Altered Shear Stress Research Articles

Similar Pathophysiological Mechanisms Between Osteoarthritis and Vascular Disease

Osteoarthritis (OA) is a prevalent, chronic joint disorder affecting millions of people worldwide, characterized by articular cartilage degradation, subchondral bone remodeling, synovial cytokine secretion, and osteophyte formation. OA primarily affects the hips, knees, hands, and spine. Patients with OA exhibit a higher prevalence of cardiovascular comorbidities and potentially important associations between OA and cardiovascular diseases have prompted investigations into potentially similar pathophysiological associations. This review explores the coexistence of atherosclerotic peripheral vascular disease (ASPVD) in OA patients, including evidence from a contemporary study suggesting associations between OA and arterial wall thickness and blood flow changes which are characteristic of early atherosclerosis, and which stimulate reactive pathology in endothelial cells. Observations from this study demonstrate elevated arterial flow volume and increased intima-media thickness in arteries ipsilateral to OA knees, suggesting a potential link between OA and arterial wall disease. We further explore the intricate relationship between the vascular system and skeletal health, highlighting bidirectional interactions among endothelial cells, inflammatory cells, and various bone cells. Mechanical endothelial cell dysfunction is discussed, emphasizing the impact of vessel wall material changes and endothelial cell responses to alterations in fluid shear stress. Inflammatory changes in OA and ASPVD are also explored, showcasing shared pathophysiological processes involving immune cell infiltration and pro-inflammatory cytokines. Additionally, the role of hypofibrinolysis in OA and ASPVD is discussed, highlighting similarities in elevations of the hypercoagulative and hypofibrinolytic factor, plasminogen activator inhibitor (PAI-1). The review suggests a provocative relationship among low-grade chronic inflammation, endothelial dysfunction, and hypofibrinolytic states in OA and ASPVD, warranting further investigation. In conclusion, this review provides an exploration of the possible associations between OA and ASPVD. While the ongoing study’s findings and other reports are observational, they suggest shared pathophysiological processes and emphasize the need for further research to elucidate additional potentially correlative linkages between these conditions. Understanding common molecular pathways may pave a way for targeted interventions that address both OA and ASPVD.

Computational analysis of cancer cell adhesion in curved vessels affected by wall shear stress for prediction of metastatic spreading.

Introduction: The dynamics of circulating tumor cells (CTCs) within blood vessels play a pivotal role in predicting metastatic spreading of cancer within the body. However, the limited understanding and method to quantitatively investigate the influence of vascular architecture on CTC dynamics hinders our ability to predict metastatic process effectively. To address this limitation, the present study was conducted to investigate the influence of blood vessel tortuosity on the behaviour of CTCs, focusing specifically on establishing methods and examining the role of shear stress in CTC-vessel wall interactions and its subsequent impact on metastasis. Methods: We computationally simulated CTC behaviour under various shear stress conditions induced by vessel tortuosity. Our computational model, based on the lattice Boltzmann method (LBM) and a coarse-grained spectrin-link membrane model, efficiently simulates blood plasma dynamics and CTC deformability. The model incorporates fluid-structure interactions and receptor-ligand interactions crucial for CTC adhesion using the immersed boundary method (IBM). Results: Our findings reveal that uniform shear stress in straight vessels leads to predictable CTC-vessel interactions, whereas in curved vessels, asymmetrical flow patterns and altered shear stress create distinct adhesion dynamics, potentially influencing CTC extravasation. Quantitative analysis shows a 25% decrease in the wall shear stress in low-shear regions and a 58.5% increase in the high-shear region. We observed high-shear regions in curved vessels to be potential sites for increased CTC adhesion and extravasation, facilitated by elevated endothelial expression of adhesion molecules. This phenomenon correlates with the increased number of adhesion bonds, which rises to approximately 40 in high-shear regions, compared to around 12 for straight vessels and approximately 5-6 in low-shear regions. The findings also indicate an optimal cellular stiffness necessary for successful CTC extravasation in curved vessels. Discussion: By the quantitative assessment of the risk of CTC extravasation as a function of vessel tortuosity, our study offers a novel tool for the prediction of metastasis risk to support the development of personalized therapeutic interventions based on individual vascular characteristics and tumor cell properties.

