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Project description:Biomechanical cues dynamically control major cellular processes but whether genetic variants actively participate in the mechano-sensing mechanisms remain unexplored. Vascular homeostasis is tightly regulated by hemodynamic forces. Exposure to disturbed blood flow at arterial sites of branching and bifurcation causes constant activation of vascular endothelium contributing to the development of atherosclerosis, the major cause of coronary artery disease (CAD). Conversely, unidirectional flow promotes the anti-inflammatory and anti-permeable endothelial phenotype resistant to atherogenesis. Genome-wide association studies (GWAS) have identified chromosome 1p32.2 as one of the loci most strongly associated with CAD susceptibility; however, the causal mechanism related to this CAD locus remain unknown. PhosphoLipid PhosPhatase 3 (PLPP3) is located at 1p32.2 and encodes a phosphatase that suppresses endothelial inflammation and promotes monolayer integrity by hydrolyzing lysophosphatidic acid. Our previous studies demonstrated that PLPP3 is significantly reduced in vascular endothelium exposed to disturbed flow while unidirectional flow significantly increases endothelial PLPP3 expression in vitro and in vivo. In addition, CAD protective allele at 1p32.2 locus is associated with increased PLPP3 in an endothelium-specific manner, shown by expression quantitative trait locus (eQTL). Using Assay for Transposase-Accessible Chromatin using Sequencing (ATAC-Seq), H3K27ac ChIP-Seq, H3K4me2 ChIP-Seq, and luciferase assays, here we identified a mechano-sensitive endothelial enhancer in PLPP3 intron 5 that is dynamically activated by unidirectional flow. Deletion of this enhancer by CRISPR/Cas9-based genome editing causatively reduces endothelial PLPP3 expression and promotes endothelial activation, and moreover, impairs endothelial PLPP3 induction by unidirectional flow. Chromatin accessibility quantitative trait locus (caQTL) mapping, allelic imbalance assay, and luciferase assays further demonstrated that CAD protective allele at rs17114036 in the PLPP3 intron 5 confers an increased enhancer activity. ChIP-PCR and luciferase assays showed that CAD protective allele C at rs17114036 creates a binding site (CACC) for mechano-sensitive transcription factor KLF2, leading to increased enhancer activity under unidirectional flow. These results elucidate the contributory role of CAD genetic predisposition in critical endothelial mechano-transduction mechanisms and suggest that human genetic variants provide a previously unappreciated layer of regulatory control in cellular mechano-sensing mechanisms.
2018-11-19 | GSE112328 | GEO
Project description:Biomechanical cues dynamically control major cellular processes but whether genetic variants actively participate in the mechano-sensing mechanisms remain unexplored. Vascular homeostasis is tightly regulated by hemodynamic forces. Exposure to disturbed blood flow at arterial sites of branching and bifurcation causes constant activation of vascular endothelium contributing to the development of atherosclerosis, the major cause of coronary artery disease (CAD). Conversely, unidirectional flow promotes the anti-inflammatory and anti-permeable endothelial phenotype resistant to atherogenesis. Genome-wide association studies (GWAS) have identified chromosome 1p32.2 as one of the loci most strongly associated with CAD susceptibility; however, the causal mechanism related to this CAD locus remain unknown. PhosphoLipid PhosPhatase 3 (PLPP3) is located at 1p32.2 and encodes a phosphatase that suppresses endothelial inflammation and promotes monolayer integrity by hydrolyzing lysophosphatidic acid. Our previous studies demonstrated that PLPP3 is significantly reduced in vascular endothelium exposed to disturbed flow while unidirectional flow significantly increases endothelial PLPP3 expression in vitro and in vivo. In addition, CAD protective allele at 1p32.2 locus is associated with increased PLPP3 in an endothelium-specific manner, shown by expression quantitative trait locus (eQTL). Using Assay for Transposase-Accessible Chromatin using Sequencing (ATAC-Seq), H3K27ac ChIP-Seq, H3K4me2 ChIP-Seq, and luciferase assays, here we identified a mechano-sensitive endothelial enhancer in PLPP3 intron 5 that is dynamically activated by unidirectional flow. Deletion of this enhancer by CRISPR/Cas9-based genome editing causatively reduces endothelial PLPP3 expression and promotes endothelial activation, and moreover, impairs endothelial PLPP3 induction by unidirectional flow. Chromatin accessibility quantitative trait locus (caQTL) mapping, allelic imbalance assay, and luciferase assays further demonstrated that CAD protective allele at rs17114036 in the PLPP3 intron 5 confers an increased enhancer activity. ChIP-PCR and luciferase assays showed that CAD protective allele C at rs17114036 creates a binding site (CACC) for mechano-sensitive transcription factor KLF2, leading to increased enhancer activity under unidirectional flow. These results elucidate the contributory role of CAD genetic predisposition in critical endothelial mechano-transduction mechanisms and suggest that human genetic variants provide a previously unappreciated layer of regulatory control in cellular mechano-sensing mechanisms.
