Contraction assessment of abdominal muscles using automated segmentation designed for wearable ultrasound applications.

PURPOSE: Wearable ultrasound devices can be used to continuously monitor muscle activity. One possible application is to provide real-time feedback during physiotherapy, to show a patient whether an exercise is performed correctly. Algorithms which automatically analyze the data can be of importance to overcome the need for manual assessment and annotations and speed up evaluations especially when considering real-time video sequences. They even could be used to present feedback in an understandable manner to patients in a home-use scenario. The following work investigates three deep learning based segmentation approaches for abdominal muscles in ultrasound videos during a segmental stabilizing exercise. The segmentations are used to automatically classify the contraction state of the muscles. METHODS: The first approach employs a simple 2D network, while the remaining two integrate the time information from the videos either via additional tracking or directly into the network architecture. The contraction state is determined by comparing measures such as muscle thickness and center of mass between rest and exercise. A retrospective analysis is conducted but also a real-time scenario is simulated, where classification is performed during exercise. RESULTS: Using the proposed segmentation algorithms, 71% of the muscle states are classified correctly in the retrospective analysis in comparison to 90% accuracy with manual reference segmentation. For the real-time approach the majority of given feedback during exercise is correct when the retrospective analysis had come to the correct result, too. CONCLUSION: Both retrospective and real-time analysis prove to be feasible. While no substantial differences between the algorithms were observed regarding classification, the networks incorporating the time information showed temporally more consistent segmentations. Limitations of the approaches as well as reasons for failing cases in segmentation, classification and real-time assessment are discussed and requirements regarding image quality and hardware design are derived.

A Novel Concept of Transperineal Focused Ultrasound Transducer for Prostate Cancer Local Deep Hyperthermia Treatments.

Design, embodiment, and experimental study of a novel concept of extracorporeal phased array ultrasound transducer for prostate cancer regional deep hyperthermia treatments using a transperineal acoustic window is presented. An optimized design of hyperthermia applicator was derived from a modelling software where acoustic and thermal fields were computed based on anatomical data. Performance tests have been experimentally conducted on gel phantoms and tissues, under 3T MRI guidance using PRFS thermometry. Feedback controlled hyperthermia (ΔT = 5 °C during 20min) was performed on two ex vivo lamb carcasses with prostate mimicking pelvic tissue, to demonstrate capability of spatio-temporal temperature control and to assess potential risks and side effects. Our optimization approach yielded a therapeutic ultrasound transducer consisting of 192 elements of variable shape and surface, pseudo randomly distributed on 6 columns, using a frequency of 700 kHz. Radius of curvature was 140 mm and active water circulation was included for cooling. The measured focusing capabilities covered a volume of 24 × 50 × 60 mm3. Acoustic coupling of excellent quality was achieved. No interference was detected between sonication and MR acquisitions. On ex vivo experiments the target temperature elevation of 5 °C was reached after 5 min and maintained during another 15 min with the predictive temperature controller showing 0.2 °C accuracy. No significant temperature rise was observed on skin and bonny structures. Reported results represent a promising step toward the implementation of transperineal ultrasound hyperthermia in a pilot study of reirradiation in prostate cancer patients.

Focused ultrasound robotic system for very small bore magnetic resonance imaging.

BACKGROUND: A magnetic resonance imaging (MRI) compatible robotic system for focused ultrasound was developed for small animal like mice or rats that fits into a 9.4 T MRI scanner (Bruker Biospec 9420, Bruker Biospin, Ettlingen, Germany). The robotic system includes two computer-controlled linear stages. MATERIALS AND METHODS: The robotic system was evaluated in a mouse-shaped, real-size agar-based mimicking material, which has similar acoustical properties as soft tissues. The agar content was 6% weight per volume (w/v), 4% w/v silica while the rest was degassed water. The transducer used has a diameter of 4 cm, operates with 2.6 MHz and focuses energy at 5 cm. RESULTS: The MRI compatibility of the robotic system was evaluated in a 9.4 T small animal scanner. The efficacy of the ultrasonic transducer was evaluated in the mimicking material using temperature measurements. CONCLUSIONS: The proposed robotic system can be utilized in a 9.4 T small animal MRI scanner. The proposed system is functional, compact and simple thus providing a useful tool for preclinical research in mice and rats.

An integrated model-based software for FUS in moving abdominal organs.

