Lymphatic endothelial cell calcium pulses are sensitive to spatial gradients in wall shear stress.
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ABSTRACT:
SUBMITTER: Surya VN
PROVIDER: S-EPMC6589782 | biostudies-literature | 2019 Mar
REPOSITORIES: biostudies-literature
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Project description:One-way valves in the lymphatic system form from lymphatic endothelial cells (LECs) during embryonic development and are required for efficient tissue drainage. Although fluid flow is thought to guide both valve formation and maintenance, how this occurs at a mechanistic level remains incompletely understood. We built microfluidic devices that reproduce critical aspects of the fluid flow patterns found at sites of valvulogenesis. Using these devices, we observed that LECs replicated aspects of the early steps in valvulogenesis: cells oriented perpendicular to flow in the region of maximum wall shear stress (WSS) and exhibited enhanced nuclear localization of FOXC2, a transcription factor required for valvulogenesis. Further experiments revealed that the cell surface protein E-selectin was required for both of these responses. Our observations suggest that spatial gradients in WSS help to demarcate the locations of valve formation, and implicate E-selectin as a component of a mechanosensory process for detecting WSS gradients.
| S-EPMC7005002 | biostudies-literature
Project description:Intracranial aneurysms tend to form at bifurcation apices, where flow impingement causes high frictional force (or wall shear stress, WSS) and flow acceleration and deceleration that create positive and negative streamwise gradients in WSS (WSSG), respectively. In vivo, intracranial aneurysms initiate under high WSS and positive WSSG. Little is known about the responses of endothelial cells (ECs) to either positive or negative WSSG under high WSS conditions. We used cDNA microarrays to profile EC gene expression exposed to positive WSSG vs. negative WSSG for 24 hours in a flow chamber with converging and diverging channels, respectively. WSS varied between 3.5 and 28.4 Pa in each gradient channel. GO and biological pathway analysis indicated that positive WSSG favored proliferation, apoptosis, and extracellular matrix processing while decreasing expression of pro-inflammatory genes. A subset of characteristic genes was validated using qPCR: Genes for ADAMTS1, CKAP2 and NCEH1 had higher expression under positive WSSG compared to negative WSSG while TAGLN, THBS1, VCAM1, CCL2, and CSF2 had lower expression. To determine if these patterns of expression are also exhibited in vivo, we tested whether the extracellular matrix related protein ADAMTS1 and proliferation were modulated by positive WSSG during intracranial aneurysm initiation. An aneurysm was induced at the basiliar terminus in rabbits by bilateral carotid ligation. WSSG at the bifurcation was determined by computational fluid dynamic simulations from 3D angiography and mapped on immunofluorescence staining for ADAMTS1 and the proliferation marker, Ki-67. Endothelial ADAMTS1 protein and Ki-67 were significantly higher in regions with positive WSSG compared to adjacent sites where WSSG was negative. Our results indicate that WSSG can elicit distinct gene expression profiles in ECs. Increased matrix processing and high levels of proliferation under positive WSSG could contribute to intracranial aneurysm initiation by causing transient gaps in the endothelium or disrupting EC signals to smooth muscle cells. Time-matched bovine aortic endothelial cells were exposed to positive wall shear stress gradient, negative wall shear stress gradient, and two no gradient samples: uniform WSS of 3.5 Pa and high WSS of 28.4 Pa for 24 hrs in an in vitro flow loop system. RNA was extracted and hybridized on Affymetrix microarrays. There were 12 samples in total, four flow conditions with three replicates each.