Mechanoresponsive ETS1 causes endothelial dysfunction and arterialization in varicose veins via NOTCH4/DLL4 signaling

Varicose veins are the most common venous disorder in humans and are characterized by hemodynamic instability due to valvular insufficiency and orthostatic lifestyle factors. It is unclear how changes in biomechanical signals cause aberrant remodeling of the vein wall. Our previous studies suggest that Notch signaling is implicated in varicose vein arterialization. In the arterial system, mechanoresponsive ETS1 is a transcriptional activator of the endothelial Notch, but its involvement in sensing disrupted venous flow and varicose vein formation has not been investigated. Here, we use human varicose veins and cultured human venous endothelial cells to show that disturbed venous shear stress activates ETS1-NOTCH4/DLL4 signaling. Notch components were highly expressed in the neointima, whereas ETS1 was upregulated in all histological layers of varicose veins. In vitro microfluidic flow-based studies demonstrate that even minute changes in venous flow patterns enhance ETS1-NOTCH4/DLL4 signaling. Uniform venous shear stress, albeit an inherently low-flow system, does not induce ETS1 and Notch proteins. ETS1 activation under altered flow was mediated primarily by MEK1/2 and, to a lesser extent, by MEK5 but was independent of p38 MAP kinase. Endothelial cell-specific ETS1 knockdown prevented disturbed flow-induced NOTCH4/DLL4 expression. TK216, an inhibitor of ETS-family, prevented the acquisition of arterial molecular identity and loss of endothelial integrity in cells exposed to the ensuing altered shear stress. We conclude that ETS1 senses blood flow disturbances and may promote venous remodeling by inducing endothelial dysfunction. Targeting ETS1 rather than downstream Notch proteins could be an effective and safe strategy to develop varicose vein therapies.

Sex-Related Differences in Vascular Function Among Older Adults with Obstructive Sleep Apnea

Obstructive sleep apnea (OSA) is a sleep disorder associated with hypertension and cardiovascular disease (CVD) risk. Endothelial dysfunction and altered shear stress are implicated in OSA-related CVD pathogenesis, however, their contributions to CVD risk in males vs females remains unclear. We obtained measures of resting blood pressure (BP), brachial artery flow-mediated dilation (FMD), brachial artery diameter, and shear stress in 71 older adults (F=30, M=41; age=66.9±7.7) diagnosed with mild, moderate, or severe OSA based on oxygen desaturation index (ODI) score. All were prospective participants in an ongoing clinical trial (NCT04932447). Group differences (mean ± SD) were assessed with two-way ANOVAs (sex X severity) and associations between ODI score, baseline diameter, relative FMD, shear stress, and BP were assessed with Pearson correlations. We hypothesized that a) regardless of sex, individuals with greater OSA severity exhibit lower relative FMD and mean shear stress, greater baseline diameter and higher BP, and b) for a given OSA severity, females have greater relative FMD and mean shear stress and lower baseline diameter and BP compared to males. In line with our hypotheses, we found greater mean shear stress in females compared to males (F=69.6±26.9 s−1, M=53.5±16.7 s−1, p=0.035) and among females, those with mild OSA had higher mean shear stress than those with severe OSA (mild=77.6±26.1 s−1, severe= 44.4±8.5 s−1, p=0.009). Contrary to expectations, mean shear stress did not differ as a function of OSA severity in male subjects, nor was there any difference in FMD or BP across OSA severity between males and females. In subjects with moderate OSA, FMD was greater in females than males (F=6.07±3.89%, M=3.07±2.39%, p=0.047). On average, baseline diameters were larger in adults with severe OSA compared to moderate OSA (moderate=3.66±0.59 mm, severe=4.33±0.61 mm, p=0.003), and females with mild (F=3.20±0.47 mm, M=4.73±0.50 mm, p<0.001) and moderate (F=3.31±0.42 mm, M=3.91±0.57 mm, p=0.007) OSA had smaller baseline diameters than their male counterparts. There were no differences in baseline diameter between males and females with severe OSA. We report a negative correlation between mean shear stress and BP (systolic BP, r=-0.48, p=0.011; diastolic BP, r=-0.55, p=0.002) and a positive correlation between ODI score and baseline diameter (r=0.54, p=0.003) in females but not males. Conversely, ODI score and relative FMD were positively correlated in male subjects (r=0.45, p=0.009), but not females. Overall, our findings highlight sex-specific vascular differences in older adults with OSA. Relative to males, females with mild OSA exhibit greater mean shear stress and lower baseline diameters, which may confer a cardiovascular advantage. In contrast, males and females with severe OSA exhibit similar vascular characteristics. Our findings provide new insights into the impact of sex and OSA severity on vascular function and CVD risk in older adults. This work is supported by NIH/NIA grant number: 1R01AG065346. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Analyzing sediment transport in dam-break-driven swash processes: Insights from laboratory experiments using a high-speed camera

