Aortic Regurgitation Dramatically Alters the Distribution of Atherosclerotic Lesions and Enhances Atherogenesis in Mice
Objective— Hemodynamics plays a critical role in atherogenesis, but the association between flow pattern and preferential localization of lesion is not fully understood. We developed a mouse model of aortic valve regurgitation (AR) to change the aortic flow pattern and observed the effects on plaque formation.
Methods and Results— High-frequency Doppler ultrasound imaging of 10 untreated C57BL/6J mice and 6 sham-treated low-density lipoprotein receptor–deficient (Ldlr−/−) mice revealed consistent antegrade blood flow throughout the aorta and oscillatory flow only along the lesser curvature of the aortic arch. Catheter-induced AR in 7 Ldlr−/− mice produced various degrees of diastolic retrograde flow throughout the aorta. After the mice were fed a cholesterol-enriched diet for 6 weeks, the burden of atherosclerotic lesions was increased 6-fold, with the naturally plaque-resistant descending aorta becoming susceptible. The AR severity correlated positively with the lesion burden in the descending thoracic and abdominal aorta but negatively with the lesions in the ascending aorta and aortic arch.
Conclusion— This AR model is valuable for elucidating the relationship between hemodynamics and predisposition of the artery wall to atherosclerosis, because of the significant alterations in local flow patterns and the conversion of large regions in the descending aorta from lesion resistant to lesion prone.
Complex and disturbed blood flow at sites of aortic curvature, branches, bifurcations, or other arterial regions is a predisposing factor for atherosclerosis.1,2 Endothelial cells of arteries sense wall shear stress (WSS), the frictional force of blood flow, and respond differently to various flow patterns by triggering unique signal transduction pathways that lead to distinct gene expression patterns. Unidirectional laminar WSS diminishes endothelial inflammatory responses and protects the artery from atherosclerosis. In contrast, low time-averaged WSS with oscillating directions correlates with increased predisposition to atherosclerosis.3,4 However, the association between flow dynamics and atherogenesis is still not fully understood, and the roles of low WSS and oscillatory WSS have never been differentiated. Useful insights have been gained from previous studies on animals of various species by altering the blood flow through a variety of surgical interventions.4 However, those models focus on specific aspects of endothelial cell biology in limited vessel segments and are unable to provide, at the systemic level, an in vivo environment with flow alteration in a morphologically normal aortic system for a comprehensive elucidation of the flow-plaque relationship.
In the last decade, mice have commonly been used in biological research because this species is most amenable to genetic manipulation. Low-density lipoprotein receptor–deficient (Ldlr−/−) and apolipoprotein E–deficient (Apoe−/−) mice are the most popular strains for studies related to atherosclerosis.5–9 A model was developed by placing a cast around the common carotid artery of Apoe−/− mice, in an attempt to induce 3 regions of lowered, increased, and oscillatory WSS.10,11 The segment facing elevated WSS was spared from lesions. However, the proximal carotid artery exposed to the lowered flow velocity did not develop lesions evenly along the entire segment as one would expect if the low WSS was the key factor for plaque initiation. The flow pattern distal to the cast showed flow instability with associated vortices rather than oscillatory flow. As seen in humans, aortic valve regurgitation (AR) produces diastolic retrograde flow that extends to the distal aorta.12,13 We reasoned that AR in mice would create oscillatory flow throughout the aorta and provide a novel model for exploring the effect of altered hemodynamics on atherogenesis.
High-frequency ultrasound imaging has been used to study cardiac morphology and flow dynamics in mice.14–16 The entire curved flow path of the proximal aorta from aortic orifice to proximal descending aorta, including the lesser curvature, which is highly predisposed to atherosclerosis, can be visualized via the right parasternal window.14 The abdominal aorta is also easily visualized. With pulsed wave Doppler and newly available Doppler color flow imaging,17 the flow patterns in the mouse aorta can be comprehensively observed.
In this study, we evaluated aortic hemodynamics in C57BL/6J mice under baseline conditions and in Ldlr−/− mice with catheter-induced AR of different severity in comparison with their sham-treated counterparts. The effects of AR-induced oscillatory flow pattern on the extent and distribution of atherosclerotic lesions in Ldlr−/− mice were assessed. AR enhanced the overall lesion burden and dramatically altered plaque distribution, with increased plaques in the descending thoracic and abdominal aorta, regions that are naturally plaque resistant. Surprisingly, along the lesser curvature of the aortic arch, which is normally lesion prone, the plaque decreased with increasing AR severity.