2013-05-01 | E-GEOD-37127 | biostudies-arrayexpress
Project description:Intracranial aneurysms tend to form at bifurcation apices, where flow impingement causes high frictional force (or wall shear stress, WSS) and flow acceleration and deceleration that create positive and negative streamwise gradients in WSS (WSSG), respectively. In vivo, intracranial aneurysms initiate under high WSS and positive WSSG. Little is known about the responses of endothelial cells (ECs) to either positive or negative WSSG under high WSS conditions. We used cDNA microarrays to profile EC gene expression exposed to positive WSSG vs. negative WSSG for 24 hours in a flow chamber with converging and diverging channels, respectively. WSS varied between 3.5 and 28.4 Pa in each gradient channel. GO and biological pathway analysis indicated that positive WSSG favored proliferation, apoptosis, and extracellular matrix processing while decreasing expression of pro-inflammatory genes. A subset of characteristic genes was validated using qPCR: Genes for ADAMTS1, CKAP2 and NCEH1 had higher expression under positive WSSG compared to negative WSSG while TAGLN, THBS1, VCAM1, CCL2, and CSF2 had lower expression. To determine if these patterns of expression are also exhibited in vivo, we tested whether the extracellular matrix related protein ADAMTS1 and proliferation were modulated by positive WSSG during intracranial aneurysm initiation. An aneurysm was induced at the basiliar terminus in rabbits by bilateral carotid ligation. WSSG at the bifurcation was determined by computational fluid dynamic simulations from 3D angiography and mapped on immunofluorescence staining for ADAMTS1 and the proliferation marker, Ki-67. Endothelial ADAMTS1 protein and Ki-67 were significantly higher in regions with positive WSSG compared to adjacent sites where WSSG was negative. Our results indicate that WSSG can elicit distinct gene expression profiles in ECs. Increased matrix processing and high levels of proliferation under positive WSSG could contribute to intracranial aneurysm initiation by causing transient gaps in the endothelium or disrupting EC signals to smooth muscle cells.
2013-05-01 | GSE37127 | GEO
Project description:Flow impingement at arterial bifurcations causes high frictional force [or wall shear stress (WSS)], and flow acceleration and deceleration in the branches create positive and negative streamwise gradients in WSS (WSSG), respectively. Intracranial aneurysms tend to form in regions with high WSS and positive WSSG. However, little is known about the responses of endothelial cells (ECs) to either positive or negative WSSG under high WSS conditions. We used cDNA microarrays to profile gene expression in cultured ECs exposed to positive or negative WSSG for 24 h in a flow chamber where WSS varied between 3.5 and 28.4 Pa. Gene ontology and biological pathway analysis indicated that positive WSSG favored proliferation, apoptosis, and extracellular matrix processing while decreasing expression of proinflammatory genes. To determine if similar responses occur in vivo, we examined EC proliferation and expression of the matrix metalloproteinase ADAMTS1 under high WSS and WSSG created at the basilar terminus of rabbits after bilateral carotid ligation. Precise hemodynamic conditions were determined by computational fluid dynamic simulations from three-dimensional angiography and mapped on immunofluorescence staining for the proliferation marker Ki-67 and ADAMTS1. Both proliferation and ADAMTS1 were significantly higher in ECs under positive WSSG than in adjacent regions of negative WSSG. Our results indicate that WSSG elicits distinct EC gene expression profiles and particular biological pathways including increased cell proliferation and matrix processing. Such EC responses may be important in understanding the mechanisms of intracranial aneurysm initiation at regions of high WSS and positive WSSG.
| S-EPMC3798684 | biostudies-literature
Project description:Local inflammation of the endothelium is associated with a plethora of cardiovascular diseases. Vascular-targeted carriers (VTCs) have been advocated to provide focal effective therapeutics to these disease sites. Here, we examine the design of functionalized nanoparticles (NPs) as VTCs that can specifically localize at an inflamed vessel wall under pathological levels of high shear stress, associated for example with clinical (or in vivo) conditions of vascular narrowing and arteriogenesis. To test this, carboxylated fluorescent 200?nm polystyrene particles were functionalized with ligands to activated endothelium, that is, an E-selectin binding peptide (Esbp), an anti ICAM-1 antibody, or using a combination of both. The functionalized NPs were investigated in vitro using microfluidic models lined with inflamed (TNF-? stimulated) and control endothelial cells (EC). Specifically, their adhesion was monitored under different relevant wall shear stresses (i.e., 40-300?dyne/cm2) via real-time confocal microscopy. Experiments reveal a significantly higher specific adhesion of the examined functionalized NPs to activated EC for the window of examined wall shear stresses. Moreover, particle adhesion correlated with the surface coating density whereby under high surface coating (i.e., ~10,000 molecule/particle), shear-dependent particle adhesion increased significantly. Altogether, our results show that functionalized NPs can be designed to target inflamed endothelial cells under high shear stress. Such VTCs underscore the potential for attractive avenues in targeting drugs to vasoconstriction and arteriogenesis sites.
