Coronary stent implantation changes 3-D vessel geometry and 3-D shear stress distribution

Mechanisms of in-stent restenosis are not fully understood. Shear stress is known to play a role in plaque and thrombus formation and is sensitive to changes in regional vessel geometry. Hence, we evaluated the regional changes in 3-D geometry and shear stress induced by stent placement in coronary arteries of pigs.Methods. 3-D reconstruction was performed, applying a combined angiographic and IVUS technique (ANGUS), from seven Wallstents (diameter 3.5 (n=3) and 5mm (n=4)), which were implanted in seven coronary arteries of five pigs. This 3-D geometry was used to calculate locally the curvature, while the shear stress distribution was obtained by computational fluid dynamics. Local changes in shear stress were obtained at the entrance and exit of the stent for baseline (0. 65+/-0.22 ml/s) and hyperemic flow (2.60+/-0.86 ml/s) conditions. Results. After stent implantation, the curvature increased by 121% at the entrance and by 100% at the exit of the stent, resulting in local changes in shear stress. In general, at the entrance of the stent local maxima in shear stress were generated, while at the exit both local maxima and minima in shear stress were observed (p<0.05). Additionally, the shear stress at the entrance and exit of the stent were correlated with the local curvature (r: 0.30-0.84).Conclusion. Stent implantation changes 3-D vessel geometry in such a way that regions with decreased and increased shear stress occur close to the stent edges. These changes might be related to the asymmetric patterns of in-stent restenosis.     

Intracoronary radiation with gamma wire inhibits recurrent in-stent restenosis

To study the safety and efficacy of intracoronary gamma radiation delivered via a new high-activity (192)Ir source wire for the treatment of in-stent restenosis. In-stent restenosis results from neointimal tissue proliferation especially in its diffused form and presents a therapeutic challenge. Gamma radiation has been shown to decrease neointima formation within stents in animal models and in initial clinical trials. A total of 26 patients with in-stent restenosis underwent successful intervention and was treated with open-label (192)Ir using a high-activity line source. The specific activity of the source wire was 372+/-51 mCi, and the dwell time was 10.8+/-1.9 min. Primary endpoints were freedom from death, myocardial infraction (MI), and repeat target lesion revascularization (TLR) at 6 months. Secondary endpoints included angiographic restenosis and intravascular ultrasound (IVUS) neointimal hyperplasia. Procedural success was high (96.2%), and in-hospital and 30-day complications were low with no deaths, MI, or requirement for repeat revascularization. At 6 months, event-free survival was 85%: one patient required repeat PTCA, one underwent bypass surgery, and two had an MI. Baseline lesion length measured 15.77 mm. Follow-up angiography was available in 21/25 (84%) patients. The binary restenosis rates were 19.0% (4/21) in-stent and 23.8% (5/21) in-lesion. Follow-up IVUS was available in 20/25 patients. There was no increase in intimal hyperplasia from postintervention to follow-up (3.11.8 vs. 3.41.8 mm(2); P=.32). Eight patients had a reduction of neointimal intimal tissue at follow-up. These results indicate that intracoronary gamma radiation with the Angiorad source wire is safe and effective in preventing in-stent restenosis.     

Inhibition of in-stent restenosis by oral copper chelation in porcine coronary arteries

Stress-induced release of IL-1alpha and fibroblast growth factor-1 is dependent on intracellular copper and is a major driver of neointimal hyperplasia. Therefore, we assessed the effect of tetrathiomolybdate (TTM), a clinically proven copper chelator, on in-stent restenosis. Nine pigs were treated with TTM (5 mg/kg po) twice daily for 2 wk before stent implantation and for 4 wk thereafter, and nine pigs served as controls. In-stent restenosis was assessed by quantitative coronary angiography (QCA), intravascular ultrasound (IVUS), and histomorphometry. Serum ceruloplasmin activity was used as a surrogate marker of copper bioavailability. In TTM-treated animals, ceruloplasmin dropped 70 +/- 10% below baseline levels. Baseline characteristics were comparable in TTM-treated and control animals. At 4-wk follow-up, all parameters relevant to in-stent restenosis were significantly reduced in TTM-treated animals: minimal lumen diameter by QCA was 2.03 +/- 0.57 and 1.47 +/- 0.45 mm in TTM-treated and control animals, respectively (P < 0.05), percent stenosis diameter was 39% less in TTM-treated animals (27.1 +/- 16.6% vs. 44.5 +/- 16.1%, P < 0.05), minimal lumen area by IVUS was 60% larger in TTM-treated animals (4.27 +/- 1.56 vs. 2.67 +/- 1.19 mm(2), P < 0.05), and neointimal volume by histomorphometry was 37% less in TTM-treated animals (34.9 +/- 11.5 vs. 55.2 +/- 19.6 mm(3), P < 0.05). We conclude that systemic copper chelation with a clinically approved chelator significantly inhibits in-stent restenosis.     