Ten C57BL/6J mice were studied using pulsed Doppler for normal local flow patterns at 15 weeks of age (24.6±0.7g). Fifteen Ldlr−/− mice were used to study the effect of aortic flow alteration on the atherosclerotic distribution. In the experimental group of 8 Ldlr−/− mice, AR was created by catheterization via the right common carotid artery at 14 weeks of age. In the control group, with 7 Ldlr−/− mice of the same age, sham catheterization was conducted without damaging the aortic valve.
Creation of AR in Ldlr−/− Mice
Under anesthesia by ketamine hydrochloride (100 mg/kg) and xylazine (20 mg/kg), a plastic catheter was introduced via the right common carotid artery and forwarded to the aortic orifice under the guidance of ultrasound imaging. A metal wire was then introduced through the catheter to penetrate the noncoronary cusp of the aortic valve until Doppler recording showed significant diastolic retrograde flow in the aorta. At the end of surgical intervention, the catheter with the central metal wire was immediately withdrawn, and the right common carotid artery was ligated in both the AR group and the control group with sham catheterization. Temgesic (100 μL/20 gm) was given to reduce pain postsurgery.
Overall Experimental Protocol
In Ldlr−/− mice, pulsed Doppler measurements were made at 1 week postcatheterization in both the AR group and the control group. These mice were then fed a defined semipurified 1.25% cholesterol high-fat diet (D12108, Research Diets, Inc.) for 6 weeks. Blood pressure was measured in Ldlr−/− mice before and 2 weeks after AR creation using a tail-cuff system, mainly for the systolic pressure because the diastolic pressure measured using this technique was less reliable. To confirm the persistence of flow alteration, the related Doppler flow parameters and the left ventricular dimensions and function were remeasured at 4 weeks of high-fat diet feeding in Ldlr−/− mice with AR. After 6 weeks of high-fat diet feeding, lesion area and distribution in Ldlr−/− mice were analyzed.
The experimental protocol was approved by the Animal Care Committee of the Hospital for Sick Children in Toronto, Ontario, Canada.
Observation of the Aortic Flow Pattern
A high-frequency ultrasound scanner (Vevo770, VisualSonics Inc., Toronto, Ontario, Canada) with a 30-MHz transducer was used. The related technical specifications, instrumental setting, and animal preparation have been described previously.14 Following isoflurane anesthesia via a mask, Doppler flow spectra were recorded at 7 levels, as shown in Figure 1A. The Doppler sample volume was 115 μm laterally and ≤170 μm axially. At each level in the ascending aorta and aortic arch, Doppler recordings were made at 3 locations: (1) the region close to the greater curvature (outer radius), (2) the middle lumen, and (3) the region close to the lesser curvature (inner radius). In the proximal abdominal aorta, where a posterior curvature exists in the sagittal plane, the Doppler flow spectrum was recorded from the anterior (lesser curvature), middle, and posterior (greater curvature) parts of the lumen. A significant effort was made to reduce the potential effect of the intercept angle between Doppler beam line and flow direction on the flow velocity measurement. The transducer orientation was very carefully adjusted to make the Doppler intercept angle as small as possible, and all the velocity measurements were angle-corrected. In no measurement did the intercept angle exceed 60°.
C57BL/6J mice and the control Ldlr−/− mice without AR showed similar Doppler flow waveforms: (1) unidirectional biphasic pattern, with a major systolic antegrade waveform followed by a diastolic antegrade waveform of low velocity (Figure 1B); and (2) triphasic flow pattern, with an extra early diastolic retrograde wave (Figure 1C). By tracing the maximal velocity, the systolic antegrade time-velocity integral (TVI) and the diastolic retrograde and antegrade TVIs were measured separately.
In the Ldlr−/− mice with AR, biphasic Doppler waveforms were usually recorded, with antegrade flow during systole and retrograde flow throughout diastole (Figure 1D). At some locations along the lesser curvature of the aortic arch, the systolic antegrade flow waveform was very small, and the retrograde flow started to occur from the early systole (Figure 1E). In that situation, the systolic antegrade TVI and systolic retrograde TVI were measured separately. In comparing the Doppler flow parameters between the Ldlr−/− groups with and without AR, the total antegrade TVI during both the systole and diastole (ie, the whole cardiac cycle) and the total retrograde TVI during both the systole and diastole were used.
