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Schematic of swine model. Adapted with permission from McDannold N, King RL, Hynynen K. MRI monitoring of heating pro- duced by ultrasound absorption in the skull: in vivo study in pigs. Magn Reson Med 2004;51:1061– 65.
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The "spot sign" or contrast extravasation is strongly associated with hematoma formation and growth. An animal model of contrast extravasation is important to test existing and novel therapeutic interventions to inform present and future clinical studies. The purpose of this study was to create an animal model of contrast extravasation in acute int...
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... hemorrhage (ICH) accounts for 10%–30% of strokes and is the most deadly and disabling stroke type with little improvement in mortality seen during the past 20 years. 1 These characteristics underscore the importance of developing a better understanding of the pathophysiology of ICH formation and growth to facilitate the development of improved therapeutic agents or interventions. 2 The causative lesion in primary ICH is yet to be elucidated, though pathologic studies demonstrate focal vessel integrity loss in association with blood extravasation into the brain parenchyma. 3 Following initial ICH formation, continuous 4,5 or delayed 6 extravasation results in hematoma expansion, 7 which is associated with early neurologic deterioration and significant mortality. 8 Several recent studies have shown an association between contrast extravasation (CE) detected on CTA, coined the CTA “spot sign,” and hematoma growth. 9-14 Prospective studies have demonstrated that contrast extravasation independently predicts a larger hematoma size and a poorer clinical outcome. 13,14 These are the first clinical studies to suggest a robust “real-time” imaging marker of hematoma expansion. Three clinical studies are presently enrolling patients dichotomized by the CTA spot sign to validate the prior study findings and to determine the therapeutic efficacy of recombinant factor VIIa or tranexamic acid. A more recent study using dynamic spot sign imaging with a biphasic CT perfusion protocol 18 has confirmed 2 patterns of contrast extravasation associated with significantly different rates of leakage. These patterns, comprising a brisker active extravasation (spot sign) and slower postcontrast leakage (PCL), 19 are also demonstrated with early and late structural imaging, 10,19 dynamic CTA/CTP, 18 and biphasic or repeat delayed CTA acquisitions. 12 Morphologic patterns and more recent studies illustrate that the spot sign is not an all-or-none phenomenon but constitutes a spectrum of extravasation. 18,19 The extravasation rate likely significantly impacts timely and clinically meaningful hemostasis. 20 A bleeding threshold likely exists beyond which prothrombotic treatment is futile, exposing patients to harmful adverse effects without hope of therapeutic benefit. 21 Increasingly, new innova- tive surgical techniques are being developed to address contrast extravasation. 22 Knowledge of the impact of the extravasation rate on therapeutic response is critical to stratify patients to the most appropriate therapies. An animal model of acute contrast extravasation in ICH could potentially inform the patient-selection process. We describe a novel MR imaging–integrated real-time swine model of acute hematoma growth and contrast extravasation. This study was conducted with the approval of the local Research Institute Animal Care Committee (Animal Use Protocol No. 12– 435) in compliance with the guidelines established by the Canadian Council on Animal Care and the Animals for Research Act. Fourteen 6- to 8-week-old (approximately 16 –18 kg) Yorkshire male swine were chemically immobilized by using xylazine HCl (2.2 mg/kg), ketamine (15 mg/kg), and atropine (0.05 mg/kg) mixed together in a syringe by intramuscular injection into the dorsal neck or gluteal muscles. Anesthesia was initially maintained with 2%–3% isoflurane in oxygen ( Ͼ 2 L/min). Percutane- ous insertion of two 10-cm 5F catheters (Pinnacle; Terumo, To- kyo, Japan) via a Seldinger technique into the femoral artery and vein was performed for continuous arterial blood pressure monitoring and central line access, respectively. Physiologic monitoring at 5-minute intervals included clinical observation, tail pulse oximetry, rectal temperature, and blood pressure measurement through the femoral artery line. Fluid balance was maintained with intravenous normal saline (10 mL/kg/h) and equivalent normal saline volume replacement of any other losses. The animal was transferred to the MR imaging suite and placed in a supine position with the head immersed in degassed water and its limbs secured (Fig 1). Degassed water served as a conduction medium for sonography emitted from an