3D reconstruction techniques of human coronary bifurcations for shear stress computations

Background

Heterogeneity in plaque composition in human coronary artery bifurcations is associated with blood flow induced shear stress. Shear stress is generally determined by combing 3D lumen data and computational fluid dynamics (CFD). We investigated two new procedures to generate 3D lumen reconstructions of coronary artery bifurcations for shear stress computations.

Methods

We imaged 10 patients with multislice computer tomography (MSCT) and intravascular ultrasound (IVUS). The 3D reconstruction of the main branch was based on the fusion of MSCT and IVUS. The proximal part of side branch was reconstructed using IVUS data or MSCT data, resulting in two different reconstructions of the bifurcation region. The distal part of the side branch was based on MSCT data alone. The reconstructed lumen was combined with CFD to determine the shear stress. Low and high shear stress regions were defined and shear stress patterns in the bifurcation regions were investigated.

Results

The 3D coronary bifurcations were successfully generated with both reconstruction procedures. The geometrical features of the bifurcation region for the two reconstruction procedures did not reveal appreciable differences. The shear stress maps showed a qualitative agreement, and the low and high shear stress regions were similar in size and average shear stress values were identical. The low and high shear stress regions showed an overlap of approximately 75%.

Conclusion

Reconstruction of the side branch with MSCT data alone is an adequate technique to study shear stress and wall thickness in the bifurcation region. The reconstruction procedure can be applied to further investigate the effect of shear stress on atherosclerosis in coronary bifurcations.     

Axial stent strut angle influences wall shear stress after stent implantation: analysis using 3D computational fluid dynamics models of stent foreshortening

Introduction  

The success of vascular stents in the restoration of blood flow is limited by restenosis. Recent data generated from computational fluid dynamics (CFD) models suggest that the vascular geometry created by an implanted stent causes local alterations in wall shear stress (WSS) that are associated with neointimal hyperplasia (NH). Foreshortening is a potential limitation of stent design that may affect stent performance and the rate of restenosis. The angle created between axially aligned stent struts and the principal direction of blood flow varies with the degree to which the stent foreshortens after implantation.     

The effect of clopidogrel on apoptosis--an in vivo study

Clopidogrel is widely used in cardiovascular medicine, and is believed to play an important role in the pathogenesis of many cardiovascular disease processes. In particular, patients undergoing coronary stenting, who are commonly treated with clopidogrel, are candidates for in-stent restenosis. This is mainly caused by neointimal hyperplasia, so it is important to consider whether clopidogrel affects neointimal hyperplasia via apoptosis. Lymphocytes, especially T-cells, are known to play a key role in the initiation and formation of atherosclerotic plaques. The aim of this study was to investigate the effect of clopidogrel on human lymphocyte apoptosis, using a DNA fragmentation assay.     

Coronary Injury Score Correlates with Proliferating Cells and Alpha-Smooth Muscle Actin Expression in Stented Porcine Coronary Arteries

Neointimal formation and cell proliferation resulting into in-stent restenosis is a major pathophysiological event following the deployment of stents in the coronary arteries. In this study, we assessed the degree of injury, based on damage to internal elastic lamina, media, external elastic lamina, and adventitia following the intravascular stenting, and its relationship with the degree of smooth muscle cell proliferation. We examined the smooth muscle cell proliferation and their phenotype at different levels of stent injury in the coronary arteries of domestic swine fed a normal swine diet. Five weeks after stent implantation, swine with and without stents were euthanized and coronaries were excised. Arteries were embedded in methyl methacrylate and sections were stained with H&E, trichrome, and Movat’s pentachrome. The expression of Ki67, α-smooth muscle actin (SMA), vimentin, and HMGB1 was evaluated by immunofluorescence. There was a positive correlation between percent area stenosis and injury score. The distribution of SMA and vimentin was correlated with the degree of arterial injury such that arteries that had an injury score >2 did not have immunoreactivity to SMA in the neointimal cells near the stent struts, but these neointimal cells were positive for vimentin, suggesting a change in the smooth muscle cell phenotype. The Ki67 and HMGB1 immunoreactivity was highly correlated with the fragmentation of the IEL and injury in the tunica media. Thus, the extent of coronary arterial injury during interventional procedure will dictate the degree of neointimal hyperplasia, in-stent restenosis, and smooth muscle cell phenotype.     

