Application of Microfluidic Technology to Evaluate the Influence of Pulsatile Blood Flow On Adhesion of Pediatric Sickle Erytrhocytes

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 3238-3238
Author(s):  
WHITE Jennell ◽  
Moira M. Lancelot ◽  
Patrick Hines ◽  
Sharada A. Sarnaik
Keyword(s):  
Blood Flow ◽  
Shear Stress ◽  
Sickle Cell ◽  
Pulsatile Flow ◽  
Contact Time ◽  
Adhesion Assay ◽  
Pulsatile Blood Flow ◽  
Flow Conditions ◽  
Adhesive Properties

Abstract Abstract 3238 Introduction: Sickle cell disease (SCD) is characterized by microvascular occlusion mediated in part by adhesion of sickle erythrocytes (SS RBCs) to the vasculature. Advanced flow adhesion (FA) technology facilitates SS RBC adhesion studies in conditions that simulate in vivo microvascular physiology. Most currently available FA systems measure SS RBC adhesion in non-pulsatile flow conditions, versus pulsatile blood flow conditions generated by the cardiac cycle. The influence of pulsatile blood flow on SS RBC adhesion may be particularly important in pediatric SS RBC adhesion, as children have a broad range of heart rates. This study compares SS RBC adhesion in non-pulsatile and pulsatile flow conditions, utilizing a commercially available, microfluidic FA system. Methods: Peripheral blood was obtained from patients with homozygous SCD (n=7) in steady state (5–18 years) from the Sickle Cell Center at the Children's Hospital of Michigan. FA assays were performed in non-pulsatile and pulsatile flow conditions, at a shear stress of 1.0 dyne/cm2, and increasing shear stress up to 20 dyne/cm2to assess avidity. A programmable control unit regulated pulse frequency, shear stress, and temperature. Adhesion was measured to immobilized human laminin and vascular cell adhesion molecule-1 (VCAM-1). A static adhesion assay was used to assess initrinsic adhesive properties of SS RBCs independent of flow dynamics. Results: Standard assays were performed with 30 mL of isolated SS RBCs (1× 107 cells/mL), and SS RBC adhesion was comparable to levels previously reported in parallel plate flow adhesion assays. FA assays showed that adhesion to both laminin and VCAM-1 was significantly increased in the context of pulsatile blood flow (1.67Hz) compared to non-pulsatile blood flow by 8-fold and 56-fold, respectively. The relationship of SS RBC adhesion to increasing pulse frequencies was variable from patient to patient, although adhesion to both laminin and VCAM-1 was uniformly greater in all pulse frequencies tested (1.0, 1.5, 1.67, and 2.0 Hz) compared to non-pulsatile blood flow. When avidity of adhesion was tested 78% of SS RBCs remained adhered to VCAM-1 at the maximum 20dyne/cm2 shear stress, whereas 6% of SS RBCs remained adhered to laminin at a shear stress of 20 dynes/cm2. Pulsatile adhesion to VCAM-1 and laminin was unaffected by protein kinase A (PKA) inhibition, although adhesion to laminin decreased by 31% in one of three patients. To determine if increased adhesion under pulse-flow conditions was due to increased contact time with the immobilized substrate versus a change in the SS RBC's intrinsic adhesive state, we measured SS RBC adhesion in a static adhesion assay following exposure to pulsatile versus non-pulsatile conditions. There was no significant difference in static adhesion to VCAM-1, however adhesion of pulse-exposed SS RBCs to laminin was more variable. Static adhesion of pulse-exposed SS RBCs to laminin was reduced by 60% in the presence of a PKA inhibitor. Conclusions: Our data demonstrate the application of a commercially available microfluidic flow adhesion assay system for efficient assessment of SS RBC adhesive properties. In the future, such advances may allow SS RBC adhesive properties to be evaluated clinically as a predictive tool for future vaso-occlusive events, and to predict individual patient response to anti-adhesive therapy. The small volume of blood required makes this system particularly attractive for studying pediatric samples. Additionally, our data demonstrate that adhesion to both an endothelial cell substrate (VCAM-1) and a subendothelial matrix substrate (laminin) is significantly influenced by the presence of pulsatile blood flow. Although PKA may play a minor role in pulsatile adhesion to laminin, increased contact time with immobilized laminin and VCAM-1 may be a greater contributor to increased adhesion under pulsatile conditions versus non-pulsatile conditions. Pediatric SS RBCs adhered to VCAM at higher levels and with more avidity compared to laminin. The pulsatile flow conditions described in this study more closely approximate in vivo microvascular conditions compared to non-pulsatile conditions commonly used to study SS RBC adhesion. Based on these differences in adhesion under pulsatile versus non-pulsatile flow, incorporating pulsatile flow in future adhesion studies may be more representative of in vivo conditions. Disclosures: No relevant conflicts of interest to declare.