Dam-break-driven experimental tests are conducted to explore the intricate dynamics of water and sediment movement during a swash process. Utilizing a combined approach of a high-speed camera and an acoustic doppler velocity profiler, detailed records of flow velocity and sediment movements are captured. This study offers novel insights regarding flow shear stress, layer thickness and velocity of sediment transport in the backwash process, as well as behaviors of individual sand particles. Linear relationships serve to depict alterations in bed shear stress over time. The changing rate of bed shear stress can be influenced by the factors such as the beach slope, sediment grain size, and location on the beach. Increasing layer thickness of sediment transport corresponds with increasing outer layer velocity, exhibiting a linear-exponential relationship. A tentative interpretation of the inflection point (of linear and exponential relationship) designates it as a division between sheet load and a conjunction of sheet load and suspended load. With the onset of a bore and under the influence of turbulence and bubbles, sediment predominantly moves upwards, leading to sediment pickup. However, during both the late uprush phase and the backwash phase, the primary direction of sediment movement reverses downwards, favoring sediment settling. Empirical equations are formulated to estimate the parameters on the bed shear stress and the sediment transport layer. This study enhances understanding of water and sediment movement in swash processes, supplying validation data for numerical models.

Patient-specific 3D in vitro modeling and fluid dynamic analysis of primary pulmonary vein stenosis.

Primary pulmonary vein stenosis (PVS) is a rare congenital heart disease that proves to be a clinical challenge due to the rapidly progressive disease course and high rates of treatment complications. PVS intervention is frequently faced with in-stent restenosis and persistent disease progression despite initial venous recanalization with balloon angioplasty or stenting. Alterations in wall shear stress (WSS) have been previously associated with neointimal hyperplasia and venous stenosis underlying PVS progression. Thus, the development of patient-specific three-dimensional (3D) in vitro models is needed to further investigate the biomechanical outcomes of endovascular and surgical interventions. In this study, deidentified computed tomography images from three patients were segmented to generate perfusable phantom models of pulmonary veins before and after catheterization. These 3D reconstructions were 3D printed using a clear resin ink and used in a benchtop experimental setup. Computational fluid dynamic (CFD) analysis was performed on models in silico utilizing Doppler echocardiography data to represent the in vivo flow conditions at the inlets. Particle image velocimetry was conducted using the benchtop perfusion setup to analyze WSS and velocity profiles and the results were compared with those predicted by the CFD model. Our findings indicated areas of undesirable alterations in WSS before and after catheterization, in comparison with the published baseline levels in the healthy in vivo tissues that may lead to regional disease progression. The established patient-specific 3D in vitro models and the developed in vitro-in silico platform demonstrate great promise to refine interventional approaches and mitigate complications in treating patients with primary PVS.

Utilization of an Artery-on-a-Chip to Unravel Novel Regulators and Therapeutic Targets in Vascular Diseases.

In this study, organ-on-chip technology is used to develop an in vitro model of medium-to-large size arteries, the artery-on-a-chip (AoC), with the objective to recapitulate the structure of the arterial wall and the relevant hemodynamic forces affecting luminal cells. AoCs exposed either to in vivo-like shear stress values or kept in static conditions are assessed to generate a panel of novel genes modulated by shear stress. Considering the crucial role played by shear stress alterations in carotid arteries affected by atherosclerosis (CAD) and abdominal aortic aneurysms (AAA) disease development/progression, a patient cohort of hemodynamically relevant specimens is utilized, consisting of diseased and non-diseased (internal control) vessel regions from the same patient. Genes activated by shear stress follow the same expression pattern in non-diseased segments of human vessels. Single cell RNA sequencing (scRNA-seq) enables to discriminate the unique cell subpopulations between non-diseased and diseased vessel portions, revealing an enrichment of flow activated genes in structural cells originating from non-diseased specimens. Furthermore, the AoC served as a platform for drug-testing. It reproduced the effects of a therapeutic agent (lenvatinib) previously used in preclinical AAA studies, therefore extending the understanding of its therapeutic effect through a multicellular structure.

