Search

Purpose:Pancreatic ductal adenocarcinoma (PDA) is a common, deadly cancer that is challenging both to diagnose and to manage. Its hallmark is an expansive, desmoplastic stroma characterized by high mechanical stiffness. In this study, we sought to leverage this feature of PDA for two purposes: differential diagnosis and monitoring of response to treatment.Experimental Design:Harmonic motion imaging (HMI) is a functional ultrasound technique that yields a quantitative relative measurement of stiffness suitable for comparisons between individuals and over time. We used HMI to quantify pancreatic stiffness in mouse models of pancreatitis and PDA as well as in a series of freshly resected human pancreatic cancer specimens.Results:In mice, we learned that stiffness increased during progression from preneoplasia to adenocarcinoma and also effectively distinguished PDA from several forms of pancreatitis. In human specimens, the distinction of tumors versus adjacent pancreatitis or normal pancreas tissue was even more stark. Moreover, in both mice and humans, stiffness increased in proportion to tumor size, indicating that tuning of mechanical stiffness is an ongoing process during tumor progression. Finally, using a brca2–mutant mouse model of PDA that is sensitive to cisplatin, we found that tissue stiffness decreases when tumors respond successfully to chemotherapy. Consistent with this observation, we found that tumor tissues from patients who had undergone neoadjuvant therapy were less stiff than those of untreated patients.Conclusions:These findings support further development of HMI for clinical applications in disease staging and treatment response assessment in PDA.

Supplementary Methods

Figure S1

Soil carbon sequestration (SCS) practices on French agricultural land are part of the portfolio of actions available to policymakers in the field of climate change mitigation and are central to the success of the “4 per 1,000” initiative, launched by France in 2015. To date, there has been limited research considering their applicability to vineyards. A survey was circulated to 506 French winegrowers to identify the adoption rate of six SCS practices in the viticultural sector (applying organic amendments, using biochar, returning pruning residues to the soil, no-tillage, cover cropping, and introducing or preserving hedges in the vineyard) and to explore motives and barriers to adoption. The survey also investigated ways of overcoming barriers to adoption and winegrowers' perception of agri-environment measures. Differences in motivations and barriers between SCS practices were found, and winegrowers themselves suggested a need for improved communication of evidence about SCS practices and better-targeted policy incentives to support adoption.

Urban agriculture can contribute to food security, food system resilience and sustainability at the city level. Whilst studies have examined urban agricultural productivity, we lack systemic knowledge of how agricultural productivity of urban systems compares to conventional agriculture and how productivity varies for different urban spaces (e.g., allotments vs. rooftops vs. indoor farming) and growing systems (e.g., hydroponics vs. soil-based agriculture). Here, we present a global meta-analysis that seeks to quantify crop yields of urban agriculture for a broad range of crops and explore differences in yields for distinct urban spaces and growing systems. We found 200 studies reporting urban crop yields, from which 2,062 observations were extracted. ‘Lettuces and chicories’ were the most studied urban grown crops. We observed high agronomic suitability of urban areas, with urban agricultural yields on par with or greater than global average conventional agricultural yields. ‘Cucumbers and gherkins’ were the category of crops for which differences in yields between urban and conventional agriculture were the greatest (17 kg m-2 cycle-1 vs. 3.8 kg m-2 cycle-1). Some urban spaces and growing systems also had a significant effect on specific crop yields (e.g., tomato yields in hydroponic systems were significantly greater than tomato yields in soil-based systems). This analysis provides a more robust, globally-relevant evidence base on the productivity of urban agriculture that can be used in future research and practice relating to urban agriculture, especially in scaling-up studies aiming to estimate the self-sufficiency of cities and towns and their potential to meet local food demand.

The adoption of soil organic carbon sequestration (SCS) practices on agricultural land offers the double advantage of offsetting greenhouse gas (GHG) emissions and improving soil quality. However, little is known about the drivers that might influence winegrowers to adopt these practices, whose uptake remains low on viticultural land. Better understanding these drivers will be crucial to evaluate the efficacy of current policies in the viticulture sector in promoting and incentivising soil organic carbon sequestration in vineyards. This paper identified the factors influencing the adoption of SCS practices by winegrowers in France. A survey of 400 winegrowers investigated current rates of adoption and winegrowers’ perceptions of the practices. A binary logistic model suggested that winegrower’s age, being an independent winegrower, farm size, the number of workers hired, vine’s age, being certified High Environmental Value (HVE), being certified organic, practising irrigation, receiving subsidies, and winegrower’s perceived resources, attitude towards SCS practices and confidence significantly influenced the decision to adopt the practices, though their influence differed depending on the practice. The findings provide insights for GHG mitigation planning targeting the viticulture sector

Purpose: Pancreatic ductal adenocarcinoma (PDA) is a common, deadly cancer that is challenging both to diagnose and to manage. Its hallmark is an expansive, desmoplastic stroma characterized by high mechanical stiffness. In this study, we sought to leverage this feature of PDA for two purposes: differential diagnosis and monitoring of response to treatment. Experimental Design: Harmonic motion imaging (HMI) is a functional ultrasound technique that yields a quantitative relative measurement of stiffness suitable for comparisons between individuals and over time. We used HMI to quantify pancreatic stiffness in mouse models of pancreatitis and PDA as well as in a series of freshly resected human pancreatic cancer specimens. Results: In mice, we learned that stiffness increased during progression from preneoplasia to adenocarcinoma and also effectively distinguished PDA from several forms of pancreatitis. In human specimens, the distinction of tumors versus adjacent pancreatitis or normal pancreas tissue was even more stark. Moreover, in both mice and humans, stiffness increased in proportion to tumor size, indicating that tuning of mechanical stiffness is an ongoing process during tumor progression. Finally, using a brca2–mutant mouse model of PDA that is sensitive to cisplatin, we found that tissue stiffness decreases when tumors respond successfully to chemotherapy. Consistent with this observation, we found that tumor tissues from patients who had undergone neoadjuvant therapy were less stiff than those of untreated patients. Conclusions: These findings support further development of HMI for clinical applications in disease staging and treatment response assessment in PDA.