upwardly positioned modified clinical MR imaging– guided focused sonography brain system (ExAblate 4000; Insightec, Tirat Carmel, Is- rael). The focused sonography system consists of a 1024-element hemispheric phased array with a diameter of 30 cm. Accurate anatomic localization of vessels within the basal ganglia was achieved by using MR imaging localization sequences. The basal ganglia represent a frequent site for ICH and are a commonly targeted location for experimental ICH models due to lower subarachnoid risk and intraventricular extension. Following this procedure, 0.08-mL/kg (1.5 mL) perflutren lipid microsphere infusion (Definity; Lantheus Medical Imaging, North Billerica, Massachusetts) and focused low-frequency sonography (230 kHz) were delivered. MR imaging was performed before and after focused sonography vessel disruption by using 3T MR imaging (Discovery MR750; GE Healthcare, Mil- waukee, Wisconsin) and a 6-channel phased array flexible coil (GE Healthcare). The following sequences were obtained at baseline: FSE T2 (TR/TE, 3000/71 ms; echo-train length, 4; FOV, 16 ϫ 16 cm; slice thickness, 2 mm; 256 ϫ 192); 3D spoiled gradient recalled-echo (GRE) T1 (TR/TE, 6/2 ms; FOV, 12 ϫ 12 cm; slice thickness, 2 mm); axial T2 FLAIR (TR/TE, 8000/127 ms; 12 ϫ 12 cm; slice thickness, 2 mm; 128 ϫ 128); and fast GRE (TR/TE, 100/13 ms; 12 ϫ 12 cm; slice thickness, 2 mm). Thirty seconds after perflutren injection and sonographic insonation, a dynamic contrast-enhanced sequence (DCE) (3D T1 spoiled GRE; TR/TE, 6/2 ms; 12 ϫ 12 cm; slice thickness, 3 mm; 128 ϫ 128; temporal sampling, 10 seconds; scan duration, 20 minutes) was performed. Gadobutrol, 1 mmol/mL (Gadovist; Bayer Schering Pharma, Toronto, Canada), was injected via a pump injector (Spectris MR injector; MedRad, Indianola, Pennsylvania; and Solaris; Bayer HealthCare, Pittsburgh, Pennsylvania) at 0.5 mL/kg and 5 mL/s followed by a 10-mL normal saline bolus. A postgadolinium T1 study was performed at the termination of the DCE sequence (20 minutes after gadolinium injection). Fast GRE, FLAIR, and FSE T2 sequences were repeated following DCE and before procedure termination. Number Power Delivery. of Experiments. On the basis Twenty-eight of prior studies hemispheres of blood-brain were bar- insonated; rier disruption 16 hemispheres 23-25 aiming were at a used temporary to determine BBB opening optimal but power re- porting delivery; complications and 12, to test of the red effect blood of burst cell extravasation, length. 2 power ranges were selected for evaluation. Sixteen hemispheres were subjected to basal ganglia insonation, with either 70-kW ( n ϭ 8 hemispheres) or 80-kW ( n ϭ 8 hemispheres) power delivery at a Power Delivery. On the basis of prior studies of blood-brain barrier disruption 23-25 aiming at a temporary BBB opening but re- porting complications of red blood cell extravasation, 2 power ranges were selected for evaluation. Sixteen hemispheres were subjected to basal ganglia insonation, with either 70-kW ( n ϭ 8 hemispheres) or 80-kW ( n ϭ 8 hemispheres) power delivery at a constant burst length (10 ms, 1-Hz pulse-repetition frequency) and sonication duration (120 seconds). Animals were compared for DCE leakage rate, final hematoma size (see below), and vascular abnormality on histopathology. The 8 hemispheres at 80 kW were then used to compare with the 3 higher burst lengths also performed at 80 kW. MR Image Processing. Hematoma and DCE contrast volume measurements were performed on GRE and DCE images, respectively, by using the auto-trace feature with Medical Image Processing, Analysis, and Visualization, Version 4.4.1, Center for Information Technology (National Institutes of Health, Bethesda, Maryland; ). The largest re- gion-of-interest volume within the temporal DCE dataset was selected as the final DCE volume. Rate of leakage (KPS) was measured within each region of contrast leakage from the permeability maps calculated using custom IDL software (Exelis Visual Information Solutions, Boulder, Colorado) by using a previously described unidirectional 2-compartment kinetic model. 28 The DCE-derived region of interest, representing the volume of contrast leakage, was used for the permeability measurement from the KPS maps. Because the contrast leakage volumes were drawn on the same sequence from which the KPS maps were derived, the ROIs were intrinsically coregistered with the KPS maps. Morphologic Characterization of Contrast Extravasation. On the basis of clinical studies, 2 distinct patterns of contrast leakage were identified. 19 Active contrast extravasation demonstrated a brisk contrast leak with well-delineated margins, visible from early within the DCE sequence. Postcontrast leakage demonstrated slower leakage on DCE with ill-defined contrast enhancement becoming more confluent on a postcontrast T1 study performed following DCE (Figs 2 and ...
