Vasculogenic assembly of 3D capillary networks remains a promising approach to vascularizing tissue-engineered grafts, a significant outstanding challenge in tissue engineering and regenerative medicine. Current approaches for vasculogenic assembly rely on the inclusion of supporting mesenchymal cells alongside endothelial cells, co-encapsulated within vasculo-conducive materials such as low-density fibrin hydrogels. Here, we established a material-based approach to circumvent the need for supporting mesenchymal cells and report that the inclusion of synthetic matrix fibers in dense (>3 mg mL-1) 3D fibrin hydrogels can enhance vasculogenic assembly in endothelial cell monocultures. Surprisingly, we found that the addition of non-cell-adhesive synthetic matrix fibers compared to cell-adhesive synthetic fibers best encouraged vasculogenic assembly, proliferation, lumenogenesis, a vasculogenic transcriptional program, and additionally promoted cell-matrix interactions and intercellular force transmission. Implanting fiber-reinforced prevascularized constructs to assess graft-host vascular integration, we demonstrate additive effects of enhanced vascular network assembly during in vitro pre-culture, fiber-mediated improvements in endothelial cell survival and vascular maintenance post-implantation, and enhanced host cell infiltration that collectively enabled graft vessel integration with host circulation. This work establishes synthetic matrix fibers as an inexpensive alternative to sourcing and expanding secondary supporting cell types for the prevascularization of tissue constructs.

Oxidative stress and endothelial dysfunction are critical drivers of atherosclerosis, but the mechanisms regulating oxidative stress under disturbed flow conditions remain incompletely understood. The ubiquitin-proteasome system, particularly E3 ubiquitin ligases, may play a pivotal role in modulating these processes. FBXO38, an E3 ligase involved in proteasomal degradation, has been implicated in various physiological pathways, but its role in regulating oxidative stress in endothelial cells is unknown. We hypothesized that FBXO38 mitigates endothelial damage induced by low oscillatory shear stress (LOSS) by promoting the ubiquitin-proteasome-dependent degradation of Nox1, a major source of reactive oxygen species (ROS). Using an in vitro LOSS model in human umbilical vein endothelial cells (HUVECs) and an in vivo mouse partial carotid ligation model, we assessed the expression of FBXO38 and Nox1 through quantitative PCR, western blotting, immunofluorescence, and immunohistochemistry. LOSS significantly reduced FBXO38 protein expression (by ~60%, p < 0.0001 at 24 h), leading to increased Nox1 protein levels (approximately two-fold, p < 0.001) and apoptosis. FBXO38 overexpression markedly attenuated Nox1 accumulation (~50% reduction, p < 0.05), reduced ROS production, and improved cell viability under LOSS conditions, whereas FBXO38 knockdown exacerbated these effects. Moreover, FBXO38 directly interacted with Nox1, suggesting a ubiquitin-dependent degradation mechanism. Our results reveal that FBXO38 regulates endothelial oxidative stress by controlling Nox1 stability under disturbed shear stress conditions. Although FBXO38 emerges as a promising candidate for therapeutic targeting, further studies are necessary to validate its potential in preclinical and clinical settings.

Endothelial cells (ECs) are arranged side-by-side to create a semi-permeable monolayer, forming the inner lining of every blood vessel (micro and macrocirculation). Serving as the first barrier for circulating molecules and cells, ECs represent the main regulators of vascular homeostasis being able to respond to environmental changes, either physical or chemical signals, by producing several factors that regulate vascular tone and cellular adhesion. Healthy endothelium has anticoagulant properties that prevent the adhesion of leukocytes and platelets to the vessel walls, contributing to resistance to thrombus formation, and regulating inflammation, and vascular smooth muscle cell proliferation. Many risk factors of cardiovascular diseases (CVDs) promote the endothelial expression of chemokines, cytokines, and adhesion molecules. The resultant endothelial activation can lead to endothelial cell dysfunction (ECD). In vitro models of ECD allow the study of cellular and molecular mechanisms of disease and provide a research platform for screening potential therapeutic agents. Even though alternative models are available, such as animal models or ex vivo models, in vitro models offer higher experimental flexibility and reproducibility, making them a valuable tool for the understanding of pathophysiological mechanisms of several diseases, such as CVDs. Therefore, this review aims to synthesize the currently available in vitro models regarding ECD, emphasizing CVDs. This work will focus on 2D cell culture models (endothelial cell lines and primary ECs), 3D cell culture systems (scaffold-free and scaffold-based), and 3D cell culture models (such as organ-on-a-chip). We will dissect the role of external stimuli-chemical and mechanical-in triggering ECD.

The translation of biomedical devices and drug research is an expensive and long process with a low probability of receiving FDA approval. Developing physiologically relevant in vitro models with human cells offers a solution to not only improving the odds of FDA approval but also to expand our ability to study complex in vivo systems in a simpler fashion. Animal models remain the standard for pre-clinical testing; however, the data from animal models is an unreliable extrapolation when anticipating a human response in clinical trials, thus contributing to the low rates of translation. In this review, we focus on in vitro vascular or angiogenic models because of the incremental role that the vascular system plays in the translation of biomedical research. The first section of this review discusses the most common angiogenic cytokines that are used in vitro to initiate angiogenesis, followed by angiogenic inhibitors where both initiators and inhibitors work to maintain vascular homeostasis. Next, we evaluate previously published in vitro models, where we evaluate capturing the physical environment for biomimetic in vitro modeling. These topics provide a foundation of parameters that must be considered to improve and achieve vascular biomimicry. Finally, we summarize these topics to suggest a path forward with the goal of engineering human in vitro models that emulate the in vivo environment and provide a platform for biomedical device and drug screening that produces data to support clinical translation.

Altered hemodynamics is a key factor for atherosclerosis. For decades, endothelial cell (EC) responses to fluid-generated wall shear stress have been the central focus for atherogenesis. However, circulating blood is not a cell-free fluid, it contains mechanosensitive red blood cells (RBCs) that are also subjected to altered hemodynamics and release a large amount of ATP, but their impact on atherosclerosis has been overlooked. The focus of this study is the role of shear stress (SS)-induced RBC-released ATP in atherosclerosis. Hypercholesterolemic mouse models with and without RBC-Pannexin 1 deletion were used for the study. Results showed that SS-induced release of ATP from RBCs was at µM concentrations, three-orders of magnitude higher than that from other cell types. Suppression of RBC-released ATP via deletion of Pannexin 1, a mechanosensitive ATP-permeable channel, reduced high-fat diet-induced aortic plaque burden by 40%-60%. Importantly, the location and the extent of aortic atherosclerotic lesions spatially matched with the ATP deposition profile at aortic wall predicted by a computational fluid dynamic (CFD) model. Furthermore, hypercholesterolemia increases EC susceptibility to ATP with potentiated increase in [Ca2+]i, an initial signaling for aortic EC barrier dysfunction, and an essential cause for lipid accumulation and inflammatory cell infiltration. The computational prediction also provides a physics-based explanation for RBC-released ATP-induced sex disparities in atherosclerosis. Our study reveals an important role of RBC-released ATP in the initiation and progression of atherosclerosis. These novel findings provide a more comprehensive view of how altered hemodynamics and systemic risk factors synergistically contribute to atherosclerosis.NEW & NOTEWORTHY This study reveals that, in addition to fluid-derived wall shear stress, the disturbed blood flow-induced release of ATP from mechanosensitive red blood cells (RBCs), the major cellular components of blood, along with hypercholesterolemia-induced increases in endothelial cell susceptibility to ATP contribute significantly to the initiation and progression of atherosclerosis. These novel findings advance our current understanding of how altered hemodynamics and hypercholesterolemia synergistically contribute to atherosclerosis for the first time with the inclusion of RBCs.