Angiogenesis, the formation of new vessels, is one of the key mechanisms in tumor development and an appealing target for therapy. Non-invasive, high-resolution, high-sensitivity, quantitative 3-D imaging techniques are required to correctly depict tumor heterogeneous vasculature over time. Ultrafast Doppler was recently introduced and provides an unprecedented combination of resolution, penetration depth and sensitivity without requiring any contrast agents. The technique was further extended to three dimensions with ultrafast Doppler tomography (UFD-T). In this work, UFD-T was applied to the monitoring of tumor angiogenesis in vivo, providing structural and functional information at different stages of development. UFD-T volume renderings revealed that our murine model's vasculature stems from pre-existing vessels and sprouts to perfuse the whole volume as the tumor grows until a critical size is reached. Then, as the network becomes insufficient, the tumor core is no longer irrigated because the vasculature is concentrated mainly in the periphery. In addition to spatial distribution and growth patterns, UFD-T allowed a quantitative analysis of vessel size and length, revealing that the diameter distribution of vessels remained relatively constant throughout tumor growth. The network is dominated by small vessels at all stages of tumor development, with more than 74% of the vessels less than 200 µm in diameter. This study also found that cumulative vessel length is more closely related to tumor radius than volume, indicating that the vascularization becomes insufficient when a critical mass is reached. UFD-T was also compared with dynamic contrast-enhanced ultrasound and found to provide complementary information regarding the link between structure and perfusion. In conclusion, UFD-T is capable of in vivo quantitative assessment of the development of tumor vasculature (vessels with blood speed >1 mm/s [sensitivity limit] assessed with a resolution limit of 80 µm) in 3 dimensions. The technique has very interesting potential as a tool for treatment monitoring, response assessment and treatment planning for optimal drug efficiency.

Purpose Noninvasive quantitative assessment of coagulated tissue during high-intensity focused ultrasound (HIFU) ablation is one of the essential steps for tumor treatment, especially in such cases as the Pancreatic Ductal Adenocarcinoma (PDA) that has low probability of diagnosis at the early stages and high probability of forming solid carcinomas resistant to chemotherapy treatment at the late stages. Methods Harmonic motion elastography (HME) is a technique for the localized estimation of tumor stiffness. This harmonic motion imaging (HMI)-based technique is designed to map the tissue Young's modulus or stiffness noninvasively. A focused ultrasound (FUS) transducer generates an oscillating, acoustic radiation force in its focal region. The two-dimensional (2D) shear wave speed, and consequently the Young's modulus maps, is generated by tracking the radio frequency (RF) signals acquired at high frame rates. By prolonging the sonication for more than 50 s using the same methodology, the 2D Young's modulus maps are reconstructed while HIFU is applied and ablation is formed on PDA murine tumors. Results The feasibility of this technique in measuring the regional Young's modulus was first assessed in tissue-mimicking phantoms. The contrast-to-noise ratio (CNR) was found to be higher than 11.7 dB for each 2D reconstructed Young's modulus map. The mean error in this validation study was found to be equal to less than 19%. Then HME was applied on two transgenic mice with pancreatic ductal adenocarcinoma tumors. The Young's modulus median value of this tumor at the start of the HIFU application was equal to 2.1 kPa while after 45 s of sonication it was found to be approximately three times stiffer (6.7 kPa). Conclusions The HME was described herein and showed its capability of measuring tissue stiffness noninvasively by measuring the shear wave speed propagation inside the tissue and reconstructing a 2D Young's modulus map. Application of the methodology in vivo and during HIFU were thus reported here for the first time.

To limit warming to well below 2°C, most scenario projections rely on greenhouse gas removal technologies (GGRTs); one such GGRT uses soil carbon sequestration (SCS) in agricultural land. In addition to their role in mitigating climate change, SCS practices play a role in delivering agroecosystem resilience, climate change adaptability and food security. Environmental heterogeneity and differences in agricultural practices challenge the practical implementation of SCS, and our analysis addresses the associated knowledge gap. Previous assessments have focused on global potentials, but there is a need among policymakers to operationalise SCS. Here, we assess a range of practices already proposed to deliver SCS, and distil these into a subset of specific measures. We provide a multidisciplinary summary of the barriers and potential incentives towards practical implementation of these measures. First, we identify specific practices with potential for both a positive impact on SCS at farm level and an uptake rate compatible with global impact. These focus on: (a) optimising crop primary productivity (e.g. nutrient optimisation, pH management, irrigation); (b) reducing soil disturbance and managing soil physical properties (e.g. improved rotations, minimum till); (c) minimising deliberate removal of C or lateral transport via erosion processes (e.g. support measures, bare fallow reduction); (d) addition of C produced outside the system (e.g. organic manure amendments, biochar addition); (e) provision of additional C inputs within the cropping system (e.g. agroforestry, cover cropping). We then consider economic and non‐cost barriers and incentives for land managers implementing these measures, along with the potential externalised impacts of implementation. This offers a framework and reference point for holistic assessment of the impacts of SCS. Finally, we summarise and discuss the ability of extant scientific approaches to quantify the technical potential and externalities of SCS measures, and the barriers and incentives to their implementation in global agricultural systems., To limit warming to well below 2°C, many scenario projections rely on greenhouse gas removal by soil carbon sequestration (SCS) in agricultural land. Here, we provide a multidisciplinary summary of the barriers and potential incentives towards practical implementation SCS. First, we identify specific practices with potential for both a positive impact on SCS and an uptake rate compatible with global impact. Then, we then consider economic and non‐cost barriers and incentives for implementing SCS, and potential externalised impacts of implementation. Finally, we discuss the ability of existing methods to model the potential of SCS in global agricultural systems.