Developing pulsatile flow in a deployed coronary stent

A major consequence of stent implantation is restenosis that occurs due to neointimal formation. This patho-physiologic process of tissue growth may not be completely eliminated. Recent evidence suggests that there are several factors such as geometry and size of vessel, and stent design that alter hemodynamic parameters, including local wall shear stress distributions, all of which influence the restenosis process. The present three-dimensional analysis of developing pulsatile flow in a deployed coronary stent quantifies hemodynamic parameters and illustrates the changes in local wall shear stress distributions and their impact on restenosis. The present model evaluates the effect of entrance flow, where the stent is placed at the entrance region of a branched coronary artery. Stent geometry showed a complex three-dimensional variation of wall shear stress distributions within the stented region. Higher order of magnitude of wall shear stress of 530 dyn/cm2 is observed on the surface of cross-link intersections at the entrance of the stent. A low positive wall shear stress of 10 dyn/cm2 and a negative wall shear stress of -10 dyn/cm2 are seen at the immediate upstream and downstream regions of strut intersections, respectively. Modified oscillatory shear index is calculated which showed persistent recirculation at the downstream region of each strut intersection. The portions of the vessel where there is low and negative wall shear stress may represent locations of thrombus formation and platelet accumulation. The present results indicate that the immediate downstream regions of strut intersections are areas highly susceptible to restenosis, whereas a high shear stress at the strut intersection may cause platelet activation and free emboli formation.     

Circumferential vascular deformation after stent implantation alters wall shear stress evaluated with time-dependent 3D computational fluid dynamics models.

The success of vascular stents in the restoration of blood flow is limited by restenosis. Recent data generated from computational fluid dynamics (CFD) models suggest that stent geometry may cause local alterations in wall shear stress (WSS) that have been associated with neointimal hyperplasia and subsequent restenosis. However, previous CFD studies have ignored histological evidence of vascular straightening between circumferential stent struts. We tested the hypothesis that consideration of stent-induced vascular deformation may more accurately predict alterations in indexes of WSS that may subsequently account for histological findings after stenting. We further tested the hypothesis that the severity of these alterations in WSS varies with the degree of vascular deformation after implantation. Steady-state and time-dependent simulations of three-dimensional CFD arteries based on canine coronary artery measurements of diameter and blood flow were conducted, and WSS and WSS gradients were calculated. Circumferential straightening introduced areas of high WSS between stent struts that were absent in stented vessels of circular cross section. The area of vessel exposed to low WSS was dependent on the degree of circumferential vascular deformation and axial location within the stent. Stents with four vs. eight struts increased the intrastrut area of low WSS in vessels, regardless of cross-sectional geometry. Elevated WSS gradients were also observed between struts in vessels with polygonal cross sections. The results obtained using three-dimensional CFD models suggest that changes in vascular geometry after stent implantation are important determinants of WSS distributions that may be associated with subsequent neointimal hyperplasia.     

内皮祖细胞在支架术后再内皮化中应用的研究进展

经皮冠状动脉介入治疗的应用改善了冠心病患者的临床症状及预后,但现在困扰人们的问题是作为其术后并发症之一的支架内再狭窄发病率仍然很高。大量的研究证实,内膜增生在支架内再狭窄的形成中起主导作用,所以提高受损内膜再内皮化的速度是防止支架内再狭窄的一个重要措施。新近的研究表明,内皮祖细胞能参与损伤后血管内皮修复,促进受损血管内膜的再内皮化,因此,在防止支架内再狭窄中将得到进一步的研究与应用。因此,本文就内皮祖细胞在支架术后再内皮化中应用的研究进展做一综述。     

Pathology of in-stent restenosis

The process of in-stent restenosis parallels wound healing responses. Stent deployment results in early thrombus deposition and acute inflammation, granulation tissue development, and ultimately smooth muscle cell proliferation and extracellular matrix synthesis. The severity of arterial injury during stent placement correlates with increased inflammation and late neointimal growth. These pathological findings provide useful targets for therapies aimed at reducing the incidence of in-stent restenosis.     

