Imaging Sensors and Applications

In past decades, various sensor technologies have been used in all areas of our lives, thus improving our quality of life. In particular, imaging sensors have been widely applied in the development of various imaging approaches such as optical imaging, ultrasound imaging, X-ray imaging, and nuclear imaging, and contributed to achieve high sensitivity, miniaturization, and real-time imaging. These advanced image sensing technologies play an important role not only in the medical field but also in the industrial field. This Special Issue covers broad topics on imaging sensors and applications. The scope range of imaging sensors can be extended to novel imaging sensors and diverse imaging systems, including hardware and software advancements. Additionally, biomedical and nondestructive sensing applications are welcome

Noninvasive Thrombolysis using Microtripsy

Thrombosis refers to blood clot formation and when pathological, is the cause of many vascular diseases. For example, deep vein thrombosis (DVT), which affects three million Americans per year, is the formation of clots in the deep veins of the legs. Current clinical treatments include thrombolytic drugs and catheter-based surgical procedures. Both methods have significant drawbacks, such as excessive bleeding, invasiveness, and long treatment time. Ultrasound has been combined with thrombolytic drugs and/or microbubbles to enhance drug delivery. However, these methods are still quite slow and share the drawbacks of thrombolytic drugs. Histotripsy is a tissue ablation method that mechanically fractionates soft tissue via well-controlled acoustic cavitation generated by microsecond-long, high-pressure ultrasound pulses. The initial feasibility and safety of using histotripsy as a noninvasive, drug-free, and image-guided thrombolysis technique has been demonstrated both in vitro and in vivo. The overriding goal of this dissertation is clinical translation of histotripsy thrombolysis. First, an integrated image-guided histotripsy thrombolysis system suitable for clinical DVT treatment are designed and constructed. Second, the recently discovered technical innovations, microtripsy and bubble-induced color Doppler (BCD), are investigated for histotripsy thrombolysis application to further improve treatment efficacy. Microtripsy is a new histotripsy approach and uses an intrinsic threshold mechanism to generate more reproducible and predictable cavitation via a single ultrasound pulse, which can minimize vessel damage by confining cavitation within vessel lumen and eliminate cavitation on vessel wall. BCD is developed to monitor tissue motion induced by histotripsy pulses and investigated as a real-time quantitative feedback for histotripsy thrombolysis. Finally, a comprehensive pre-clinical study in a large animal DVT model is conducted to validate the safety and efficacy of this clinically designed system incorporating these technical innovations. It is our hope that this dissertation work will establish a foundation for the translation of this noninvasive thrombolysis technology into relevant clinical applications.PHDBiomedical EngineeringUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/135813/1/xizh_1.pd

Histotripsy for Pediatric Cardiac Applications.

Medicine continues to move towards less invasive techniques for many cardiac conditions, especially for high-risk patients that may not tolerate the alternative, more invasive approach. For instance, patients born with the congenital heart defect hypoplastic left heart syndrome often require emergent creation of a perforation through the atrial septum for survival prior to palliative surgery. However, most approaches are catheter based, still invasive, and continue to have significant challenges, limitations, and complications. A completely non-invasive technique such as histotripsy may provide the same result in a faster, safer, and more efficient manner. Using high-pressure ultrasound pulses applied outside the body and focused to the targeted tissue, histotripsy generates a cluster of cavitating micro-bubbles that fractionate the target tissue. The goal of this work is to investigate the safety and efficacy of histotripsy for neonatal cardiac applications. To aid in this goal, therapy guidance and monitoring techniques are developed, and an integrated histotripsy therapy system, optimized for the human neonate with congenital heart disease, was designed and constructed. In this dissertation, histotripsy is first demonstrated to be capable of generating targeted intra-cardiac communications when positioned outside the body in an intact neonatal animal model with minimal collateral damage or systemic side-effects. Second, to mitigate the possibility of unintended injury due to heart motion, real-time motion correction using ultrasound imaging is developed and integrated into a histotripsy therapy system. The performance of the motion correction is quantified in vitro and a validated in a single in vivo experiment. Third, to maximize therapy efficacy, novel bubble-induced color Doppler feedback to monitor the degree of tissue damage during histotripsy treatment is developed and validated in vitro. Finally, a histotripsy therapy transducer with appropriate physical dimensions and acoustic parameters to precisely ablate cardiac tissue non-invasively in a human neonate is developed and integrated into an ultrasound guided histotripsy therapy system. The data and the integrated system accomplished from this dissertation form the essential foundation to a pioneering clinical trial for histotripsy cardiac therapy in infants, which will position histotripsy for application on a broad range of cardiac disorders in patients of all ages.PHDBiomedical EngineeringUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/108732/1/millerrm_1.pd

How sonoporation disrupts cellular structural integrity: morphological and cytoskeletal observations