Hemodynamic functional ultrasound imaging (fUS) of neural activity provides a unique combination of spatial coverage, spatiotemporal resolution and compatibility with freely moving animals. However, deep and transcranial monitoring of brain activity and the imaging of dynamics in slow-flowing blood vessels remains challenging. To enhance fUS capabilities, we introduce biomolecular hemodynamic enhancers based on gas vesicles (GVs), genetically encodable ultrasound contrast agents derived from buoyant photosynthetic microorganisms. We show that intravenously infused GVs enhance ultrafast Doppler ultrasound contrast and visually-evoked hemodynamic contrast in transcranial fUS of the mouse brain. This hemodynamic contrast enhancement is smoother than that provided by conventional microbubbles, allowing GVs to more reliably amplify neuroimaging signals.

Background and aims: Poor specificity and predictive values of current cross-sectional radiological imaging methods in evaluation of pancreatic adenocarcinoma (PDAC) limit the clinical capability to accurately stage the tumor pre-operatively and provide optimal surgical treatment and improve patient outcomes. Methods: In this study, we applied Harmonic Motion Elastography (HME), a quantitative ultrasound-based imaging method to calculate Young's modulus (YM) in PDAC mouse models (n = 30) and human pancreatic resection specimens of PDAC (n=32). We compared the YM to the collagen assessment by Picrosirius red (PSR) stain on corresponding histologic sections. Results: HME is capable of differentiating between different levels of fibrosis in transgenic mice. In mice without pancreatic fibrosis, the measured YM was 4.2 ± 1.3 kPa, in fibrotic murine pancreata, YM was 5.5 ± 2.0 kPa and in murine PDAC tumors, YM was 11.3 ± 1.7 kPa. The corresponding PSR values were 2.0 ± 0.8 %, 9.8 ± 3.4 %, and 13.2 ± 1.2%, respectively. In addition, three regions within each human surgical PDAC specimen were assessed: tumor, which had both the highest Young's modulus (YM > 40 kPa) and collagen density (PSR > 40 %); non-neoplastic adjacent pancreas, which had the lowest Young's modulus (YM < 15 kPa) and collagen density (PSR < 10%) and a transitional peri-lesional region between the tumor and non-neoplastic pancreas with an intermediate value of measured Young's modulus (15 kPa < YM < 40 kPa) and collagen density (15% < PSR < 35 %). Conclusion: In conclusion, a non-invasive, quantitative imaging tool for detecting, staging and delineating PDAC tumor margins based on the change in collagen density was developed.

The majority of disease processes involves changes in the micro-structure of the affected tissue, which can translate to changes in the mechanical properties of the corresponding tissue. Harmonic motion imaging (HMI) is an elasticity imaging technique that allows the study of the mechanical parameters of tissue by detecting the tissue response by a harmonic motion field, which is generated by oscillatory acoustic radiation force. HMI has been demonstrated in tumor detection and characterization as well as monitoring of ablation procedures. In this study, an analytical HMI model is demonstrated and compared with a finite element model (FEM), allowing rapid and accurate computation of the displacement, strain, and shear wave velocity (SWV) at any location in a homogenous linear elastic material. Average absolute differences between the analytical model and the FEM were respectively 1.2% for the displacements and 0.5% for the strains for 41 940 force voxels at 0.22 s per displacement evaluation. A convergence study showed that the average difference could be further decreased to 1.0% and 0.15% for the displacements and strains, respectively, if force resolution is increased. SWV fields, as calculated with the FEM and the analytical model, have regional differences in velocities up to 0.57 m s−1 with an average absolute difference of 0.11 ± 0.07 m s−1, primarily due to imperfections in the non-reflecting FEM boundary conditions. The apparent SWV differed from the commonly used plane-wave approximation by up to 1.2 m s−1 due to near and intermediate field effects. Maximum displacement amplitudes for a model with an inclusion stabilize within 10% of the homogenous model at an inclusion radius of 10 mm while the maximum strain reacts faster, stabilizing at an inclusion radius of 3 mm. In conclusion, an analytical model for HMI stiffness estimation is presented in this paper. The analytical model has advantages over FEM as the full-field displacements do not need to be calculated to evaluate the model at a single measurement point. This advantage, together with the computational speed, makes the analytical model useful for real-time imaging applications. However, the analytical model was found to have restrictive assumptions on tissue homogeneity and infinite dimensions, while the FEM approaches were shown adaptable to variable geometry and non-homogenous properties.

Sensitivity of contrast-enhanced ultrasound (CEUS) to microvascular flow modifications can be limited by intra-injection variability (injected dose, rate, volume). To evaluate the effect of injection variability on microvascular flow evaluation, CEUS was compared between controlled and manual injections where enhancement was assessed in vitro within a flow phantom, in normal murine kidney (N = 12) and in murine ectopic tumors (N = 10). For both in vitro and in vivo measurements in the renal cortex, controlled injections significantly improved reproducibility of functional parameter estimation. Their coefficient of variation (CV) in the renal cortex ranged from 4 to 19 % for controlled injection vs. 5 to 43 % for manual injections. For measurements in tumors, controlled injection only decreased the CV significantly for the mean transit time. In tumors, multiple injections of contrast agent with a 15-min delay between each were shown to strongly modify contrast uptake by facilitating penetration of microbubbles. Improved reproducibility of CEUS assessments in murine models should provide more robust quantification of flow parameters and more sensitive evaluation of tumor modifications in therapeutic models.