The diastolic retrograde TVI recorded at the center of the abdominal aorta was used to measure the overall AR severity. All parameters were averaged for 3 cardiac cycles.
In the supplemental experiments, the aortic flow pattern in C57BL/6J mice and in Ldlr−/− mice with and without AR was evaluated using Doppler color flow imaging, which visualizes the 2-dimensional flow distribution across the aortic lumen throughout cardiac cycle and provides further evidence for the local flow patterns observed by pulsed Doppler. In another group of C57BL/6J mice with and without AR, the aortic diameters and leukocyte count were measured.
Assessment of Atherosclerotic Lesion
After 6 weeks of high-fat diet feeding, blood was collected for measuring plasma cholesterol and white blood cells. Mice were perfusion-fixed with 4% paraformaldehyde. The entire aorta was harvested and stained with oil red O and cut into 3 segments: (1) the proximal aorta to the first pair of intercostal arteries, which was opened longitudinally along the greater curvature; (2) the descending thoracic aorta to the level of diaphragm; and (3) the abdominal aorta to its bifurcation. The descending thoracic aorta and abdominal aorta were opened along the frontmost wall. The aortic segments were pinned flat onto a black silicon dish for photography, and the relative surface area occupied by oil red O–stained lesions was quantified using software developed by Scion Co. (Frederick, Md). In this analysis, the proximal aortic segment was further divided to 3 regions: the ascending aorta from the distal margin of aortic sinuses to the level proximal to innominate artery, the proximal aortic arch proximal to the left subclavian artery, and the rest as the distal aortic arch.
All parameters were expressed as the mean±SEM. The Student t test was used to compare the plaque burdens between the Ldlr−/− mice with and without AR. Linear regression was used to evaluate the correlation between the AR severity and the plaque burden. P<0.05 was chosen as the level of statistical significance.
The success rate of AR creation was higher than 90%. Only 1 AR mouse died from heart failure. One Ldlr−/− control mouse was excluded because of anatomic variations of the aorta.
Aortic Flow Pattern in C57BL/6J Mice
In the aortic orifice (L0), the velocity distribution was uniform across the lumen (Figure 2A). From the proximal ascending aorta to the distal aortic arch (L1 to L5), unidirectional biphasic flow spectrum was recorded along the greater curvature, with a major systolic antegrade waveform immediately followed by a diastolic antegrade wave of low velocity. In contrast, a triphasic or biphasic spectrum was found along the lesser curvature. A major systolic antegrade waveform was followed by a retrograde waveform at the early diastole and then by a small antegrade waveform or no flow in the rest of diastole (Figure 1B and 1C). Figure 2 shows the asymmetrical systolic antegrade flow with higher TVI along the greater curvature, the early diastolic retrograde flow detected mainly along the lesser curvature, and the mild antegrade flow in the rest of diastole between the proximal ascending aorta and the distal aortic arch. In the proximal abdominal aorta (L6), unidirectional biphasic flow waveform, with a major antegrade waveform during systole and a considerable antegrade waveform during diastole, was always found, with no retrograde flow throughout the cardiac cycle.
Altered Aortic Flow Pattern in Ldlr−/− Mice With AR
Control Ldlr−/− mice demonstrated a similar aortic flow pattern to that seen in C57BL/6J mice. In the Ldlr−/− mice with AR, the aortic flow pattern significantly changed, with diastolic retrograde flow occurring in the central aorta. In the ascending aorta and aortic arch, the systolic antegrade flow velocity along the greater curvature increased significantly (up to ≈3-fold) (Figure 1D) because of the compensation in left ventricular systolic function. In contrast, along the lesser curvature of the distal ascending aorta (L2), the systolic antegrade flow was significantly reduced in amplitude and also shortened in duration, appearing only during very early systole. As a consequence, the retrograde flow became much more temporally dominant, being present during the mid- to late systole and the whole diastole, especially in mice with severe AR (Figure 1E). Figure 3 compares the antegrade and retrograde TVIs along the greater and lesser curvatures of the ascending aorta and aortic arch (L1 to L5) between the Ldlr−/− groups with and without AR.