Transcriptional Profiling of Human Endothelial Cells Unveils PIEZO1 and Mechanosensitive Gene Regulation by Prooxidant and Inflammatory Inputs.

PIEZO1 is a mechanosensitive cation channel implicated in shear stress-mediated endothelial-dependent vasorelaxation. Since altered shear stress patterns induce a pro-inflammatory endothelial environment, we analyzed transcriptional profiles of human endothelial cells to determine the effect of altered shear stress patterns and subsequent prooxidant and inflammatory conditions on PIEZO1 and mechanosensitive-related genes (MRG). In silico analyses were validated in vitro by assessing PIEZO1 transcript levels in both the umbilical artery (HUAEC) and vein (HUVEC) endothelium. Transcriptional profiling showed that PIEZO1 and some MRG associated with the inflammatory response were upregulated in response to high (15 dyn/cm2) and extremely high shear stress (30 dyn/cm2) in HUVEC. Changes in PIEZO1 and inflammatory MRG were paralleled by p65 but not KLF or YAP1 transcription factors. Similarly, PIEZO1 transcript levels were upregulated by TNF-alpha (TNF-α) in diverse endothelial cell types, and pre-treatment with agents that prevent p65 translocation to the nucleus abolished PIEZO1 induction. ChIP-seq analysis revealed that p65 bonded to the PIEZO1 promoter region, an effect increased by the stimulation with TNF-α. Altogether this data showed that NF-kappa B activation via p65 signaling regulates PIEZO1 expression, providing a new molecular link for prooxidant and inflammatory responses and mechanosensitive pathways in the endothelium.

Biomechanical determinants of endothelial permeability assessed in standard and modified hollow-fibre bioreactors.

Effects of mechanical stress on the permeability of vascular endothelium are important to normal physiology and in the development of atherosclerosis. Here we elucidate novel effects using commercially available and modified hollow-fibre bioreactors, in which endothelial cells form confluent monolayers lining plastic capillaries with porous walls, contained in a cartridge. The capillaries were perfused with a near-aortic waveform, and permeability was assessed by the movement of rhodamine-labelled albumin from the intracapillary to the extracapillary space. Permeability was increased by acute application of shear stress and decreased by chronic shear stress compared with a static control: this has previously been shown only for multidirectional flows. Increasing viscosity reduced permeability under both acute and chronic shear; since shear rate remained unchanged, these effects resulted from altered shear stress. Reducing pulsatility increased permeability, contrary to the widely held assumption that flow which is highly oscillatory causes endothelial dysfunction. Chronic convection across the monolayer increased effective permeability more than could be explained by the addition of advective transport, contrary to results from previous acute experiments. The off-the-shelf and modified bioreactors provide an excellent tool for investigating the biomechanics of endothelial permeability and have revealed novel effects of flow duration, viscosity, pulsatility and transmural flow.

Detection of Early Endothelial Dysfunction by Optoacoustic Tomography.

Variations in vascular wall shear stress are often presumed to result in the formation of atherosclerotic lesions at specific arterial regions, where continuous laminar flow is disturbed. The influences of altered blood flow dynamics and oscillations on the integrity of endothelial cells and the endothelial layer have been extensively studied in vitro and in vivo. Under pathological conditions, the Arg-Gly-Asp (RGD) motif binding integrin αvβ3 has been identified as a relevant target, as it induces endothelial cell activation. Animal models for in vivo imaging of endothelial dysfunction (ED) mainly rely on genetically modified knockout models that develop endothelial damage and atherosclerotic plaques upon hypercholesterolemia (ApoE-/- and LDLR-/-), thereby depicting late-stage pathophysiology. The visualization of early ED, however, remains a challenge. Therefore, a carotid artery cuff model of low and oscillating shear stress was applied in CD-1 wild-type mice, which should be able to show the effects of altered shear stress on a healthy endothelium, thus revealing alterations in early ED. Multispectral optoacoustic tomography (MSOT) was assessed as a non-invasive and highly sensitive imaging technique for the detection of an intravenously injected RGD-mimetic fluorescent probe in a longitudinal (2-12 weeks) study after surgical cuff intervention of the right common carotid artery (RCCA). Images were analyzed concerning the signal distribution upstream and downstream of the implanted cuff, as well as on the contralateral side as a control. Subsequent histological analysis was applied to delineate the distribution of relevant factors within the carotid vessel walls. Analysis revealed a significantly enhanced fluorescent signal intensity in the RCCA upstream of the cuff compared to the contralateral healthy side and the downstream region at all time points post-surgery. The most obvious differences were recorded at 6 and 8 weeks after implantation. Immunohistochemistry revealed a high degree of αv-positivity in this region of the RCCA, but not in the left common carotid artery (LCCA) or downstream of the cuff. In addition, macrophages could be detected by CD68 immunohistochemistry in the RCCA, showing ongoing inflammatory processes. In conclusion, MSOT is capable of delineating alterations in endothelial cell integrity in vivo in the applied model of early ED, where an elevated expression of integrin αvβ3 was detected within vascular structures.