The air-blood barrier protects the lung from blood/serum entering the air spaces, i.e., from "drowning in your own fluids". Failure leads to pulmonary edema, a regularly fatal complication during the Covid-19 pandemic which claimed 7 million lives worldwide. Finding no mathematical models for the underlying fluid mechanics, we created the first. Governing flow equations for alveolar capillary, interstitium, and alveolus are coupled by crossflows at the capillary and epithelial membranes and end-exit flows to the lymphatics. Case examples include normal/recovery, cardiogenic pulmonary edema, acute respiratory distress syndrome, effects of positive end expiratory pressure, and a wide range of parameter values for permeability of the membranes and interstitial matrix. Previously unknown membrane fluid shear stresses calculate to values that affect cell function in many systems. We add active epithelial reabsorption which has two effects: shifting streamlines to favor alveolar-lymphatic clearance and adding to the direct alveolar-capillary clearance. Simple algebraic equations are derived for the interstitial fluid pressure, pi, membrane crossflow velocities and the critical capillary pressure, pcrit, above which edema occurs. For validation, the pcrit predictions fit clinical definitions and flow calculations of lymphatic vs capillary clearance match animal experimental data. For decades the value of pi has been imposed as an input, whereas we calculate the value as an output. They don't agree. Since the space is too small for measurements, the ability to calculate pi and pcrit offers new insights, questions long-held beliefs, and opens applications from physiological studies to personalized clinical care.

A longstanding challenge in microfluidics has been the efficient delivery of fluids from macro-scale pumping systems into microfluidic devices, known as the "world-to-chip" problem. Thus far, the entire industry has accepted the use of imperfect, rigid tubing and connectors as the ecosystem within which to operate, which, while functional, are often cumbersome, labor-intensive, prone to errors, and ill-suited for high-throughput experimentation. In this paper, we introduce TapeTech microfluidics, a flexible and scalable solution designed to address the persistent "world-to-chip" problem in microfluidics, particularly in organ-on-a-chip (OoC) applications. TapeTech offers a streamlined alternative, utilizing adhesive tape and thin-film polymers to create adaptable, integrated multi-channel ribbon connectors that simplify fluidic integration with pumps and reservoirs. Key features of TapeTech include reduced pressure surges, easy priming, rapid setup, easy multiplexing, and broad compatibility with existing devices and components, which are essential for maintaining stable fluid dynamics and protecting sensitive cell cultures. Furthermore, TapeTech is designed to flex around the lids of Petri dishes, enhancing sterility and transportability by enabling easy transfer between incubators, biosafety cabinets (BSCs), and microscopes. The rapid design-to-prototype iteration enabled by TapeTech allows users to quickly develop connectors for a wide range of microfluidic devices. Importantly, we showcase the utility of TapeTech in OoC cultures requiring fluid flow. We also highlight other utilities, such as real-time microscopy and a well-plate medium exchanger. The accessibility of this technology should enable more laboratories to simplify design and setup of microfluidic experiments, and increase technology adoption.

Proteins that allow water to move in and out of cells help shape the development of new blood vessels.

Modulation of endothelial traction is critical for the responses of endothelial cells to fluid shear stress (FSS), which has profound implications for vascular health and atherosclerosis. Previously, we demonstrated that under high FSS, endothelial cells rapidly increase traction forces, followed by relaxation, with traction aligning in the flow direction. In contrast, low shear preconditioning induces a modest short-term increase in traction (<30 min), followed by a secondary long-term (>14 hr) rise, with traction/cells aligning perpendicular to the flow. The upstream mechanosensors driving these responses, however, remain unknown. Here, we sought the roles of Piezo1 and TRPV4 ion channels in shear-induced traction modulation. We report that HUVECs with Piezo1 silencing reduced the initial traction rise in half under high FSS compared to those by WT cells, while not affecting the traction modulation in response to low FSS or traction/cell alignment to the flow direction. Conversely, cells with siTRPV4 fully abrogated the initial traction rise, as well as alignment of traction and cells, in response to both high and low FSS conditions. Dual inhibition of Piezo1 and TRPV4 further impaired both initial and long-term traction under high FSS. Interestingly, dual-inhibited cells displayed larger initial traction responses to low FSS compared to control cells, suggesting the involvement of alternative calcium-independent pathways that become dominant when both ion channels are nonfunctional. Additionally, either ion channel inhibition led to secondary long-term traction increase even under high FSS condition. These findings suggest that while both Piezo1 and TRPV4 channels contribute to shear mechanotransduction, TRPV4 plays more dominant role than Piezo1 in mediating the initial traction rise and sustaining long-term relaxation under high or low shear stress, highlighting their critical and distinct contributions to endothelial mechanotransduction and remodeling.

A novel microfluidic platform was designed to study the cellular architecture of endothelial cells (ECs) in an environment replicating the 3D organization and flow of blood vessels. In particular, the platform was constructed to investigate EC defects in slow-flow venous malformations (VMs) under varying shear stress and flow conditions. The platform featured a standard microtiter plate footprint containing 32 microfluidic units capable of replicating wall shear stress (WSS) in normal veins and enabling precise control of shear stress and flow directionality without the need for complex pumping systems. Using genetically engineered human umbilical vein endothelial cells (HUVECs) and induced pluripotent stem cell (iPSC)-derived ECs (iECs) to express the recurrent TIE2L914F VM mutation we assessed responses on EC orientation and area, actin organization, and Golgi polarization to uni- and bidirectional flow and varying WSS. Comparison of control and TIE2L914F expressing ECs showed differential cellular responses to flow and WSS in terms of cell shape elongation, orientation of F-actin, and Golgi polarization, indicating altered mechanosensory or mechanotransduction signaling pathways in the presence of the VM causative mutation. The data also revealed significant differences in how the primary and iPSC-derived iECs responded to flow. As a conclusion, the developed microfluidic platform allowed simulation of multiple flow conditions in a scalable and pumpless format. The design made it a desirable tool for studying different EC types as well as cellular changes in vascular disease. The platform should offer new opportunities for biomechanical research by providing a controlled environment to analyze the flow-dependent mechanosensory pathways in ECs.