The Effect of Asymmetry in Abdominal Aortic Aneurysms Under Physiologically Realistic Pulsatile Flow Conditions

2003 ◽  
Vol 125 (2) ◽  
pp. 207-217 ◽  
Author(s):  
E. A. Finol ◽  
K. Keyhani ◽  
C. H. Amon
Keyword(s):  
Blood Flow ◽  
Shear Stress ◽  
Wall Shear Stress ◽  
Pulsatile Flow ◽  
Reynolds Numbers ◽  
Secondary Flows ◽  
Aortic Aneurysms ◽  
Pulsatile Blood Flow ◽  
Abdominal Aortic ◽  
Wall Shear

In the abdominal segment of the human aorta under a patient’s average resting conditions, pulsatile blood flow exhibits complex laminar patterns with secondary flows induced by adjacent branches and irregular vessel geometries. The flow dynamics becomes more complex when there is a pathological condition that causes changes in the normal structural composition of the vessel wall, for example, in the presence of an aneurysm. This work examines the hemodynamics of pulsatile blood flow in hypothetical three-dimensional models of abdominal aortic aneurysms (AAAs). Numerical predictions of blood flow patterns and hemodynamic stresses in AAAs are performed in single-aneurysm, asymmetric, rigid wall models using the finite element method. We characterize pulsatile flow dynamics in AAAs for average resting conditions by means of identifying regions of disturbed flow and quantifying the disturbance by evaluating flow-induced stresses at the aneurysm wall, specifically wall pressure and wall shear stress. Physiologically realistic abdominal aortic blood flow is simulated under pulsatile conditions for the range of time-average Reynolds numbers 50⩽Rem⩽300, corresponding to a range of peak Reynolds numbers 262.5⩽Repeak⩽1575. The vortex dynamics induced by pulsatile flow in AAAs is depicted by a sequence of four different flow phases in one period of the cardiac pulse. Peak wall shear stress and peak wall pressure are reported as a function of the time-average Reynolds number and aneurysm asymmetry. The effect of asymmetry in hypothetically shaped AAAs is to increase the maximum wall shear stress at peak flow and to induce the appearance of secondary flows in late diastole.

Factors Influencing Leukocyte Adherence in Microvessels

1977 ◽  
Vol 38 (04) ◽  
pp. 0823-0830 ◽  
Author(s):  
Mayrovttz N. Harvey ◽  
Wiedeman P. Mary ◽  
Ronald F. Tuma
Keyword(s):  
Blood Flow ◽  
Shear Stress ◽  
Tissue Injury ◽  
Threshold Value ◽  
In Vivo Studies ◽  
Leukocyte Adherence ◽  
Rolling Velocity ◽  
Subsequent Behavior ◽  
Flow Stasis

SummaryIn vivo studies of the microcirculation of an untraumatized and unanesthetized animal preparation has shown that leukocyte adherence to vascular endothelium is an extremely rare occurrence. Induction of leukocyte adherence can be produced in a variety of ways including direct trauma to the vessels, remote tissue injury via laser irradiation, and denuding the epithelium overlying the observed vessels. The role of blood flow and local hemodynamics on the leukocyte adherence process is quite complex and still not fully understood. From the results reported it may be concluded that blood flow stasis will not produce leukocyte adherence but will augment pre-existing adherence. Studies using 2 quantitative measures of adherence, leukocyte flux and leukocyte velocity have shown these parameters to be affected differently by local hemodynamics. Initial adherence appears to be critically dependent on the magnitude of the blood shear stress at the vessel wall as evidenced by the lack of observable leukocyte flux above some threshold value. Subsequent behavior of the leukocytes as characterized by their average rolling velocity shows no apparent relationship to shear stress but, for low velocities, may be related to the linear blood velocity.

