽  
Weiyu Chen ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Vascular Endothelial Cells ◽  
Cell Model ◽  
Tube Formation ◽  
Luciferase Assay ◽  
Endothelial Cell Proliferation ◽  
Vascular Endothelial ◽  
Atherosclerotic Lesions ◽  
17Β Estradiol ◽  
Cell Adhesion Molecule 1

Abstract Context: Endothelial microRNA 126 (miR-126) attenuates the development of atherosclerosis (AS). However, there is no evidence showing the role of miR-126 in estrogen’s antiatherogenic effects. Objective: We hypothesized that 17β-estradiol (E2) modulates miR-126 expression and thus may improve endothelial function and retard AS development. Design/Setting/Participants: This was a prospective cohort study of 12 healthy regularly menstruating female volunteers. ApoE−/− mice were used as the atherosclerosis model and human umbilical vascular endothelial cells (HUVECs) were cultured as the cell model. Main Outcome Measures: Serum hormones and miR-126-3p levels were measured up to 3 times for 1 cycle. Real-time polymerase chain reaction, histology for atherosclerotic lesions, immunofluorescence, luciferase assay, transfection experiments, cell proliferation, migration and tube formation assay, and western blot were performed. Results: Serum concentrations of miR-126-3p in cycling women were higher at the ovulatory and luteal phases than in the follicular phase, and they were positively correlated with E2 values. Administration of miR-126-3p mimics to ApoE−/− mice-attenuated atherogenesis, and antagomir-126-3p partially reversed the protective effect of E2 on atherogenesis. In HUVECs, E2 increased miR-126-3p expression via upregulation of Ets-1 (a transcription factor for miR-126). c-Src/Akt signaling was important for E2-mediated expression of Ets-1/miR-126. E2 decreased expression of miR-126-3p target Spred1 (a protein that inhibits mitogenic signaling). Overexpression of Spred1 partially blocked enhancement of endothelial cell proliferation, migration, and tube formation by E2. Additionally, E2 regulates miR-126-3p–mediated expression of vascular cell adhesion molecule-1 to inhibit monocyte adhesion into HUVECs. Conclusions: E2 protection against atherogenesis is possibly mediated by Ets-1/miR-126.

scholarly journals CircRNA RSF1 regulated ox-LDL induced vascular endothelial cells proliferation, apoptosis and inflammation through modulating miR-135b-5p/HDAC1 axis in atherosclerosis

2021 ◽  
Vol 54 (1) ◽  
Author(s):  
Xiaohao Zhang ◽  
Junying Lu ◽  
Qinghua Zhang ◽  
Qiang Luo ◽  
Bin Liu
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cells ◽  
Cell Adhesion ◽  
Adhesion Molecule ◽  
Cell Adhesion Molecule ◽  
Luciferase Reporter ◽  
Endothelial Cell Proliferation ◽  
Vascular Endothelial ◽  
Apoptosis And Inflammation ◽  
Cell Adhesion Molecule 1