A biphasic flow pattern with diastolic retrograde flow was also observed in the proximal abdominal aorta (L6) of AR mice, which was not found in the control Ldlr−/− mice. A systolic antegrade flow with parabolic but slightly skewed velocity distribution was observed, with the velocity along the posterior wall (greater curvature) being significantly higher than that along the anterior wall (lesser curvature). The diastolic retrograde flow was symmetrically parabolic. In terms of the Doppler spectral bandwidth, the flow spectrum in AR mice was similar to that in the Ldlr−/− control group, indicating laminar flow in both groups (Figure 4).
Compared with the Ldlr−/− control group, the AR group showed significant increases in the left ventricular chamber dimensions, wall thickness, stroke volume, and cardiac output at 1 week postcatheterization (Table). During 4 weeks of follow-up observation, the AR group showed further increases in the left ventricular chamber dimensions (the end-diastolic diameter increased from 4.3±0.1 to 4.7±0.2 mm, P<0.05; the end-systolic diameter increased from 3.0±0.1 to 3.5±0.2 mm, P<0.05) and in the posterior wall thickness at end-diastole (increased from 0.82±0.02 to 0.98±0.05 mm, P<0.05) but not in the left ventricular stroke volume (32.5±1.8 versus 34.8±3.5 μL) or the cardiac output (14.5±1.1 versus 16.0±2.4 mL/min). The severity of AR (measured as the TVI of the diastolic retrograde flow at the center of abdominal aorta) did not change over time (0.8±0.2 versus 0.9±0.2 cm). In Ldlr−/− mice with AR, the systolic blood pressure did not show any significant difference before and 2 weeks after AR induction (110±3 versus 109±2 mm Hg).
At the end of the study, the AR group displayed a significant increase in blood leukocyte count (16.8×106±1.8×106 versus 7.1×106±0.8×106/mL, P<0.01) and in heart weight (0.18±0.04 versus 0.12±0.02 g, P<0.05), but no difference in body weight compared with the control group. There was no difference in serum cholesterol level between the AR and control groups (cholesterol, 30.3±1.7 versus 32.4±0.6 mmol/L).
Altered Distribution of Atherosclerosis in the Ldlr−/− Mice With AR
Lipid-laden lesions in the control Ldlr−/− mice were found primarily along the lesser curvature of the ascending aorta and proximal aortic arch, but not beyond the left subclavian artery (Figure 5A). The rest of the aorta was almost free of lesions, except for very small lesions at the ostia of branches. In Ldlr−/− mice with AR, the plaque burden of the whole aorta was significantly higher relative to the control group, largely because of a dramatic increase in lesions in the descending thoracic and abdominal aorta (Figure 5A and 5B). A positive correlation was found between plaque burden in the descending thoracic and abdominal aorta and the AR severity (Figure 5C). Moreover, the pattern of plaque distribution was closely related to the curvatures in the descending aorta, which mainly exist in the sagittal plane, as illustrated in Figure 1A. As seen from the dissected aortic specimen, plaques were always located along the lesser curvatures, which were the posterior wall of the midthoracic descending aorta and the anterior wall of the proximal abdominal aorta (Figure 5A).
In the ascending aorta and the proximal aortic arch, the plaque burden was comparable between the control and AR groups in terms of the average values. However, to our surprise, within the AR group the plaque burden along the lesser curvature tended to decrease with the increasing AR severity. Negative correlations (r=0.81 and ≈0.83; P<0.05) were found between the plaque burden in these 2 segments and the AR severity. In the distal aortic arch beyond the left subclavian artery, the plaque burden was significantly higher in the AR group than in the control group.
The Doppler color flow imaging in C57BL/6J mice and the Ldlr−/− mice with AR and the aortic diameter measurements and leukocyte count in C57BL/6J mice with AR are presented in the Supplemental Materials, available online at http://atvb.ahajournals.org.
In this study, the aortic flow pattern in C57BL/6J mice serves as baseline physiological data for normal mice. The creation of AR in Ldlr−/− mice significantly alters the aortic flow pattern and fundamentally changes the atherosclerotic distribution in a consistent manner. To our knowledge, such a change in plaque distribution as a direct result of flow alteration has not been previously reported. The wide range of the changes in flow pattern and lesion severity enables a correlative analysis between the degree of oscillatory flow and the severity of plaque formation. We believe that this novel model with a controllable degree of flow alteration but no direct injury to the aortic wall is ideal for in-depth exploration of the interaction between the local flow dynamics and the aortic endothelium in early atherogenesis.