Harmonic Motion Imaging (HMI) is a radiation-force-based ultrasound elasticity imaging technique, which is designed for both tissue relative stiffness imaging and reliable high intensity focused ultrasound (HIFU) treatment monitoring. The objective of this study is to develop and demonstrate the feasibility of 2D focused ultrasound (FUS) beam steering for HMI using a 93-element, FUS phased array. HMI with steered FUS beam was acquired in tissue-mimicking phantoms. The HMI displacement was imaged within the steering range of +/−1.7 mm laterally and +/−2 mm axially. Using the steered FUS beam, HMI can be used to image a larger tissue volume with higher efficiency and without requiring mechanical movement of the transducer.

Vineyards are usually cultivated in soils characterised by low soil organic carbon (SOC) content and have high risks of soil erosion and degradation. Increasing SOC stocks in these cropping systems has the potential to contribute to climate change mitigation through SOC sequestration and to enhance soil quality. We conducted a meta-analysis and compared the SOC stock response ratio, the SOC stock rate of change, and the SOC sequestration rate in vineyards under different SOC sequestration (SCS) practices relative to conventional management. SCS practices included organic amendments (OA), biochar amendments (BC), returning pruning residues to the soil (PR), no-tillage (NT), cover cropping (CC), and several combinations of these practices. The average SOC sequestration rate of SCS management was 7.53 Mg CO2-eq. ha−1 yr−1 to a 30-cm soil depth. The highest SOC sequestration rate (11.06 Mg CO2-eq. ha−1 yr−1) was achieved under a combination of OA+NT and the lowest (2.82 Mg CO2-eq. ha−1 yr−1) was observed under PR treatments. Field experiments performed in particularly hot and dry bioclimatic zones were associated with lower SOC sequestration rates relative to those performed in more temperate areas. The high SOC sequestration rates obtained for many SCS practices, and the large land area dedicated to viticulture worldwide (7.45 Mha), imply that the adoption of SCS practices in vineyards can contribute to the global efforts to offset atmospheric greenhouse gas concentrations via SOC sequestration to mitigate climate change.

International audience; The aim of this study was to evaluate the capacity of BR55, an ultrasound contrast agent specifically targeting vascular endothelial growth factor receptor 2 (VEGFR2), to distinguish the specific anti-VEGFR2 therapy effect of sunitinib from other anti-angiogenic effects of a therapy (imatinib) that does not directly inhibit VEGFR2. Sunitinib, imatinib and placebo were administered daily for 11 d (264 h) to 45 BalbC mice bearing ectopic CT26 murine colorectal carcinomas. During the course of therapy, B-mode ultrasound, contrast-enhanced ultrasound and VEGFR2-targeted contrast-enhanced ultrasound were performed to assess tumor morphology, vascularization and VEGFR2 expression, respectively. The angiogenic effects on these three aspects were characterized using tumor volume, contrast-enhanced area and differential targeted enhancement. Necrosis, microvasculature and expression of VEGFR2 were also determined by histology and immunostaining. B-Mode imaging revealed that tumor growth was significantly decreased in sunitinib-treated mice at day 11 (p < 0.05), whereas imatinib did not affect growth. Functional evaluation revealed that the contrast-enhanced area decreased significantly (p < 0.02) and by similar amounts under both anti-angiogenic treatments by day 8 (192 h): −23% for imatinib and −21% for sunitinib. No significant decrease was observed in the placebo group. Targeted contrast-enhanced imaging revealed lower differential targeted enhancement, that is, lower levels of VEGFR2 expression, in sunitinib-treated mice relative to placebo-treated mice from 24 h (p < 0.05) and relative to both placebo- and imatinib-treated mice from 48 h (p < 0.05). Histologic assessment of tumors after the final imaging indicated that necrotic area was significantly higher for the sunitinib group (21%) than for the placebo (8%, p < 0.001) and imatinib (11%, p < 0.05) groups. VEGFR2-targeted ultrasound was able to sensitively differentiate the anti-VEGFR2 effect from the reduced area of tumor with functional flow produced by both anti-angiogenic agents. BR55 molecular imaging was, thus, able both to detect early therapeutic response to sunitinib in CT26 tumors as soon as 24 h after the beginning of the treatment and to provide early discrimination (48 h) between tumor response during anti-angiogenic therapy targeting VEGFR2 expression and response during anti-angiogenic therapy not directly acting on this receptor.

Pancreatic Ductal Adenocarcinoma (PDA) is one of the most lethal cancers with a 5-year survival rate of 8% and 2% for late stage diagnosis. This is due to the dense stroma of PDA at distal stages and increasing drug resistance. Cisplatin, a platinum-based chemotropic drug, is used to treat PDA by damaging DNA of cells through mutation of the Brca gene. In this study, Harmonic Motion Imaging (HMI) which is an all ultrasound based elastography imaging technique, is used to monitor the mechanical effect of Cisplatin on genetically-engineered transgenic mice. HMI generates oscillatory displacements at the focus and is used to measure the viscoelastic properties of wide range of tissues. The high power output of HMI makes it appropriate for characterizing PDA due to the highly stiff nature of PDA and the fact that it is deeply laid in the body.

Recently, we have shown that Harmonic Motion Imaging (HMI) can be used to differentiate relative stiffness and monitor HIFU ablations in small lumpectomy human breast specimens. The objective of this study is to apply HMI on post-surgical mastectomy breast specimen to mimic the in vivo environment and characterize tumor at different depth for better tumor localization and identification before and after HIFU treatment. 2D HMI displacement maps could be generated to map the induced peak-to-peak displacement covering the entire target area within two minutes indicating lower displacement in the tumor region and higher displacement in the peripheral tissue. A Student's paired t-test across all specimens showed a significant difference in the HMI displacement (P < 0.0001) between the tumor and the perilesional tissue. HMI was shown to successfully map the relative stiffness at variable depths in post-surgical human breast mastectomy specimens. This study therefore lays the foundation for upcoming clinical application of HMI on the breast for tumor detection and subsequent thermal ablative treatment.