Preservation of sickle cell blood-flow patterns during MR imaging: an in vivo study

1988 ◽  
Vol 151 (1) ◽  
pp. 139-141 ◽  
Author(s):  
AS Brody ◽  
SH Embury ◽  
WC Mentzer ◽  
ML Winkler ◽  
CA Gooding
Keyword(s):  
Blood Flow ◽  
Mr Imaging ◽  
Sickle Cell ◽  
Flow Patterns ◽  
In Vivo Study ◽  
Blood Flow Patterns

scholarly journals Endothelial cell monolayer formation on a small-diameter vascular graft surface under pulsatile flow conditions

2021 ◽  
Vol 23 (3) ◽  
pp. 101-114
Author(s):  
M. Yu. Khanova ◽  
E. A. Velikanova ◽  
V. G. Matveeva ◽  
E. O. Krivkina ◽  
T. V. Glushkova ◽  
...  
Keyword(s):  
Shear Stress ◽  
Endothelial Cell ◽  
Pulsatile Flow ◽  
Vascular Graft ◽  
Cell Monolayer ◽  
Small Diameter ◽  
Flow Conditions ◽  
Endothelial Cell Culture ◽  
Monolayer Formation ◽  
Whole Transcriptome

Objective: to create a cell-populated small-diameter vascular graft (SDVG) using autologous endothelial cells and extracellular matrix proteins, and to evaluate the efficiency of endothelial cell monolayer formation during shear stress preconditioning in a SDVG.Materials and methods. PHBV/PCL tubular scaffolds of vascular grafts were made by electrospinning from a mixture of polyhydroxybutyrate-valerate (PHBV) copolymer and polycaprolactone (PCL) and modified with fibrin. To populate the graft, an endothelial cell culture was isolated from the blood of patients with coronary heart disease. Phenotyping of endothelial colony-forming cell (ECFC) culture was performed by flow cytometry and immunofluorescence microscopy. Cell proliferative and angiogenic activity were also studied. Cell-populated vascular scaffolds were cultured in a pulsatile flow setup with a final shear stress of 2.85 dyne/cm2. The effect of pulsatile flow on monolayer formation was assessed by immunofluorescence, scanning electron microscopy, atomic force microscopy, and whole-transcriptome RNA sequencing.Results. Under the influence of pulsatile flow, endothelial cells that were seeded into the tubular scaffold showed an increase in the expression level of endothelial profile proteins, focal adhesion and cytoskeleton. In contrast to endothelial cell culture on a vascular graft surface under static conditions, when cultured under pulsatile flow with 2.85 dyne/ cm2 shear stress, endothelial lining cells have an increased ability to adhere and are oriented along the pulsatile flow path. Whole-transcriptome RNA sequencing showed that induced shear stress increased expression levels of differentially expressed genes encoding proteins that ensure vascular development, endothelial integrity, and endothelial metabolism. A protocol for fabrication of a personalized cell-populated biodegradable SDVG under pulsatile flow conditions was developed.Conclusion. The use of autologous fibrin and ECFC culture, as well as shear stress preconditioning, allow to obtain a personalized cell-populated SDVG with continuous functional endothelial monolayer adapted to the flow.

scholarly journals In vivo blood flow abnormalities in the transgenic knockout sickle cell mouse

1999 ◽  
Vol 103 (6) ◽  
pp. 915-920 ◽  
Author(s):  
Stephen H. Embury ◽  
Narla Mohandas ◽  
Chris Paszty ◽  
Philip Cooper ◽  
Anthony T.W. Cheung
Keyword(s):  
Blood Flow ◽  
Sickle Cell

Pulsatile Flow in Fusiform Models of Abdominal Aortic Aneurysms: Flow Fields, Velocity Patterns and Flow-Induced Wall Stresses

