Hemodynamic response to thermal stress varies with sex and age: a murine MRI study.

PURPOSE: The cardiovascular (CV) system plays a vital role in thermoregulation because of its influence on heat transfer via forced convection and conduction by changes in blood distribution, blood velocity, and proximity of vessels to surrounding tissues. To fully understand the cardiovascular system's role in thermoregulation, blood distribution (influenced by cardiac output, vessel size, blood flow, and pressure) must be quantified, ideally across sex and age. Additionally, wall shear stress is quantified because it is an important metric in cardiovascular disease localization and progression. By investigating the effect of thermal conditions on wall shear stress at a healthy baseline, researchers can begin to study the confluence of thermal condition with pathology or exercise. The purpose of this study is to determine the influence of sex and age on the CV response to temperature. In this work, the effect of core body temperature on hemodynamics of the murine arterial and venous systems has been studied non-invasively, at multiple locations across age and sex. METHODS: Male and female, adult and aged, mice (n = 20) were anesthetized and underwent MRI at 7 T. Data were acquired from four co-localized vessel pairs (the neck [carotid/jugular], torso [suprarenal and infrarenal aorta/inferior vena cava (IVC)], periphery [femoral artery/vein]) at core temperatures of 35, 36, 37, and 38 °C. Sixteen CINE, ECG-gated, phase contrast frames with one-directional velocity encoding (through plane) were acquired perpendicular to each vessel. Each frame was analyzed to quantify blood velocity and volumetric flow using a semi-automated in-house MATLAB script. Wall shear stress (WSS) was calculated using the Hagen-Poiseulle formula. A multivariable regression for WSS in the femoral artery was fitted with temperature, sex, age, body weight, and heart rate as variables. RESULTS: Blood velocity and volumetric flow were quantified in eight vessels at four core body temperatures. Flow in the infrarenal IVC linearly increased with temperature for all groups (p = .002; adjusted means of slopes: male vs. female, 0.37 and 0.28 cm/(s × °C); adult vs. aged, 0.22 and 0.43 cm/(s × °C)). Comparing average volumetric flow response to temperature, groups differed for the suprarenal aorta (adult < aged, p < .05), femoral artery (adult < aged, p < .05), and femoral vein (adult male < aged male, p < .001). The two-way interaction terms of temperature and body weight and temperature and sex had the largest effect on wall shear stress. CONCLUSIONS: Age, in particular, had a significant impact on hemodynamic response as measured by volumetric flow (e.g., aged males > adult males) and WSS at peak-systole (e.g., aged males < adult males). The hemodynamic data can provide physiologically-relevant parameters, including sex and age difference, to computational fluid dynamics models and provide baseline data for the healthy murine vasculature to use as a benchmark for investigations of a variety of physiological (thermal stress) and pathophysiological conditions of the cardiovascular system.

Anatomical location, sex, and age influence murine arterial circumferential cyclic strain before and during dobutamine infusion.

BACKGROUND: One of the primary biomechanical factors influencing arterial health is their deformation across the cardiac cycle, or cyclic strain, which is often associated with arterial stiffness. Deleterious changes in the cardiovascular system, e.g., increased arterial stiffness, can remain undetected until the system is challenged, such as under a cardiac stressor like dobutamine. PURPOSE: To quantify cyclic strain in mice at different locations along the arterial tree prior to and during dobutamine infusion, while evaluating the effects of sex and age. STUDY TYPE: Control/cohort study. ANIMAL MODEL: Twenty C57BL/6 mice; male, female; ∼12 and 24 weeks of age; n = 5 per group. FIELD STRENGTH/SEQUENCE: 7T; CINE MRI with 12 frames, velocity compensation, and prospective cardiac gating. ASSESSMENT: Prior to and during the infusion of dobutamine, Green-Lagrange circumferential cyclic strain was calculated from perimeter measurements derived from CINE data acquired at the carotid artery, suprarenal and infrarenal abdominal aorta, and iliac artery. STATISTICAL TESTS: Analysis of variance (ANOVA) followed by post-hoc tests was used to evaluate the influence of dobutamine, anatomical location, sex, and age. RESULTS: Heart rates did not differ between groups prior to or during dobutamine infusion (P = 0.87 and P = 0.08, respectively). Dobutamine increased cyclic strain in each group. Within a group, increases in strain were similar across arteries. At the suprarenal aorta, strain was reduced in older mice at baseline (young 27.6 > mature 19.3%, P = 0.01) and during dobutamine infusion (young 53.0 > mature 36.2%, P = 0.005). In the infrarenal aorta, the response (dobutamine - baseline) was reduced in older mice (young 21.9 > mature 13.5%, P = 0.04). DATA CONCLUSION: Dobutamine infusion increases circumferential cyclic strain throughout the arterial tree of mice. This effect is quantifiable using CINE MRI. The results demonstrate that strain prior to and during dobutamine is influenced by anatomical location, sex, and age. LEVEL OF EVIDENCE: 3 Technical Efficacy: Stage 2 J. Magn. Reson. Imaging 2019;49:69-80.

