In Vivo Analysis Of Angiotensin II-Induced Hypertension And Vascular Disease In Rats

Murine models of abdominal aortic aneurysms (AAA) are commonly used to study the pathogenesis of this disease. Rats are often used in laboratory studies because of their size and because they are more physiologically similar to humans than mice are. In this study, we subcutaneously implanted 9-week-old apolipoprotein E-deficient (apoE KO) rats with angiotensin II-filled pumps to study the effects of this vasoconstricting hormone on aneurysm development. Deletion of the apoE gene in mice causes excess lipid accumulation in the blood vessels, thereby increasing the likelihood of atherosclerotic lesions in the aorta. However, atherosclerotic plaque buildup was not evident when the rats consumed either normal chow or high fat diets, and no dissecting aneurysms were identified via ultrasound. We observed an average of 32.5 +/- 28.8 mmHg increase in systolic blood pressures as early as 4 days post-pump implantation and an increase of 54.2 +/- 25.9 mmHg from baseline 6 weeks after angiotensin II infusion began. This study shows that apoE KO rats can be valuable models for the physiological development of hypertension but not for dissecting suprarenal aneurysms. Future studies will focus on studying the effects of angiotensin II on the hearts of these genetically modified animals

Angiotensin II-Induced Hypertension in Apolipoprotein E-Deficient Rats

Abdominal aortic aneurysms (AAAs) are characterized by a weakened vessel wall and a diameter 50% greater than normal. AAA are usually asymptomatic until they are near rupturing, which can be fatal if not treated immediately. Apolipoprotein E-deficient (ApoE) mice are commonly used as a model to study aneurysm growth. Our lab has created a similar model using rats, which are more similar to humans. This study focuses on the analysis of blood pressures collected from ApoE rats for comparison with a known mouse model. Five ApoE rats (1 female, 4 males) received subcutaneous implants of osmotic mini pumps that released a continuous flow of angiotensin II (AngII) at 200 ng/kg/min. AngII is a protein known to increase blood pressure by acting on the renin-angiotensin system. Systolic, diastolic, and mean arterial pressures were measured using a non-invasive tail cuff system (CODA, Kent Scientific). Measurements were taken before pump implantation and on days 3, 7, 14, 21, and 28 after implantation. Mean arterial pressure increased from 133.8 ± 21.2 mmHg before pump implantation to 169.4 ± 20.3 mmHg on day 28. Systolic and diastolic pressures rose in a similar manner. Although the blood pressure increased in a manner similar to the mice, no aneurysms were observed in any of the rats. This may be due to species differences that affect vessel thickness and metabolic rate. Further investigations will be needed to determine why ApoE rats become hypertensive due to AngII, but do not develop suprarenal dissecting aortic aneurysms

3D Modeling of Murine Abdominal Aortic Aneurysms: Quantification of Segmentation and Volumetric Reconstruction

Abdominal Aortic Aneurysms (AAA) cause 5,900 deaths in the United States each year. Surgical intervention is clinically studied by non-invasive techniques such as computed tomography and magnetic resonance imaging. However, three-dimensional (3D) ultrasound imaging has become an inexpensive alternative and useful tool to characterize aneurysms, allowing for reconstruction of the vessel, quantification of hemodynamics through computational fluid dynamics (CFD) simulation, and possible prediction of aortic expansion and rupture. However, current analysis techniques for these images require the use of multiple software platforms for either modeling or simulation, prompting the need for alternatives to improve data processing. This study monitors the development of AAAs in apolipoprotein E-deficient mice infused with Angiotensin II using 3D ultrasound imaging with the purpose of evaluating the accuracy of SimVascular, a semi-automated specialized open source simulation software; for image reconstruction. The total volume to length ratio of the suprarenal aorta was obtained for 7 mice and compared to software that allows only segmentation and volume quantification (VevoLAB; FUJIFILM VisualSonics). We found that the volume per length measurements obtained with SimVascular (10.57 ± 6.96 mm2) were very similar to those obtained by VevoLAB (10.55 ± 6.95 mm2, p=0.77). In conclusion, SimVascular is an optimal tool for reconstructing vessel geometries from 3D ultrasound data due to its robust accuracy, efficiency, and semi-automatic computational processing capabilities used for modeling that will allow for future CFD simulation