Preeclampsia (PE) is a challenge in maternal healthcare due to its complex nature, characterized by high blood pressure, protein in the urine, and damage to various organs. There is evidence linking PE to endothelial dysfunction (ED), triggered by substances released from an oxygen-deprived placenta. Previous in vitro studies have not considered the impact of in vivo elements, such as the different patterns of blood flow, and laminar (LSS) vs. oscillatory (OSS) shear stress, on the development of ED. We investigated the impact of plasma from healthy pregnant women (HP), subjects with gestational hypertension (GH), and PE patients on global gene expression of human coronary endothelial cells (HCAECs) under LSS and OSS. Our findings revealed a unique transcriptional profile of endothelial cells induced by plasma incubation in LSS. Notably, OSS resulted in similar transcriptomes irrespective of plasma treatment. Under LSS, GH plasma resulted in a proliferative profile, whereas PE plasma was linked to pro-inflammatory and antioxidant profiles compared to HP plasma. Our findings demonstrate that shear stress levels influence the endothelial cell transcriptome in response to plasma from hypertensive pregnancy patients. Both PE and GH can induce endothelial dysfunction under atheroprotective LSS, with a more significant effect observed with PE-derived plasma.

In this work, we present a cost effective and open-source modular cone-and-plate (MoCAP) device that incorporates shear stress in the popular Transwell® insert system. This system acts as a lid that incorporates flow into 24-well Transwell® inserts while preserving the ability to conduct molecular profiling assays. Moreover, the MoCAP device can be rapidly reconfigured to test multiple shear stress profiles within a single device. To demonstrate the utility of the MoCAP, we conducted select assays on several different brain microvascular endothelial cell (BMEC) lines that comprise models of the blood-brain barrier (BBB), since shear stress can play an important role in BBB function. Our results characterize how shear stress modulates passive barrier function and GLUT1 expression across the different BMEC lines. Overall, we anticipate this low cost mechanofluidic device will be useful to the mechanobiology community.

The aberrant vascular response associated with tendon injury results in circulating immune cell infiltration and a chronic inflammatory feedback loop leading to poor healing outcomes. Studying this dysregulated tendon repair response in human pathophysiology has been historically challenging due to the reliance on animal models. To address this, our group developed the human tendon-on-a-chip (hToC) to model cellular interactions in the injured tendon microenvironment; however, this model lacked the key element of physiological flow in the vascular compartment. Here, we leveraged the modularity of our platform to create a fluidic hToC that enables the study of circulating immune cell and vascular crosstalk in a tendon injury model. Under physiological shear stress consistent with postcapillary venules, we found a significant increase in the endothelial leukocyte activation marker intercellular adhesion molecule 1 (ICAM-1), as well as enhanced adhesion and transmigration of circulating monocytes across the endothelial barrier. The addition of tissue macrophages to the tendon compartment further increased the degree of circulating monocyte infiltration into the tissue matrix. Our findings demonstrate the importance of adding physiological flow to the human tendon-on-a-chip, and more generally, the significance of flow for modeling immune cell interactions in tissue inflammation and disease.

Hemodynamic forces play a crucial role in modulating endothelial cell (EC) behavior, significantly influencing blood vessel responses. While traditional in vitro studies often explore ECs under static conditions, ECs are exposed to various hemodynamic forces in vivo. This study investigates how wall shear stress (WSS) influences EC metabolism, focusing on the interplay between WSS and key metabolic pathways.

The aim of this study is to examine the effects of WSS on EC metabolism, specifically evaluating its impact on central carbon metabolism and glycolysis using transcriptomics and tracer metabolomics approaches.

ECs were exposed to WSS, and transcriptomic analysis was performed to assess gene expression changes related to metabolic pathways. Tracer metabolomics was used to track metabolic fluxes, focusing on glutamine and glycolytic metabolism. Additionally, chemical inhibition of glutamate dehydrogenase was conducted to evaluate its role in EC fitness under WSS.

Transcriptomic data revealed upregulation of glutamine and glutamate pathways, alongside downregulation of glycolytic activity in ECs exposed to WSS. Tracer metabolomics confirmed that WSS promotes glutamine anaplerosis into the Krebs cycle, while decreasing glycolytic metabolism. Suppression of glutamate dehydrogenase impaired EC fitness under WSS conditions.

Our findings illuminate that ECs subjected to WSS exhibit a preference for glutamine as a key nutrient source for central carbon metabolism pathways, indicating diminished reliance on glycolysis. This study elucidates the nutritional predilections and regulatory mechanisms governing EC metabolism under WSS in vitro, underscoring the pivotal role of physical stimuli in shaping EC metabolic responses.

Cardiac rehabilitation, a comprehensive exercise-based lifestyle and medical management, is effective in decreasing morbidity and improving life quality in patients with coronary heart disease. Endothelial function, an irreplaceable indicator in coronary heart disease progression, is measured by various methods in traditional cardiac rehabilitation pathways, including medicinal treatment, aerobic training, and smoking cessation. Nevertheless, studies on the effect of some emerging cardiac rehabilitation programs on endothelial function are limited. This article briefly reviewed the endothelium-beneficial effects of different cardiac rehabilitation pathways, including exercise training, lifestyle modification and psychological intervention in patients with coronary heart disease, and related experimental models, and summarized both uncovered and potential cellular and molecular mechanisms of the beneficial roles of various cardiac rehabilitation pathways on endothelial function. In exercise training and some lifestyle interventions, the enhanced bioavailability of nitric oxide, increased circulating endothelial progenitor cells (EPCs), and decreased oxidative stress are major contributors to preventing endothelial dysfunction in coronary heart disease. Moreover, the preservation of endothelial-dependent hyperpolarizing factors and inflammatory suppression play roles. On the one hand, to develop more endothelium-protective rehabilitation methods in coronary heart disease, adequately designed and sized randomized multicenter clinical trials should be advanced using standardized cardiac rehabilitation programs and existing assessment methods. On the other hand, additional studies using suitable experimental models are warranted to elucidate the relationship between some new interventions and endothelial protection in both macro- and microvasculature.

Microvascular brain endothelial cells tightly limit the entry of blood components and peripheral cells into the brain by forming the blood-brain barrier (BBB). The BBB is regulated by a cascade of mechanical and chemical signals including shear stress and elasticity of the adjacent endothelial basement membrane (BM). During physiological aging, but especially in neurological diseases including multiple sclerosis (MS), stroke, small vessel disease, and Alzheimer's disease (AD), the BBB is exposed to inflammation, rigidity changes of the BM, and disturbed cerebral blood flow (CBF). These altered forces lead to increased vascular permeability, reduced endothelial reactivity to vasoactive mediators, and promote leukocyte transmigration. Whereas the molecular players involved in leukocyte infiltration have been described in detail, the importance of mechanical signalling throughout this process has only recently been recognized. Here, we review relevant features of mechanical forces acting on the BBB under healthy and pathological conditions, as well as the endothelial mechanosensory elements detecting and responding to altered forces. We demonstrate the underlying complexity by focussing on the family of transient receptor potential (TRP) ion channels. A better understanding of these processes will provide insights into the pathogenesis of several neurological disorders and new potential leads for treatment.