Assessing structural and functional response of murine vasculature to acute ß-adrenergic stimulation in vivo during hypothermic and hyperthermic conditions.

Background: Because of the importance of adrenoreceptors in regulating the cardiovascular (CV) system and the role of the CV system in thermoregulation, understanding the response to these two stressors is of interest. The purpose of this study was to assess changes of arterial geometry and function in vivo during thermal and ß-adrenergic stress induced in mice and quantified by MRI.Methods: Male mice were anesthetized and imaged at 7 T. Anatomical and functional data were acquired from the neck (carotid artery), torso (suprarenal and infrarenal aorta and iliac artery) and periphery (femoral artery). Intravenous dobutamine (tail vein catheter, 40 µg/kg/min, 0.12 mL/h) was used as ß-adrenergic stressor. Baseline and dobutamine data were acquired at minimally hypothermic (35 °C) and minimally hyperthermic (38 °C) core temperatures. Cross-sectional vessel area and maximum cyclic strain were measured across the cardiac cycle.Results: Vascular response varied by location and by core temperature. For minimally hypothermic conditions (35 °C), average, maximum and minimum areas decreased with dobutamine only at the suprarenal aorta (avg: -17.9%, max: -13.5%, min: -21.4%). For minimally hyperthermic conditions (38 °C), vessel areas decreased between baseline and dobutamine at the carotid (avg: -19.6%, max: -15.5%, min: -19.3%) and suprarenal aorta (avg: -24.2%, max: -17.4%, min: -17.3%); whereas, only the minimum vessel area decreased for the iliac artery (min: -14.4%). Maximum cyclic strain increased between baseline and dobutamine at the iliac artery for both conditions and at the suprarenal aorta at hyperthermic conditions.Conclusions: At hypothermic conditions, the vessel area response to dobutamine is diminished compared to hyperthermic conditions where the vessel area response mimics normothermic dobutamine conditions. The varied response emphasizes the need to monitor and control body temperature during medical conditions or treatments that may be accompanied by hypothermia, especially when vasoactive agents are used.

Cross-sectional area of the murine aorta linearly increases with increasing core body temperature.

PURPOSE: The cardiovascular (CV) system plays a vital role in thermoregulation. To date, the response of core vasculature to increasing core temperature has not been adequately studied in vivo. Our objective was to non-invasively quantify the arterial response in murine models due to increases in body temperature, with a focus on core vessels of the torso and investigate whether responses were dependent on sex or age. METHODS: Male and female, adult and aged mice were anaesthetised and underwent magnetic resonance imaging (MRI). Data were acquired from the circle of Willis (CoW), heart, infrarenal aorta and peripheral arteries at core temperatures of 35, 36, 37 and 38 °C (±0.2 °C). RESULTS: Vessels in the CoW did not change. Ejection fraction decreased and cardiac output (CO) increased with increasing temperature in adult female mice. Cross-sectional area of the aorta increased significantly and linearly with temperature for all groups, but at a diminished rate for aged animals (p < 0.01; male and female: adult, 0.019 and 0.024 mm2/°C; aged, 0.017 and 0.011 mm2/°C). Aged male mice had a diminished response in the periphery (% increase in femoral artery area from 35 to 38 °C, male and female: adult, 67 and 65%; aged, 0.1 and 57%). CONCLUSION: Previously unidentified increases in aortic area due to increasing core temperature are biologically important because they may affect conductive and convective heat transfer. Leveraging non-invasive methodology to quantify sex and age dependent vascular responses due to increasing core temperature could be combined with bioheat modelling in order to improve understanding of thermoregulation.