Effects of freeze–thaw cycles on the shear stress induced on the cemented sand–structure interface

Freeze–thaw alternation and freeze–thaw cycle erosion are the main reasons for the failure of the cemented soil–structure interface in cold regions. In this study, the direct shear and cyclic direct shear tests of cemented sand–structure interface were conducted in the frozen state (−10 °C) and thawed state (25 °C) at a 300 kPa normal stress under different freeze–thaw cycles (i.e., 0, 1, 5, 10, 15, 20, 25, 30). The experimental results for the shear stresses of the cemented sand–structure interface are presented, and the mechanism of shear stress formation and change was examined through mesoscopic analysis. Variations in shear stress with different freeze–thaw cycles were consistent. Typically, 10 was identified as the critical freeze–thaw cycle number for adfreeze strength, and the hyperbolic model of shear stress–displacement curve in the thawed state was obtained. The peak shear stresses of different freeze–thaw cycles gradually decreased as the shearing cycle number increased until entering a stable stage in frozen state and were close to each other in the thawed state. The internal damage of cemented sand and the formation of ice crystals on the surface increased with the increase in freeze–thaw cycle number. Hence, internal change in cemented sand under freeze–thaw cycles is the fundamental reason for alterations in shear stress. The results of this study shed light on shear stress complexity at the cemented sand–structure interface under freeze–thaw cycles and provide references for practical engineering.

Activation of Notch signaling in endothelium cause upregulation of N-terminal acetylated histone 1

Highlights. Notch signaling is known to be important regulator of endothelium homeostasis and cardiovascular disease. Particularly, Notch seems to be associated with pathological changes in endothelium epigenome although no such Notch effects have been found. We have discovered that activation of Notch signaling alters histone 1 repertoire in the human endothelial cells and this is the first example of epigenomic Notch targets.Aim. The disturbance of blood flow and alteration of physiological shear stress is one of the main reasons for endothelial dysfunction. Mechanosensitive and dosedependent Notch pathway is assumed to be an important player of endothelial dysfunction progression, but the molecular mechanisms of the influence of Notch dysregulation on endothelium are still not understood. In particular, there is no data about possible targets of Notch in the endothelial epigenome.Methods. Here we focused on the analysis of changes in histone code of human umbilical vein endothelial cells (HUVEC) after activation of Notch. For this purpose, we transduced cells by lentiviruses with construction for Notch 1 intracellular domain (N1ICD) overexpression or by empty vector (control). Then we isolated histone enriched fraction and secretome proteins and performed their shotgun proteomics analysis on timsToF Pro instrument. Proteomics data are available via ProteomeXchange with identifier PXD032978.Results. We found the shift in proteomics profile of HUVEC caused by Notch activation and, particularly, the increase in the levels of N-terminal acetylated forms of histone 1: H1-0, H1-3, H1-4, H1-5, H1-10. We also found changes in the cell secretome profile which are associated with the decrease in proangiogenic effect of HUVEC secretome.Conclusion. Our data identified epigenomic Notch targets and we assume that changes in H1 repertoire might be associated with cardiovascular disease progression in vivo.