Wall shear stresses remain elevated in mature arteriovenous fistulas: a case study

Maintaining vascular access (VA) patency continues to be the greatest challenge for dialysis patients. VA dysfunction, primarily due to venous neointimal hyperplasia development and stenotic lesion formation, is mainly attributed to complex hemodynamics within the arteriovenous fistula (AVF). The effect of VA creation and the subsequent geometrical remodeling on the hemodynamics and shear forces within a mature patient-specific AVF is investigated. A 3D reconstructed geometry of a healthy vein and a fully mature patient-specific AVF was developed from a series of 2D magnetic resonance image scans. A previously validated thresholding technique for region segmentation and lumen cross section contour creation was conducted in MIMICS 10.01, allowing for the creation of a 3D reconstructed geometry. The healthy vein and AVF computational models were built, subdivided, and meshed in GAMBIT 2.3. The computational fluid dynamic (CFD) code FLUENT 6.3.2 (Fluent Inc., Lebanon, NH) was employed as the finite volume solver to determine the hemodynamics and shear forces within the healthy vein and patient-specific AVF. Geometrical alterations were evaluated and a CFD analysis was conducted. Substantial geometrical remodeling was observed, following VA creation with an increase in cross-sectional area, out of plane curvature (maximum angle of curvature in AVF=30?deg), and angle of blood flow entry. The mean flow velocity entering the vein of the AVF is dramatically increased. These factors result in complex three-dimensional hemodynamics within VA junction (VAJ) and efferent vein of the AVF. Complex flow patterns were observed and the maximum and mean wall shear stress (WSS) magnitudes are significantly elevated. Flow reversal was found within the VAJ and efferent vein. Extensive geometrical remodeling during AVF maturation does not restore physiological hemodynamics to the VAJ and venous conduit of the AVF, and high WSS and WSS gradients, and flow reversal persist. It is theorized that the vessel remodelling and the continued non-physiological hemodynamics within the AVF compound to result in stenotic lesion development.     

Inhibition of interleukin-1β convertase is associated with decrease of neointimal hyperplasia after coronary artery stenting in pigs

Inhibition of IL-1 convertase has been shown to decrease inflammation and apoptosis, which are features of the neointimal development after vascular interventions. The aim of our study was to reduce neointimal proliferation after stenting of the porcine coronary artery, using the irreversible IL-1 convertase and caspase-1 inhibitor acetyl-tyrosinyl-valyl-alanyl-aspartyl-chloromethyl-ketone (Ac-YVAD-cmk). Before coronary stent implantation, 8 pigs received an intracoronary infusion of 50 mg Ac-YVAD-cmk into the left coronary artery (group 1, n - 8), while 8 animals served as untreated controls (group 2). After 4 weeks, coronary angiography and intracoronary ultrasound (IVUS) with 3D measurements were performed. IVUS revealed a smaller in-stent intimal volume (27.3 ± 11.6 vs. 75.8 ± 18.4 mm³, p < 0.005) and a decreased maximal percentage area stenosis (36.1 ± 8.5 vs. 69.0 ± 8.2%, p < 0.001) in group 1 vs. group 2. A smaller maximal neointimal thickness (0.63 ± 0.28 vs. 1.75 ± 0.94 mm, p < 0.005) and a decreased maximal neointimal area (2.14 ± 1.29 vs. 5.03 ± 1.92 mm², p < 0.005), assessed by computerized planimetry, were found in group 1 vs. group 2. Lower apoptotic indices of the neointimal cells were observed in the treated animals (3.0 vs. 13.4% of total intimal cells, p < 0.05). The coronary arterial tissue IL-1 level was significantly decreased in the animals treated with Ac-YVAD-cmk (0.254 ± 0.162 vs. 0.463 ± 0.307 pg/mg protein, p < 0.05), and exhibited a positive linear correlation (r = 0.581, p = 0.013) with the in-stent plaque volume. In conclusion, intracoronary administration of Ac-YVAD-cmk before coronary artery stenting results in significantly decreased neointimal hyperplasia due to the inhibition of local IL-1 production and decreased neointimal apoptosis.     