Posters: no. 1Control ID: 1672429OBJECTIVES: In considering sonoporation for drug delivery applications, it is essential to understand how living cells respond to this puncturing force. Here we seek to investigate the effects of sonoporation on cellular structural integrity. We hypothesize that the membrane morphology and cytoskeletal behavior of sonoporated cells under recovery would inherently differ from that of normal viable cells. METHODS: A customized and calibrated exposure platform was developed for this work, and the ZR-75-30 breast carcinoma cells were used as the cell model. The cells were exposed to either single or multiple pulses of 1 MHz ultrasound (pulse length: 30 or 100 cycles; PRF: 1kHz; duration: up to 60s) with 0.45 MPa spatial-averaged peak negative pressure and in the presence of lipid-shelled microbubbles. Confocal microscopy was used to examine insitu the structural integrity of sonoporated cells (identified as ones with exogenous fluorescent marker internalization). For investigations on membrane morphology, FM 4-64 was used as the membrane dye (red), and calcein was used as the sonoporation marker (green); for studies on cytoskeletal behavior, CellLight (green) and propidium iodide (red) were used to respectively label actin filaments and sonoporated cells. Observation started from before exposure to up to 2 h after exposure, and confocal images were acquired at real-time frame rates. Cellular structural features and their temporal kinetics were quantitatively analyzed to assess the consistency of trends amongst a group of cells. RESULTS: Sonoporated cells exhibited membrane shrinkage (decreased by 61% in a cell’s cross-sectional area) and intracellular lipid accumulation (381% increase compared to control) over a 2 h period. The morphological repression of sonoporated cells was also found to correspond with post-sonoporation cytoskeletal processes: actin depolymerization was observed as soon as pores were induced on the membrane. These results show that cellular structural integrity is indeed disrupted over the course of sonoporation. CONCLUSIONS: Our investigation shows that the biophysical impact of sonoporation is by no means limited to the induction of membrane pores: e.g. structural integrity is concomitantly affected in the process. This prompts the need for further fundamental studies to unravel the complex sequence of biological events involved in sonoporation.postprin

Developmental delays and subcellular stress as downstream effects of sonoporation

Posters: no. 2Control ID: 1672434OBJECTIVES: The biological impact of sonoporation has often been overlooked. Here we seek to obtain insight into the cytotoxic impact of sonoporation by gaining new perspectives on anti-proliferative characteristics that may emerge within sonoporated cells. We particularly focused on investigating the cell-cycle progression kinetics of sonoporated cells and identifying organelles that may be stressed in the recovery process. METHODS: In line with recommendations on exposure hardware design, an immersion-based ultrasound platform has been developed. It delivers 1 MHz ultrasound pulses (100 cycles; 1 kHz PRF; 60 s total duration) with 0.45 MPa peak negative pressure to a cell chamber that housed HL-60 leukemia cells and lipid-shelled microbubbles at a 10:1 cell-tobubble ratio (for 1e6/ml cell density). Calcein was used to facilitate tracking of sonoporated cells with enhanced uptake of exogenous molecules. The developmental trend of sonoporated cells was quantitatively analyzed using BrdU/DNA flow cytometry that monitors the cell population’s DNA synthesis kinetics. This allowed us to measure the temporal progression of DNA synthesis of sonoporated cells. To investigate whether sonoporation would upset subcellular homeostasis, post-exposure cell samples were also assayed for various proteins using Western blot analysis. Analysis focus was placed on the endoplasmic reticulum (ER): an important organelle with multi-faceted role in cellular functioning. The post-exposure observation time spanned between 0-24 h. RESULTS: Despite maintaining viability, sonoporated cells were found to exhibit delays in cell-cycle progression. Specifically, their DNA synthesis time was lengthened substantially (for HL-60 cells: 8.7 h for control vs 13.4 h for the sonoporated group). This indicates that sonoporated cells were under stress: a phenomenon that is supported by our Western blot assays showing upregulation of ER-resident enzymes (PDI, Ero1), ER stress sensors (PERK, IRE1), and ER-triggered pro-apoptotic signals (CHOP, JNK). CONCLUSIONS: Sonoporation, whilst being able to facilitate internalization of exogenous molecules, may inadvertently elicit a cellular stress response. These findings seem to echo recent calls for reconsideration of efficiency issues in sonoporation-mediated drug delivery. Further efforts would be necessary to improve the efficiency of sonoporation-based biomedical applications where cell death is not desirable.postprin