Transcriptomic analysis of a 3D blood–brain barrier model exposed to disturbed fluid flow

Cerebral aneurysms are more likely to form at bifurcations in the vasculature, where disturbed fluid is prevalent due to flow separation at sufficiently high Reynolds numbers. While previous studies have demonstrated that altered shear stress exerted by disturbed flow disrupts endothelial tight junctions, less is known about how these flow regimes alter gene expression in endothelial cells lining the blood–brain barrier. Specifically, the effect of disturbed flow on expression of genes associated with cell–cell and cell–matrix interaction, which likely mediate aneurysm formation, remains unclear. RNA sequencing of immortalized cerebral endothelial cells isolated from the lumen of a 3D blood–brain barrier model reveals distinct transcriptional changes in vessels exposed to fully developed and disturbed flow profiles applied by both steady and physiological waveforms. Differential gene expression, validated by qRT-PCR and western blotting, reveals that lumican, a small leucine-rich proteoglycan, is the most significantly downregulated gene in endothelial cells exposed to steady, disturbed flow. Knocking down lumican expression reduces barrier function in the presence of steady, fully developed flow. Moreover, adding purified lumican into the hydrogel of the 3D blood–brain barrier model recovers barrier function in the region exposed to fully developed flow. Overall, these findings emphasize the importance of flow regimes exhibiting spatial and temporal heterogeneous shear stress profiles on cell–matrix interaction in endothelial cells lining the blood–brain barrier, while also identifying lumican as a contributor to the formation and maintenance of an intact barrier.

Patients with Bicuspid Aortopathy and Aortic Dilatation.

(1) Background: Bicuspid aortic valve (BAV) is the most frequent congenital cardiac disease. Alteration of ascending aorta diameter is a consequence of shear stress alterations due to haemodynamic abnormalities developed from inadequate valve cusp coaptation. (2) Objective: This narrative review aims to discuss anatomical, pathophysiological, genetical, ultrasound, and radiological aspects of BAV disease, focusing on BAV classification related to imaging patterns and flux models involved in the onset and developing vessel dilatation. (3) Methods: A comprehensive search strategy was implemented in PubMed from January to May 2022. English language articles were selected independently by two authors and screened according to the following criteria. (4) Key Contents and Findings: Ultrasound scan is the primary step in the diagnostic flowchart identifying structural and doppler patterns of the valve. Computed tomography determines aortic vessel dimensions according to the anatomo-pathology of the valve. Magnetic resonance identifies hemodynamic alterations. New classifications and surgical indications derive from these diagnostic features. Currently, indications correlate morphological results, dissection risk factors, and genetic alterations. Surgical options vary from aortic valve and aortic vessel substitution to aortic valve repair according to the morphology of the valve. In selected patients, transcatheter aortic valve replacement has an even more impact on the treatment choice. (5) Conclusions: Different imaging approaches are an essential part of BAV diagnosis. Morphological classifications influence the surgical outcome.

Circulating Notch1 in response to altered vascular wall shear stress in adults.

What is the central question of this study? Is the plasma concentration of Notch1 extracellular domain altered in response to decreased and increased vascular wall shear stress in the forearm in humans? What is the main finding and its importance? Notch1 extracellular domain is increased with acute increases in antegrade shear rate but does not change with 20min of decreased shear rate caused by distal forearm occlusion. A novel and integral endothelial mechanosensor in humans that can help explain vascular endothelial adjustments in response to increases in antegrade shear stress was characterized. Notch1 has been proposed as a novel endothelial mechanosensor that is central for signalling adjustments in response to changes in vascular wall shear stress. However, there remains no controlled in vivo study in humans. Accordingly, we sought to address the question of whether plasma concentrations of Notch1 extracellular domain (ECD) is altered in response to transient changes in vascular wall shear stress. In 10 young healthy adults (6M/4F), alterations in shear stress were induced by supra-systolic cuff inflation around the wrist. The opposite arm was treated as a time control with no wrist cuff inflation. Plasma was collected from an antecubital vein of both arms at baseline, 20min of wrist cuff inflation (low shear), as well as 1-2min (high shear) and 15min following (recovery) wrist cuff release. The Notch1 ECD was quantified using a commercially available ELISA. Duplex ultrasound was used to confirm alterations in shear stress. In the experimental arm, concentrations of Notch1 ECD remained statistically similar to baseline at all time points except for immediately following cuff release where it was elevated by ∼50% (P=0.033), coinciding with the condition of high antegrade shear rate. Concentrations of Notch1 ECD remained unchanged in the control arm through all time points. These data indicate that Notch1 is a viable biomarker for quantifying mechanotransduction in response to increased shear stress in humans, and it may underlie the vascular adaptations or mal-adaptations associated with conditions that impact antegrade shear.