Stenosis or narrowing of arteries due to the buildup of plaque is a common occurrence in atherosclerosis and coronary artery disease (CAD), limiting blood flow to the heart and posing substantial cardiovascular risk. While the role of geometric irregularities in arterial stenosis is well-documented, the complex interplay between the abnormal hemorheology and asymmetric shape in flow characteristics remains unexplored.

This study investigates the influence of varying hematocrit (Hct) levels, often caused by conditions such as diabetes and anemia, on flow patterns in an idealized eccentric stenotic artery using computational fluid dynamics simulations. We consider three physiological levels of Hct, 25%, 45%, and 65%, representing anemia, healthy, and diabetic conditions, respectively. The numerical simulations are performed for different combinations of shape eccentricity and blood rheological parameters, and hemodynamic indicators such as wall shear stress (WSS), oscillatory shear index (OSI), are relative residence time (RRT) are calculated to assess the arterial health.

Our results reveal the significant influence of Hct level on stenosis progression. CAD patients with anemia are exposed to lower WSS and higher OSI, which may increase the propensity for plaque progression and rupture. However, for CAD patients with high Hct level - as is often the case in diabetes - the WSS at the minimal lumen area increases rapidly, which may also lead to plaque rupture and cause adverse events such as heart attacks. These disturbances promote endothelial dysfunction, inflammation, and thrombus formation, thereby intensifying cardiovascular risk.

Our findings underscore the significance of incorporating hemorheological parameters, such as Hct, into computational models for accurate assessment of flow dynamics. We envision that insights gained from this study will inform the development of tailored treatment strategies and interventions in CAD patients with common comorbidities such as diabetes and anemia, thus mitigating the adverse effects of abnormal hemorheology and reducing the ever-growing burden of cardiovascular diseases.

Vascular tissue engineering faces significant challenges in creating in vitro vascular disease models, implantable vascular grafts, and vascularized tissue/organ constructs due to limitations in manufacturing precision, structural complexity, replicating the composited architecture, and mimicking the mechanical properties of natural vessels. Light-based 3D bioprinting, leveraging the unique advantages of light including high resolution, rapid curing, multi-material adaptability, and tunable photochemistry, offers transformative solutions to these obstacles. With the emergence of diverse light-based 3D bioprinting techniques and innovative strategies, the advances in vascular tissue engineering have been significantly accelerated. This review provides an overview of the human vascular system and its physiological functions, followed by an in-depth discussion of advancements in light-based 3D bioprinting, including light-dominated and light-assisted techniques. We explore the application of these technologies in vascular tissue engineering for creating in vitro vascular disease models recapitulating key pathological features, implantable blood vessel grafts, and tissue analogs with the integration of capillary-like vasculatures. Finally, we provide readers with insights into the future perspectives of light-based 3D bioprinting to revolutionize vascular tissue engineering.

Shear stress plays a critical role in regulating physiological processes within microcirculatory systems. While particle imaging velocimetry is a standard technique for quantifying shear flow, uncertainty near boundaries and low resolution remain severe restrictions. Additionally, shear stress determination is particularly challenging in biofluids due to their significant non-Newtonian behaviors. The present study develops a shearmetry technique in physiological settings using a biomimetic fluid containing rare earth-doped luminescent nanorods acting in two roles. First, they are used as colloidal additives adjusting rheological properties in physiological media. Their anisotropic morphology and interparticle interaction synergistically induce a non-Newtonian shear-thinning effect emulating real biofluids. Second, they can probe shear stress due to the shear-induced alignment. The polarized luminescence of the nanorods allows for quantifying their orientational order parameter and thus correlated shear stress. Using scanning confocal microscopy, we demonstrate the tomographic mapping of the shear stress distribution in microfluidics. High shear stress is evident near the constriction and the cellular periphery, in which non-Newtonian effects can have a significant impact. This emerging shearmetry technique is promising for implementation in physiological and rheological environments of biofluids.

Non-coding RNAs (ncRNAs) have key roles in the etiology of many illnesses, including heart failure, myocardial infarction, stroke, and in physiological processes like angiogenesis. In transcriptional regulatory circuits that control heart growth, signaling, and stress response, as well as remodeling in cardiac disease, ncRNAs have become important players. Studies on ncRNAs and cardiovascular disease have made great progress recently. Here, we go through the functions of non-coding RNAs (ncRNAs) like circular RNAs (circRNAs), and microRNAs (miRNAs) as well as long non-coding RNAs (lncRNAs) in modulating cardiovascular disorders.

Carotid plaque vulnerability is a significant factor in the risk of cardiocerebrovascular events, with intraplaque neovascularization (IPN) being a crucial characteristic of plaque vulnerability. This study investigates the value of ultrasound vector flow imaging (V-flow) for measuring carotid plaque wall shear stress (WSS) in predicting the extent of IPN.

We enrolled 140 patients into three groups: 53 in the plaque group (72 plaques), 23 in the stenosis group (27 plaques), and 64 in the control group. V-flow was used to measure WSS parameters, including the average WSS (WSS mean) and the maximum WSS (WSS max), across three plaque locations: mid-upstream, maximum thickness, and mid-downstream. Contrast-enhanced ultrasound examination was used in 76 patients to analyze IPN and its correlation with WSS parameters.

WSS max in the stenosis group was significantly higher than that in the control and plaque groups at the maximum thickness part (P < .05) and WSS mean in the stenosis group was significantly lower than that in the control group at the mid-upstream and mid-downstream segments (P < .05). WSS mean in the plaque group was significantly lower than that of the control group at all three locations (P < .05). Contrast-enhanced ultrasound examination revealed that plaques with neovascularization enhancement exhibited significantly higher WSS values (P < .05), with a positive correlation between WSS parameters and IPN enhancement grades, particularly WSS max at the thickest part (r = 0.508). Receiver operating characteristic curve analysis of WSS parameters for evaluating IPN showed that the efficacy of WSS max in evaluating IPN was better than that of WSS mean (P < .05), with an area under the curve of 0.7762 and 0.6973 (95% confidence intervals, 0.725-0.822 and 0.642-0.749, respectively). The cut-offs were 4.57 Pa and 1.12 Pa, sensitivities were 74.03% and 63.64%, and specificities were 75.00% and 68.18%.

V-flow effectively measures WSS in carotid plaques. WSS max provides a promising metric for assessing IPN, offering potential insights into plaque characteristics and showing some potential in predicting plaque vulnerability.