Abstract Background Atherosclerosis (AS) is the most common type in cardiovascular disease. Due to its complex pathogenesis, the exact etiology of AS is unclear. circRNA has been shown to play an essential role in most diseases. However, the underlying mechanism of circRNA in AS has been not understood clearly. Methods Quantitative Real-Time PCR assay was used to detect the expression of circRSF1, miR-135b-5p and histone deacetylase 1 (HDAC1). Western blot was applied to the measure of protein expression of HDAC1, B-cell lymphoma-2 (Bcl-2), BCL2-associated X (Bax), cleaved-caspase-3, vascular cell adhesion molecule 1 (VCAM1), intercellular cell adhesion molecule-1 (ICAM1) and E-selectin. MTT assay and flow cytometry were used to detect cell proliferation and apoptosis, respectively. Dual luciferase reporter assay and RIP assay was used to determine the relationship among circRSF1, miR-135b-5p and HDAC1. Besides, an ELISA assay was performed to measure the levels of IL-1β, IL-6, TNF-α and IL-8. Results In this study, ox-LDL inhibited circRSF1 and HDAC1 expression while upregulated miR-135b-5p expression in Human umbilical vein endothelial cells (HUVECs). Importantly, ox-LDL could inhibit HUVECs growth. Moreover, promotion of circRSF1 or inhibition of miR-135b-5p induced cell proliferation while inhibited apoptosis and inflammation of ox-LDL-treated HUVECs, which was reversed by upregulating miR-135b-5p or downregulating HDCA1 in ox-LDL-treated HUVECs. More than that, we verified that circRSF1 directly targeted miR-135b-5p and HDAC1 was a target mRNA of miR-135b-5p in HUVECs. Conclusion CircRSF1 regulated ox-LDL-induced vascular endothelial cell proliferation, apoptosis and inflammation through modulating miR-135b-5p/HDAC1 axis in AS, providing new perspectives and methods for the treatment and diagnosis of AS.

scholarly journals An In Vitro Model of Diabetic Retinal Vascular Endothelial Dysfunction and Neuroretinal Degeneration

2021 ◽  
Vol 2021 ◽  
pp. 1-12
Author(s):  
Qiyun Wang ◽  
Xinyuan Zhang ◽  
Kaiyue Wang ◽  
Ling Zhu ◽  
Bingjie Qiu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Endothelial Dysfunction ◽  
Vascular Endothelial Cells ◽  
Tube Formation ◽  
Cell Counting ◽  
Vascular Endothelial ◽  
Migration Assay ◽  
Tube Formation Assay ◽  
Wide Range

Background. Diabetic retinopathy (DR) is a leading cause of blindness in working-age populations. Proper in vitro DR models are crucial for exploring pathophysiology and identifying novel therapeutic targets. This study establishes a rational in vitro diabetic retinal neuronal-endothelial dysfunction model and a comprehensive downstream validation system. Methods. Human retinal vascular endothelial cells (HRMECs) and retinal ganglion cells (RGCs) were treated with different glucose concentrations with mannitol as matched osmotic controls. Cell proliferation and viability were evaluated by the Cell Counting Kit-8. Cell migration was measured using a transwell migration assay. Cell sprouting was assessed by a tube formation assay. The VEGF expression was assessed by ELISA. RGCs were labeled by neurons and RGC markers TUJ1 and BRN3A for quantitative and morphological analysis. Apoptosis was detected using PI/Hoechst staining and TUNEL assay and quantified by ImageJ. Results. Cell proliferation and migration in HRMECs were significantly higher in the 25 mM glucose-treated group ( p < 0.001 ) but lower in the 50 mM and 100 mM groups ( p < 0.001 ). The permeability and the apoptotic index in HRMECs were statistically higher in the 25 mM, 50 mM, and 100 mM groups ( p < 0.05 ). The tube formation assay found that all the parameters were significantly higher in the 25 mM and 50 mM groups ( p < 0.001 ) concomitant with the elevated VEGFA expression in HRMECs ( p = 0.016 ). Cell viability was significantly lower in the 50 mM, 100 mM, and 150 mM groups in RGCs ( p 50 mM = 0.013 , p 100 mM = 0.019 , and p 150 mM = 0.002 ). Apoptosis was significantly elevated, but the proportion of RGCs with neurite extension was significantly lower in the 50 mM, 100 mM, and 150 mM groups ( p 50 mM < 0.001 , p 100 m M < 0.001 , and p 150 mM < 0.001 ). Conclusions. We have optimized glucose concentrations to model diabetic retinal endothelial (25-50 mM) or neuronal (50-100 mM) dysfunction in vitro, which have a wide range of downstream applications.

scholarly journals Hepatitis B Virus preS2 Domain Promotes Angiogenesis in Hepatocellular Carcinoma via Transcriptional Activation of VEGFA Promoter