2004 ◽  
Vol 126 (4) ◽  
pp. 438-446 ◽  
Author(s):  
Robert A. Peattie ◽  
Tiffany J. Riehle ◽  
Edward I. Bluth
Keyword(s):  
Shear Stress ◽  
Pulsatile Flow ◽  
Maximum Shear Stress ◽  
Wall Stress ◽  
Abdominal Aortic Aneurysms ◽  
Flow Fields ◽  
Aortic Aneurysms ◽  
Abdominal Aortic ◽  
Risk Of Rupture

As one important step in the investigation of the mechanical factors that lead to rupture of abdominal aortic aneurysms, flow fields and flow-induced wall stress distributions have been investigated in model aneurysms under pulsatile flow conditions simulating the in vivo aorta at rest. Vortex pattern emergence and evolution were evaluated, and conditions for flow stability were delineated. Systolic flow was found to be forward-directed throughout the bulge in all the models, regardless of size. Vortices appeared in the bulge initially during deceleration from systole, then expanded during the retrograde flow phase. The complexity of the vortex field depended strongly on bulge diameter. In every model, the maximum shear stress occurred at peak systole at the distal bulge end, with the greatest shear stress developing in a model corresponding to a 4.3 cm AAA in vivo. Although the smallest models exhibited stable flow throughout the cycle, flow in the larger models became increasingly unstable as bulge size increased, with strong amplification of instability in the distal half of the bulge. These data suggest that larger aneurysms in vivo may be subject to more frequent and intense turbulence than smaller aneurysms. Concomitantly, increased turbulence may contribute significantly to wall stress magnitude and thereby to risk of rupture.

Design and Analysis of a Shear Stress Sensor for Microcirculation Investigations (Invited Paper)

2003 ◽  
Author(s):  
Risa Robinson ◽  
Lynn Fuller ◽  
Harvey Palmer ◽  
Mary Frame
Keyword(s):  
Blood Flow ◽  
Shear Stress ◽  
Wall Shear Stress ◽  
Computational Technique ◽  
Flow Modification ◽  
Stress Sensors ◽  
Shear Stress Sensors ◽  
Wall Shear

Blood flow regulation in the microvascular network has been investigated by means of computational fluid dynamics, in vivo particle tracking and microchannel models. It is evident from these studies that shear stress along the wall is a key factor in the communication network that results in blood flow modification, yet current methods for shear stress determination are acknowledged to be imprecise. Micromachining technology allows for the development of implantable shear stress sensors that will enable us to monitor wall shear stress at multiple locations in arteriole bifurcations. In this study, a microchannel was employed as an in vitro model of a microvessel. Thermal shear stress sensors were used to mimic the endothelial cells that line the vessel wall. A three dimensional computational model was created to simulate the system’s thermal response to the constant temperature control circuit and related wall shear stress. The model geometry included a silicon wafer section with all the fabrication layers — silicon dioxide, poly silicon resistor, silicon nitride — and a microchannel with cross section 17 μm × 17 μm. This computational technique was used to optimize the dimensions of the system for a 0.01 Reynolds number flow at room temperature in order to reduce the amount of heat lost to the substrate and to predict and maximize the signal response. Results of the design optimization are presented and the fabrication process discussed.

Effects of GMI-1070, a Pan-Selectin Inhibitor, on Leukocyte Adhesion In Sickle Cell Disease: Results From a Phase 1/2 Study

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 262-262 ◽  
Author(s):  
Ted Wun ◽  
Laura M. De Castro ◽  
Lori Styles ◽  
Anthony Cheung ◽  
Shannon Chase ◽  
...  
Keyword(s):  
Clinical Trial ◽  
Blood Flow ◽  
Sickle Cell ◽  
Ex Vivo ◽  
Leukocyte Adhesion ◽  
Statistical Significance ◽  
Phase 1 ◽  
Microvascular Blood Flow ◽  
Bulbar Conjunctiva

