Caveolin-1 Deficiency Induces Spontaneous Endothelial-to-Mesenchymal Transition in Murine Pulmonary Endothelial Cells in Vitro

Rationale: Endothelial-to-mesenchymal transition (EndMT) is a dynamic biological process involved in pathological vascular remodelling. However, the molecular mechanisms that govern this transition remain largely unknown, including the contribution of long non-coding RNAs (lncRNAs). Objective: To investigate the role of lncRNAs in EndMT and their relevance to vascular remodelling. Methods and Results: To study EndMT in vitro, primary endothelial cells (EC) were treated with transforming growth factor-β2 and interleukin-1β. Single-cell and bulk RNA-sequencing were performed to investigate the transcriptional architecture of EndMT and identify regulated lncRNAs. The functional contribution of seven lncRNAs during EndMT was investigated based on a DsiRNA screening assay. The loss of lncRNA MIR503HG was identified as a common signature across multiple human EC types undergoing EndMT in vitro. MIR503HG depletion induced a spontaneous EndMT phenotype, while its overexpression repressed hallmark EndMT changes, regulating 29% of its transcriptome signature. Importantly, the phenotypic changes induced by MIR503HG were independent of miR-424 and miR-503, which overlap the lncRNA locus. The pathological relevance of MIR503HG down-regulation was confirmed in vivo using Sugen/Hypoxia (SuHx)-induced pulmonary hypertension (PH) in mouse, as well as in human clinical samples, in lung sections and blood outgrowth endothelial cells (BOECs) from pulmonary arterial hypertension (PAH) patients. Overexpression of human MIR503HG in SuHx mice led to reduced mesenchymal marker expression, suggesting MIR503HG therapeutic potential. We also revealed that MIR503HG interacts with the Polypyrimidine Tract Binding Protein 1 (PTB1) and regulates its protein level. PTBP1 regulation of EndMT markers suggests that the role of MIR503HG in EndMT might be mediated in part by PTBP1. Conclusions: This study reports a novel lncRNA transcriptional profile associated with EndMT and reveals the crucial role of the loss of MIR503HG in EndMT and its relevance to pulmonary hypertension.

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Pyrogallol-Phloroglucinol-6 6-Bieckol on Attenuates High-Fat Diet-Induced Hypertension by Modulating Endothelial-to-Mesenchymal Transition in the Aorta of Mice

Oxidative Medicine and Cellular Longevity ◽

10.1155/2021/8869085 ◽

2021 ◽

Vol 2021 ◽

pp. 1-12

Author(s):

Myeongjoo Son ◽

Seyeon Oh ◽

Ji Tae Jang ◽

Kuk Hui Son ◽

Kyunghee Byun

Keyword(s):

Endothelial Cells ◽

High Fat Diet ◽

Oxidized Ldl ◽

Intima Media Thickness ◽

Mesenchymal Cell ◽

High Fat ◽

Mesenchymal Transition ◽

Induced Hypertension ◽

Endothelial To Mesenchymal Transition

Endothelial-to-mesenchymal transition (EndMT), which is involved in the development of various cardiovascular diseases, is induced by dyslipidemia or obesity. In dyslipidemia, the increased levels of oxidized low-density lipoproteins (oxLDL) upregulated the lectin-type oxidized LDL receptor 1 (Lox-1), which then upregulated the down signaling pathways of PKC-α/MMPs/TGF-β/SMAD2 or 3 and increased the EndMT. In this study, we investigated the effect of pyrogallol-phloroglucinol-6,6-bieckol (PPB), which is a compound of Ecklonia cava (E. cava), on decreased blood pressure (BP) by attenuating the EndMT in a high-fat diet- (HFD-) fed animal model. We also investigated PPB’s attenuation effect on EndMT in oxLDL-treated mouse endothelial cells as an in vitro model. The results indicated that, in the aorta or endothelial cells of mice, the HFD or oxLDL treatment significantly increased the expression of Lox-1/PKC-α/MMP9/TGF-β/SMAD2/SMAD3. The PPB treatment significantly decreased its expression. In contrast, the HFD or oxLDL treatment significantly decreased the expression of the EC markers (PECAM-1 and vWF) while the PPB treatment significantly increased them. Moreover, the HFD or oxLDL treatment significantly increased the expression of the mesenchymal cell markers (α-SMA and vimentin) while PPB treatment significantly decreased them. PPB decreased the intima-media thickness and extracellular matrix amount of the aorta and attenuated the BP, which was increased by the HFD. In conclusion, PPB attenuated the upregulation of Lox-1/PKC-α/MMP9/TGF-β/SMAD2 and 3 and restored the EndMT in HFD-fed animals. Moreover, PPB showed a restoring effect on HFD-induced hypertension.