Pancreatic ductal adenocarcinoma (PDA) has one of the lowest prognosis due to non-specific symptoms leading to late diagnosis. The tumor is characterized by an unusually dense stroma limiting chemotherapeutic perfusion. Harmonic Motion Imaging (HMI) assesses tissue mechanical properties by inducing localized oscillation resulting from a periodic acoustic radiation force. The amplitude of the displacement is inversely related to the underlying tissue stiffness. The objective of this study was to evaluate the utility of HMI in post-surgical human pancreatic cancer specimens to detect the tumor, and assess the tumor response to neoadjuvant treatments.

The successful clinical application of high intensity focused ultrasound (HIFU) ablation depends on reliable monitoring of the lesion formation. Harmonic motion imaging guided focused ultrasound (HMIgFUS) is an ultrasound-based elasticity imaging technique, which monitors HIFU ablation based on the stiffness change of the tissue instead of the echo intensity change in conventional B-mode monitoring, rendering it potentially more sensitive to lesion development. Our group has shown that predicting the lesion location based on the radiation force-excited region is feasible during HMIgFUS. In this study, the feasibility of a fast lesion mapping method is explored to directly monitor the lesion map during HIFU. The harmonic motion imaging (HMI) lesion map was generated by subtracting the reference HMI image from the present HMI peak-to-peak displacement map, as streamed on the computer display. The dimensions of the HMIgFUS lesions were compared against gross pathology. Excellent agreement was found between the lesion depth (r 2 = 0.81, slope = 0.90), width (r 2 = 0.85, slope = 1.12) and area (r 2 = 0.58, slope = 0.75). In vivo feasibility was assessed in a mouse with a pancreatic tumor. These findings demonstrate that HMIgFUS can successfully map thermal lesions and monitor lesion development in real time in vitro and in vivo. The HMIgFUS technique may therefore constitute a novel clinical tool for HIFU treatment monitoring.

Longitudinal imaging techniques are needed that can meaningfully probe the tumor microenvironment and its spatial heterogeneity. Contrast-enhanced ultrasound, shear wave elastography and quantitative ultrasound are ultrasound-based techniques that provide information on the vascular function and micro-/macroscopic tissue structure. Modifications of the tumor microenvironment induced by cytotoxic and anti-angiogenic molecules in ectopic murine Lewis lung carcinoma tumors were monitored. The most heterogenous structures were found in tumors treated with anti-angiogenic drug that simultaneously accumulated the highest levels of necrosis and fibrosis. The anti-angiogenic group presented the highest number of correlations between parameters related to vascular function and those related to the micro-/macrostructure of the tumor microenvironment. Results suggest how patterns of multiparametric ultrasound modifications can be related to provide a more insightful marker of changes occurring within tumors during therapy.

Recently, ultrasonic imaging of soft tissue mechanics has been increasingly studied to image otherwise undetectable pathologies. However, many underlying mechanisms of tissue stiffening remain unknown, requiring small animal studies and adapted elasticity mapping techniques. Harmonic motion imaging (HMI) assesses tissue viscoelasticity by inducing localized oscillation from a periodic acoustic radiation force. The objective of this study was to evaluate the feasibility of HMI for in vivo elasticity mapping of abdominal organs in small animals. Pathological cases, i.e. chronic pancreatitis and pancreatic cancer, were also studied in vivo to assess the capability of HMI for detection of the change in mechanical properties. A 4.5 MHz focused ultrasound transducer (FUS) generated an amplitude-modulated beam resulting in 50 Hz harmonic tissue oscillations at its focus. Axial tissue displacement was estimated using 1D-cross-correlation of RF signals acquired with a 7.8 MHz diagnostic transducer confocally aligned with the FUS. In vitro results in canine liver and kidney showed the correlation between HMI displacement and Young's moduli measured by rheometry compression testing. HMI was capable of providing reproducible elasticity maps of the mouse abdominal region in vivo allowing the identification of, from stiffest to softest, the murine kidney, pancreas, liver, and spleen. Finally, pancreata affected by pancreatitis and pancreatic cancer showed HMI displacements 1.7 and 2.2 times lower than in the control case, respectively, indicating higher stiffness. The HMI displacement amplitude was correlated with the extent of fibrosis as well as detecting the very onset of stiffening even before fibrosis could be detected on H&E. This work shows that HMI can produce reliable elasticity maps of mouse abdominal region in vivo, thus providing a potentially critical tool to assess pathologies affecting organ elasticity.

Harmonic Motion Imaging for Focused Ultrasound (HMIFU) is a technique that can perform and monitor high-intensity focused ultrasound (HIFU) ablation. An oscillatory motion is generated at the focus of a 93-element and 4.5 MHz center frequency HIFU transducer by applying a 25 Hz amplitude-modulated signal using a function generator. A 64-element and 2.5 MHz imaging transducer with 68kPa peak pressure is confocally placed at the center of the HIFU transducer to acquire the radio-frequency (RF) channel data. In this protocol, real-time monitoring of thermal ablation using HIFU with an acoustic power of 7 W on canine livers in vitro is described. HIFU treatment is applied on the tissue during 2 min and the ablated region is imaged in real-time using diverging or plane wave imaging up to 1,000 frames/second. The matrix of RF channel data is multiplied by a sparse matrix for image reconstruction. The reconstructed field of view is of 90° for diverging wave and 20 mm for plane wave imaging and the data are sampled at 80 MHz. The reconstruction is performed on a Graphical Processing Unit (GPU) in order to image in real-time at a 4.5 display frame rate. 1-D normalized cross-correlation of the reconstructed RF data is used to estimate axial displacements in the focal region. The magnitude of the peak-to-peak displacement at the focal depth decreases during the thermal ablation which denotes stiffening of the tissue due to the formation of a lesion. The displacement signal-to-noise ratio (SNRd) at the focal area for plane wave was 1.4 times higher than for diverging wave showing that plane wave imaging appears to produce better displacement maps quality for HMIFU than diverging wave imaging.