A study on the change in plasma membrane potential during sonoporation

Posters: no. 4Control ID: 1680329OBJECTIVES: There has been validated that the correlation of sonoporation with calcium transients is generated by ultrasound-mediated microbubbles activity. Besides calcium, other ionic flows are likely involved in sonoporation. Our hypothesis is the cell electrophysiological properties are related to the intracellular delivery by ultrasound and microbubbles. In this study, a real-time live cell imaging platform is used to determine whether plasma membrane potential change is related to the sonoporation process at the cellular level. METHODS: Hela cells were cultured in DMEM supplemented with 10% FBS in Opticell Chamber at 37 °C and 5% CO2, and reached 80% confluency before experiments. The Calcein Blue-AM, DiBAC4(3) loaded cells in the Opticell chamber filled with PI solution and Sonovue microbubbles were immerged in a water tank on a inverted fluorescence microscope. Pulsed ultrasound (1MHz freq., 20 cycles, 20Hz PRF, 0.2-0.5MPa PNP) was irradiated at the angle of 45° to the region of interest for 1s.The real-time fluorescence imaging for different probes was acquired by a cooled CCD camera every 20s for 10min. The time-lapse fluorescence images were quantitatively analyzed to evaluate the correlation of cell viability, intracellular delivery with plasma membrane potential change. RESULTS: Our preliminary data showed that the PI fluorescence, which indicated intracellular delivery, was immediately accumulated in cells adjacent to microbubbles after exposure, suggesting that their membranes were damaged by ultrasound-activated microbubbles. However, the fluorescence reached its highest level within 4 to 6 minutes and was unchanged thereafter, indicating the membrane was gradually repaired within this period. Furthermore, using DIBAC4(3), which detected the change in the cell membrane potential, we found that the loss of membrane potential might be associated with intracellular delivery, because the PI fluorescence accumulation was usually accompanied with the change in DIBAC4 (3) fluorescence. CONCLUSIONS: Our study suggests that there may be a linkage between the cell membrane potential change and intracellular delivery mediated by ultrasound and microbubbles. We also suggest that other ionic flows or ion channels may be involved in the cell membrane potential change in sonoporation. Further efforts to explore the cellular mechanism of this phenomenon will improve our understanding of sonoporation.postprin

Real-time imaging of cellular dynamics during low-intensity pulsed ultrasound exposure

Control ID: 1671584Oral Session 5 - Bioeffects of therapeutic ultrasoundOBJECTIVE: Although the therapeutic potential of low-intensity pulsed ultrasound is unquestionable, the wave-matter interactions involved in the process remain to be vaguely characterized. Here we seek to undertake a series of in-situ cellular imaging studies that aim to analyze the mechanical impact of low-intensity pulsed ultrasound on attached fibroblasts from three different aspects: membrane, cytoskeleton, and nucleus. METHODS: Our experimental platform comprised an in-house ultrasound exposure hardware that was coupled to a confocal microscopy system. The waveguided ultrasound beam was geometrically aligned to the microscope’s fieldof-view that corresponds to the center of a polystyrene dish containing fibroblasts. Short ultrasound pulses (5 cycles; 2 kHz PRF) with 0.8 MPa peak acoustic pressure (0.21 W/cm2 SPTA intensity) were delivered over a 10 min period. Live imaging was performed on both membrane (CellMask) and cytoskeleton (actin-GFP, tubulin-RFP) over the entire observation period (up to 30 min after end of exposure). Also, pre- and post-exposure fixed-cell imaging was conducted on the nucleus (Hoechst 33342) and two cytoskeleton components related to stress fibers: F-actin (phalloidin-FITC) and vincullin (Alexa Fluor 647 conjugated). To study whether mechanotransduction was responsible in mediating ultrasound-cell interactions, some experiments were conducted with the addition of gadolinium that blocks stretch-sensitive ion channels. RESULTS: Cell shrinkage was evident over the course of low-intensity pulsed ultrasound exposure. This was accompanied with contraction of actin and tubulin. Also, an increase in central stress fibers was observed at the end of exposure, while the nucleus was found to have decreased in size. Interestingly, after the exposure, a significant rebound in cell volume was observed over a 30 min. period. These effects were not observed in cases with gadolinium blockage of mechanosensitive ion channels. CONCLUSIONS: Our results suggest that low-intensity pulsed ultrasound would transiently induce remodeling of a cell’s membrane and cytoskeleton, and it will lead to repression of nucleus. This indicates that ultrasound after all represents a mechanical stress on cellular membrane. The post-exposure outgrowth phenomenon is also of practical relevance as it may be linked to the stimulatory effects that have been already observed in low-intensity pulsed ultrasound treatments.postprin

Development of positioning devices for MRI-guided high intensity focused ultrasound (HIFU) for abdominal, thyroid and brain, tumours

High intensity focused ultrasound (HIFU) is a promising technology for a variety of therapeutic applications. This concept initiated in 1942 by Lynn Zwemer [1]. HIFU has long been known as a minimal invasive or non-invasive procedure that destroys tissue through ablation. However, it is only in recent years that clinical applications are becoming feasible, with the development of high power ultrasound transducers compatible with the MRI scanner which is used to monitor these non-invasive HIFU applications. New technologies, combined with more sophisticated treatment methods and monitoring methods allow non-invasive procedures in many areas such as the brain, eye, breast, kidney, liver, pancreas, thyroid, uterine fibroids and pancreas. Meanwhile, new investigations are underway for treading cardiac arithmia, strokes, palliative pain treatment of bone metastases and brain disorders such as Parkinson’s disease, essential tremor, and neuropathic pain. These optimistic investigations have encouraged physicians and provided them new valuable tools for medical research

Precise Lesion Formation in Histotripsy Therapy Using Strategic Pulsing Methods..