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Trajectory modeling of endothelial-to-mesenchymal transition reveals galectin-3 as a mediator in pulmonary fibrosis

Cell Death and Disease ◽

10.1038/s41419-021-03603-0 ◽

2021 ◽

Vol 12 (4) ◽

Author(s):

Wangyue Jia ◽

Zhaoyan Wang ◽

Ceshu Gao ◽

Jian Wu ◽

Qiong Wu

Keyword(s):

Endothelial Cells ◽

Pulmonary Fibrosis ◽

Vascular Remodeling ◽

Mesenchymal Transition ◽

Upstream Regulator ◽

Galectin 3 ◽

Endothelial To Mesenchymal Transition ◽

Highly Correlated ◽

Stimulation Of

AbstractThe endothelial-to-mesenchymal transition (EndMT) is an important source of fibrotic cells in idiopathic pulmonary fibrosis (IPF). However, how endothelial cells (ECs) are activated and how EndMT impact IPF remain largely elusive. Here, we use unsupervised pseudotemporal analysis to recognize the heterogeneity of ECs and reconstruct EndMT trajectory of bleomycin (BLM)-treated Tie2creER/+;Rosa26tdTomato/+ IPF mice. Genes like C3ar1 and Lgals3 (protein name galectin-3) are highly correlated with the transitional pseudotime, whose expression is gradually upregulated during the fate switch of ECs from quiescence to activation in fibrosis. Inhibition of galectin-3 via siRNA or protein antagonists in mice could alleviate the pathogenesis of IPF and the transition of ECs. With the stimulation of human pulmonary microvascular endothelial cells (HPMECs) by recombinant proteins and/or siRNAs for galectin-3 in vitro, β-catenin/GSK3β signaling and its upstream regulator AKT are perturbed, which indicates they mediate the EndMT progress. These results suggest that EndMT is essential to IPF process and provide potential therapeutic targets for vascular remodeling.

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Abstract MP02: Angiotensin II Mediates Microvascular Rarefaction In Vivo and Exacerbates Endothelial-To-Mesenchymal Transition In Vitro

Hypertension ◽

10.1161/hyp.66.suppl_1.mp02 ◽

2015 ◽

Vol 66 (suppl_1) ◽

Author(s):

Katrin Nather ◽

Mónica Flores-Muñoz ◽

Rhian M Touyz ◽

Christopher M Loughrey ◽

Stuart A Nicklin

Keyword(s):