Harmonic Motion Imaging for Focused Ultrasound (HMIFU) is a technique that can perform and monitor high-intensity focused ultrasound (HIFU) ablation. An oscillatory motion is generated at the focus of a 93-element and 4.5 MHz center frequency HIFU transducer by applying a 25 Hz amplitude-modulated signal using a function generator. A 64-element and 2.5 MHz imaging transducer with 68kPa peak pressure is confocally placed at the center of the HIFU transducer to acquire the radio-frequency (RF) channel data. In this protocol, real-time monitoring of thermal ablation using HIFU with an acoustic power of 7 W on canine livers in vitro is described. HIFU treatment is applied on the tissue during 2 min and the ablated region is imaged in real-time using diverging or plane wave imaging up to 1,000 frames/second. The matrix of RF channel data is multiplied by a sparse matrix for image reconstruction. The reconstructed field of view is of 90° for diverging wave and 20 mm for plane wave imaging and the data are sampled at 80 MHz. The reconstruction is performed on a Graphical Processing Unit (GPU) in order to image in real-time at a 4.5 display frame rate. 1-D normalized cross-correlation of the reconstructed RF data is used to estimate axial displacements in the focal region. The magnitude of the peak-to-peak displacement at the focal depth decreases during the thermal ablation which denotes stiffening of the tissue due to the formation of a lesion. The displacement signal-to-noise ratio (SNRd) at the focal area for plane wave was 1.4 times higher than for diverging wave showing that plane wave imaging appears to produce better displacement maps quality for HMIFU than diverging wave imaging.

Harmonic motion imaging (HMI) is a radiationforce- based elasticity imaging technique that tracks oscillatory tissue displacements induced by sinusoidal ultrasonic radiation force to assess the resulting oscillatory displacement denoting the underlying tissue stiffness. The objective of this study was to evaluate the feasibility of HMI in pancreatic tumor detection and high-intensity focused ultrasound (HIFU) treatment monitoring. The HMI system consisted of a focused ultrasound transducer, which generated sinusoidal radiation force to induce oscillatory tissue motion at 50 Hz, and a diagnostic ultrasound transducer, which detected the axial tissue displacements based on acquired radio-frequency signals using a 1-D cross-correlation algorithm. For pancreatic tumor detection, HMI images were generated for pancreatic tumors in transgenic mice and normal pancreases in wild-type mice. The obtained HMI images showed a high contrast between normal and malignant pancreases with an average peak-to-peak HMI displacement ratio of 3.2. Histological analysis showed that no tissue damage was associated with HMI when it was used for the sole purpose of elasticity imaging. For pancreatic tumor ablation monitoring, the focused ultrasound transducer was operated at a higher acoustic power and longer pulse length than that used in tumor detection to simultaneously induce HIFU thermal ablation and oscillatory tissue displacements, allowing HMI monitoring without interrupting tumor ablation. HMI monitoring of HIFU ablation found significant decreases in the peak-to-peak HMI displacements before and after HIFU ablation with a reduction rate ranging from 15.8% to 57.0%. The formation of thermal lesions after HIFU exposure was confirmed by histological analysis. This study demonstrated the feasibility of HMI in abdominal tumor detection and HIFU ablation monitoring.

Harmonic motion imaging (HMI) is a radiation force-based elasticity imaging technique that tracks tissue harmonic displacements induced by periodic ultrasonic radiation force to assess tissue stiffness. The objective of this study was to evaluate the feasibility of HMI in pancreatic tumor detection and high-intensity focused ultrasound (HIFU) treatment monitoring. The HMI system consisted of a focused ultrasound transducer (FUS), which generated periodic radiation force to induce harmonic tissue motion at 50 Hz, and a diagnostic ultrasound transducer, which detected the axial tissue displacement within the targeted region using 1D cross-correlation of acquired radiofrequency signals of ultrasound echoes. For pancreatic tumor detection, HMI displacement images were generated for pancreatic tumors in transgenic mice and healthy pancreases of wild-type mice. For pancreatic tumor ablation monitoring, FUS was used to induce HIFU thermal ablation and tissue motion at the same time, allowing HMI monitoring without interrupting tumor ablation. All pancreases were excised immediately after sonication for histological evaluation. The obtained HMI displacement images showed a high contrast between normal and malignant tissue. The HMI monitoring of HIFU ablation displayed consistent pancreatic stiffening after 2 min ablation, and the formation of thermal lesions was confirmed by the histological analysis. This study demonstrated for the first time the feasibility of HMI in pancreatic tumor detection and HIFU ablation monitoring. It was also the first application of a radiation-force based imaging technique in abdominal tumor HIFU treatment monitoring.

Assessing structural organization and dynamics of tumor blood supply is of highest interest since numerous cancer treatments rely on antiangiogenic effects. Dynamic Contrast Enhanced Ultrasound (DCE-US) is widely used in cancer research to extract quantitative parameters reflecting tumor perfusion. However, the underlying vascular structure is not revealed and local spatial fluctuations in DCE parametric maps remain unexplained. The recently introduced Ultrafast Doppler Tomography (UFD-T) has been shown able to reconstruct the 3D vascular network of a rat brain with a 100µm resolution. It is applied here to tumor vasculature imaging and quantitative hemodynamics assessment along with DCE-US to show the complementarity between the techniques.

Robust, novel information on tumor microenvironment would improve therapeutic follow-up in oncology. Shear wave elastography (SWE) evaluates Young's modulus (stiffness) based on shear wave velocity. Dynamic contrast-enhanced ultrasound (DCE-US) traces microvascular flow with intravascular microbubbles. In this study, an ectopic tumor model was modified using two different types of drugs a cytotoxic (cyclophosphamide) and an anti-angiogenic (sunitinib). Both DCE-US and SWE imaging were sensitive to functional, physiological and mechanical modifications of the tumor and provided complementarity information to describe these modifications.