Wall shear stress (WSS) is a critical factor in vascular biology, and both high and low WSS are implicated in atherosclerosis. Fibronectin (FN) is a key extracellular matrix protein that plays an important role in cell activities. Under high shear stress, plasma FN undergoes fibrillogenesis; however, its behavior under low shear stress remains unclear. This study aimed to investigate the formation ofin vitrocell-free fibrillar FN (FFN) under low shear rate conditions and its effect on bovine aortic endothelial cell behavior. FN (500µg ml-1) was perfused through slide chambers at three flow rates (0.16 ml h-1, 0.25 ml h-1, and 0.48 ml h-1), corresponding to low shear rates of 0.35 s-1, 0.55 s-1, and 1.05 s-1, respectively, for 4 h at room temperature. The formed FN matrices were observed using fluorescence microscopy and scanning electron microscopy. Under low shear rates, distinct FN matrix structures were observed. FFN0.48 formed immense fibrils with smooth surfaces, FFN0.25 formed a matrix with a rough surface, and FFN16 exhibited nodular structures. FFN0.25 supported cell activities to a greater extent than native FN and other FFN surfaces. Our study suggests that abnormally low shear conditions impact FN structure and function and enhance the understanding of FN fibrillogenesis in vascular biology, particularly in atherosclerosis.

The growth of new blood vessels through angiogenesis is a highly coordinated process, which is initiated by chemokine gradients that activate endothelial cells within a perfused parent vessel to sprout into the surrounding 3D tissue matrix. While both biochemical signals from pro-angiogenic factors, as well as mechanical cues originating from luminal fluid flow that exerts shear stress on the vessel wall, have individually been identified as major regulators of endothelial cell sprouting, it remains unclear whether and how both types of cues synergize. To fill this knowledge gap, here, we created a 3D biomimetic model of chemokine gradient-driven angiogenic sprouting, in which a micromolded tube inside a hydrogel matrix is seeded with endothelial cells and connected to a perfusion system to control fluid flow rates and resulting shear forces on the vessel wall. To allow for the formation of chemokine gradients despite the presence of luminal flow, a nanoporous synthetic hydrogel that supports angiogenesis but limits the interstitial flow proved crucial. Using this system, we find that luminal flow and resulting shear stress is a major regulator of the speed and morphogenesis of angiogenic sprouting, whose action is mediated through changes in vascular permeability.

Liver is responsible for the metabolization processes of up to 90 % of compounds and toxins in the body. Therefore liver-on-a-chip systems, as an in vitro promising cell culture platform, have great importance for fundamental science and drug development. In most of the liver-on-a-chip studies, seeding cells on both sides of a porous membrane, which represents the basement membrane, fail to resemble the native characteristics of biochemical, biophysical, and mechanical properties. In this study, polycarbonate (PC) and polyethylene terephthalate (PET) membranes were coated with gelatin to address this issue by accurately mimicking the native basement membrane present in the space of Disse. Various coating methods were used, including doctor blade, gel micro-injection, electrospinning, and spin coating. Spin coating was demonstrated to be the most effective technique owing to the ability to produce thin gel thickness with desirable surface roughness for cell interactions on both sides of the membrane. HepG2 and EA.HY926 cells were seeded on the upper and bottom sides of the gelatin-coated PET membrane and cultured on-chip for 7 days. Cell viability increased from 90 % to 95 %, while apoptotic index decreased. Albumin secretion notably rose between days 1-7 and 4-7, while GST-α secretion decreased from day 1 to day 7. In conclusion, the optimized spin coating process reported here can effectively modify the membranes to better mimic the native basement membrane niche characteristics.

Vasculature-on-chip (VoC) models have become a prominent tool in the study of microvasculature functions because of their cost-effective and ethical production process. These models typically use a hydrogel in which the three-dimensional (3D) microvascular structure is embedded. Thus, VoCs are directly impacted by the physical and chemical cues of the supporting hydrogel. Endothelial cell (EC) response in VoCs is critical, especially in organ-specific vasculature models, in which ECs exhibit specific traits and behaviors that vary between organs. Many studies customize the stimuli ECs perceive in different ways; however, customizing the hydrogel composition accordingly to the target organ's extracellular matrix (ECM), which we believe has great potential, has been rarely investigated. We explored this approach to organ-specific VoCs by fabricating microvessels (MVs) with either human umbilical vein ECs or human brain microvascular ECs in a 3D cylindrical VoC using a collagen hydrogel alone or one supplemented with laminin and hyaluronan, components found in the brain ECM. We characterized the physical properties of these hydrogels and analyzed the barrier properties of the MVs. Barrier function and tight junction (ZO-1) expression improved with the addition of laminin and hyaluronan in the composite hydrogel.

To evaluate the association between the pulmonary vein (PV) entry site morphology after total anomalous pulmonary vein repair (TAPVC) and postoperative pulmonary vein stenosis (PVS).

Computed tomography (CT) examination was performed to determine the PV entry site morphology. The width of the PV confluence was divided by the width of the left atrium (LA) to obtain the cPV/LA index. The cPV/LA index was compared between patients with and without postoperative PVS.

Fifty-one patients who had undergone CT after TAPVC repair were included, with a median cPV/LA index of 0.5 (interquartile range (IQR) = 0.349-0.654). Among them, 27 patients developed postoperative PVS. The median cPV/LA index after primary TAPVC repair was significantly lower in patients with PVS compared to those without PVS (0.367, IQR = 0.308-0.433 vs. 0.657, IQR = 0.571-0.783, P < 0.0001). Additionally, the cPV/LA index after surgical re-intervention for PVS was significantly smaller in patients who developed recurrent stenosis compared to those who remained free-from re-stenosis after surgical relief (0.459, IQR = 0.349-0.556; vs. 0.706, IQR = 0.628-0.810, P = 0.0045).

A small PV confluence width is associated with the development of postoperative PVS and recurrent stenosis after surgical relief of PVS. Our results suggest that adequate bilateral pulmonary vein lateralization during TAPVC surgery is crucial.

Peripheral artery disease (PAD) is the manifestation of atherosclerosis characterized by the accumulation of plaques in the arteries of the lower limbs. Interestingly, growing evidence suggests that the pathology of PAD is multifaceted and encompasses both vascular and skeletal muscle dysfunctions, which contributes to blunted physical capabilities and diminished quality of life. Importantly, it has been suggested that many of these pathological impairments may stem from blunted reduction-oxidation (redox) handling. Of note, in those with PAD, excessive production of reactive oxygen species (ROS) outweighs antioxidant capabilities resulting in oxidative damage, which may have systemic consequences. It has been suggested that antioxidant supplementation may be able to assist in handling ROS. However, the activation of various ROS production sites makes it difficult to determine the efficacy of these antioxidant supplements. Therefore, this review focuses on the common cellular mechanisms that facilitate ROS production and discusses how excessive ROS may impair vascular and skeletal muscle function in PAD. Furthermore, we provide insight for current and potential antioxidant therapies, specifically highlighting activation of the Kelch-like ECH-associated protein 1 (Keap1) - Nuclear Factor Erythroid 2-related factor 2 (Nrf2) pathway as a potential pharmacological therapy to combat ROS accumulation and aid in vascular function, and physical performance in patients with PAD. Altogether, this review provides a better understanding of excessive ROS in the pathophysiology of PAD and enhances our perception of potential therapeutic targets that may improve vascular function, skeletal muscle function, walking capacity, and quality of life in patients with PAD.