2018-11-19 | GSE112336 | GEO
Project description:Differential epigenetic profiling of human aortic endothelial cells (HAEC) in response to shear stress forces
| PRJNA445703 | ENA
Project description:Differential epigenetic profiling of human aortic endothelial cells (HAEC) in response to shear stress forces using ATAC-Seq
| PRJNA445695 | ENA
Project description:Differential epigenetic profiling of human aortic endothelial cells (HAEC) in response to shear stress forces using ChIP-Seq of H3K27ac
| PRJNA445765 | ENA
Project description:Most acute coronary events occur in the upstream region of stenotic atherosclerotic plaques that experience laminar shear stress (LSS) elevated above normal physiological levels. Many studies have described the atheroprotective effect on endothelial behavior of normal physiological LSS (approximately 15 dynes/cm(2)) compared to static or oscillatory shear stress (OSS), but it is unknown whether the levels of elevated shear stress imposed by a stenotic plaque would preserve, enhance or reverse this effect. Therefore we used transcriptomics and related functional analyses to compare human endothelial cells exposed to laminar shear stress of 15 (LSS15-normal) or 75 dynes/cm(2) (LSS75-elevated). LSS75 upregulated expression of 145 and downregulated expression of 158 genes more than twofold relative to LSS15. Modulation of the metallothioneins (MT1-G, -M, -X) and NADPH oxidase subunits (NOX2, NOX4, NOX5, and p67phox) accompanied suppression of reactive oxygen species production at LSS75. Shear induced changes in dual specificity phosphatases (DUSPs 1, 5, 8, and 16 increasing and DUSPs 6 and 23 decreasing) were observed as well as reduced ERK1/2 but increased p38 MAP kinase phosphorylation. Amongst vasoactive substances, endothelin-1 expression decreased whereas vasoactive intestinal peptide (VIP) and prostacyclin expression increased, despite which intracellular cAMP levels were reduced. Promoter analysis by rVISTA identified a significant over representation of ATF and Nrf2 transcription factor binding sites in genes upregulated by LSS75 compared to LSS15. In summary, LSS75 induced a specific change in behavior, modifying gene expression, reducing ROS levels, altering MAP kinase signaling and reducing cAMP levels, opening multiple avenues for future study.
| S-EPMC3412226 | biostudies-literature
Project description:The goal of this study was to find longitudinal transcriptional response of human aortic endothelial cells (HAECs) to pulsatile shear (PS) and oscillatory shear (OS) and compare them with the responses in human umbilical vein endothelial cells (HUVECs) [GSE103672]. PS is associated with an atheroprotective endothelial phenotype, while OS is associated with an atheroprone endothelial phenotype. Using RNASeq method (single-ended 75-bp sequencing on Illumina Hi-seq 4000 instrument), we measured the transcriptional response at 3 time-points (1, 4, and 24 hrs) under PS and OS conditions. Measurements were also taken under static condition (ST, no flow) at t = 0 hr. Three replicates were used for each condition/time-point. Our results indicate that the responses of HAECs and HUVECs are qualitatively similar for endothelial-function relevant genes and several important pathways with a few exceptions, thus demonstrating that HUVECs can be used as a model to investigate the effects of shear on arterial ECs, with some reservations. Our findings show that HAECs exhibit an earlier response or faster kinetics as compared to HUVECs. The comparative analysis in this study offers new insights into the mechanisms of common and disparate stress responses across these two endothelial cell types.
2021-01-13 | GSE160611 | GEO
Project description:Endothelial cells are constantly exposed to fluid shear stresses that regulate vascular morphogenesis, homeostasis, and disease. The mechanical responses of endothelial cells to relatively high shear flow such as that characteristic of arterial circulation has been extensively studied. Much less is known about the responses of endothelial cells to slow shear flow such as that characteristic of venous circulation, early angiogenesis, atherosclerosis, intracranial aneurysm, or interstitial flow. Here we used a novel, to our knowledge, microfluidic technique to measure traction forces exerted by confluent vascular endothelial cell monolayers under slow shear flow. We found that cells respond to flow with rapid and pronounced increases in traction forces and cell-cell stresses. These responses are reversible in time and do not involve reorientation of the cell body. Traction maps reveal that local cell responses to slow shear flow are highly heterogeneous in magnitude and sign. Our findings unveil a low-flow regime in which endothelial cell mechanics is acutely responsive to shear stress.
| S-EPMC4624165 | biostudies-literature
Project description:Porcine endothelial cells were preconditioned by a basal level shear stress of 15 ± 15 dynes/cm2 at 1 Hz for 24 hours, and an acute increase in shear stress frequency (2 Hz) was then applied. The transcriptomics studies using microarray identified genes that were sensitive to the elevated shear frequency. keywords: adaptation, shear stress, frequency, microarray, gene expression Porcine endothelial cells were preconditioned by a basal level shear stress of 15 ± 15 dynes/cm2 at 1 Hz for 24 hours, and an acute increase in shear stress magnitude (2 Hz) was then applied. Gene expression at multiple time points was measured using microarray.
2011-12-01 | E-GEOD-26513 | biostudies-arrayexpress
Project description:Porcine endothelial cells were preconditioned by a basal level shear stress of 15 ± 15 dynes/cm2 at 1 Hz for 24 hours, and an acute increase in shear stress frequency (2 Hz) was then applied. The transcriptomics studies using microarray identified genes that were sensitive to the elevated shear frequency. keywords: adaptation, shear stress, frequency, microarray, gene expression
2011-12-01 | GSE26513 | GEO