In Vitro Flow Chamber Design for the Study of Endothelial Cell (patho)physiology

2021 ◽  
Author(s):  
Meghan E Fallon ◽  
Rick Mathews ◽  
Monica T Hinds
Keyword(s):  
Shear Stress ◽  
Endothelial Cell ◽  
Complex Geometry ◽  
Flow Chamber ◽  
Arterial System ◽  
Design Parameters ◽  
Cell Phenotype ◽  
Local Geometry ◽  
Stress Environments

Abstract In the native vasculature, flowing blood produces a frictional force on vessel walls that directly effects endothelial cell phenotype and function. In the arterial system, the vasculature's local geometry directly influences variations in flow profiles and shear stress magnitudes. Straight arterial sections with pulsatile shear stress have been shown to promote an athero-protective endothelial phenotype. Conversely, areas with a more complex geometry, such as arterial bifurcations and branch points with disturbed flow patterns and a lower, oscillatory shear stress, typically lead to endothelial dysfunction and the pathogenesis of cardiovascular diseases. Many studies have investigated the regulation of endothelial responses to various shear stress environments. Importantly, the accurate in vitro simulation of in vivo hemodynamics is critical to the deeper understanding of mechano-transduction through the proper use and design of flow chamber devices. In this review, we describe several flow chamber apparatuses and their fluid mechanics design parameters, including parallel plate flow chambers, cone-and plate devices, and microfluidic devices. In addition, chamber-specific design criteria and relevant equations are defined in detail for the accurate simulation of shear stress environments to study endothelial cell responses

Abstract W P407: Shear Stress and Co-culture with Astrocytes Determine Brain Microvascular Endothelial Cell Phenotype

Stroke ◽  
2015 ◽  
Vol 46 (suppl_1) ◽  
Author(s):  
Sina Salehi Omran ◽  
Fernando Garcia Polite ◽  
Elazer Edelman ◽  
Mercedes Balcells-Camps
Keyword(s):  
Endothelial Cells ◽  
Shear Stress ◽  
Endothelial Cell ◽  
Immunocytochemical Staining ◽  
Microvascular Endothelial Cells ◽  
Cell Phenotype ◽  
Brain Microvascular Endothelial Cells ◽  
Glut 1 ◽  
Microvascular Endothelial

Introduction: Dementia has classically been identified to be of either vascular or neural origin. These domains overlap and are complementary, thus we consider dementia a disease of the single cerebrovascular unit. Our objective was to generate a modular platform for co-culture of brain microvascular endothelial cells and astrocytes that also incorporates mechanical stresses, and to then use this model of the cerebrovascular unit microenvironment to study the unit in vitro . Hypothesis: We assessed the hypothesis that endothelial health and blood-brain barrier integrity are modulated by shear stress and co-culture with astrocytes. Methods: We lithographed polydimethylsiloxane substrate on Teflon negative molds with subjacent rectangular channels of 0.45 and 2 mm depth for seeding of human brain microvascular cells and astrocytes, respectively, separated by a polytetrafluoroethylene (0.45 μm pore size) membrane under no flow or physiologic flow (6.2 dynes/cm 2 ) for one week. Immunocytochemical staining for glial fibrillary acidic protein and CD31 was simultaneously visualized by confocal microscopy. Cells from each channel were detached via trypsinization, and expression of transport proteins P-glycoprotein (P-gp) and glucose transporter-1 (GLUT-1), in addition to junction proteins zona occludens-1 (ZO-1) and CD31, was measured by Western Blot. Results: We stably co-cultured brain microvascular endothelial cells and astrocytes with no chamber leakage or mixing. CD31 staining revealed endothelial cell alignment to direction of flow. Expression of ZO-1 by endothelial cells increased in presence of flow and co-culture independently, by 1.6-fold in combined conditions relative to static monoculture (p<0.05). For P-gp, the increase in combined conditions was 5.5-fold (p<0.05). GLUT-1 and CD31 levels did not change significantly with co-culture or flow. Conclusion: Cell biology devoid of microenvironmental cues provides limited insight, especially when considering whole tissues, on the impact of disease. A co-culture system that introduces multiple cells, flow, controlled stress and independent visualization and sampling of each cell domain adds deeper understanding and greater value to in vitro biological models and tissue biology.

scholarly journals Turbulent fluid shear stress induces vascular endothelial cell turnover in vitro.