Changes in Vessel Properties During Early Progression of Murine Abdominal Aortic Aneurysms from In Vivo Ultrasound

Abdominal aortic aneurysms (AAA) are a common and frequently fatal disease characterized by the weakening and dilation of the aorta. The larger the aneurysm, the higher the chances are of rupturing and life-threatening hemorrhage. The aim of this study is to apply the angiotensin II (AngII) model of AAAs in male apolipoprotein-E-deficient mice (apoE-/- C57Bl/6J), in order to analyze, quantify, and understand the pathologies and characteristics associated with early AAA development. To date, many studies focusing on the evaluation of AAA characteristics have been performed ex vivo. Therefore, we focused on in vivo assessment, through the use of high frequency ultrasound technology, to measure parameters such as aortic diameter, volume, circumferential cyclic strain, blood flow velocity, and thrombus volume. Data analysis from this 7-day study suggests an increase in aortic diameter and volume, a decrease in aortic blood flow velocity and strain, and large variations in volumes of the thrombotic volume within the aneurysm’s false lumen. Our analysis of these AAA features has shown that there are a variety of patterns, which may provide insight into further understanding AAA pathology in this model. Future work will focus on comparing the relationship between the features during early AAA formation

In Vivo Flow Measurements of Murine Renal Arteries and Veins with High Frequency Ultrasound

The number of glomeruli in the kidneys has been shown to have an effect on the decline in renal function over time (Brenner, Garcia, Anderson 1988). Furthermore, flow in the renal arteries and veins may depend on the number of glomeruli in the kidney. Consistent in vivo measurements of volumetric flow in the renal arteries and veins are difficult to obtain. Thus, the purpose of this study was to develop non-invasive imaging techniques capable of estimating arterial and venous flow to kidneys. A high-frequency small animal ultrasound system was chosen based upon its excellent spatial and temporal resolution when imaging mice (Vevo 2100, VisualSonics, Inc.). Velocity profiles of the renal arteries and veins in C57BL/6 male mice (n=4) were measured. Motion, color Doppler, and pulsed wave Doppler data were acquired and used to determine renal diameter, maximum velocity, mean velocity, and volumetric flow for both kidneys. For the renal artery the average volumetric flow was 33.31±7.16 mm3/s and for the renal vein it was 30.23±4.58 mm3/s. The next step will be imaging the same animals multiple times to ensure that these measurements are consistent over prolonged periods of time. Then data will be collected from different breeds of mice to conclude whether or not differences in glomeruli number affect renal flow. Measurement of volumetric flow in the renal arteries and veins can lead to important insights into how the glomeruli density in kidneys relates to renal flow and function

Characterization of Left-Ventricular Thrombus Formation Using High Frequency Ultrasound

Heart failure is a leading cause of death in the United States, and cardiac thrombus, a common morbidity associated with heart failure, significantly increases a patient’s risk of embolic events. The objective of this project is to characterize left-ventricular (LV) thrombus development using high frequency ultrasound imaging in a murine model. C57BL/6J wild-type mice (n=6) were injected intraperitoneally with iron dextran five times a week for six weeks to increase oxidative stress in the heart. Granulocyte-colony stimulating factor (G-CSF) was subcutaneously injected daily during the second week to initiate stem cell migration and stimulate endothelial cell activation, thus increasing the hypercoagulability state of the blood. A high-frequency, small animal ultrasound system (Vevo2100, VisualSonics FUJIFILM Inc.) and a 40 MHz central frequency transducer were used to track LV thrombus progression and evaluate LV function weekly. Four out of six mice developed thrombus, but no significant differences in LV performance were observed when compared to mice that did not form a thrombus. Further investigation is necessary to study the role of attenuated heart function on thrombus formation. Future work will incorporate a murine model of myocardial infarction to investigate if a severely compromised heart increases the risk of or accelerates LV thrombus formation. This study will aid in identifying patients who are predisposed to thrombus formation following a heart attack, leading to more effective prevention and treatment methods

Visualization of Complex Flow Patterns in Angiotensin II-Induced Dissecting Murine Abdominal Aortic Aneurysms with High Frequency Ultrasound