Abstract 550: Magnetic Nanoparticle-mediated Targeting Of Endothelium To Address Restenosis In A Bioprinted In Vitro Model Of Pulmonary Arteries

Vascular restenosis is a major complication in recanalized arteries. Nanoparticles (NPs) have shown great promise as delivery systems in advancing strategies to treat such vascular anomalies. By enabling precise targeting, NPs can overcome the challenges of low drug efficacy and off-target effects. Here we present a biomimetic in vitro platform comprised of 3D bioprinting, nanomaterials, and perfusion technologies, to study the use of NP targeting to address endothelial overgrowth. We bioprinted 3D vascular channels at high fidelity, using gelatin methacrylate as bioink, with artery-like stiffness. Human endothelial cells (ECs) were used to endothelialize the printed channels. GFP-labelled superparamagnetic iron oxide NPs (SPIONs), loaded with the Rapamune anti-proliferative drug, were perfused through the bifurcated artery model at physiological rate. Computational modeling predicted greatest level of alterations in wall shear stress in the conduit’s junction with the artery, identifying this region prone to restenosis. A neodymium disc magnet was embedded in the printed tissue to attract the therapeutic SPIONs to the region of high risk. In vitro dynamic culture was conducted for 2 wks. We assessed cell viability, proliferation, and function using AlamarBlue and immunohistochemistry. Results showed significant targeted effect of NP delivery in reducing EC overgrowth. This platform enables design of precise targeting of therapeutics to treat a variety of cardiovascular diseases at a high spatial and temporal control.

Wall Shear Stress Alteration: a Local Risk Factor of Atherosclerosis.

Wall shear stress describes the mechanical influence of blood flow on the arterial wall. In this review, we discuss the role of the wall shear stress in the development of atherosclerosis and its complications. Areas with chronically low, oscillating wall shear stress are most prone to plaque development and include outer bifurcation walls and inner walls of arches. In some diseases, patients have lower wall shear stress even in straight arterial segments; also, these findings were associated with atherosclerosis. High wall shear stress develops in the distal part (shoulder) of a stenosis and contributes to plaque destabilization. Wall shear stress changes are involved in the development of atherosclerosis. They are not fully understood yet and act in concert with tangential wall stress.

Simultaneous Upregulation of Elastolytic and Elastogenic Factors Are Necessary for Regulated Collateral Diameter Expansion.

Background: During arteriogenesis, outward remodeling of the arterial wall expands luminal diameter to produce increased conductance in developing collaterals. We have previously shown that diameter expansion without loss of internal elastic lamina (IEL) integrity requires both degradation of elastic fibers and LOX-mediated repair. The aim of this study was to investigate the expression of genes involved in remodeling of the extracellular matrix (ECM) using a model of arteriogenesis.Methods: Sprague-Dawley rats underwent femoral artery ligation with distal arteriovenous fistula (FAL + AVF) placement. Profunda femoral arteries (PFA) were harvested for analysis at various time points. Serum desmosine, an amino acid found exclusively in elastin, was evaluated with enzyme-linked immunosorbent assay (ELISA) as a marker of tissue elastolysis. Tissue mRNA isolated from FAL + AVF exposed PFAs was compared to the contralateral sham-operated using qPCR. HCAECs were cultured under low shear stress (8 dyn·s/cm2) for 24 h and then exposed to high shear stress (40 dyn·s/cm2) for 2–6 h. Primers used included FBN-1, FBN-2, Timp-2, LOX-1, Trop-E, Cath-K, Cath-S, MMP-2, MMP-9, FBLN-4, and FBLN-5 and were normalized to GAPDH. mRNA fold changes were quantified using the 2-ΔΔCq method. Comparisons between time points were made with non-parametric ANOVA analysis with Bonferroni adjustment.Results: PFAs showed IEL reorganization during arteriogenesis. Serum desmosine levels are significantly elevated at 2 days and one week, with a return to baseline thereafter (p < 0.01). Expression of ECM structural proteins (FBN-1, FBN-2, FBLN-4, FBLN-5, Tropoelastin, TIMP-2, LOX-1) and elastolytic proteins (MMP-2, MMP-9, Cathepsin S, Cathepsin K) exhibited an early peak (p < 0.05) relative to sham PFAs. After two weeks, expression returned to baseline. HCAECs demonstrated upregulation of FBN-2, FBLN-5, LOX-1 and Trop-E at 4 h of high shear stress, as well as elastolytic protein MMP-2.Conclusions: Elastin degradation begins early in arteriogenesis and is mediated by local upregulation of elastolytic genes. Elastolysis appears to be simultaneously balanced by production of elastic fiber components which may facilitate stabilization of the IEL. Endothelial cells are central to initiation of arteriogenesis and begin ECM remodeling in response to altered shear stress.