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Fang Luan ◽  
Bin Liu ◽  
Xiangrui Guo ◽  
Wang Yong
Keyword(s):  
Hepatocellular Carcinoma ◽  
Endothelial Cells ◽  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Vascular Endothelial Cells ◽  
Luciferase Assay ◽  
Luciferase Reporter ◽  
Endothelial Cell Proliferation ◽  
Vascular Endothelial ◽  
B Virus

: Angiogenesis is closely related to the development and progression of hepatocellular carcinoma (HCC). Angiogenic factors have been confirmed to be overexpressed in HCC. The hepatitis B virus preS2 domain is a transactivator that plays an important role in hepatitis B virus (HBV)-related HCC. Here, we aimed to investigate the potential of the preS2 domain in inducing angiogenesis in HCC. A total of 25 cases of pathologically confirmed HCC were screened. The levels of preS2, CD34, and vascular endothelial growth factor A (VEGFA) in HCC samples were evaluated by immunohistochemistry (IHC). The proliferation of vascular endothelial cells was detected by CCK-8. Besides, VEGFA was analyzed by Western blot in HCC cells. The effect of preS2 on the VEGFA promoter was measured by dual-luciferase reporter assays. We found that preS2 domain-positive HCCs had significantly higher microvessel density (MVD) and VEGFA expression than preS2 domain-negative HCCs. Overexpression of preS2 upregulated VEGFA expression in HepG2 and activated vascular endothelial cell proliferation. However, blocking preS2 expression reduced VEGFA expression in HepG2.2.15 and inhibited the proliferation of vascular endothelial cells. In addition, a dual-luciferase assay indicated that the preS2 domain could activate VEGFA promoter activity. In conclusion, we showed that the expression of the preS2 domain promotes angiogenesis by transactivating the VEGFA promoter in HCC.

scholarly journals Regulation of Endothelial Cell Proliferation and Vascular Assembly through Distinct mTORC2 Signaling Pathways

2015 ◽  
Vol 35 (7) ◽  
pp. 1299-1313 ◽  
Author(s):  
Shan Wang ◽  
Katherine R. Amato ◽  
Wenqiang Song ◽  
Victoria Youngblood ◽  
Keunwook Lee ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Vascular Endothelial Cells ◽  
Endothelial Cell Proliferation ◽  
Vascular Endothelial ◽  
Relative Contribution ◽  
Protein Kinase Cα

Mammaliantargetofrapamycin (mTOR) is a serine/threonine kinase that regulates a diverse array of cellular processes, including cell growth, survival, metabolism, and cytoskeleton dynamics. mTOR functions in two distinct complexes, mTORC1 and mTORC2, whose activities and substrate specificities are regulated by complex specific cofactors, including Raptor and Rictor, respectively. Little is known regarding the relative contribution of mTORC1 versus mTORC2 in vascular endothelial cells. Using mouse models of Raptor or Rictor gene targeting, we discovered that Rictor ablation inhibited vascular endothelial growth factor (VEGF)-induced endothelial cell proliferation and assemblyin vitroand angiogenesisin vivo, whereas the loss of Raptor had only a modest effect on endothelial cells (ECs). Mechanistically, the loss of Rictor reduced the phosphorylation of AKT, protein kinase Cα (PKCα), and NDRG1 without affecting the mTORC1 pathway. In contrast, the loss of Raptor increased the phosphorylation of AKT despite inhibiting the phosphorylation of S6K1, a direct target of mTORC1. Reconstitution of Rictor-null cells with myristoylated AKT (Myr-AKT) rescued vascular assembly in Rictor-deficient endothelial cells, whereas PKCα rescued proliferation defects. Furthermore, tumor neovascularizationin vivowas significantly decreased upon EC-specific Rictor deletion in mice. These data indicate that mTORC2 is a critical signaling node required for VEGF-mediated angiogenesis through the regulation of AKT and PKCα in vascular endothelial cells.

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