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Citations
... Extravasation on MRI was found to be closely correlated with HE, indicating ongoing bleeding. Aviv et al. (89) later developed an animal model of contrast extravasation (SpS) in acute ICH based on MRI, but no significant correlation was found between SpS and HE. Since there was no corresponding MRI marker for SpS at the time, Katharina et al. (90) conducted further research and found that SpS could be detected using post-contrast T1-weighted and dynamic T1-weighted MRI images. ...
Introduction
Stroke is a major global health concern and is ranked as the second leading cause of death worldwide, with the third highest incidence of disability. Intracerebral hemorrhage (ICH) is a devastating form of stroke that is responsible for a significant proportion of stroke-related morbidity and mortality worldwide. Hematoma expansion (HE), which occurs in up to one-third of ICH patients, is a strong predictor of poor prognosis and can be potentially preventable if high-risk patients are identified early. In this review, we provide a comprehensive summary of previous research in this area and highlight the potential use of imaging markers for future research studies.
Recent advances
Imaging markers have been developed in recent years to aid in the early detection of HE and guide clinical decision-making. These markers have been found to be effective in predicting HE in ICH patients and include specific manifestations on Computed Tomography (CT) and CT Angiography (CTA), such as the spot sign, leakage sign, spot-tail sign, island sign, satellite sign, iodine sign, blend sign, swirl sign, black hole sign, and hypodensities. The use of imaging markers holds great promise for improving the management and outcomes of ICH patients.
Conclusion
The management of ICH presents a significant challenge, and identifying high-risk patients for HE is crucial to improving outcomes. The use of imaging markers for HE prediction can aid in the rapid identification of such patients and may serve as potential targets for anti-HE therapies in the acute phase of ICH. Therefore, further research is needed to establish the reliability and validity of these markers in identifying high-risk patients and guiding appropriate treatment decisions.
... The premise of MB-mediated therapy is that MBs entering the localized region of the FUS beam undergo pressure-dependent oscillations and interact with tissue at time-averaged acoustic power levels that are over two orders of magnitude lower than the threshold without them [1,2], and at more predictable pressures [3]. Compelling studies have highlighted the ability to elicit enhanced microvascular permeability [4,5], inflammation [6], apoptosis [7], necrosis [8], thrombolysis [9,10], thrombogenesis [11], angiogenesis [12], vessel rupture [13], and vascular shutdown [14,15]. ...
Rationale: Focused ultrasound-stimulated microbubbles have been shown to be capable of inducing blood flow shutdown and necrosis in a range of tissue types in an approach termed antivascular ultrasound or nonthermal ablation. In oncology, this approach has demonstrated tumor growth inhibition, and profound synergistic antitumor effects when combined with traditional platforms of chemo-, radiation- and immune-therapies. However, the exposure schemes employed have been broad and underlying mechanisms remain unclear with fundamental questions about exposures, vessel types and sizes involved, and the nature of bubble behaviors and their acoustic emissions resulting in vascular damage - impeding the establishment of standard protocols.
Methods: Here, ultrasound transmitters and receivers are integrated into a murine dorsal window chamber tumor model for intravital microscopy studies capable of real-time visual and acoustic monitoring during antivascular ultrasound. Vessel type (normal and tumor-affected), caliber, and viability are assessed under higher pressure conditions (1, 2, and 3 MPa), and cavitation signatures are linked to the biological effects.