Abdominal aortic aneurysm (AAA) rupture is a common cause of mortality in the United States. Current treatments are only employed once the risk of rupture outweighs the risks associated with surgery. Murine models have been developed to characterize AAA pathogenesis in the hope that new treatments will be developed. For this study, angiotensin II (AngII) was infused subcutaneously into apolipoprotein E-deficient (ApoE-/-) mice using an osmotic mini-pump over 28 days. ApoE-/- mice (16-week-old, 3 females, 2 males) were imaged using a VisualSonics Vevo 2100 high frequency ultrasound before pump implantation and 3, 7, 14, 21, and 27 days following implantation. Images were acquired in the transverse and longitudinal planes from the suprarenal region of the aorta. Blood pressure measurements were taken using a tail-cuff system (CODA, Kent Scientific). Three mice (1 female, 2 male) developed aneurysms within the first 14 days of infusion. Pre-study abdominal aortas had a diastolic diameter of 0.84±0.09 mm and a systolic diameter of 0.96±0.08 mm. By day 21, AAAs had a diastolic diameter of 1.51±0.59 mm and a systolic diameter of 1.56±0.59 mm. Initially, mice had a systolic blood pressure of 111.94±6.53 mmHg and a diastolic pressure of 82.38±5.13 mmHg. These pressures steadily elevated but eventually began to plateau. By day 27, systolic pressure had risen to 154.92±11.43 mmHg and diastolic pressure to 115.77±10.25 mmHg. Color Doppler images revealed complex, recirculating flow within the aneurysms, a phenomenon which could affect vessel remodeling. In conclusion, this study utilized in vivo sonographic methods to characterize AAA development

Transport of nitrite from large arteries modulates regional blood flow during stress and exercise

Background: Acute cardiovascular stress increases systemic wall shear stress (WSS)–a frictional force exerted by the flow of blood on vessel walls–which raises plasma nitrite concentration due to enhanced endothelial nitric oxide synthase (eNOS) activity. Upstream eNOS inhibition modulates distal perfusion, and autonomic stress increases both the consumption and vasodilatory effects of endogenous nitrite. Plasma nitrite maintains vascular homeostasis during exercise and disruption of nitrite bioavailability can lead to intermittent claudication. Hypothesis: During acute cardiovascular stress or strenuous exercise, we hypothesize enhanced production of nitric oxide (NO) by vascular endothelial cells raises nitrite concentrations in near-wall layers of flowing blood, resulting in cumulative NO concentrations in downstream arterioles sufficient for vasodilation. Confirmation and implications: Utilizing a multiscale model of nitrite transport in bifurcating arteries, we tested the hypothesis for femoral artery flow under resting and exercised states of cardiovascular stress. Results indicate intravascular transport of nitrite from upstream endothelium could result in vasodilator-active levels of nitrite in downstream resistance vessels. The hypothesis could be confirmed utilizing artery-on-a-chip technology to measure NO production rates directly and help validate numerical model predictions. Further characterization of this mechanism may improve our understanding of symptomatic peripheral artery occlusive disease and exercise physiology

Stable Thermally-Modulated Nanodroplet Ultrasound Contrast Agents

Liquid perfluorocarbon-based nanodroplets are stable enough to be used in extravascular imaging, but provide limited contrast enhancement due to their small size, incompressible core, and small acoustic impedance mismatch with biological fluids. Here we show a novel approach to overcoming this limitation by using a heating–cooling cycle, which we will refer to as thermal modulation (TM), to induce echogenicity of otherwise stable but poorly echogenic nanodroplets without triggering a transient phase shift. We apply thermal modulation to high-boiling point tetradecafluorohexane (TDFH) nanodroplets stabilized with a bovine serum albumin (BSA) shell. BSA-TDFH nanodroplets with an average diameter under 300 nanometers showed an 11.9 ± 5.4 mean fold increase in echogenicity on the B-mode and a 13.9 ± 6.9 increase on the nonlinear contrast (NLC) mode after thermal modulation. Once activated, the particles maintained their enhanced echogenicity (p \u3c 0.001) for at least 13 h while retaining their nanoscale size. Our data indicate that thermally modulated nanodroplets can potentially serve as theranostic agents or sensors for various applications of contrast-enhanced ultrasound