Focused ultrasound surgery (FUS) is a non-invasive method for tissue ablation that has the potential for complete and controlled local tumour destruction with minimal side effects. The treatment of abdominal organs such as the liver, however, requires particular technological support in order to enable a safe, efficient and effective treatment. As FUS is applied from outside the patient's body, suitable imaging methods, such as magnetic resonance imaging or diagnostic ultrasound, are needed to guide and track the procedure. To facilitate an efficient FUS procedure in the liver, the organ motion during breathing and the partial occlusion by the rib cage need to be taken into account in real time, demanding a continuous patient-specific adaptation of the treatment configuration. Modelling the patient's respiratory motion and combining this with tracking data improves the accuracy of motion predictions. Modelling and simulation of the FUS effects within the body allows the use of treatment planning and has the potential to be used within therapy to increase knowledge about the patient status. This article describes integrated model-based software for patient-specific modelling and prediction for FUS treatments of moving abdominal organs.

Pharmacodynamics of streptavidin-coated cyanoacrylate microbubbles designed for molecular ultrasound imaging.

OBJECTIVES: To assess the pharmacodynamic behavior of cyanoacrylate, streptavidin-coated microbubbles (MBs) and to investigate their suitability for molecular ultrasound imaging. MATERIALS AND METHODS: Biodistribution of MBs was analyzed in tumor-bearing mice using gamma-counting, immunohistochemistry, flow cytometry, and ultrasound. Further, vascular endothelial growth factor receptor 2-antibody coupled MBs were used to image tumor neovasculature. RESULTS: After 1 minute >90% of MBs were cleared from the blood and pooled in the lungs, liver, and spleen. Subsequently, within 1 hour a decent reincrease of MB-concentration was observed in the blood. The remaining MBs were removed by liver and spleen macrophages. About 30% of the phagocytosed MBs were intact after 48 hours. Shell fragments were found in the kidneys only. No relevant MB-accumulation was observed in tumors. In contrast, vascular endothelial growth factor receptor 2-specific MBs accumulated significantly within the tumor vasculature (P < 0.05). CONCLUSIONS: The pharmacokinetic behavior of streptavidin-coated cyanoacrylate MBs has been studied. In this context, the low amount of MBs in tumors after >5 minutes is beneficial for specific targeting of angiogenesis.

Molecular profiling of angiogenesis with targeted ultrasound imaging: early assessment of antiangiogenic therapy effects.

Molecular ultrasound is capable of elucidating the expression of angiogenic markers in vivo. However, the capability of the method for volumetric "multitarget quantification" and for the assessment of antiangiogenic therapy response has rather been investigated. Therefore, we generated cyanoacrylate microbubbles linked to vascular endothelial growth factor receptor 2 (VEGFR2) and alphavbeta3 integrin binding ligands and quantified their accumulation in squamous cell carcinoma xenografts (HaCaT-ras-A-5RT3) in mice with the quantitative volumetric ultrasound scanning technique, sensitive particle acoustic quantification. Specificity of VEGFR2 and alphavbeta3 integrin binding microbubbles was shown, and changes in marker expression during matrix metalloproteinase inhibitor treatment were investigated. In tumors, accumulation of targeted microbubbles was significantly higher compared with nonspecific ones and could be inhibited competitively by addition of the free ligand in excess. Also, multimarker imaging could successfully be done during the same imaging session. Molecular ultrasound further indicated a significant increase of VEGFR2 and alphavbeta3 integrin expression during tumor growth and a considerable decrease in both marker densities after matrix metalloproteinase inhibitor treatment. Histologic data suggested that the increasing VEGFR2 and alphavbeta3 integrin concentrations in tumors during growth are related to an up-regulation of its expression by the endothelial cells, whereas its decrease under therapy is more related to the decreasing relative vessel density. In conclusion, targeted ultrasound appears feasible for the longitudinal molecular profiling of tumor angiogenesis and for the sensitive assessment of therapy effects in vivo.

BioShuttle-mediated plasmid transfer.

An efficient gene transfer into target tissues and cells is needed for safe and effective treatment of genetic diseases like cancer. In this paper, we describe the development of a transport system and show its ability for transporting plasmids. This non-viral peptide-based BioShuttle-mediated transfer system consists of a nuclear localization address sequence realizing the delivery of the plasmid phNIS-IRES-EGFP coding for two independent reporter genes into nuclei of HeLa cells. The quantification of the transfer efficiency was achieved by measurements of the sodium iodide symporter activity. EGFP gene expression was measured with Confocal Laser Scanning Microscopy and quantified with biostatistical methods by analysis of the frequency of the amplitude distribution in the CLSM images. The results demonstrate that the "BioShuttle"-Technology is an appropriate tool for an effective transfer of genetic material carried by a plasmid.

Ultrasound exposure can increase the membrane permeability of human neutrophil granulocytes containing microbubbles without causing complete cell destruction.