Results: Vascular events occurred preferentially in tumor-affected vessels with greater incidence in smaller vessels and with more severity as a function of increasing pressure. Vascular blood flow shutdown was found to be due to a combination of focal disruption events and network-related flow changes. Acoustic emissions displayed elevated broadband noise and distinct sub- and ultra-harmonics and their associated third-order peaks with increasing pressure.
Conclusions: The observed vascular events taken collectively with identified cavitation signatures provide an improved mechanistic understanding of antivascular ultrasound at the microscale, with implications for establishing a specific treatment protocol and control platform.
... The premise of MB-mediated therapy is that MBs entering the localized region of the FUS beam undergo pressure-dependent oscillations and interact with tissue at time-averaged acoustic power levels that are over two orders of magnitude lower than the threshold without them [1,2], and at more predictable pressures [3]. Compelling studies have highlighted the ability to elicit enhanced microvascular permeability [4,5], inflammation [6], apoptosis [7], necrosis [8], thrombolysis [9,10], thrombogenesis [11], angiogenesis [12], vessel rupture [13], and vascular shutdown [14,15]. ...
Focused ultrasound stimulated microbubbles have been shown to be capable of inducing blood flow shutdown and necrosis in a range of tissue types in an approach termed antivascular ultrasound or mechanical ablation. In oncology, this approach has demonstrated tumor growth inhibition, and profound synergistic antitumor effects when combined with traditional platforms of chemo, radiation and immune therapies. However, the exposure schemes employed have been broad and underlying mechanisms remain unclear with fundamental questions about exposures, vessel types and sizes involved, and the nature of bubble behaviors and their acoustic emissions resulting in vascular damage, impeding the establishment of standard protocols. Here, ultrasound transmitters and receivers are integrated into a murine dorsal window chamber tumor model for intravital microscopy studies capable of real time visual and acoustic monitoring during antivascular ultrasound. Vessel type (normal and tumor affected), caliber, and viability are assessed under higher pressure conditions (1, 2, and 3 MPa), and cavitation signatures are linked to the biological effects. Vascular events occurred preferentially in tumor affected vessels with greater incidence in smaller vessels and with more severity as a function of increasing pressure. Vascular blood flow shutdown was found to be due to a combination of focal disruption events and network related flow changes. Acoustic emissions displayed elevated broadband noise and distinct sub and ultra harmonics and their associated third order peaks with increasing pressure. The observed vascular events taken collectively with identified cavitation signatures provide an improved mechanistic understanding of antivascular ultrasound at the microscale, with implications for establishing a specific treatment protocol and control platform.
... The CTA spot sign and contrast extravasation were considered to be independent risks for hematoma expansion and poor prognosis (Schindlbeck et al. 2016). In an experimental ICH model of swine constructed by the focused sonography technique, Aviv et al. quantified the contrast extravasation rate in real-time, indicating that the values were consistent with the contrast extravasation rate reported in ICH patients (Aviv et al. 2014;D'Esterre et al. 2011). Conceivably, by constructing such experimental animal models and evaluating the core neuroimaging markers affecting hematoma expansion, a transformation from basic research to medical guidelines for ICH patients is expected. ...
Stroke has become the second leading cause of death in people aged higher than 60 years, with cancer being the first. Intracerebral hemorrhage (ICH) is the most lethal type of stroke. Using imaging techniques to evaluate the evolution of intracranial hematomas in patients with hemorrhagic stroke is worthy of ongoing research. The difficulty in obtaining ultra-early imaging data and conducting intensive dynamic radiographic imaging in actual clinical settings has led to the application of experimental animal models to assess the evolution of intracranial hematomas. Herein, we review the current knowledge on primary intracerebral hemorrhage mechanisms, focus on the progress of animal studies related to hematoma development and secondary brain injury, introduce preclinical therapies, and summarize related challenges and future directions.
... Still, several reports have examined the association between MRI biomarkers and ICH growth yielding to date two important contributions to ICH expansion research. First, fast-paced MRI sequences have allowed to visualize the dynamics of active intracerebral contrast leakage in MRI both in animal models [47], and anecdotally in human [48], providing pathophysiological insight into the phenomena underlying HE. Second, MRI allows to assess the association between pre-existing cerebral small vessel disease (cSVD) parenchymal damage, and the acute growth and final volume of ICH ( Figure 3). ...