Shear Stress Alterations Activate BMP4/pSMAD5 Signaling and Induce Endothelial Mesenchymal Transition in Varicose Veins.

Chronic venous diseases, including varicose veins, are characterized by hemodynamic disturbances due to valve defects, venous insufficiency, and orthostatism. Veins are physiologically low shear stress systems, and how altered hemodynamics drives focal endothelial dysfunction and causes venous remodeling is unknown. Here we demonstrate the occurrence of endothelial to mesenchymal transition (EndMT) in human varicose veins. Moreover, the BMP4-pSMAD5 pathway was robustly upregulated in varicose veins. In vitro flow-based assays using human vein, endothelial cells cultured in microfluidic chambers show that even minimal disturbances in shear stress as may occur in early stages of venous insufficiency induce BMP4-pSMAD5-based phenotype switching. Furthermore, low shear stress at uniform laminar pattern does not induce EndMT in venous endothelial cells. Targeting the BMP4-pSMAD5 pathway with small molecule inhibitor LDN193189 reduced SNAI1/2 expression in venous endothelial cells exposed to disturbed flow. TGFβ inhibitor SB505124 was less efficient in inhibiting EndMT in venous endothelial cells exposed to disturbed flow. We conclude that disturbed shear stress, even in the absence of any oscillatory flow, induces EndMT in varicose veins via activation of BMP4/pSMAD5-SNAI1/2 signaling. The present findings serve as a rationale for the possible use of small molecular mechanotherapeutics in the management of varicose veins.

Abstract 13336: Investigating the Effects of Physiological Shear Stress on Monocyte Phenotype for Therapeutic Development in Atherosclerosis

Introduction: Monocytes are highly plastic immune cells which play a significant role in atherosclerosis development and progression. Monocyte phenotype is modulated by stimuli including physical parameters like the shear stress induced by blood flow. Atherosclerotic plaque formation impedes blood flow thereby impacting the shear stress applied to monocytes. The altered shear stress could contribute to the abnormal monocyte phenotype populations in atherosclerotic patients. Aim: To understand the relationship between monocyte phenotypic response to varying physiological shear stresses. Methods: To investigate the dynamics of monocytes under inflammatory stresses, THP-1wt cells were exposed to 40 dyne/cm 2 and 5 dyne/cm 2 which represent above normal and below-normal, atherosclerotic shear stress conditions respectively. Monocytes were either untreated or stimulated with lipopolysaccharide (LPS) to simulate low-grade inflammation. To evaluate the phenotypic response, ELISA and qPCR measurements of pro-inflammatory (TNF-α IL-12, NOS2, MAC-1, CCL2, CD40), anti-inflammatory (IL-10), and adhesion molecules (ICAM-1, VCAM-1, CCR2) were taken directly after, and four hours post shearing. Results: Shear stresses of 5 dyne/cm 2 or 40 dyne/cm 2 show differential response profiles in monocytes. At both stresses, THP-1wt cells downregulated expression of inflammatory and adhesion molecules after 4hrs. However, pre-stimulation with LPS resulted in either maintained or increased levels of expression. Notably, LPS stimulation for both shear stresses saw a 40-fold or greater increase in TNF-α, CCL2, CD40, and VCAM-1 after 4-hour incubation. In all conditions angiotensin converting enzyme 2 (ACE2), a molecule important for blood pressure regulation, was downregulated. Conclusion: The data concludes that shear stress has an impact on the activation of monocytes in both unstimulated and stimulated conditions. LPS data suggests that monocytes which are already in proinflammatory conditions are more susceptible to sustained activation from shear stress.