Brain Distribution and Efficacy of the Brain Penetrant PI3K Inhibitor GDC-0084 in Orthotopic Mouse Models of Human Glioblastoma.

Glioblastoma multiforme (GBM) is the most common primary brain tumor in adults. Limited treatment options have only marginally impacted patient survival over the past decades. The phophatidylinositol 3-kinase (PI3K) pathway, frequently altered in GBM, represents a potential target for the treatment of this glioma. 5-(6,6-Dimethyl-4-morpholino-8,9-dihydro-6H-[1,4]oxazino[4,3-e]purin-2-yl)pyrimidin-2-amine (GDC-0084) is a PI3K inhibitor that was specifically optimized to cross the blood-brain barrier. The goals of our studies were to characterize the brain distribution, pharmacodynamic (PD) effect, and efficacy of GDC-0084 in orthotopic xenograft models of GBM. GDC-0084 was tested in vitro to assess its sensitivity to the efflux transporters P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) and in vivo in mice to evaluate its effects on the PI3K pathway in intact brain. Mice bearing U87 or GS2 intracranial tumors were treated with GDC-0084 to assess its brain distribution by matrix-assisted laser desorption ionization (MALDI) imaging and measure its PD effects and efficacy in GBM orthotopic models. Studies in transfected cells indicated that GDC-0084 was not a substrate of P-gp or BCRP. GDC-0084 markedly inhibited the PI3K pathway in mouse brain, causing up to 90% suppression of the pAkt signal. MALDI imaging showed GDC-0084 distributed evenly in brain and intracranial U87 and GS2 tumors. GDC-0084 achieved significant tumor growth inhibition of 70% and 40% against the U87 and GS2 orthotopic models, respectively. GDC-0084 distribution throughout the brain and intracranial tumors led to potent inhibition of the PI3K pathway. Its efficacy in orthotopic models of GBM suggests that it could be effective in the treatment of GBM. GDC-0084 is currently in phase I clinical trials.

Monovalent antibody design and mechanism of action of onartuzumab, a MET antagonist with anti-tumor activity as a therapeutic agent.

Binding of hepatocyte growth factor (HGF) to the receptor tyrosine kinase MET is implicated in the malignant process of multiple cancers, making disruption of this interaction a promising therapeutic strategy. However, targeting MET with bivalent antibodies can mimic HGF agonism via receptor dimerization. To address this limitation, we have developed onartuzumab, an Escherichia coli-derived, humanized, and affinity-matured monovalent monoclonal antibody against MET, generated using the knob-into-hole technology that enables the antibody to engage the receptor in a one-to-one fashion. Onartuzumab potently inhibits HGF binding and receptor phosphorylation and signaling and has antibody-like pharmacokinetics and antitumor activity. Biochemical data and a crystal structure of a ternary complex of onartuzumab antigen-binding fragment bound to a MET extracellular domain fragment, consisting of the MET Sema domain fused to the adjacent Plexins, Semaphorins, Integrins domain (MET Sema-PSI), and the HGF ß-chain demonstrate that onartuzumab acts specifically by blocking HGF α-chain (but not ß-chain) binding to MET. These data suggest a likely binding site of the HGF α-chain on MET, which when dimerized leads to MET signaling. Onartuzumab, therefore, represents the founding member of a class of therapeutic monovalent antibodies that overcomes limitations of antibody bivalency for targets impacted by antibody crosslinking.

Influences of aortic motion and curvature on vessel expansion in murine experimental aneurysms.