1986 ◽  
Vol 83 (7) ◽  
pp. 2114-2117 ◽  
Author(s):  
P. F. Davies ◽  
A. Remuzzi ◽  
E. J. Gordon ◽  
C. F. Dewey ◽  
M. A. Gimbrone
Keyword(s):  
Shear Stress ◽  
Endothelial Cell ◽  
Fluid Shear Stress ◽  
Vascular Endothelial Cell ◽  
Vascular Endothelial ◽  
Cell Turnover ◽  
Fluid Shear ◽  
Turbulent Fluid

Endothelial albumin permeability is shear dependent, time dependent, and reversible

1991 ◽  
Vol 260 (6) ◽  
pp. H1992-H1996 ◽  
Author(s):  
H. Jo ◽  
R. O. Dull ◽  
T. M. Hollis ◽  
J. M. Tarbell
Keyword(s):  
Shear Stress ◽  
Endothelial Cell ◽  
Fluorescein Isothiocyanate ◽  
Vascular Wall ◽  
Laminar Shear Stress ◽  
Aortic Endothelial Cells ◽  
Transendothelial Permeability ◽  
Steady Laminar Shear Stress ◽  
Laminar Shear

Altered permeability of vascular endothelium to macromolecules may play a role in vascular disease as well as vascular homeostasis. Because the shear stress of flowing blood on the vascular wall is known to influence many endothelial cell properties, an in vitro system to measure transendothelial permeability (Pe) to fluorescein isothiocyanate conjugated bovine serum albumin under defined physiological levels of steady laminar shear stress was developed. Bovine aortic endothelial cells grown on polycarbonate filters pretreated with gelatin and fibronectin constituted the model system. Onset of 1 dyn/cm2 shear stress resulted in a Pe rise from 5.1 +/- 1.3 x 10(-6) cm/s to 21.9 +/- 4.6 X 10(-6) cm/s at 60 min (n = 6); while 10 dyn/cm2 shear stress increased Pe from 4.8 +/- 1.5 X 10(-6) cm/s to 50.2 +/- 6.8 X 10(-6) cm/s at 30 min and 49.6 +/- 8.9 X 10(-6) cm/s at 60 (n = 9). Pe returned to preshear values within 120 and 60 min after removal of 1 and 10 dyn/cm2 shear stress, respectively. The data show that endothelial cell Pe in vitro is acutely sensitive to shear stress.

scholarly journals Modeling cellular crosstalk and organotypic vasculature development with human iPSC-derived endothelial cells and cardiomyocytes

2020 ◽  
Author(s):  
Emmi Helle ◽  
Minna Ampuja ◽  
Alexandra Dainis ◽  
Laura Antola ◽  
Elina Temmes ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Shear Stress ◽  
Endothelial Cell ◽  
Single Cell ◽  
Rna Sequencing ◽  
Tissue Growth ◽  
Cell Phenotype ◽  
Cell Populations ◽  
Pump System ◽  
Single Cell Rna Sequencing

AbstractRationaleCell-cell interactions are crucial for the development and function of the organs. Endothelial cells act as essential regulators of tissue growth and regeneration. In the heart, endothelial cells engage in delicate bidirectional communication with cardiomyocytes. The mechanisms and mediators of this crosstalk are still poorly known. Furthermore, endothelial cells in vivo are exposed to blood flow and their phenotype is greatly affected by shear stress.ObjectiveWe aimed to elucidate how cardiomyocytes regulate the development of organotypic phenotype in endothelial cells. In addition, the effects of flow-induced shear stress on endothelial cell phenotype were studied.Methods and resultsHuman induced pluripotent stem cell (hiPSC) -derived cardiomyocytes and endothelial cells were grown either as a monoculture or as a coculture. hiPS-endothelial cells were exposed to flow using the Ibidi-pump system. Single-cell RNA sequencing was performed to define cell populations and to uncover the effects on their transcriptomic phenotypes. The hiPS-cardiomyocyte differentiation resulted in two distinct populations; atrial and ventricular. Coculture had a more pronounced effect on hiPS-endothelial cells compared to hiPS-cardiomyocytes. Coculture increased hiPS-endothelial cell expression of transcripts related to vascular development and maturation, cardiac development, and the expression of cardiac endothelial cell -specific genes. Exposure to flow significantly reprogrammed the hiPS-endothelial cell transcriptome, and surprisingly, promoted the appearance of both venous and arterial clusters.ConclusionsSingle-cell RNA sequencing revealed distinct atrial and ventricular cell populations in hiPS-cardiomyocytes, and arterial and venous-like cell populations in flow exposed hiPS-endothelial cells. hiPS-endothelial cells acquired cardiac endothelial cell identity in coculture. Our study demonstrated that hiPS-cardiomoycytes and hiPS-endothelial cells readily adapt to coculture and flow in a consistent and relevant manner, indicating that the methods used represent improved physiological cell culturing conditions that potentially are more relevant in disease modelling. In addition, novel cardiomyocyte-endothelial cell crosstalk mediators were revealed.

Hemodynamic Shear Stress And Endothelial Cell Function: In Vitro Studies

1981 ◽  
Author(s):  
M A Gimbrone ◽  
C F Dewey ◽  
P F Davies ◽  
S R Bussolari
Keyword(s):  
Shear Stress ◽  
Endothelial Cell ◽  
Fluid Shear Stress ◽  
Cell Structure ◽  
Structure And Function ◽  
Cellular Migration ◽  
Shear Stresses ◽  
Endothelial Cell Function ◽  
And Function