International audience; Ultrasound (US) scanners typically apply lossy, non-linear modifications to the US data for visualization purposes. The resulting images are then stored as compressed video data. Some system manufacturers provide dedicated software for quantification purposes to eliminate such processing distortions, at least partially. This is currently the recommended approach for quantitatively assessing changes in contrast-agent concentration from clinical data. However, the machine-specific access to US data and the limited set of analysis functionalities offered by each dedicated-software package make it difficult to perform comparable analyses with different US systems. The objective of this work was to establish if linearization of compressed video images obtained with an arbitrary US system can provide an alternative to dedicated-software analysis of machine-specific files for the estimation of echo-power. For this purpose, an Aplio 50 system (Toshiba Medical Systems, Tochigi, Japan), coupled with dedicated CHI-Q (Contrast Harmonic Imaging Quantification) software by Toshiba Medical Systems, was used. Results were compared with two approaches that apply algorithms to estimate relative echo-power from compressed video images: commercially available VueBox software by Bracco Suisse SA (Geneva, Switzerland) and in-laboratory software called PixPower. The echo-power estimated by CHI-Q analysis indicated a strong linear relationship versus agent concentration in vitro (R(2) ≥ 0.9996) for dynamic range (DR) settings of DR60 and DR80, with slopes between 9.22 and 9.57 dB/decade (p = 0.05). These values approach the theoretically predicted dependence of 10.0 dB/decade (equivalent to 3 dB for each concentration doubling). Echo-power estimations obtained from compressed video images with VueBox and PixPower also exhibited strong linear proportionality with concentration (R(2) ≥ 0.9996), with slopes between 9.30 and 9.68 dB/decade (p = 0.05). On an independent in vivo data set (N = 24), the difference in echo-power estimation between CHI-Q and each of the other two approaches was calculated after excluding regions that contain pixels affected by saturated or thresholded pixel values. The mean difference in estimates (expressed in decibels) was -0.25 dB between VueBox and CHI-Q (95% confidence interval: -0.75 to 0.26 dB) and -0.17 dB between PixPower and CHI-Q (95% confidence interval: -0.67 to 0.13 dB). To achieve linearization of data, one of the approaches (VueBox) requires calibration files provided by the software manufacturer for each machine type and setting. The other (PixPower) requires empirical correction of the imaging dynamic range based on ground truth data. These requirements could potentially be removed if US system manufacturers were willing to make relevant information on the applied processing publically available. Reliable echo-power estimation from linearized data would facilitate inclusion of different US systems in multicentric studies and more widespread implementation of emerging techniques for quantitative analysis of contrast ultrasound.

DCE-US, which allows follow up of tumor vasculature during therapy, is potentially a very useful tool to predict early response to therapy. In this study we act on the vascularization of two different tumor models, colorectal (CT26) and Lewis lung carcinomas (3LL), using sunitinib, an anti-angiogenic drug. Time-dependent perfusion parameters are explored to characterize tumor perfusion. Significant differences are highlighted in term of flow pattern for treated versus control mice in one tumor model. Differences are also identified for tumor filling between the two tumor models. For 3LL mice under therapy, modifications of functional vascularization begin at day 7 as characterized by slower filling compared to 3LL control mice. Results also exhibit slower filling patterns for CT26 relative to 3LL tumor controls. Histology results reveal smaller vessels (diameter

Dynamic contrast-enhanced ultrasonography (DCEUS) can detect perfusion modifications during anti-angiogenic therapy preceding tumor size regression. In particular, perfusion parameters can be useful to predict the early response of a patient to a therapy. However, assessment is sensitive to injection variability. In this work, we propose a method for controlled injection of ultrasound contrast agent and demonstrate the improved evaluation of contrast enhancement obtained relative to manual injections for both in vitro and in vivo experiments. Perfusion parameters of the area under the curve for the wash-in and rise time gained the most in terms of reproducibility, improvement of CV values were from 24% to 5% and 25% to 7%, respectively. This study, demonstrates the benefit of using a controlled injection system to improve reproducibility of UCA injection in pre-clinical models.

To estimate perfusion using Dynamic Contrast-Enhanced Ultrasound (DCE-US), some manufacturers provide software that analyses DICOM Raw Data formatted files. In those files, most of the information cannot be correctly interpreted except by the manufacturer software. This approach when available is often recommended over use of more accessible DICOM JPEG files which encapsulate ultrasound system images as compressed JPEG images. However, the manufacturers' dedicated software is machine specific with limited functionalities which hinders multicentric studies using different ultrasound machines. The objective of this work is to develop techniques and criteria defining conditions under which compressed video images obtained with the Aplio 50 from Toshiba can be used to accurately estimate contrast concentration. Dedicated CHIQ software from Toshiba was compared to two approaches based on compressed images: commercially available VueBox™ software from Bracco and in-laboratory software PixPower. Proportionality between echo-power estimates and dose of contrast agents in suspension was validated in vitro for the three approaches. VueBox™ and PixPower echo-power estimations from DCE-US sequences acquired in an in vivo tumor model were in agreement with those obtained with the CHIQ reference. This work shows that, for the dynamic ranges considered in this study, DICOM JPEG files can be used to accurately estimate the echo power from video compressed images using linearizing equations based on rigorous calibration steps.