Endothelial Cells ◽

Smooth Muscle ◽

Cardiac Fibrosis ◽

Smooth Muscle Actin ◽

Muscle Actin ◽

Mesenchymal Transition ◽

Endothelial To Mesenchymal Transition

Cardiac fibrosis accompanies numerous cardiovascular diseases (CVD) such as hypertension and myocardial infarction and increases myocardial stiffness leading to contractile dysfunction. Recently, endothelial-to-mesenchymal transition (EndMT) has been shown to contribute to myocardial fibrosis. EndMT describes a process by which endothelial cells transform into mesenchymal cells such as fibroblasts and has been implicated in many fibrotic diseases. Angiotensin II (AngII) plays a key role in myocardial fibrosis and has been associated with the activation of fibroblasts to myofibroblasts and an increase in myocardial collagen deposition. Here, we assessed the role of AngII in capillary loss and EndMT in vivo and in vitro . C57BL/6J mice were infused with H 2 O (control) or 24μg/kg/hr AngII for 4 weeks. Mice infused with AngII developed significant cardiac fibrosis characterised by the deposition of collagen I (2.5-fold vs. control; p<0.05) and III (1.9-fold vs. control; p<0.05). Capillary density was assessed by CD31 immunohistochemistry and revealed significant vascular rarefaction (control 2161±111 vs . AngII 838±132 capillaries/mm 2 ; p<0.05). To investigate whether AngII can induce EndMT in vitro , human coronary artery endothelial cells were stimulated with 10ng/mL TGFβ 1 alone or in combination with 1μM AngII for 10 days. AngII significantly enhanced TGFβ 1 -induced gene expression of α-smooth muscle actin (TGFβ 1 1.8-fold; TGFβ 1 ±AngII 4.3-fold vs . control; p<0.05) and collagen I (TGFβ 1 9.2-fold; TGFβ 1 +AngII 30.2-fold vs . control; p<0.05). Concomitantly, AngII significantly increased α-smooth muscle actin protein expression (TGFβ 1 3.9-fold; TGFβ 1 +AngII 23.6-fold vs . control; p<0.05) and significantly decreased CD31 expression (TGFβ 1 0.9-fold; TGFβ 1 +AngII 0.7-fold vs . control; p<0.05), suggesting AngII acts in concert with TGFβ 1 to enhance conversion of endothelial cells to myofibroblasts. Further studies investigating the underlying mechanism, including the role of the Smad pathway, are ongoing. These results demonstrate that AngII induces vascular rarefaction in vivo and potentiates TGFβ 1 -induced EndMT in vitro. Understanding the molecular basis for these observations may help to identify new therapeutic options in CVD.

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DDRE-06. REGULATION OF TUMOR MICROENVIRONMENT VIA ENDOTHELIAL-TO-MESENCHYMAL TRANSITION BLOCKADE IN GLIOBLASTOMA-ASSOCIATED BRAIN ENDOTHELIAL CELLS

Neuro-Oncology Advances ◽

10.1093/noajnl/vdab024.028 ◽

2020 ◽

Vol 3 (Supplement_1) ◽

pp. i7-i7

Author(s):

Nagore I Marin’ Ramos ◽

Niyati Jhaveri ◽

Thu Zan Thein ◽

Thomas C Chen

Keyword(s):

Endothelial Cells ◽

Tumor Microenvironment ◽

Tumor Recurrence ◽

Brain Endothelial Cells ◽

Clinical Value ◽

Mesenchymal Transition ◽

Novel Treatments ◽

Endothelial To Mesenchymal Transition

Abstract Glioblastoma multiforme (GBM) is a malignant brain tumor noted for its extensive vascularity, aggressiveness, and highly invasive nature. Glioma stem cells (GSC) are a subpopulation of cells resistant to treatments and considered responsible for tumor recurrence. GSC are found in the vascular niches of the tumors, where endothelial cells (EC) secrete factors that stimulate GSC self-renewal. There are several studies regarding the effects of the vasculature on CSC and tumorigenesis, but little is known about how GSC affects the vasculature. Resistance to therapies and tumor recurrence greatly rely on the pro-angiogenic nature and aberrant vasculature of GBM. The endothelial-to-mesenchymal transition (EndMT) supports the pro-angiogenic and invasive characteristics of GBM. Hence, blocking the EndMT would be a promising approach to inhibit tumor progression and recurrence. We have examined the dynamic cross-talk between GSC and EC during EndMT. We demonstrate that GSC induce EndMT in brain endothelial cells (BEC), through a collaboration between TGF-β and Notch pathways, nicotinamide N-methyltransferase upregulation and other key signaling routes. Elucidating the cells and molecular pathways responsible for this process represents a milestone in the understanding of the tumor microenvironment and will help develop novel treatments in glioma therapy. One promising treatment, developed by our research group, is the conjugate of temozolomide and perillyl alcohol (POH), NEO212. This drug blocks EndMT induction in BEC and reverts the mesenchymal phenotype of tumor-associated BEC (TuBEC), reducing the invasiveness and pro-angiogenic properties of GBM in vitro and in vivo. We are currently performing Investigational New Drug (IND)-enabling studies, and we foresee that NEO212 will be of great clinical value for the treatment of GBM.