OBJECTIVE: To quantitatively compare aortic curvature and motion with resulting aneurysm location, direction of expansion, and pathophysiological features in experimental abdominal aortic aneurysms (AAAs). METHODS AND RESULTS: MRI was performed at 4.7 T with the following parameters: (1) 3D acquisition for vessel geometry and (2) 2D cardiac-gated acquisition to quantify luminal motion. Male 24-week-old mice were imaged before and after AAA formation induced by angiotensin II (AngII)-filled osmotic pump implantation or infusion of elastase. AngII-induced AAAs formed near the location of maximum abdominal aortic curvature, and the leftward direction of expansion was correlated with the direction of suprarenal aortic motion. Elastase-induced AAAs formed in a region of low vessel curvature and had no repeatable direction of expansion. AngII significantly increased mean blood pressure (22.7 mm Hg, P<0.05), whereas both models showed a significant 2-fold decrease in aortic cyclic strain (P<0.05). Differences in patterns of elastin degradation and localization of fluorescent signal from protease-activated probes were also observed. CONCLUSIONS: The direction of AngII aneurysm expansion correlated with the direction of motion, medial elastin dissection, and adventitial remodeling. Anterior infrarenal aortic motion correlated with medial elastin degradation in elastase-induced aneurysms. Results from both models suggest a relationship between aneurysm pathological features and aortic geometry and motion.

Stereotactic Transcranial Focused Ultrasound Targeting System for Murine Brain Models.

An inexpensive, accurate focused ultrasound stereotactic targeting method guided by pretreatment magnetic resonance imaging (MRI) images for murine brain models is presented. An uncertainty of each sub-component of the stereotactic system was analyzed. The entire system was calibrated using clot phantoms. The targeting accuracy of the system was demonstrated with an in vivo mouse glioblastoma (GBM) model. The accuracy was quantified by the absolute distance difference between the prescribed and ablated points visible on the pre treatment and posttreatment MR images, respectively. A precalibration phantom study ( N = 6 ) resulted in an error of 0.32 ± 0.31, 0.72 ± 0.16, and 1.06 ± 0.38 mm in axial, lateral, and elevational axes, respectively. A postcalibration phantom study ( N = 8 ) demonstrated a residual error of 0.09 ± 0.01, 0.15 ± 0.09, and 0.47 ± 0.18 mm in axial, lateral, and elevational axes, respectively. The calibrated system showed significantly reduced ( ) error of 0.20 ± 0.21, 0.34 ± 0.24, and 0.28 ± 0.21 mm in axial, lateral, and elevational axes, respectively, in the in vivo GBM tumor-bearing mice ( N = 10 ).

In vivo quantification of murine aortic cyclic strain, motion, and curvature: implications for abdominal aortic aneurysm growth.

PURPOSE: To develop methods to quantify cyclic strain, motion, and curvature of the murine abdominal aorta in vivo. MATERIALS AND METHODS: C57BL/6J and apoE(-/-) mice underwent three-dimensional (3D) time-of-flight MR angiography to position cardiac-gated 2D slices at four locations along the abdominal aorta where circumferential cyclic strain and lumen centroid motion were calculated. From the 3D data, a centerline through the aorta was created to quantify geometric curvature at 0.1-mm intervals. Medial elastin content was quantified with histology postmortem. The location and shape of abdominal aortic aneurysms (AAAs), created from angiotensin II infusion, were evaluated qualitatively. RESULTS: Strain waveforms were similar at all locations and between groups. Centroid motion was significantly larger and more leftward above the renal vessels than below (P < 0.05). Maximum geometric curvature occurred slightly proximal to the right renal artery. Elastin content was similar around the circumference of the vessel. AAAs developed in the same location as the maximum curvature and grew in the same direction as vessel curvature and motion. CONCLUSION: The methods presented provide temporally and spatially resolved data quantifying murine aortic motion and curvature in vivo. This noninvasive methodology will allow serial quantification of how these parameters influence the location and direction of AAA growth.

In Vivo MRI Assessment of Blood Flow in Arteries and Veins from Head-to-Toe Across Age and Sex in C57BL/6 Mice.