Robust, novel image-based information on the tumor microenvironment would improve therapeutic follow-up in oncology. SWE evaluates tissue stiffness, CEUS assesses microvascular flow using intravascular microbubbles and QUS measures tissue microstructure. Our goal was to evaluate sensitivity and complementarity of these techniques during cytotoxic and antiangiogenic therapies in ectopic, murine tumors. A multi-targeted receptor tyrosine kinase antiangiogenic drugs inhibiting vascularization (sunitinib, N = 24, 40mg/kg/day), an alkylating cytotoxic agent that adds an alkyl group to DNA to interfere with DNA replication (cyclophosphamide, N = 26, 150mg/kg for 3 days) and placebo (N = 26) were administered in murine Lewis Lung Carcinomas bearing C57BL6 mice. Six days after implantation, tumors were imaged (SWE, CEUS and QUS) prior to therapy (Day 0) and on Days 1, 3, 7, 9 and 13. Tumors were harvested at Day 9 (N = 2 per group) and Day 13 (N = 10 per group) and stained to evaluate% necrosis and fibrosis. Table. 1 summarizes the% variation of each measured parameter from the start to the end of therapy. Imaging parameters for microvascular function and stiffness were most strongly modified for tumors treated with sunitinib compared to cytotoxic and placebo therapies. At Day 13, tumors treated with sunitinib presented a heterogeneous structure with more fibrosis and necrosis than for other treatment groups. This heterogeneous structure may explain why the standard deviation of stiffness was highest for the antiangiogenic group. The sensitivity of CEUS to microvascular modifications and SWE to biomarkers such as necrosis and fibrosis, in this model, underline potential for monitoring complementarity aspects of the tumor microenvironment during therapy. CytotoxicAnti-angiogenicPlaceboVariation(%)P-valueVariation(%)P-valueVariation(%)P-valueSWEStiffness20*0.03680* Table 1: Variation between Day 0 and Day 13 of mean values of each parameter for each group. Citation Format: Alexandre Dizeux, Thomas Payen, Guillaume Barrois, Michele Lamuraglia, Capucine Baldini, Delphine Le Guillou, Eva Comperat, Jean-Luc Gennisson, Mickael Tanter, Michael Oelze, S. Lori Bridal. In vivo discrimination of tumor modifications during antiangiogenic and cytotoxic therapy using ultrasonography modalities: Shear Wave Elastography (SWE), Contrast Enhanced Ultrasound (CEUS) and Quantitative Ultrasound (QUS). [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1497. doi:10.1158/1538-7445.AM2015-1497

Background: Anti-angiogenic drugs are a pillar of the therapeutic approach against cancer. Contrast ultrasound (CUS) can detect perfusion modifications during anti-angiogenic therapy preceding tumor size regression. However, contrast imaging would benefit from better standardization of the technique. This work compares discrimination of tumors in Sorafenib-treated and placebo mice using CUS evaluation of relative blood volume (rBV), mean transit time (mTT) and wash in rate (WiR). Methods: Ewing's sarcoma tumors were grafted orthotopically by sub-capsular injection of 100 µL of SK-NEP-1 cells (5 x 106cells/mL) in the left kidney of nude fifteen mice (Elevage Janvier, Le Genest-St-Isle, France). Contrast enhanced ultrasonography was performed using a clinical ultrasound system Acuson Sequoia 512 with a 14 MHz probe (Siemens Medical Solutions, Mountain View). The tumor area (A) was calculated according to A = (≥ x L x w)/2. CUS (Sonovue, Bracco SA, Milan, Italy) with destruction-reperfusion was performed using cadence pulse-sequence (CPS). Data were acquired from each mouse at 14 days after tumor-cell injection. The treatment was started at day 14 just after the CUS baseline. At day 14 (day of start therapy) the 12 mice were randomized into two groups. One group received (6 mice) the anti-angiogenic treatment Sorafenib daily (10mg/kg/jr) per os and the other group (6 mice) received corresponding doses of placebo by the same route for 8 days. Image sequences were recorded in DICOM format and analyzed with prototype software from Bracco Suisse SA (Geneva, Switzerland) allowing selection of regions of interest, approximate linearization of the data and fitting of flow models to time-intensity data to estimate functional parameters within the tumor. All parameters were normalized with respect to normal cortex perfusion. Statistical analysis was performed using Wilcoxon Signed-Rank tests. Results: Ultrasound estimates of the tumor size increased with time for both groups, but tumor size was never significantly different between the two groups. Before the normalization with the adjacent renal cortex perfusion, the functional parameters did not present a significant difference between the two groups. Therefore after normalization: the rBV shows a significant difference Sorafenib vs placebo (p=0.008) at 8 days, the mTT also becomes significant after 8 days of treatment (p=0.02). The WiR shows a significant difference between groups at 8 days (p=0.005), but also at 6 days (p=0.04). Conclusions: functional CUS was an effective technique for monitoring the early response of anti-angiogenic treatment. The parameters obtained by fitting flow models of destruction-reperfusion demonstrated the capacity to standardize this angiogenesis analysis. Also the normalization with respect to adjacent normal tissue appears to reduce estimate variability and improve functional contrast assessment. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2428. doi:1538-7445.AM2012-2428

International audience; Robust, novel information on tumor microenvironment would improve therapeutic follow-up in oncology. Shear wave elastography (SWE) evaluates Young's modulus (stiffness) based on shear wave velocity. Dynamic contrast-enhanced ultrasound (CEUS) traces microvascular flow with intravascular microbubbles. In this study, an ectopic tumor model was modified using two different types of drugs a cytotoxic (cyclophosphamide) and an anti-angiogenic (sunitinib). Both CEUS and SWE imaging were sensitive to functional, physiological and mechanical modifications of the tumor and provided complementarity information to describe these modifications.

Harmonic motion imaging (HMI) is a radiation force-based elasticity imaging technique that estimates tissue harmonic displacements induced by an oscillatory ultrasonic radiation force to assess tissue stiffness. The objective of this study was to evaluate the feasibility of applying HMI on pancreatic tumor detection and high-intensity focused ultrasound (HIFU) treatment monitoring.