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Biological Effects of XyloCore, a Glucose Sparing PD Solution, on Mesothelial Cells: Focus on Mesothelial-Mesenchymal Transition, Inflammation and Angiogenesis

Nutrients ◽

10.3390/nu13072282 ◽

2021 ◽

Vol 13 (7) ◽

pp. 2282

Author(s):

Valentina Masola ◽

Mario Bonomini ◽

Maurizio Onisto ◽

Pietro Manuel Ferraro ◽

Arduino Arduini ◽

...

Keyword(s):

Endothelial Cell ◽

Cell Viability ◽

Biological Effects ◽

Membrane Integrity ◽

Glucose Content ◽

Mesenchymal Transition ◽

Low Glucose ◽

Endothelial To Mesenchymal Transition ◽

Osmotic Agents

Glucose-based solutions remain the most used osmotic agents in peritoneal dialysis (PD), but unavoidably they contribute to the loss of peritoneal filtration capacity. Here, we evaluated at a molecular level the effects of XyloCore, a new PD solution with a low glucose content, in mesothelial and endothelial cells. Cell viability, integrity of mesothelial and endothelial cell membrane, activation of mesothelial and endothelial to mesenchymal transition programs, inflammation, and angiogenesis were evaluated by several techniques. Results showed that XyloCore preserves mesothelial and endothelial cell viability and membrane integrity. Moreover XyloCore, unlike glucose-based solutions, does not exert pro-fibrotic, -inflammatory, and -angiogenic effects. Overall, the in vitro evidence suggests that XyloCore could represent a potential biocompatible solution promising better outcomes in clinical practice.

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Endothelial-to-mesenchymal transition in lipopolysaccharide-induced acute lung injury drives a progenitor cell-like phenotype

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00074.2016 ◽

2016 ◽

Vol 310 (11) ◽

pp. L1185-L1198 ◽

Cited By ~ 12

Author(s):

Toshio Suzuki ◽

Yuji Tada ◽

Rintaro Nishimura ◽

Takeshi Kawasaki ◽

Ayumi Sekine ◽

...

Keyword(s):

Bone Marrow ◽

Endothelial Cells ◽

Acute Lung Injury ◽

Lung Injury ◽

Vascular Endothelial Cells ◽

Repair Process ◽

Vascular Endothelial ◽

Mesenchymal Transition ◽

Oxidase Inhibition ◽

Endothelial To Mesenchymal Transition

Pulmonary vascular endothelial function may be impaired by oxidative stress in endotoxemia-derived acute lung injury. Growing evidence suggests that endothelial-to-mesenchymal transition (EndMT) could play a pivotal role in various respiratory diseases; however, it remains unclear whether EndMT participates in the injury/repair process of septic acute lung injury. Here, we analyzed lipopolysaccharide (LPS)-treated mice whose total number of pulmonary vascular endothelial cells (PVECs) transiently decreased after production of reactive oxygen species (ROS), while the population of EndMT-PVECs significantly increased. NAD(P)H oxidase inhibition suppressed EndMT of PVECs. Most EndMT-PVECs derived from tissue-resident cells, not from bone marrow, as assessed by mice with chimeric bone marrow. Bromodeoxyuridine-incorporation assays revealed higher proliferation of capillary EndMT-PVECs. In addition, EndMT-PVECs strongly expressed c- kit and CD133. LPS loading to human lung microvascular endothelial cells (HMVEC-Ls) induced reversible EndMT, as evidenced by phenotypic recovery observed after removal of LPS. LPS-induced EndMT-HMVEC-Ls had increased vasculogenic ability, aldehyde dehydrogenase activity, and expression of drug resistance genes, which are also fundamental properties of progenitor cells. Taken together, our results demonstrate that LPS induces EndMT of tissue-resident PVECs during the early phase of acute lung injury, partly mediated by ROS, contributing to increased proliferation of PVECs.