Although widely used as a preclinical model for studying cardiovascular diseases, there is a scarcity of in vivo hemodynamic measurements of the naïve murine system in multiple arterial and venous locations, from head-to-toe, and across sex and age. The purpose of this study is to quantify cardiovascular hemodynamics in mice at different locations along the vascular tree while evaluating the effects of sex and age. Male and female, adult and aged mice were anesthetized and underwent magnetic resonance imaging. Data were acquired from four co-localized vessel pairs (carotid/jugular, suprarenal and infrarenal aorta/inferior vena cava (IVC), femoral artery/vein) at normothermia (core temperature 37 ± 0.2 °C). Influences of age and sex on average velocity differ by location in arteries. Average arterial velocities, when plotted as a function of distance from the heart, decrease nearly linearly from the suprarenal aorta to the femoral artery (adult and aged males: - 0.33 ± 0.13, R2 = 0.87; - 0.43 ± 0.10, R2 = 0.95; adult and aged females: - 0.23 ± 0.07, R2 = 0.91; - 0.23 ± 0.02, R2 = 0.99). Average velocity of aged males and average volumetric flow of aged males and females tended to be larger compared to adult comparators. With cardiovascular disease as the leading cause of death and with the implications of cardiovascular hemodynamics as important biomarkers for health and disease, this work provides a foundation for sex and age comparisons in pathophysiology by collecting and analyzing hemodynamic data for the healthy murine arterial and venous system from head-to-toe, across sex and age.

Effects of Histotripsy on Local Tumor Progression in an in vivo Orthotopic Rodent Liver Tumor Model.

OBJECTIVE AND IMPACT STATEMENT: This is the first longitudinal study investigating the effects of histotripsy on local tumor progression in an in vivo orthotopic, immunocompetent rat hepatocellular carcinoma (HCC) model. INTRODUCTION: Histotripsy is the first noninvasive, nonionizing, nonthermal, mechanical ablation technique using ultrasound to generate acoustic cavitation to liquefy the target tissue into acellular debris with millimeter accuracy. Previously, histotripsy has demonstrated in vivo ablation of noncancerous liver tissue. METHODS: N1-S1 HCC tumors were generated in the livers of immunocompetent rats (n = 6, control; n = 15, treatment). Real-time ultrasound-guided histotripsy was applied to ablate either 100% tumor volume + up to 2mm margin (n = 9, complete treatment) or 50-75% tumor volume (n = 6, partial treatment) by delivering 1-2 cycle histotripsy pulses at 100 Hz PRF (pulse repetition frequency) with p - ≥30MPa using a custom 1MHz transducer. Rats were monitored weekly using MRI (magnetic resonance imaging) for 3 months or until tumors reached ~25mm. RESULTS: MRI revealed effective post-histotripsy reduction of tumor burden with near-complete resorption of the ablated tumor in 14/15 (93.3%) treated rats. Histopathology showed <5mm shrunken, non-tumoral, fibrous tissue at the treatment site at 3 months. Rats with increased tumor burden (3/6 control and 1 partial treatment) were euthanized early by 2-4 weeks. In 3 other controls, histology revealed fibrous tissue at original tumor site at 3 months. There was no evidence of histotripsy-induced off-target tissue injury. CONCLUSION: Complete and partial histotripsy ablation resulted in effective tumor removal for 14/15 rats, with no evidence of local tumor progression or recurrence.

PET of glial metabolism using 2-18F-fluoroacetate.

UNLABELLED: Imaging of the glial activation that occurs in response to central nervous system trauma and inflammation could become a powerful technique for the assessment of several neuropathologies. The selective uptake and metabolism of 2-(18)F-fluoroacetate ((18)F-FAC) in glia may represent an attractive strategy for imaging glial metabolism. METHODS: We have evaluated the use of (18)F-FAC as a specific PET tracer of glial cell metabolism in rodent models of glioblastoma, stroke, and ischemia-hypoxia. RESULTS: Enhanced uptake of (18)F-FAC was observed (6.98 +/- 0.43 percentage injected dose per gram [%ID/g]; tumor-to-normal ratio, 1.40) in orthotopic U87 xenografts, compared with healthy brain tissue. The lesion extent determined by (18)F-FAC PET correlated with that determined by MRI (R(2) = 0.934, P = 0.007). After transient middle cerebral artery occlusion in the rat brain, elevated uptake of (18)F-FAC (1.00 +/- 0.03 %ID/g; lesion-to-normal ratio, 1.90) depicted the ischemic territory and correlated with infarct volumes as determined by 2,3,5-triphenyltetrazolium chloride staining (R(2) = 0.692, P = 0.010) and with the presence of activated astrocytes detected by anti-glial fibrillary acidic protein. Ischemia-hypoxia, induced by permanent ligation of the common carotid artery with transient hypoxia, resulted in persistent elevation of (18)F-FAC uptake within 30 min of the induction of hypoxia. CONCLUSION: Our data support the further evaluation of (18)F-FAC PET for the assessment of glial cell metabolism associated with neuroinflammation.