The apolipoprotein A‐I mimetic peptide,D‐4F,restrains neointimal formation through heme oxygenase‐1 up‐regulation

D‐4F, an apolipoprotein A‐I (apoA‐I) mimetic peptide, possesses distinctly anti‐atherogenic effects. However, the biological functions and mechanisms of D‐4F on the hyperplasia of vascular smooth muscle cells (VSMCs) remain unclear. This study aimed to determine its roles in the proliferation and migration of VSMCs. In vitro, D‐4F inhibited VSMC proliferation and migration induced by ox‐LDL in a dose‐dependent manner. D‐4F up‐regulated heme oxygenase‐1 (HO‐1) expression in VSMCs, and the PI3K/Akt/AMP‐activated protein kinase (AMPK) pathway was involved in these processes. HO‐1 down‐regulation with siRNA or inhibition with zinc protoporphyrin (Znpp) impaired the protective effects of D‐4F on the oxidative stress and the proliferation and migration of VSMCs. Moreover, down‐regulation of ATP‐binding cassette transporter A1 (ABCA1) abolished the activation of Akt and AMPK, the up‐regulation of HO‐1 and the anti‐oxidative effects of D‐4F. In vivo, D‐4F restrained neointimal formation and oxidative stress of carotid arteries in balloon‐injured Sprague Dawley rats. And inhibition of HO‐1 with Znpp decreased the inhibitory effects of D‐4F on neointimal formation and ROS production in arteries. In conclusion, D‐4F inhibited VSMC proliferation and migration in vitro and neointimal formation in vivo through HO‐1 up‐regulation, which provided a novel prophylactic and therapeutic strategy for anti‐restenosis of arteries.     

Blockade of both alpha1A- and alpha1B-adrenergic receptor subtype signaling is required to inhibit neointimal formation in the mouse femoral artery

Attenuation of early restenosis after percutaneous coronary intervention (PCI) is important for the successful treatment of coronary artery disease. Some clinical studies have shown that hypertension is a risk factor for early restenosis after PCI. These findings suggest that alpha(1)-adrenergic receptors (alpha(1)-ARs) may facilitate restenosis after PCI because of alpha(1)-AR's remarkable contribution to the onset of hypertension. In this study, we examined the neointimal formation after vascular injury in the femoral artery of alpha(1A)-knockout (alpha(1A)-KO), alpha(1B)-KO, alpha(1D)-KO, alpha(1A)-/alpha(1B)-AR double-KO (alpha(1AB)-KO), and wild-type mice to investigate the functional role of each alpha(1)-AR subtype in neointimal formation, which is known to promote restenosis. Neointimal formation 4 wk after wire injury was significantly (P < 0.05) smaller in alpha(1AB)-KO mice than in any other group of mice, while blood pressures were not altered in any of the groups of mice after wire injury compared with those before it. These results suggest that lack of both alpha(1A)- and alpha(1B)-ARs could be necessary to inhibit neointimal formation in the mouse femoral artery.     

Intravascular ultrasound observations of atherosclerotic lesion formation and restenosis in patients with diabetes mellitus

Coronary artery disease is more aggressive in diabetic patients than in nondiabetics; they have more diffuse disease, higher mortality rates and worse clinical outcomes after coronary interventions. Intravascular ultrasound (IVUS) produces transmural tomographic images of the coronary arteries in vivo. Recent IVUS studies have provided new insights into the mechanisms of stenosis formation and restenosis in both nondiabetic and diabetic patients. Arterial remodeling is defined as a change in arterial area. During atherogenesis, an increase in arterial area usually accompanies plaque accumulation to delay lumen compromise. Stenosis formation is related to: (a) the rate of plaque accumulation versus the rate of positive remodeling; and (b) the limits and ultimate failure of positive remodeling. However, there is a marked variability in remodeling. IVUS studies have suggested that remodeling may be impaired in some diabetic patients during atherogenesis. Following non-stent catheter-based interventions, serial (post-intervention and follow-up) IVUS studies have shown that the change in lumen area correlates better with the change in arterial area (remodeling) than with the change in plaque area (neointimal hyperplasia). In some patients, a positive remodeling response mitigates against the increase in plaque area to limit late lumen loss and restenosis. Neointimal hyperplasia is exaggerated in diabetic patients. Despite this, there is a reduced frequency of positive remodeling, potentially similar to the impaired positive remodeling in some diabetic patients during atherogenesis. Failed or inadequate arterial remodeling may contribute to the pathogenesis and natural history of atherosclerotic coronary artery disease in diabetic patients.     