This study strongly suggests that oscillatory flow is a key factor in the initiation of atherosclerosis. First of all, in C57BL/6J and the Ldlr−/− mice without AR, the oscillatory flow pattern is observed only along the lesser curvature of the ascending aorta and the aortic arch, spatially corresponding to the predilection sites of plaques as found in this and previous studies.8,9 Second, the present data confirm the consistent antegrade flow throughout cardiac cycle in the proximal abdominal aorta of normal mice, as was predicted by computational fluid simulations18 and also observed in vivo by MRI.19 Consistent with this, plaques are not abundant in the mouse abdominal aorta. This differs from the human abdominal aorta, which experiences significant diastolic flow reversal2,20–22 and is a preferential site for plaque formation.23 Furthermore, when AR was induced in Ldlr−/− mice, the descending thoracic aorta and abdominal aorta become plaque-prone. The only significant change in the Doppler flow recording was the presence of diastolic retrograde flow, whereas the systolic antegrade velocity and the properties of the flow spectrum (such as bandwidth) changed minimally.
Another possibility is that, even with the presence of oscillatory flow throughout the aorta, the elevated systolic antegrade velocity may be protective and prevent the aortic wall from developing plaque. As previously reported, endothelial cells subjected to elevated WSS tend to elongate and align in the direction of flow and undergo cytoskeletal remodeling. High WSS also promotes the release of vasodilators from endothelial cells that inhibit coagulation, adhesion, and migration of leukocytes and smooth muscle proliferation, while simultaneously promoting endothelial cell survival.4 All these factors reduce the probability of plaque formation. In this study, the greater curvatures of the ascending aorta, the aortic arch, and other curved segments along the descending aorta facing the oscillatory flow pattern are spared from atherosclerotic lesions. The elevated systolic antegrade velocity at these locations may, to a certain extent, be responsible for the absence of plaque.
In mice with severe AR, plaques decrease significantly along the lesser curvature of the distal ascending aorta and proximal aortic arch, a region that is naturally prone to atherogenesis. A possible reason is that the dramatically elevated antegrade flow velocity along the greater curvature causes a much more asymmetrical flow velocity distribution during systole, and consequently a very early flow reversal on the opposite side of the lumen because of flow separation. Thus, the flow pattern along the lesser curvature changes from oscillatory to predominantly retrograde. Similar flow separation and early reversal during systole along the lesser curvature was also observed in humans.24
AR may result in higher pulse blood pressure and to some extent promote plaque formation.25 However, normal systolic blood pressure was observed in Ldlr−/− mice with AR, whereas the significant systolic expansion of the aorta with unchanged diastolic diameter was found in C57BL/6J mice with AR (Supplemental Table). We speculate that the pulse pressure in the AR mice studied did not significantly increase because of the normal compliance of the aorta in young adult mice as compared with the increased stiffness of the aorta in aged/hypertensive human patients.25
A number of systemic factors, such as heart failure and the consequent activation of the sympathetic nervous system and invocation of systemic mediators of inflammation, could contribute to the increased plaque formation. However, they would act in a systemic manner. The present data demonstrate a very distinctive, consistent, and nonuniform pattern of plaque redistribution in AR mice, with increased lesion burden in the descending aorta but comparable or decreased lesions in the ascending aorta and arch. These data strongly suggest a direct relationship between the AR-induced changes in local flow (associated with vascular morphology) and the lesion formation. In addition, the cardiac function in AR mice maintained a compensatory status during the experimental period, as evidenced by the normal left ventricular fractional shortening and the increased cardiac output. The fact that the leukocytosis was observed only in hypercholesterolemic Ldlr−/− mice with AR suggests a synergistic influence of the AR-induced flow alteration and the hypercholesterolemia on circulating leukocyte level. The enhanced atherogenesis with elevated circulating cytokines/growth factors may account for this.
Because of the limited acoustic window for ultrasound imaging, some aortic segments, such as most of the descending thoracic aorta, were not well visualized. For the accessible aortic segments, only a single imaging section was applied, and the flow pattern in 3-dimensional space was not demonstrated. However, the imaging section used in this study allowed visualization of the aortic arch with the most significant curvature. The plane with the most asymmetrical flow patterns and the aortic walls with the most asymmetrical plaque distribution were observed.
Only the axial flow velocity was measured, and the lateral flow could not be evaluated. Helical velocity has been described in the human aorta.24 However, as seen from the orientation of the plaques in mice, the flow in the axial direction plays the dominant role in plaque formation. The plaques are usually limited in a relatively narrow region in the axial direction, with little lateral extension.