In recent years, research on organ-on-a-chip technology has been flourishing, particularly for drug screening and disease model development. Fibroblasts and vascular endothelial cells engage in crosstalk through paracrine signaling and direct cell-cell contact, which is essential for the normal development and function of the heart. Therefore, to faithfully recapitulate cardiac function, it is imperative to incorporate fibroblasts and vascular endothelial cells into a heart-on-a-chip model. Here, we report the development of a human heart-on-a-chip composed of induced pluripotent stem cell (iPSC)-derived cardiomyocytes, fibroblasts, and vascular endothelial cells. Vascular endothelial cells cultured on microfluidic channels responded to the flow of culture medium mimicking blood flow by orienting themselves parallel to the flow direction, akin to in vivo vascular alignment in response to blood flow. Furthermore, the flow of culture medium promoted integrity among vascular endothelial cells, as evidenced by CD31 staining and lower apparent permeability. The tri-culture condition of iPSC-derived cardiomyocytes, fibroblasts, and vascular endothelial cells resulted in higher expression of the ventricular cardiomyocyte marker IRX4 and increased contractility compared to the bi-culture condition with iPSC-derived cardiomyocytes and fibroblasts alone. Such tri-culture-derived cardiac tissues exhibited cardiac responses similar to in vivo hearts, including an increase in heart rate upon noradrenaline administration. In summary, we have achieved the development of a heart-on-a-chip composed of cardiomyocytes, fibroblasts, and vascular endothelial cells that mimics in vivo cardiac behavior.

Multiscale agent-based modeling frameworks have recently emerged as promising mechanobiological models to capture the interplay between biomechanical forces, cellular behavior, and molecular pathways underlying restenosis following percutaneous transluminal angioplasty (PTA). However, their applications are mainly limited to idealized scenarios. Herein, a multiscale agent-based modeling framework for investigating restenosis following PTA in a patient-specific superficial femoral artery (SFA) is proposed. The framework replicates the 2-month arterial wall remodeling in response to the PTA-induced injury and altered hemodynamics, by combining three modules: (i) the PTA module, consisting in a finite element structural mechanics simulation of PTA, featuring anisotropic hyperelastic material models coupled with a damage formulation for fibrous soft tissue and the element deletion strategy, providing the arterial wall damage and post-intervention configuration, (ii) the hemodynamics module, quantifying the post-intervention hemodynamics through computational fluid dynamics simulations, and (iii) the tissue remodeling module, based on an agent-based model of cellular dynamics. Two scenarios were explored, considering balloon expansion diameters of 5.2 and 6.2 mm. The framework captured PTA-induced arterial tissue lacerations and the post-PTA arterial wall remodeling. This remodeling process involved rapid cellular migration to the PTA-damaged regions, exacerbated cell proliferation and extracellular matrix production, resulting in lumen area reduction up to 1-month follow-up. After this initial reduction, the growth stabilized, due to the resolution of the inflammatory state and changes in hemodynamics. The similarity of the obtained results to clinical observations in treated SFAs suggests the potential of the framework for capturing patient-specific mechanobiological events occurring after PTA intervention.

In order to treat most vascular diseases, arterial grafts are commonly employed for replacing small-diameter vessels, yet they often cause thrombosis. The growth of endothelial cells along the interior surfaces of these grafts (substrates) is critical to mitigate thrombosis. Typically, endothelial cells are cultured inside these grafts under laminar flow conditions to emulate the native environment of blood vessels and produce an endothelium. Alternatively, the substrate structure could have a similar influence on endothelial cell behavior as laminar flow conditions. In this study, we investigated whether substrates with aligned fiber structures could induce responses in human umbilical vein endothelial cells (HUVECs) akin to those elicited by laminar flow. Our observations revealed that HUVECs on aligned substrates displayed significant morphological changes, aligning parallel to the fibers, similar to effects reported under laminar flow conditions. Conversely, HUVECs on random substrates maintained their characteristic cobblestone appearance. Notably, cell migration was more significant on aligned substrates. Also, we observed that while vWF expression was similar between both substrates, the HUVECs on aligned substrates showed more expression of platelet/endothelial cell adhesion molecule-1 (PECAM-1/CD31), laminin, and collagen IV. Additionally, these cells exhibited increased gene expression related to critical functions such as proliferation, extracellular matrix production, cytoskeletal reorganization, autophagy, and antithrombotic activity. These findings indicated that aligned substrates enhanced endothelial growth and behavior compared to random substrates. These improvements are similar to the beneficial effects of laminar flow on endothelial cells, which are well-documented compared to static or turbulent flow conditions.

The increasing prevalence of type 2 diabetes mellitus (T2DM) is a significant worldwide health concern caused by sedentary lifestyles and unhealthy diets. Beyond glycemic control, T2DM impacts multiple organ systems, leading to various complications. While traditionally associated with cardiovascular and microvascular complications, emerging evidence indicates significant effects on pulmonary health. Pulmonary vascular dysfunction and fibrosis, characterized by alterations in vascular tone and excessive extracellular matrix deposition, are increasingly recognized in individuals with T2DM. The onset of T2DM is often preceded by prediabetes, an intermediate hyperglycemic state that is associated with increased diabetes and cardiovascular disease risk. This review explores the relationship between T2DM, pulmonary vascular dysfunction and pulmonary fibrosis, with a focus on potential links with prediabetes. Pulmonary vascular function, including the roles of nitric oxide (NO), prostacyclin (PGI2), endothelin-1 (ET-1), thromboxane A2 (TxA2) and thrombospondin-1 (THBS1), is discussed in the context of T2DM and prediabetes. Mechanisms linking T2DM to pulmonary fibrosis, such as oxidative stress, dysregulated fibrotic signaling, and chronic inflammation, are explained. The impact of prediabetes on pulmonary health, including endothelial dysfunction, oxidative stress, and dysregulated vasoactive mediators, is highlighted. Early detection and intervention during the prediabetic stage may reduce respiratory complications associated with T2DM, emphasizing the importance of management strategies targeting blood glucose regulation and vascular health. More research that looks into the mechanisms underlying pulmonary complications in T2DM and prediabetes is needed.