Intracerebral hemorrhage (ICH) accounts for 10% to 20% of all strokes worldwide and is associated with high morbidity and mortality. Neuroimaging is clinically important for the rapid diagnosis of ICH and underlying etiologies, but also for identification of ICH expansion, often as-sociated with an increased risk for poor outcome. In this context, rapid assessment of early hema-toma expansion risk is both an opportunity for therapeutic intervention and a potential hazard for hematoma evacuation surgery. In this review, we provide an overview of the current literature surrounding the use of multimodal neuroimaging of ICH for etiological diagnosis, prediction of early hematoma expansion, and prognostication of neurological outcome. Specifically, we discuss standard imaging using computed tomography, the value of different vascular imaging modalities to identify underlying causes and present recent advances in magnetic resonance imaging and computed tomography perfusion.
... Still, several reports have examined the association between MRI biomarkers and ICH growth yielding to date two important contributions to ICH expansion research. First, fast-paced MRI sequences have allowed to visualize the dynamics of active intracerebral contrast leakage in MRI both in animal models [47], and anecdotally in human [48], providing pathophysiological insight into the phenomena underlying HE. Second, MRI allows to assess the association between pre-existing cerebral small vessel disease (cSVD) parenchymal damage, and the acute growth and final volume of ICH ( Figure 3). ...
... The initial report of sonographic disruption of the blood-brain barrier in swine was performed by Aviv et al. in 2014. In this study, they targeted the vessels within the basal ganglia of young swine using MRI-guided sonography with the objective of generating a model of ICH that allowed them to quantify hematoma through the MRI detection of extravasated contrast [166]. Rupture of intracerebral vessels is thus performed with the magnetic resonance-guided focused ultrasound (MRgFUS), a technique used to perform minimally invasive targeted tissue surgeries based on the thermal ablation caused by focused ultrasounds at determined frequencies. ...
In the search of animal stroke models providing translational advantages for biomedical research, pigs are large mammals with interesting brain characteristics and wide social acceptance. Compared to rodents, pigs have human-like highly gyrencephalic brains. In addition, increasingly through phylogeny, animals have more sophisticated white matter connectivity; thus, ratios of white-to-gray matter in humans and pigs are higher than in rodents. Swine models provide the opportunity to study the effect of stroke with emphasis on white matter damage and neuroanatomical changes in connectivity, and their pathophysiological correlate. In addition, the subarachnoid space surrounding the swine brain resembles that of humans. This allows the accumulation of blood and clots in subarachnoid hemorrhage models mimicking the clinical condition. The clot accumulation has been reported to mediate pathological mechanisms known to contribute to infarct progression and final damage in stroke patients. Importantly, swine allows trustworthy tracking of brain damage evolution using the same non-invasive multimodal imaging sequences used in the clinical practice. Moreover, several models of comorbidities and pathologies usually found in stroke patients have recently been established in swine. We review here ischemic and hemorrhagic stroke models reported so far in pigs. The advantages and limitations of each model are also discussed.
... The biological outcomes resulting from ultrasound-stimulated microbubble activity in vivo are dependent on several factors related to the sonication scheme (i.e., frequency, peak negative pressure, burst length, burst repetition frequency, exposure duration), the microbubble contrast agent (e.g., gas core and shell composition, size distribution, administration method), and the local environment (e.g., tissue/tumor vascularity, blood-oxygen level, blood viscosity, ambient over-pressure) [37]. Moreover, the range of exposure conditions over which the desired bioeffects can be induced within the targeted region(s) without causing excessive and/or off-target cavitation effects is relatively narrow [28,29,[38][39][40][41], with potential for blood vessel rupture in cases of overexposure [42]. Furthermore, FUS parameter selection is challenging during brain applications as substantial variability in transcranial ultrasound transmission efficiency [43,44] and imperfect trans-skull focusing via current aberration correction approaches [6] complicate estimation of the pressure fields generated in situ from a fixed transducer output. ...