| S-EPMC7237145 | biostudies-literature
Project description:The alignment of arterial endothelial cells (ECs) with the mean wall shear stress (WSS) vector is the prototypical example of their responsiveness to flow. However, evidence for this behaviour rests on experiments where many WSS metrics had the same orientation or where they were incompletely characterized. In the present study, we tested the phenomenon more rigorously. Aortic ECs were cultured in cylindrical wells on the platform of an orbital shaker. In this system, orientation would differ depending on the WSS metric to which the cells aligned. Variation in flow features and hence in possible orientations was further enhanced by altering the viscosity of the medium. Orientation of endothelial nuclei was compared with WSS characteristics obtained by computational fluid dynamics. At low mean WSS magnitudes, ECs aligned with the modal WSS vector, while at high mean WSS magnitudes they aligned so as to minimize the shear acting across their long axis (transverse WSS). Their failure to align with the mean WSS vector implies that other aspects of endothelial behaviour attributed to this metric require re-examination. The evolution of a mechanism for minimizing transverse WSS is consistent with it having detrimental effects on the cells and with its putative role in atherogenesis.
| S-EPMC7879765 | biostudies-literature
Project description:A striking feature of atherosclerosis is its patchy distribution within the vascular system; certain arteries and certain locations within each artery are preferentially affected. Identifying the local risk factors underlying this phenomenon may lead to new therapeutic strategies. The large variation in lesion prevalence in areas of curvature and branching has motivated a search for haemodynamic triggers, particular those related to wall shear stress (WSS). The fact that lesions are rich in blood-derived lipids has motivated studies of local endothelial permeability. However, the location of lesions, the underlying haemodynamic triggers, the role of permeability, the routes by which lipids cross the endothelium, and the mechanisms by which WSS affects permeability have all been areas of controversy. This review presents evidence for and against the current consensus that lesions are triggered by low and/or oscillatory WSS and that this type of shear profile leads to elevated entry of low density lipoprotein (LDL) into the wall via widened intercellular junctions; it also evaluates more recent evidence that lesion location changes with age, that multidirectional shear stress plays a key role, that LDL dominantly crosses the endothelium by transcytosis, and that the link between flow and permeability results from hitherto unrecognised shear-sensitive mediators.
| S-EPMC8948426 | biostudies-literature
Project description:BACKGROUND:To explore the possibility of creating three-dimensional (3D) estimation models for patient-specific wall thickness (WT) maps using patient-specific and cohort-averaged WT, wall shear stress (WSS), and vessel diameter maps in asymptomatic atherosclerotic carotid bifurcations. METHODS:Twenty subjects (aged 75?±?6 years [mean?±?standard deviation], eight women) underwent a 1.5-T MRI examination. Non-gated 3D phase-contrast gradient-echo images and proton density-weighted echo-planar images were retrospectively assessed for WSS, diameter estimation, and WT measurements. Spearman's ? and scatter plots were used to determine correlations between individual WT, WSS, and diameter maps. A bootstrapping technique was used to determine correlations between 3D cohort-averaged WT, WSS, and diameter maps. Linear regression between the cohort-averaged WT, WSS, and diameter maps was used to predict individual 3D WT. RESULTS:Spearman's ? averaged over the subjects was -?0.24?±?0.18 (p?<?0.001) and 0.07?±?0.28 (p?=?0.413) for WT versus WSS and for WT versus diameter relations, respectively. Cohort-averaged ?, averaged over 1000 bootstraps, was -?0.56 (95% confidence interval [-?0.74,-?0.38]) for WT versus WSS and 0.23 (95% confidence interval [-?0.06, 0.52]) for WT versus diameter. Scatter plots did not reveal relationships between individual WT and WSS or between WT and diameter data. Linear relationships between these parameters became apparent after averaging over the cohort. Spearman's ? between the original and predicted WT maps was 0.21?±?0.22 (p?<?0.001). CONCLUSIONS:With a combination of bootstrapping and cohort-averaging methods, 3D WT maps can be predicted from the individual 3D WSS and diameter maps. The methodology may help to elucidate pathological processes involving WSS in carotid atherosclerosis.