Cell-free tissue-engineered vascular grafts provide a promising alternative to treat cardiovascular disease, but timely endothelialization is essential for ensuring patency and proper functioning post-implantation. Recent studies from our lab showed that blood cells like monocytes (MCs) and macrophages (Mϕ) may contribute directly to cellularization and regeneration of bioengineered arteries in small and large animal models. While MCs and Mϕ are leucocytes that are part of the innate immune response, they share common developmental origins with endothelial cells (ECs) and are known to play crucial roles during vessel formation (angiogenesis) and vessel repair after inflammation/injury. They are highly plastic cells that polarize into pro-inflammatory and anti-inflammatory phenotypes upon exposure to cytokines and differentiate into other cell types, including EC-like cells, in the presence of appropriate chemical and mechanical stimuli. This review focuses on the developmental origins of MCs and ECs; the role of MCs and Mϕ in vessel repair/regeneration during inflammation/injury; and the role of chemical signalling and mechanical forces in Mϕ inflammation that mediates vascular graft regeneration. We postulate that comprehensive understanding of these mechanisms will better inform the development of strategies to coax MCs/Mϕ into endothelializing the lumen and regenerate the smooth muscle layers of cell-free bioengineered arteries and veins that are designed to treat cardiovascular diseases and perhaps the native vasculature as well.

The purpose of this study is to investigate the morphometric properties of the internal carotid artery (ICA) by measuring the diameters and angles of its segments and exploring variations related to sex and the presence of aneurysms.

Digital subtraction angiography (DSA) images were utilized from 130 aneurysm patients and 75 non-aneurysm individuals to create 3D ICA models using 3D Slicer software. Segment diameters were measured via Autodesk Meshmixer 3.5.474 and angles were evaluated using ImageJ software.

In total, DSA images of 130 aneurysm patients and 75 individuals with normally reported carotid systems were evaluated. It was found that the intracranial aneurysms (IAs) were predominantly formed on the anterior cerebral artery (ACA) in males (%43), whereas in females IAs were frequently localized in the C6 segment (31.7%) and middle cerebral artery (MCA) (30.2%). In the control group, the evaluation of gender differences in segment diameters and angles revealed that males had significantly larger C4 and C5 segment diameters (4.62 vs. 4.32 mm and 4.41 vs. 4.09 mm, respectively) and a greater C6 angle (146.9° vs. 139.7°) compared to females. Comparisons between patients with an aneurysm at the anterior cerebral artery (ACA) and the control group revealed that the ACA group had wider diameters in the C1 (4.88 vs. 4.53 mm), C3 (4.65 vs. 4.4 mm), C5 (4.51 vs. 4.25 mm), and ACA (2.36 vs. 2.06 mm) segments. Additionally, the ACA group had wider angles in the ACA (104.1° vs. 94.1°) and C6 segments (147.7° vs. 143.3°), whereas the control group exhibited wider angles in the middle cerebral artery (MCA) segment (141.5° vs. 135.5°) compared to the ACA aneurysm group. Patients with anterior cerebral artery (ACA) aneurysms exhibited larger diameters in C1, C3, C5, C6, and ACA segments compared to the control group. Additionally, while the control group had larger MCA angle, patients with ACA aneurysms had larger angles in C6 segment and ACA.

Our results demonstrated that formation of aneurysms is affected by anatomical configuration of the ICA as well as sex characteristics, particularly regarding the ACA and MCA bifurcation angles, which showed associations with aneurysms in the respective branches.

Epigenetic changes, particularly histone compaction modifications, have emerged as critical regulators in the epigenetic pathway driving endothelial cell phenotype under constant exposure to laminar forces induced by blood flow. However, the underlying epigenetic mechanisms governing endothelial cell behavior in this context remain poorly understood. To address this knowledge gap, we conducted in vitro experiments using human umbilical vein endothelial cells subjected to various tensional forces simulating pathophysiological blood flow shear stress conditions, ranging from normotensive to hypertensive forces. Our study uncovers a noteworthy observation wherein endothelial cells exposed to high shear stress demonstrate a decrease in the epigenetic marks H3K4ac and H3K27ac, accompanied by significant alterations in the levels of HDAC (histone deacetylase) proteins. Moreover, we demonstrate a negative regulatory effect of increased shear stress on HOXA13 gene expression and a concomitant increase in the expression of the long noncoding RNA, HOTTIP, suggesting a direct association with the suppression of HOXA13. Collectively, these findings represent the first evidence of the role of histone-related epigenetic modifications in modulating chromatin compaction during mechanosignaling of endothelial cells in response to elevated shear stress forces. Additionally, our results highlight the importance of understanding the physiological role of HOXA13 in vascular biology and hypertensive patients, emphasizing the potential for developing small molecules to modulate its activity. These findings warrant further preclinical investigations and open new avenues for therapeutic interventions targeting epigenetic mechanisms in hypertensive conditions.

Endothelial cells (ECs) within the vascular system encounter fluid shear stress (FSS). High, laminar FSS promotes vasodilation and anti-inflammatory responses, whereas low or disturbed FSS induces dysfunction and inflammation. However, the adaptation of endothelial cells (ECs) to dynamically changing FSS patterns remains underexplored. Here, by combining traction force microscopy with a custom flow chamber, we examined human umbilical vein endothelial cells adapting their traction during transitions from short-term low shear to long-term high shear stress. We discovered that the initial low FSS elevates the traction by only half of the amount in response to direct high FSS even after flow changes to high FSS. However, in the long term under high FSS, the flow started with low FSS triggers a substantial second rise in traction for over 10 h. In contrast, the flow started directly with high FSS results in a quick traction surge followed by a huge reduction below the baseline traction in <30 min. Importantly, we find that the orientation of traction vectors is steered by initial shear exposure. Using Granger causality analysis, we show that the traction that aligns in the flow direction under direct high FSS functionally causes cell alignment toward the flow direction. However, EC traction that orients perpendicular to the flow that starts with temporary low FSS functionally causes cell orientation perpendicular to the flow. Taken together, our findings elucidate the significant influence of initial short-term low FSS on lasting changes in endothelial traction that induces EC alignment.NEW & NOTEWORTHY In our study, we uncover that preconditioning with low shear stress yields enduring impacts on endothelial cell traction and orientation, persisting even after transitioning to high-shear conditions. Using Granger causality analysis, we demonstrate a functional link between the direction of cell traction and subsequent cellular alignment across varying shear environments.