Abstract Abstract 262 Adhesion molecules are critically involved in the pathophysiology of sickle cell disease (SCD). A growing body of evidence from animal models and humans supports the role of selectin-mediated cell adhesion in the pathophysiology of vaso-occlusion. Leukocyte adhesion has been demonstrated to reduce microvascular blood flow in a sickle cell mouse model, and there is evidence to suggest leukocyte adhesion in humans contributes to vaso-occlusion (Stuart et al, Lancet 2004; Turhan et al, PNAS 2002). GMI-1070 is a pan-selectin inhibitor that targets E-, P-, and L-selectins and has previously been shown to restore blood flow and improve survival in a mouse model of vaso-occlusion (Chang et al, Blood 2010). Also, it is potent inhibitor of adhesion of human neutrophils to immobilized E-selectin and ICAM-1 under flow conditions in vitro. As part of a Phase 1/2 study of GMI-1070 in patients with SCD we determined the effects of the drug on biomarkers of adhesion in vivo and ex vivo. Methods: An open-label phase 1/2 study of the safety, PK, and activity of GMI-1070 was performed, enrolling adults with SCD at steady state. Here we are reporting data on GMI-1070 anti-adhesion activity. GMI-1070 was administered in two IV doses given on the same day: 20 mg/kg as a loading dose, followed 10 hours later by 10 mg/kg. Serial WBC count with differential (all 15 subjects), computer-assisted intravital microscopy (CAIM) (4 subjects), and ex vivo activity of GMI-1070 in plasma (4 subjects) were measured. CAIM is a non-invasive technique for quantitative measurement of microvascular blood flow in vessels of the bulbar conjunctiva. In this study it was used to measure RBC velocity before and after dosing (at 30 minutes, 2, 4, 8, and 24 hours) with GMI-1070. Ex vivo evaluation of plasma GMI-1070 activity in a cell adhesion assay was also performed, with samples taken at 0, 8, 24, and 48 hours after first infusion. Results: Adults were enrolled at three centers: 13 with HbSS, 2 with HbSB0thal. All were African-American, and 9 were male. All subjects received both doses of study drug. The t1/2 was 7.7 hours. Mean baseline WBC was 10 K/mm3, and baseline absolute neutrophil count (ANC) was 5.4 K/mm3. The mean WBC was 10.4, 11.6, 10.2, and 8.7 K/mm3 at 8, 24, 48 hours and 7 days, respectively. The ANC mean was 5.5, 7.5, 5.6, and 4.2 K/mm3 at the same time points. ANC % change from baseline was significant at 24 and 48 hours (p=0.001, 0.025 mixed effects model) (Figure). There were no significant changes in absolute monocyte or lymphocyte counts. There was no correlation with any clinical adverse events. In one subject, the WBC rose from 10.4 to a peak of 28, with no clinical symptoms or significant changes in other lab values. CAIM (n=4) evaluating microvascular blood flow in the bulbar conjunctiva (measured in screen pixels/sec), showed mean RBC velocity at baseline was 335 (SD 70) pixels/sec, with mean values at 30 min., 2, 4, and 8 hours of 368 (59), 345 (63), 341 (59), and 338 (83) pixels/sec respectively, and returned to baseline of 317 (82) pixels/sec at 24 h. These differences did not reach statistical significance (mixed effects model). In ex vivo evaluation of neutrophil adhesion to matrix proteins (as quantified by # of bound neutrophils per 50× field) from samples 0, 4, 8, 24, and 48 hours after the first infusion revealed reduction in adhesions at 4 and 8 hours; mean adhered neutrophils were 33 (17), 20 (11), 18 (9), 47 (49), and 42 (15) respectively. However, these differences did not reach statistical significance. In conclusion, GMI-1070 infusion resulted in neutrophilia, a trend towards increased RBC velocity in the vessels of the bulbar conjunctiva immediately after infusion, and reduced leukoctye adhesion in an ex vivo assay despite neutrophilia. All of these findings are consistent with an anti-adhesive effect on leukocyte adhesion in vivo and suggest that the findings in sickle mouse studies can be translated into SCD patients. This study supports further evaluation of GMI-1070 for the treatment of vaso-occlusive episodes in SCD.Figure:Observed (mean/SE) absolute WBC and ANC over timeFigure:. Observed (mean/SE) absolute WBC and ANC over time Disclosures: Wun: GlycoMimetics: Clinical Trial Sponsorship, Consultancy; Eli Lilly: Clinical Trial Sponsorship, Consultancy. Off Label Use: This drug (GMI-1070) has not been approved for any clinical indication. De Castro:GlycoMimetics: clinical trial sponsorship. Styles:GlycoMimetics: Clinical Trial Sponsorship, Consultancy. Cheung:GlycoMimetics: clinical trial sponsorship. Chase:GlycoMimetics: clinical trial sponsorship. Simon:GlycoMimetics: Research Funding. Magnani:GlycoMimetics: Employment, Equity Ownership. Thackray:GlycoMimetics: Employment, Equity Ownership.