The vascular endothelial lining in vivo is constantly subjected to hemodynamic shear stresses resulting from normal and altered patterns of blood flow. To facilitate the study of effects of fluid shear stress on endothelial cell structure and function, we have developed an in vitro system, utilizing a cone-plate apparatus, to subject coverslip cultures of bovine aortic endothelial cells (BAEC) to controlled levels of shear (up to 102 dynes/cm2) in either laminar or turbulent flow. The magnitude and direction of shear stress within the system are accurately known from both theory and experimental measurements. The data reported here are for laminar flow. Subconfluent BAEC cultures continuously exposed to 1-5 dynes/cm2 shear proliferated at a rate comparable to that of static cultures, and postconfluent monolayers appeared unaltered morphologically for up to 1 week. In contrast, BAEC cultures (both postconfluent and subconfluent) exposed to 8 dynes/cm2 developed dramatic, time-dependent morphological changes. By 48 hrs, cells uniformly assumed an ellipsoidal configuration, with their major axes aligned in the direction of flow. Exposure to >10 dynes/cm2 caused variable cell detachment from plain glass substrates. Cellular migration into linear “wounds”, created in confluent areas, was influenced by both the direction and amplitude of applied shear. Exposure to 8 dynes/ cm2 induced functional alterations, including increased fluid (bulk phase) endocytosis, prostaglandin production and platelet reactivity. These observations indicate that fluid mechanical forces can directly influence endothelial cell structure and function. Hemodynamic modulation of endothelial cell behavior may be relevant to normal vessel wall physiology, as well as the pathogenesis of atherosclerosis and thrombosis.

scholarly journals Atorvastatin Reduces the Proadhesive and Prothrombotic Endothelial Cell Phenotype Induced by Cocaine and Plasma From Cocaine Consumers In Vitro

2014 ◽  
Vol 34 (11) ◽  
pp. 2439-2448 ◽  
Author(s):  
Claudia G. Sáez ◽  
Karla Pereira-Flores ◽  
Roberto Ebensperger ◽  
Olga Panes ◽  
Teresa Massardo ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Cell Phenotype

Effect of shear stress on microvessel network formation of endothelial cells with in vitro three-dimensional model

2004 ◽  
Vol 287 (3) ◽  
pp. H994-H1002 ◽  
Author(s):  
Akinori Ueda ◽  
Masaki Koga ◽  
Mariko Ikeda ◽  
Susumu Kudo ◽  
Kazuo Tanishita
Keyword(s):  
Endothelial Cells ◽  
Shear Stress ◽  
Network Formation ◽  
Three Dimensional ◽  
Collagen Gel ◽  
Flow Chamber ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Collagen Gels

Shear stress stimulus is expected to enhance angiogenesis, the formation of microvessels. We determined the effect of shear stress stimulus on three-dimensional microvessel formation in vitro. Bovine pulmonary microvascular endothelial cells were seeded onto collagen gels with basic fibroblast growth factor to make a microvessel formation model. We observed this model in detail using phase-contrast microscopy, confocal laser scanning microscopy, and electron microscopy. The results show that cells invaded the collagen gel and reconstructed the tubular structures, containing a clearly defined lumen consisting of multiple cells. The model was placed in a parallel-plate flow chamber. A laminar shear stress of 0.3 Pa was applied to the surfaces of the cells for 48 h. Promotion of microvessel network formation was detectable after ∼10 h in the flow chamber. After 48 h, the length of networks exposed to shear stress was 6.17 (±0.59) times longer than at the initial state, whereas the length of networks not exposed to shear stress was only 3.30 (±0.41) times longer. The number of bifurcations and endpoints increased for networks exposed to shear stress, whereas the number of bifurcations alone increased for networks not exposed to shear stress. These results demonstrate that shear stress applied to the surfaces of endothelial cells on collagen gel promotes the growth of microvessel network formation in the gel and expands the network because of repeated bifurcation and elongation.

Computational Prediction of Fluid Induced Stress States in Dynamically Conditioned Engineered Heart Valve Tissues

2012 ◽  
Author(s):  
M. Salinas ◽  
D. Schmidt ◽  
R. Lange ◽  
M. Libera ◽  
S. Ramaswamy
Keyword(s):  
Shear Stress ◽  
Steady Flow ◽  
Heart Valve ◽  
Heart Valves ◽  
Critical Role ◽  
Computational Prediction ◽  
Mechanical Conditioning ◽  
Custom Made ◽  
Stress Environments

There is extensive documented evidence that mechanical conditioning plays a significant role in the development of tissue grown in-vitro for heart valve scaffolds [1–3]. Modern custom made bioreactors have been used to study the mechanobiology of engineered heart valve tissues [1]. Specifically fluid-induced shears stress patterns may play a critical role in up-regulating extracellular matrix secretion by progenitor cell sources such as bone marrow derived stem cells (BMSCs) [2] and increasing the possibility of cell differentiation towards a heart valve phenotype. We hypothesize that specific biomimetic fluid induced shear stress environments, particularly oscillatory shear stress (OSS), have significant effects on BMSCs phenotype and formation rates. As a first step here, we attempt to quantify and delineate the entire 3-D flow field by developing a CFD model to predict the fluid induced shear stress environments on engineered heart valves tissue under quasi-static steady flow and dynamic steady flow conditions.

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