With high-frequency ultrasound, this study convincingly demonstrates the spatial association between the local flow pattern and the atherosclerotic distribution in mice. In C57BL/6J and Ldlr−/− mice with unchanged flow dynamics, the oscillatory flow is found only along the lesser curvature of ascending aorta and aortic arch, corresponding to the plaque-prone region in Ldlr−/− mice. In the Ldlr−/− mice with catheter-induced AR, the oscillatory flow pattern is detected all over the central aorta, and the atherosclerotic distribution throughout the aorta is fundamentally changed. The descending thoracic and abdominal aorta, which normally face consistently antegrade flow and are naturally lesion-free, become plaque-prone, with the plaque burden positively correlated to the AR severity. However, even with the retrograde flow, the greater curvature of aorta, where the antegrade velocity is elevated, shows a decreased likelihood of developing plaques. This novel mouse model with wide variety of local flow patterns but no direct injury to arterial wall is ideal for studying the role of flow dynamics in atherogenesis in vivo at cellular and molecular levels.
The authors thank Dr S. Lee Adamson for the tail-cuff system for blood pressure measurement.
Sources of Funding
This work is part of the Mouse Imaging Centre at the Hospital for Sick Children and the University of Toronto. The infrastructure was funded by the Canada Foundation for Innovation and Ontario Innovation Trust. The research was funded by an Ontario Research and Development Challenge Fund, and the Heart and Stroke Foundation of Ontario (grants T6107 and T6060). Dr Henkelman and Dr Foster hold Canada Research Chairs. Dr Cybulsky is a Career Investigator of the Heart and Stroke Foundation of Ontario.
F.S.F. has a financial interest in VisualSonics Inc.
Dr Zhou and Dr Zhu contributed equally to this work.
Received on: December 26, 2009; final version accepted on: March 4, 2010.
Ku DN, Giddens DP, Zarins CK, Glagov S. Pulsatile flow and atherosclerosis in the human carotid bifurcation: positive correlation between plaque location and low oscillating shear stress. Arteriosclerosis. 1985; 5: 293–302.
Traub O, Berk BC. Laminar shear stress: mechanisms by which endothelial cells transduce an atheroprotective force. Arterioscler Thromb Vasc Biol. 1998; 18: 677–685.
Zadelaar S, Kleemann R, Verschuren L, de Vries-Van der Weij J, van der Hoorn J, Princen HM, Kooistra T. Mouse models for atherosclerosis and pharmaceutical modifiers. Arterioscler Thromb Vasc Biol. 2007; 27: 1706–1721.
Won D, Zhu SN, Chen M, Teichert AM, Fish JE, Matouk CC, Bonert M, Ojha M, Marsden PA, Cybulsky MI. Relative reduction of endothelial nitric-oxide synthase expression and transcription in atherosclerosis-prone regions of the mouse aorta and in an in vitro model of disturbed flow. Am J Pathol. 2007; 171: 1691–1704.
Cheng C, Tempel D, van Haperen R, van der Baan A, Grosveld F, Daemen MJ, Krams R, de Crom R. Atherosclerotic lesion size and vulnerability are determined by patterns of fluid shear stress. Circulation. 2006; 113: 2744–2753.
Ambrosi P, Faugère G, Desfossez L, Habib G, Bory M, Luccioni R, Bernard P. Assessment of aortic regurgitation severity by magnetic resonance imaging of the thoracic aorta. Eur Heart J. 1995; 16: 406–409.
Zhou YQ, Foster FS, Nieman BJ, Davidson L, Chen XJ, Henkelman RM. Comprehensive transthoracic cardiac imaging in mice using ultrasound biomicroscopy with anatomical confirmation by magnetic resonance imaging. Physiol Genomics. 2004; 18: 232–244.
Amirbekian S, Long RC Jr, Consolini MA, Suo J, Willett NJ, Fielden SW, Giddens DP, Taylor WR, Oshinski JN. In vivo assessment of blood flow patterns in abdominal aorta of mice with MRI: implications for AAA localization. Am J Physiol Heart Circ Physiol. 2009; 297: H1290–H1295.
Moore JE Jr, Maier SE, Ku DN, Boesiger P. Hemodynamics in the abdominal aorta: a comparison of in vitro and in vivo measurements. J Appl Physiol. 1994; 76: 1520–1527.