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LncRNA AERRIE Is Required for Sulfatase 1 Expression, but Not for Endothelial-to-Mesenchymal Transition

International Journal of Molecular Sciences ◽

10.3390/ijms22158088 ◽

2021 ◽

Vol 22 (15) ◽

pp. 8088

Author(s):

Tan Phát Pham ◽

Anke S. van Bergen ◽

Veerle Kremer ◽

Simone F. Glaser ◽

Stefanie Dimmeler ◽

...

Keyword(s):

Endothelial Cells ◽

Pulmonary Hypertension ◽

Transcription Factors ◽

Mesenchymal Phenotype ◽

Mesenchymal Transition ◽

Inflammatory Signals ◽

Pathological Conditions ◽

Non Coding Rna ◽

Endothelial To Mesenchymal Transition ◽

Long Non Coding Rna

Endothelial cells can acquire a mesenchymal phenotype through a process called Endothelial-to-Mesenchymal transition (EndMT). This event is found in embryonic development, but also in pathological conditions. Blood vessels lose their ability to maintain vascular homeostasis and ultimately develop atherosclerosis, pulmonary hypertension, or fibrosis. An increase in inflammatory signals causes an upregulation of EndMT transcription factors, mesenchymal markers, and a decrease in endothelial markers. In our study, we show that the induction of EndMT results in an increase in long non-coding RNA AERRIE expression. JMJD2B, a known EndMT regulator, induces AERRIE and subsequently SULF1. Silencing of AERRIE shows a partial regulation of SULF1 but showed no effect on the endothelial and mesenchymal markers. Additionally, the overexpression of AERRIE results in no significant changes in EndMT markers, suggesting that AERRIE is marginally regulating mesenchymal markers and transcription factors. This study identifies AERRIE as a novel factor in EndMT, but its mechanism of action still needs to be elucidated.

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Endothelial-to-Mesenchymal Transition in Human Adipose Tissue Vasculature Alters the Particulate Secretome and Induces Endothelial Dysfunction

Arteriosclerosis Thrombosis and Vascular Biology ◽

10.1161/atvbaha.119.312826 ◽

2019 ◽

Vol 39 (10) ◽

pp. 2168-2191 ◽

Cited By ~ 3

Author(s):

Bronson A. Haynes ◽

Li Fang Yang ◽

Ryan W. Huyck ◽

Eric J. Lehrer ◽

Joshua M. Turner ◽

...

Keyword(s):

Adipose Tissue ◽

Endothelial Dysfunction ◽

Smooth Muscle Actin ◽

Phenotypic Change ◽

Alpha Smooth Muscle Actin ◽

Mesenchymal Transition ◽

Molecular Features ◽

Endothelial To Mesenchymal Transition

Objective: Endothelial cells (EC) in obese adipose tissue (AT) are exposed to a chronic proinflammatory environment that may induce a mesenchymal-like phenotype and altered function. The objective of this study was to establish whether endothelial-to-mesenchymal transition (EndoMT) is present in human AT in obesity and to investigate the effect of such transition on endothelial function and the endothelial particulate secretome represented by extracellular vesicles (EV). Approach and Results: We identified EndoMT in obese human AT depots by immunohistochemical co-localization of CD31 or vWF and α-SMA (alpha-smooth muscle actin). We showed that AT EC exposed in vitro to TGF-β (tumor growth factor-β), TNF-α (tumor necrosis factor-α), and IFN-γ (interferon-γ) undergo EndoMT with progressive loss of endothelial markers. The phenotypic change results in failure to maintain a tight barrier in culture, increased migration, and reduced angiogenesis. EndoMT also reduced mitochondrial oxidative phosphorylation and glycolytic capacity of EC. EVs produced by EC that underwent EndoMT dramatically reduced angiogenic capacity of the recipient naïve ECs without affecting their migration or proliferation. Proteomic analysis of EV produced by EC in the proinflammatory conditions showed presence of several pro-inflammatory and immune proteins along with an enrichment in angiogenic receptors. Conclusions: We demonstrated the presence of EndoMT in human AT in obesity. EndoMT in vitro resulted in production of EV that transferred some of the functional and metabolic features to recipient naïve EC. This result suggests that functional and molecular features of EC that underwent EndoMT in vivo can be disseminated in a paracrine or endocrine fashion and may induce endothelial dysfunction in distant vascular beds.

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