Histotripsy is a noninvasive, cavitation-based ultrasound therapy that can create mechanical tissue ablation through dense energetic clouds of microbubbles generated by high-pressure and short ultrasound pulses. Histotripsy therapy has been shown capable of 1) creating intracardiac flow channels for congenital heart disease treatment, 2) fractionating blood clots for treating deep vein thrombosis, 3) fractionating prostatic tissue for benign prostatic hyperplasia (BPH) treatments, and 4) fragmenting model renal calculi for treating kidney stone. The overall objective of this dissertation is to develop ultrasound pulsing techniques that can lead to more precise and controlled bubble cloud generation in histotripsy therapy. Three strategic pulsing methods have been developed and characterized in this dissertation. 1) Bubble cloud formation using the intrinsic threshold mechanism: when ultrasound pulses shorter than 3 cycles are applied, the generation of bubble clouds only depends on one or two negative half cycles exceeding an intrinsic threshold of the medium. This intrinsic threshold is highly repeatable and has a very sharp transition zone. 2) Dual-beam histotripsy: a low-frequency pump pulse is applied to enable a high-frequency probe pulse to exceed the intrinsic threshold. The high-frequency probe pulse provides precision in lesion formation, while the low-frequency pump pulse, which is more resistant to attenuation and aberration, raises the pressure level of the targeted treatment region. 3) Frequency compounding: a near half-cycle (monopolar) pulse is synthesized using an array transducer composed of elements with various resonant frequencies. Histotripsy using a negative-polarity half-cycle pulse can limit the influence of positive phases on bubble cloud generation, leading to a more precise and controlled lesion formation. These three techniques were realized using custom design ultrasound array transducers and examined in red-blood-cell tissue-mimicking phantoms, and the first two techniques were further validated in ex vivo tissues. Additionally, an application in metastatic lymph node ablation is studied in vivo using supra-intrinsic-threshold pulses. In conclusion, this dissertation demonstrates three strategic ultrasound pulsing methods that can lead to precise lesion formation in histotripsy therapy. Future work involves examining the applicability of these pulsing methods in in vivo experiments and studying potential applications for monopolar pulses in ultrasound diagnostic imaging.PHDBiomedical EngineeringUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/107092/1/kwlin_1.pd

Noninvasive Thrombolysis Using Histotripsy Pulsed Ultrasound Cavitation Therapy.

Histotripsy is a noninvasive ultrasound therapy that utilizes short, high-amplitude, focused ultrasound pulses to mechanically reduce targeted tissue structures to liquid debris by acoustic cavitation. In this work, the physical mechanisms of histotripsy and its application as a method of thrombolysis were investigated. Cavitation activity which causes tissue breakdown during histotripsy was studied by high-speed photography. It was found that cavitation clouds form due to scattering of shock waves in a focused ultrasound pulse from individual inertial cavitation bubbles. The scattered shock is a large tensile wave which expands clusters of cavitation bubbles when the tensile pressure is greater than a measured threshold of approximately 30 MPa. The interaction of this cavitation with tissue and cells was explored with a phantom containing agarose and red blood cells to measure cavitation-based mechanical damage. The observations indicated that cell lysis may be achieved by bubble-induced tensile strain upon expansion, causing membrane rupture. Based on these studies, focused histotripsy therapy transducers were designed to controllably generate cavitation clouds in the vasculature for performing thrombolysis. Transducers were integrated with ultrasound imagers to provide feedback for targeting and monitoring progress of treatment. Rapid thrombolysis was observed when histotripsy was applied to clots in-vitro, and the resulting debris was mainly subcellular and unlikely to cause embolism. Additionally, it was observed that histotripsy can attract, trap, and destroy free clot fragments in a vessel phantom. Based on these observations, a noninvasive embolus trap (NET) was developed, acting as a filter to prevent embolism during the thrombolysis procedure. An in-vivo porcine model of deep-vein thrombosis was used to evaluate the safety and efficacy of the histotripsy thrombolysis technique. These experiments demonstrated the feasibility of the treatment and suggest histotripsy can achieve rapid clot breakdown in a controlled manner.Ph.D.Biomedical EngineeringUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/91496/1/adamdm_1.pd