Transcranial magnetic resonance imaging (MRI)-guided focused ultrasound (FUS) thermal ablation is under clinical investigation for non-invasive neurosurgery, though its use is restricted to central brain targets due primarily to skull heating effects. The combination of FUS and contrast agent microbubbles greatly reduces the ultrasound exposure levels needed to ablate brain tissue and may help facilitate the use of transcranial FUS ablation throughout the brain. However, sources of variability exist during microbubble-mediated FUS procedures that necessitate the continued development of systems and methods for online treatment monitoring and control, to ensure that excessive and/or off-target bioeffects are not induced from the exposures.
Methods: Megahertz-rate three-dimensional (3D) microbubble imaging in vivo was performed during nonthermal ablation in rabbit brain using a clinical-scale prototype transmit/receive hemispherical phased array system.
Results: In-vivo volumetric acoustic imaging over microsecond timescales uncovered spatiotemporal microbubble dynamics hidden by conventional whole-burst temporal averaging. Sonication-aggregate ultrafast 3D source field intensity data were predictive of microbubble-mediated tissue damage volume distributions measured post-treatment using MRI and confirmed via histopathology. Temporal under-sampling of acoustic emissions, which is common practice in the field, was found to impede performance and highlighted the importance of capturing adequate data for treatment monitoring and control purposes.
Conclusion: The predictive capability of ultrafast 3D microbubble imaging, reported here for the first time, will enable future microbubble-mediated FUS treatments with unparalleled precision and accuracy, and will accelerate the clinical translation of nonthermal tissue ablation procedures both in the brain and throughout the body.
... Since our study was a small case series, we need more clinical experience using DSC-MRI for detecting the spot sign in acute ICH, and further imaging sequences such as CE-T1 may be required to complement DSC-MRI, especially if TTI is more delayed than 4-6 hours when ICH may have dark SI on DSC-MRI. In T1WI, hyperacute ICH usually has iso-SI, and a GBCA leak may appear as a bright spot on CE-T1 (9)(10)(11)(12). Finally, in terms of the radiation hazards, MRI may be preferable to CTA or another combined CT Although an MRI examination requires a longer acquisition time than does a CT examination, and may not be a possible option for an unstable patient, a combination of limited sequences, such as DSC-MRI, T2*-GRE, and CE-T1WI with a fast imaging technique, can reduce the acquisition time to less than 5 minutes. ...
... (Schook et al. 2005) The swine ICH model has been reported in many non-RNAseq investigations. (Aviv et al. 2014;Brunberg et al. 2013;Gu et al. 2011;Loftspring et al. 2007;Orakcioglu et al. 2015;Orakcioglu et al. 2012;Wagner et al. 2006;Xi et al. 1998;Zhou et al. 2014) and has advantages versus rodents such as the larger size of the pig brain, approximately 40 times larger than a rat, its gyrencephalic nature, larger white matter composition with better evaluation of damage and edema, (Adeoye et al. 2011;Wagner 2007;Wagner et al. 1996) and ease of experimental reproducibility. (Hua and Xi 2009). ...
Intracerebral hemorrhage (ICH) is a severe neurological disorder with no proven treatment. Our prior research identified a significant association with monocyte level and ICH mortality. To advance our understanding, we sought to identify gene expression after ICH using a swine model to test the hypothesis that ICH would induce peripheral blood mononuclear cell (PBMC) gene expression. In 10 pigs with ICH, two PBMC samples were drawn from each with the first immediately prior to ICH induction and the second six hours later. RNA-seq was performed with subsequent bioinformatics analysis using Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and Ingenuity® Pathway Analysis (IPA). There were 182 significantly upregulated and 153 significantly down-regulated differentially expressed genes (DEGs) after ICH. Consistent with findings in humans, significant GO and KEGG pathways were primarily related to inflammation and the immune response. Five genes, all upregulated post-ICH and known to be associated with monocyte activation, were repeatedly DEGs in the significant KEGG pathways: CD14, TLR4, CXCL8, IL-18, and CXCL2. In IPA, the majority of upregulated disease/function categories were related to inflammation and immune cell activation. TNF and LPS were the most significantly activated upstream regulators, and ERK was the most highly connected node in the top network. ICH induced changes in PBMC gene expression within 6 h of onset related to inflammation, the immune response, and, more specifically, monocyte activation. Further research is needed to determine if these changes affect outcomes and may represent new therapeutic targets.