Collateral vein dynamics in mouse models of venous thrombosis: Pathways consistent with humans.

INTRODUCTION: Prolific collateralization in the venous system has been associated with more severe disease. However, there is a scarcity of information on venogenesis and collateral vessel progression over time. Further, little is understood regarding the relevance of the most common preclinical model-the mouse-for studying venous collateralization. The purpose of this work was to non-invasively and quantitatively characterize collateral vein development and progression in two murine models of deep vein thrombosis using magnetic resonance imaging (MRI). METHODS: Venous thrombosis (VT) was induced in 12-14-week-old male C57BL/6 mice using either the inferior vena cava (IVC) ligation model (n = 5) or the electrolytic IVC model (n = 5). Magnetic Resonance Imaging (MRI) methods optimized for small venous imaging were used on days 2, 6, 14, and 21 following venous thrombosis induction to quantify collateral development and thrombus volume. RESULTS: Collateral veins ~150-200 µm in diameter could be tracked in three dimensions. Collateral pathways were influenced by pre-existing anatomy; mice with bilateral IVC branches showed a predominant superficial collateral pathway (superficial and internal epigastric veins), whereas mice with no lateral branches exhibited a strong intermediate collateral pathway (gonadal and periureteric veins) and were less likely to develop ascending lumbar collaterals. The degree of venogenesis showed a positive correlation with thrombus volume in both models (combined R2 = 0.64, p < 0.0001). CONCLUSIONS: Venous collateral pathways in C57BL/6 mice are consistent with those described in humans. Collateral pathways are influenced by pre-existing anatomy, and the degree of collateralization correlates with thrombus volume.

Minimally invasive gas embolization using acoustic droplet vaporization in a rodent model of hepatocellular carcinoma.

Hepatocellular carcinoma is the third leading cause of cancer-related deaths worldwide. Many patients are not eligible for curative therapies, such as surgical resection of the tumor or a liver transplant. Transarterial embolization is one therapy clinically used in these cases; however, this requires a long procedure and careful placement of an intraarterial catheter. Gas embolization has been proposed as a fast, easily administered, more spatially selective, and less invasive alternative. Here, we demonstrate the feasibility and efficacy of using acoustic droplet vaporization to noninvasively generate gas emboli within vasculature. Intravital microscopy experiments were performed using the rat cremaster muscle to visually observe the formation of occlusions. Large gas emboli were produced within the vasculature in the rat cremaster, effectively occluding blood flow. Following these experiments, the therapeutic efficacy of gas embolization was investigated in an ectopic xenograft model of hepatocellular carcinoma in mice. The treatment group exhibited a significantly lower final tumor volume (ANOVA, p = 0.008) and growth rate than control groups - tumor growth was completely halted. Additionally, treated tumors exhibited significant necrosis as determined by histological analysis. To our knowledge, this study is the first to demonstrate the therapeutic efficacy of gas embolotherapy in a tumor model.

Adenosine receptor agonism protects against NETosis and thrombosis in antiphospholipid syndrome.