Pulsatile blood flow in the entire coronary arterial tree: theory and experiment

2006 ◽  
Vol 291 (3) ◽  
pp. H1074-H1087 ◽  
Author(s):  
Yunlong Huo ◽  
Ghassan S. Kassab
Keyword(s):  
Blood Flow ◽  
Pulsatile Flow ◽  
Pulse Wave ◽  
Coronary Circulation ◽  
Low Frequency ◽  
Flow Distribution ◽  
Pulsatile Blood Flow ◽  
Arterial Tree ◽  
Cross Section Area ◽  
Theory And Experiment

The pulsatility of coronary circulation can be accurately simulated on the basis of the measured branching pattern, vascular geometry, and material properties of the coronary vasculature. A Womersley-type mathematical model is developed to analyze pulsatile blood flow in diastole in the absence of vessel tone in the entire coronary arterial tree on the basis of previously measured morphometric data. The model incorporates a constitutive equation of pressure and cross-section area relation based on our previous experimental data. The formulation enables the prediction of the impedance, the pressure distribution, and the pulsatile flow distribution throughout the entire coronary arterial tree. The model is validated by experimental measurements in six diastolic arrested, vasodilated porcine hearts. The agreement between theory and experiment is excellent. Furthermore, the present pulse wave results at low frequency agree very well with previously published steady-state model. Finally, the phase angle of flow is seen to decrease along the trunk of the major coronary artery and primary branches toward the capillary vessels. This study represents the first, most extensive validated analysis of Womersley-type pulse wave transmission in the entire coronary arterial tree down to the first segment of capillaries. The present model will serve to quantitatively test various hypotheses in the coronary circulation under pulsatile flow conditions.

scholarly journals CD44 regulates blood-brain barrier integrity in response to fluid shear stress

2020 ◽  
Author(s):  
Brandon J. DeOre ◽  
Paul P. Partyka ◽  
Fan Fan ◽  
Peter A. Galie
Keyword(s):  
Blood Flow ◽  
Shear Stress ◽  
Blood Brain Barrier ◽  
Fluid Shear Stress ◽  
Small Gtpase ◽  
Brain Barrier ◽  
Fluid Shear ◽  
Blood Brain

AbstractFluid shear stress is an important mediator of vascular permeability, yet the molecular mechanisms underlying the response of the blood-brain barrier to shear have yet to be studied in cerebral vasculature despite its importance for brain homeostasis. The goal of this study is to probe components of shear mechanotransduction within the blood-brain barrier to gain a better understanding of pathologies associated with changes in cerebral blood flow including ischemic stroke. Interrogating the effects of shear stress in vivo is complicated by the complexity of factors in the brain parenchyma and the difficulty associated with modulating blood flow regimes. Recent advances in the ability to mimic the in vivo microenvironment using three-dimensional in vitro models provide a controlled setting to study the response of the blood-brain barrier to shear stress. The in vitro model used in this study is compatible with real-time measurement of barrier function using transendothelial electrical resistance as well as immunocytochemistry and dextran permeability assays. These experiments reveal that there is a threshold level of shear stress required for barrier formation and that the composition of the extracellular matrix, specifically the presence of hyaluronan, dictates the flow response. Gene editing to modulate the expression of CD44, a receptor for hyaluronan that previous studies have identified to be mechanosensitive, demonstrates that the receptor is required for the endothelial response to shear stress. Manipulation of small GTPase activity reveals CD44 activates Rac1 while inhibiting RhoA activation. Additionally, adducin-γ localizes to tight junctions in response to shear stress and RhoA inhibition and is required to maintain the barrier. This study identifies specific components of the mechanosensing complex associated with the blood-brain barrier response to fluid shear stress, and therefore illuminates potential targets for barrier manipulation in vivo.

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