Activated polymorphonuclear neutrophil (PMN) granulocytes can bind and subsequently phagocytose microbubbles used as ultrasound (US) contrast agents. The purpose of the present study was to assess insonation effects on cell membrane integrity and metabolic activity of activated PMN. Furthermore, we investigated whether or not there is an acoustic threshold at which insonation of PMN results in increase of membrane permeability without causing complete cell destruction. PMN isolated from healthy volunteers were activated with phorbol myristate acetate (PMA) for 15 min to allow phagocytosis of albumin and lipid microbubbles and were subsequently exposed to US with a mechanical index between 0.15 and 1.8. Apoptosis, loss of membrane integrity and formation of cell fragments were evaluated by measurement of lactate dehydrogenase leakage and by double staining with annexin V and propidium iodide, using flow cytometry. Neutrophil superoxide anion generation was measured photometrically. Insonation of activated PMN in the presence of microbubbles amplified apoptosis and lactate dehydrogenase leakage and induced loss of membrane integrity and complete cell destruction with increasing acoustic pressures. The bioeffects observed by insonation with high mechanical indices (1.0 to 1.8), and particularly the formation of cell fragments, were significantly more pronounced in the presence of albumin microbubbles. Insonation in the presence of lipid microbubbles increased cell membrane permeability, but caused significantly less cell destruction and left the metabolic activity of activated PMN uninfluenced. Thus, both albumin and lipid microbubbles induce apoptosis and membrane injury during insonation of activated PMN. However, insonation in the presence of lipid microbubbles seems to influence cell viability to a smaller extent. This could be of advantage in the setting of US-guided local drug delivery. In this setting, increase of membrane permeability may allow bioactive substances to enter into cells, which survive the US treatment, and specifically modify their function.

Focal gene induction in the liver of rats by a heat-inducible promoter using focused ultrasound hyperthermia: preliminary results.

OBJECTIVE: We sought to examine high-intensity focused ultrasound (HIFU)-induced hyperthermia in the liver of a rat model to focally induce green-fluorescent protein (GFP). MATERIALS AND METHODS: A total of 25 Copenhagen rats were included in this study. Rats were divided into groups treated with an adenovirus coding for green fluorescent protein (GFP) under the control of a hsp70B promoter and a CMV promoter. Ad-CMV-GFP-treated rats served as positive control. Untreated controls only subjected to MRI +/- HIFU-treatment served to find out optimal power of HIFU in the target area of the liver. Temperature was noninvasively monitored by temperature sensitive magnetic resonance imaging (MRI). RESULTS: Rats treated with Ad-hsp70B-GFP demonstrated localized gene induction within the liver parenchyma, in good correlation with MRI and histology. Applying an acoustic power of 1.92 W a relatively uniform focal temperature up to 42 +/- 5 degrees C within the liver parenchyma could be documented. 3 x 10(9) plaque-forming units proved to account for a very homogeneous liver infection. Number of fluorescent cells in the region of hyperthermia was similar to the control group treated with Ad-CMV-GFP. CONCLUSION: Using the introduced parameters spatially controlled gene induction within a parenchymal organ such as the liver in rats using HIFU under control of MRI is feasible.

Apoptosis signals in lymphoblasts induced by focused ultrasound.

We investigated the effects of focused ultrasound (FUS) on specific molecular signaling and cellular response in three closely related human Tk6 lymphoblast cell lines that differed only in their p53 status. The applied ultrasound parameters fell between the physical dose range, which is safely used in medical diagnostics (peak pressure<0.1 MPa) and that used for high-energy FUS thermal ablation therapy (peak pressure>10 MPa). Based on cDNA microarrays and protein analysis, we found that FUS at the intermediate peak pressure of 1.5 MPa induced a complex signaling cascade with upregulation of proapoptotic genes [e.g., p53, p21, Thy1 (CD 90)]. Simultaneously, FUS downregulated cellular survival components (e.g., bcl-2, SOD). The p53 status was important for the reaction of the cells to ultrasound. Apoptosis and G1 arrest were induced primarily in p53+ cells, while p53- cells showed less apoptosis but exhibited G2 arrest. Likewise, the proliferation of lymphoblasts was much more strongly inhibited in p53+ than in p53- cells. Microarray analysis further demonstrated an upregulation of genes involved in oxidative stress (e.g., ferritin), suggesting that indirect sonochemical effects via reactive oxygen species play a causative role in the interaction of ultrasound with lymphoblasts. An important characteristic of FUS in therapeutic ultrasound applications is its ability to be administered to the human body in a targeted manner while sparing intermediate tissues. Therefore, our data indicate that this noninvasive, mechanical wave transmission, which is free of ionizing radiation, has the potential to specifically induce localized cell signals and apoptosis.