Potentiation of neutrophil extracellular trap (NET) release is one mechanism by which antiphospholipid antibodies (aPL Abs) effect thrombotic events in patients with antiphospholipid syndrome (APS). Surface adenosine receptors trigger cyclic AMP (cAMP) formation in neutrophils, and this mechanism has been proposed to regulate NETosis in some contexts. Here we report that selective agonism of the adenosine A2A receptor (CGS21680) suppresses aPL Ab-mediated NETosis in protein kinase A-dependent fashion. CGS21680 also reduces thrombosis in the inferior vena cavae of both control mice and mice administered aPL Abs. The antithrombotic medication dipyridamole is known to potentiate adenosine signaling by increasing extracellular concentrations of adenosine and interfering with the breakdown of cAMP. Like CGS21680, dipyridamole suppresses aPL Ab-mediated NETosis via the adenosine A2A receptor and mitigates venous thrombosis in mice. In summary, these data suggest an anti-inflammatory therapeutic paradigm in APS, which may extend to thrombotic disease in the general population.

The temporal correlation of dynamic contrast-enhanced magnetic resonance imaging with tumor angiogenesis in a murine glioblastoma model.

OBJECTIVE: Glioblastoma multiforme (GBM) is a WHO grade IV malignant brain tumor with poor prognosis, despite advances in surgical and adjuvant therapy. GBM is characterized by areas of central necrosis and high levels of angiogenesis, during which increased vascular permeability allows for the extravasation of endothelial progenitor cells to support blood vessel and tumor growth. The purpose of this study was to characterize changes in tumor vascular permeability, vascular density and vessel morphology in vivo during angiogenesis. METHODS: An orthotropic murine (GL26) glioblastoma model was used in this study. in vivo serial dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) in combination with histologic and molecular genetic analyses was performed to correlate in vivo imaging of vascular development. RESULTS: DCE-MRI revealed a significant change in tumor vessel permeability dependent upon tumor progression and size. Time to max signal intensity displayed a stepwise increase between days 21 and 24 (p<0.05), a critical period before exponential tumor growth during which a significant increase in tumor vascular density and vessel caliber is observed on histology. Furthermore, quantitative real-time PCR revealed a corollary increase in angiogenic signaling molecules before the observed changes on DCE-MRI. DISCUSSION: In vivo changes of orthotopic glioma blood vessel permeability as shown by DCE-MRI correlates with histologic quantification of vascular density and vessel caliber as well as with the molecular expression of angiogenic factors. DCE-MRI is a useful tool for non-invasive in vivo monitoring of angiogenesis in pre-clinical tumor models.

Cross-sectional areas of deep/core veins are smaller at lower core body temperatures.

The cardiovascular system plays a crucial role in thermoregulation. Deep core veins, due to their large size and role in returning blood to the heart, are an important part of this system. The response of veins to increasing core temperature has not been adequately studied in vivo. Our objective was to noninvasively quantify in C57BL/6 mice the response of artery-vein pairs to increases in body temperature. Adult male mice were anesthetized and underwent magnetic resonance imaging. Data were acquired from three colocalized vessel pairs (the neck [carotid/jugular], torso [aorta/inferior vena cava (IVC)], periphery [femoral artery/vein]) at core temperatures of 35, 36, 37, and 38°C. Cross-sectional area increased with increasing temperature for all vessels, excluding the carotid. Average area of the jugular, aorta, femoral artery, and vein linearly increased with temperature (0.10, 0.017, 0.017, and 0.027 mm2 /°C, respectively; P < 0.05). On average, the IVC has the largest venous response for area (18.2%/°C, vs. jugular 9.0 and femoral 10.9%/°C). Increases in core temperature from 35 to 38 °C resulted in an increase in contact length between the aorta/IVC of 29.3% (P = 0.007) and between the femoral artery/vein of 28.0% (P = 0.03). Previously unidentified increases in the IVC area due to increasing core temperature are biologically important because they may affect conductive and convective heat transfer. Vascular response to temperature varied based on location and vessel type. Leveraging noninvasive methodology to quantify vascular responses to temperature could be combined with bioheat modeling to improve understanding of thermoregulation.

Update on the electrolytic IVC model for pre-clinical studies of venous thrombosis.