Hepatic physiology depends on the liver's complex structural composition which among others, provides high oxygen supply rates, locally differential oxygen tension, endothelial paracrine signaling, as well as residual hemodynamic shear stress to resident hepatocytes. While functional improvements were shown by implementing these factors into hepatic culture systems, direct cause-effect relationships are often not well characterized-obfuscating their individual contribution in more complex microphysiological systems. By comparing increasingly complex hepatic in vitro culture systems that gradually implement these parameters, we investigate the influence of the cellular microenvironment to overall hepatic functionality in pharmacological applications. Here, hepatocytes were modulated in terms of oxygen tension and supplementation, endothelial coculture, and exposure to fluid shear stress delineated from oxygen influx. Results from transcriptomic and metabolomic evaluation indicate that particularly oxygen supply rates are critical to enhance cellular functionality-with cellular drug metabolism remaining comparable to physiological conditions after prolonged static culture. Endothelial signaling was found to be a major contributor to differential phenotype formation known as metabolic zonation, indicated by WNT pathway activity. Lastly, oxygen-delineated shear stress was identified to direct cellular fate towards increased hepatic plasticity and regenerative phenotypes at the cost of drug metabolic functionality - in line with regenerative effects observed in vivo. With these results, we provide a systematic evaluation of critical parameters and their impact in hepatic systems. Given their adherence to physiological effects in vivo, this highlights the importance of their implementation in biomimetic devices, such as organ-on-a-chip systems. Considering recent advances in basic liver biology, direct translation of physiological structures into in vitro models is a promising strategy to expand the capabilities of pharmacological models.

Despite significant progress in human medicine, certain diseases remain challenging to promptly diagnose and treat. Hence, the imperative lies in the development of more exhaustive criteria and tools. Tissue and cellular mechanics exhibit distinctive traits in both normal and pathological states, suggesting that "force" represents a promising and distinctive target for disease diagnosis and treatment. Atomic force microscopy (AFM) holds great promise as a prospective clinical medical device due to its capability to concurrently assess surface morphology and mechanical characteristics of biological specimens within a physiological setting. This review presents a comprehensive examination of the operational principles of AFM and diverse mechanical models, focusing on its applications in investigating tissue and cellular mechanics associated with prevalent diseases. The findings from these studies lay a solid groundwork for potential clinical implementations of AFM. RESEARCH HIGHLIGHTS: By examining the surface morphology and assessing tissue and cellular mechanics of biological specimens in a physiological setting, AFM shows promise as a clinical device to diagnose and treat challenging diseases.

The cellular helical structure is well known for its crucial role in development and disease. Nevertheless, the underlying mechanism governing this phenomenon remains largely unexplored, particularly in recapitulating it in well-controlled engineering systems. Leveraging advanced microfluidics, we present compelling evidence of the spontaneous emergence of helical endothelial tubes exhibiting robust right-handedness governed by inherent cell chirality. To strengthen our findings, we identify a consistent bias toward the same chirality in mouse vascular tissues. Manipulating endothelial cell chirality using small-molecule drugs produces a dose-dependent reversal of the handedness in engineered vessels, accompanied by non-monotonic changes in vascular permeability. Moreover, our three-dimensional cell vertex model provides biomechanical insights into the chiral morphogenesis process, highlighting the role of cellular torque and tissue fluidity in its regulation. Our study unravels an intriguing mechanism underlying vascular chiral morphogenesis, shedding light on the broader implications and distinctive perspectives of tubulogenesis within biological systems.

As an effective cell assembly method, three-dimensional bioprinting has been widely used in building organ models and tissue repair over the past decade. However, different shear stresses induced throughout the entire printing process can cause complex impacts on cell integrity, including reducing cell viability, provoking morphological changes and altering cellular functionalities. The potential effects that may occur and the conditions under which these effects manifest are not clearly understood. Here, we review systematically how different mammalian cells respond under shear stress. We enumerate available experimental apparatus, and we categorise properties that can be affected under disparate stress patterns. We also summarise cell damaging mathematical models as a predicting reference for the design of bioprinting systems. We concluded that it is essential to quantify specific cell resistance to shear stress for the optimisation of bioprinting systems. Besides, as substantial positive impacts, including inducing cell alignment and promoting cell motility, can be generated by shear stress, we suggest that we find the proper range of shear stress and actively utilise its positive influences in the development of future systems.

Within the vascular system, endothelial cells (ECs) are exposed to fluid shear stress (FSS), a mechanical force exerted by blood flow that is critical for regulating cellular tension and maintaining vascular homeostasis. The way ECs react to FSS varies significantly; while high, laminar FSS supports vasodilation and suppresses inflammation, low or disturbed FSS can lead to endothelial dysfunction and increase the risk of cardiovascular diseases. Yet, the adaptation of ECs to dynamically varying FSS remains poorly understood. This study focuses on the dynamic responses of ECs to brief periods of low FSS, examining its impact on endothelial traction-a measure of cellular tension that plays a crucial role in how endothelial cells respond to mechanical stimuli. By integrating traction force microscopy (TFM) with a custom-built flow chamber, we analyzed how human umbilical vein endothelial cells (HUVECs) adjust their traction in response to shifts from low to high shear stress. We discovered that initial exposure to low FSS prompts a marked increase in traction force, which continues to rise over 10 hours before slowly decreasing. In contrast, immediate exposure to high FSS causes a quick spike in traction followed by a swift reduction, revealing distinct patterns of traction behavior under different shear conditions. Importantly, the direction of traction forces and the resulting cellular alignment under these conditions indicate that the initial shear experience dictates long-term endothelial behavior. Our findings shed light on the critical influence of short-lived low-shear stress experiences in shaping endothelial function, indicating that early exposure to low FSS results in enduring changes in endothelial contractility and alignment, with significant consequences for vascular health and the development of cardiovascular diseases.

The NLRP3 inflammasome is a crucial component of the innate immune system, playing a pivotal role in initiating and regulating the body's inflammatory response to various pathogens and cellular damage. Environmental stimuli, such as temperature, pH level, and nutrient availability, can influence the behavior and functions of innate immune cells, including immune cell activity, proliferation, and cytokine production. However, there is limited understanding regarding how mechanical forces, like shear stress, govern the intrinsic inflammatory reaction, particularly the activation of the NLRP3 inflammasome, and how shear stress impacts NLRP3 inflammasome activation through its capacity to induce alterations in gene expression and cytokine secretion. Here, we investigated how shear stress can act as a priming signal in NLRP3 inflammasome activation by exposing immortalized bone marrow-derived macrophages (iBMDMs) to numerous physiologically relevant magnitudes of shear stress before chemically inducing inflammasome activation. We demonstrated that shear stress of large magnitudes was able to prime iBMDMs more effectively for inflammasome activation compared to lower shear stress magnitudes, as quantified by the percentage of cells where ASC-CFP specks formed and IL-1β secretion, the hallmarks of inflammasome activation. Testing this in NLRP3 and caspase-1 knockout iBMDMs showed that the NLRP3 inflammasome was primarily primed for activation due to shear stress exposure. Quantitative polymerase chain reaction (qPCR) and a small-molecule inhibitor study mechanistically determined that shear stress regulates the NLRP3 inflammasome by upregulating Piezo1, IKKβ, and NLRP3. These findings offer insights into the mechanistic relationship among physiological shear stresses, inflammasome activation, and their impact on the progression of inflammatory diseases and their interconnected pathogenesis.