| S-EPMC6177500 | biostudies-literature
Project description:BackgroundLymphatic endothelial cells form an important barrier necessary for normal lymph formation and propulsion. However, little is known about how physical forces within lymphatic vessels affect endothelial barrier function. The purpose of this study was to characterize how laminar flow affects lymphatic endothelial barrier function and to test whether endothelial cells respond to flow changes by activating the intracellular actin cytoskeleton to enhance barrier function.Methods and resultsCultured adult human dermal microlymphatic endothelial cells (HMLEC-d) were grown on small gold electrodes arranged within a flow channel, and transendothelial electrical resistance (TER), an index of barrier function, was determined. Laminar flow was applied to the cells at a baseline shear stress of 0.5 dynes/cm(2), and was increased to 2.5, 5.0, or 9.0 dynes/cm(2), causing a magnitude-dependent increase in barrier function that was reversed 30 min later when the shear stress was returned to baseline. This response was abolished by blockade of actin dynamics with 10 microM phalloidin, and significantly inhibited by blockade of Rac1 activity with 50 microM NSC23766. Blockade of protein kinase A (10 microM H-89) did not inhibit the response. Mathematical modeling based on our impedance data showed that the flow-induced changes in TER were primarily due to altered current flow between cells and not beneath cells.ConclusionsThese results suggest that lymphatic endothelial cells dynamically alter their morphology and barrier function in response to changes in shear stress by a mechanism dependent upon Rac1-mediated actin dynamics.
| S-EPMC2883493 | biostudies-literature
Project description:Objective: The vulnerability of atherosclerotic plaques is among the leading cause of ischemic stroke. High wall shear stress (WSS) promotes the instability of atherosclerotic plaques by directly imparting mechanical stimuli, but the specific mechanisms remain unclear. We speculate that modulation of mechanosensitive genes may play a vital role in accelerating the development of plaques. The purpose of this study was to find mechanosensitive genes in vascular endothelial cells (ECs) through combining microarray data with bioinformatics technology and further explore the underlying dynamics-related mechanisms that cause the progression and destabilization of atherosclerotic plaques. Methods: Microarray data sets for human vascular ECs under high and normal WSS were retrieved from the Gene Expression Omnibus (GEO) database. Differentially expressed genes (DEGs) were identified through the R language. The performance of enrichment analysis and protein-protein interaction (PPI) network presented the biological function and signaling pathways of the DEGs. Hub genes were identified based on the PPI network and validated by GEO data sets. Predicted transcription factor (TF) genes and miRNAs interaction with potential mechanosensitive genes were identified by NetworkAnalyst. Results: A total of 260 DEGs, 121 upregulated and 139 downregulated genes, were screened between high and normal WSS from GSE23289. A total of 10 hub genes and four cluster modules were filtered out based on the PPI network. The enrichment analysis showed that the biological functions of the hub genes were mainly involved in responses to unfolded protein and topologically incorrect protein, and t to endoplasmic reticulum stress. The significant pathways associated with the hub genes were those of protein processing in the endoplasmic reticulum, antigen processing, and presentation. Three out of the 10 hub genes, namely, activated transcription factor 3 (ATF3), heat shock protein family A (Hsp70) member 6 (HSPA6), and dual specificity phosphatase 1 (DUSP1, also known as CL100, HVH1, MKP-1, PTPN10), were verified in GSE13712. The expression of DUSP1 was higher in the senescent cell under high WSS than that of the young cell. The TF-miRNA-mechanosensitive gene coregulatory network was constructed. Conclusion: In this work, we identified three hub genes, ATF3, HSPA6, and DUSP1, as the potential mechanosensitive genes in the human blood vessels. DUSP1 was confirmed to be associated with the senescence of vascular ECs. Therefore, these three mechanosensitive genes may have emerged as potential novel targets for the prediction and prevention of ischemic stroke. Furthermore, the TF-miRNA-mechanosensitive genes coregulatory network reveals an underlying regulatory mechanism and the pathways to control disease progression.
| S-EPMC8787366 | biostudies-literature