BACKGROUND: The electrolytic inferior vena cava model (EIM) is a murine venous thrombosis (VT) model that produces a non-occlusive thrombus. The thrombus forms in the direction of blood flow, as observed in patients. The EIM is valuable for investigations of therapeutics due to the presence of continuous blood flow. However, the equipment used to induce thrombosis in the original model description was expensive and has since been discontinued. Further, the fibrinolytic system had not been previously studied in the EIM. OBJECTIVES: We aimed to provide an equipment alternative. Additionally, we further characterized the model through mapping the current and time dependency of thrombus resolution dynamics, and investigated the fibrinolytic system from acute to chronic VT. RESULTS: A voltage to current converter powered by a direct current power supply was constructed and validated, providing an added benefit of significantly reducing costs. The current and time dependency of thrombus volume dynamics was assessed by MRI, demonstrating the flexibility of the EIM to investigate both pro-thrombotic and anti-thrombotic conditions. Additionally, the fibrinolytic system was characterized in EIM. Centripetal distribution of plasminogen was observed over time, with peak staining at day 6 post thrombus induction. Both active circulating plasminogen activator inhibitor-1 (PAI-1) and vein wall gene expression of PAI-1 peaked at day 2, coinciding with a relative decrease in tissue plasminogen activator and urokinase plasminogen activator. CONCLUSIONS: The EIM is a valuable model of VT that can now be performed at low cost and may be beneficial in investigations of the fibrinolytic system.

Gas Embolization in a Rodent Model of Hepatocellular Carcinoma Using Acoustic Droplet Vaporization.

Trans-arterial embolization is a commonly used therapy in unresectable hepatocellular carcinoma. Current methods involve the careful placement of an intraarterial catheter and the deposition of embolizing particles. Gas embolotherapy has been proposed as an embolization method with the potential for high spatial resolution without the need for a catheter. This method involves vaporizing intravenouslyadministered droplets into gas bubbles using focused ultrasound - a process termed acoustic droplet vaporization. The bubbles can become lodged in the vasculature, thereby creating an embolus. Here, we initially demonstrate the feasibility of achieving significant targeted embolization with this method in the rat cremaster using intravital microscopy. The therapy was then tested in an ectopic xenograft mouse model of hepatocellular carcinoma. Gas embolotherapy was shown to maintain the tumor volume at baseline over a twoweek treatment course while control groups showed significant tumor growth. These preliminary results demonstrate thatgas embolotherapy could serve as an effective noninvasive method for the management of unresectable hepatocellular carcinoma.

Histotripsy for Non-Invasive Ablation of Hepatocellular Carcinoma (HCC) Tumor in a Subcutaneous Xenograft Murine Model.

Histotripsy fractionates tissue through a mechanical, non-invasive ultrasonic ablation process that precisely controls acoustic cavitation while utilizing real-time ultrasound (US) imaging guidance. This study investigates the potential, feasibility and tumor volume reduction effects of histotripsy for liver cancer ablation in a subcutaneous in vivo murine Hepatocellular Carcinoma (HCC) model. Hep3B tumors were generated in the right flanks of 14 NSG and 7 NOD-SCID mice. The mice were grouped as follows: A (acute, NSG with n=9 treatment and n=1 control), B (chronic, NSG with n=2 treatment and n=2 control) and C (chronic NODSCID, with n=6 treatment and n=1 control). Treatment was performed when the tumor diameters reached >5 mm. 1-2 cycle histotripsy pulses at 100 Hz PRF (p- >30 MPa) were delivered using a custom built 1 MHz therapy transducer attached to a motorized positioner, which scanned the transducer focus to traverse the targeted tumor volume, guided by real-time US imaging. Tumor ablation effectiveness was assessed by obtaining T1, T2 and T2* weighted MR images. Post euthanasia, treated tumor, brain, and lung tissue samples were harvested for histology. Histology of acute group A showed fractionation of targeted region with a sharp boundary separating it from untreated tissue. Groups B and C demonstrated effective tumor volume reduction post treatment on MRI as the homogenate and edema were resorbed within 23 weeks. However, as the tumor was subcutaneous, it was not possible to set adequate treatment margin and since the mice were immune-compromised, residual viable tumor cells eventually developed into tumor regrowth at 3-9 weeks after histotripsy. Groups B and C showed no signs of metastasis in the lung and brain. Our study successfully demonstrated the potential of histotripsy for non-invasive HCC ablation in a subcutaneous murine model. Additional work is ongoing to study the response of histotripsy in immune-competent orthotopic liver tumor models.