Hemodynamics in stented vertebral artery ostial stenosis based on computational fluid dynamics simulations

Hemodynamic factors may affect the potential occurrence of in-stent restenosis (ISR) after intervention procedure of vertebral artery ostial stenosis (VAOS). The purpose of the present study is to investigate the influence of stent protrusion length in implantation strategy on the local hemodynamics of the VAOS. CTA images of a 58-year-old female patient with posterior circulation transient ischemic attack were used to perform a 3D reconstruction of the vertebral artery. Five models of the vertebral artery before and after the stent implantation were established. Model 1 was without stent implantation, Model 2–5 was with stent protruding into the subclavian artery for 0, 1, 2, 3 mm, respectively. Computational fluid dynamics simulations based on finite element analysis were employed to mimic the blood flow in arteries and to assess hemodynamic conditions, particularly the blood flow velocity and wall shear stress (WSS). The WSS and the blood flow velocity at the vertebral artery ostium were reduced by 85.33 and 35.36% respectively after stents implantation. The phenomenon of helical flow disappeared. Hemodynamics comparison showed that stent struts that protruded 1 mm into the subclavian artery induced the least decrease in blood speed and WSS. The results suggest that stent implantation can improve the hemodynamics of VAOS, while stent struts that had protruded 1 mm into the subclavian artery would result in less thrombogenesis and neointimal hyperplasia and most likely decrease the risk of ISR.     

Fabrication of drug-eluting covered stents with micropores and differential coating of heparin and FK506

To reduce in-stent restenosis rates, we developed a novel drug-eluting covered stent with a microporous elastometric covered film, in which its luminal surface was flat and immobilized with heparin for anticoagulation and its outer surface immobilized with FK506 to prevent neointimal hyperplasia. One month after implantation into the bilateral common carotid arteries, all stented arteries were patent and the luminal surfaces were fully covered with a confluent of endothelial cells irrespective of the drug immobilization. In the control group, which consisted of covered stents without drug immobilization, intensive inflammatory cells adjacent to the stents and neointimal hyperplasia, indicating vascular injury, were observed. In contrast, in the developed drug-eluting stents, only a few inflammatory cells around the stent strut and covered film were observed, and there was no significant neointimal thickening.     

Measurement of hemodynamics in human carotid artery using ultrasound and computational fluid dynamics.

The objective of the study was to investigate the feasibility of using computational fluid dynamic modeling (CFD) with noninvasive ultrasound measurements to determine time-variant three-dimensional (3D) carotid arterial hemodynamics in humans in vivo. The effects of hyperoxia and hypoxic hypercapnia on carotid artery local hemodynamics were examined by use of this approach. Six normotensive volunteers followed a double-blind randomized crossover design. Blood pressure, heart rate, and carotid blood flow were measured while subjects breathed normal air, a mixture of 5% CO(2) and 15% O(2) (hypoxic hypercapnia), and 100% O(2) (hyperoxia). Carotid artery geometry was reconstructed on the basis of B-mode ultrasound images by using purpose-built image processing software. Time-variant 3D carotid hemodynamics were estimated by using finite volume-based CFD. Systemic blood pressure was not significantly affected by hyperoxia or hypoxic hypercapnia, but heart rate decreased significantly with hyperoxia. There was an increase in diastolic flow velocity in the external carotid artery after hypoxic hypercapnia, but otherwise carotid blood flow velocities did not change significantly. Compared with normal air, hyperoxic conditions were associated with a decrease in the width of the region of flow separation in the external carotid artery. During hyperoxia, there was also an increase in the minimum and a decrease in maximum shear stress in the bifurcation and hence a reduction in cyclic variation in shear stress. Hypoxic hypercapnia was associated with a reduced duration of flow separation in the external carotid artery and an increase in the minimum shear stress without affecting the cyclic variation in shear stress. This study demonstrates the feasibility of using noninvasive ultrasound techniques in conjunction with CFD to describe time-variant 3D hemodynamics in the human carotid arterial bifurcation in vivo.