An inverse finite-element model of heel-pad indentation

A numerical-experimental approach has been developed to characterize heel-pad deformation at the material level. Left and right heels of 20 diabetic subjects and 20 nondiabetic subjects matched for age, gender and body mass index were indented using force-controlled ultrasound. Initial tissue thickness and deformation were measured using M-mode ultrasound; indentation forces were recorded simultaneously. An inverse finite-element analysis of the indentation protocol using axisymmetric models adjusted to reflect individual heel thickness was used to extract nonlinear material properties describing the hyperelastic behavior of each heel. Student's t-tests revealed that heel pads of diabetic subjects were not significantly different in initial thickness nor were they stiffer than those from nondiabetic subjects. Another heel-pad model with anatomically realistic surface representations of the calcaneus and soft tissue was developed to estimate peak pressure prediction errors when average rather than individualized material properties were used. Root-mean-square errors of up to 7% were calculated, indicating the importance of subject-specific modeling of the nonlinear elastic behavior of the heel pad. Indentation systems combined with the presented numerical approach can provide this information for further analysis of patient-specific foot pathologies and therapeutic footwear designs.     

Determination of lesion size by ultrasound during radiofrequency catheter ablation.

The catheter tip temperature that is used to control the radiofrequency generator output poorly correlates to lesion size. We, therefore, evaluated lesions created in vitro using a B-mode ultrasound imaging device as a potential means to assess lesion generation during RF applications non-invasively. Porcine ventricular tissue was immersed in saline solution at 37 degrees C. The catheter was fixed in a holder and positioned in a parallel orientation to the tissue with an array transducer (7.5 MHz) app. 3 cm above the tissue. Lesions were produced either in a temperature controlled mode with a 4-mm tip catheter with different target temperatures (50, 60, 70 and 80 degrees C, 80 W maximum output) or in a power controlled mode (25, 50 and 75 W, 20 ml/min irrigation flow) using an irrigated tip catheter. Different contact forces (0.5 N, 1.0 N) were tested, and RF was delivered for 60 s. A total of 138 lesions was produced. Out of these, 128 could be identified on the ultrasound image. The lesion depth and volume was on average 4.1 +/- 1.6 mm and 52 +/- 53 mm3 as determined by ultrasound and 3.9 +/- 1.7 mm and 52 +/- 55 mm3 as measured thereafter, respectively. A linear correlation between the lesion size determined by ultrasound and that measured thereafter was demonstrated with a correlation coefficient of r = 0.87 for lesion depth and r = 0.93 for lesion volume. We conclude that lesions can be assessed by B-mode ultrasound imaging.     

Single Molecule Probe Scanning Optical Force Imaging Microscopefor Viewing Live Cells

We have developed an imaging system that combines the soft compliance of an optical trap with the sensitivity of single particle tracking to image forces on/in live cells using a single molecule probe. The probe used is a single (or few) molecule of interest that is conjugated with a single 40 nm colloidalgold probe. The colloidal gold/membrane protein complex, freely diffusing on a live cell, is held in a laser trap while the cell is scanned underneath. Computer control allows for synchronization of the cell scan and capture of the probe position. Resistance to the dragging of the probe images a fine structure of barriers in the membrane of live cells.     

An in situ calibration of an ultrasound transducer: a potential application for an ultrasonic indentation test of articular cartilage

A change in mechanical properties of articular cartilage would be considered one of the most reliable signs of cartilage degeneration. While an indentation method has the potential to measure the cartilage properties in vivo, an accurate measurement of cartilage thickness in situ is technically difficult. An ultrasound transducer has often been used to measure the cartilage thickness. However, its accuracy is limited by the lack of an accurate measurement of the ultrasound speed of cartilage, for the ultrasound speed varies according to the pathological conditions of the tissue. Therefore, the objective of this study is to develop an in situ calibration method of predicting the true ultrasound speed of cartilage and thus allow the ultrasound transducer to measure the thickness of the tissue with great accuracy. By simultaneously implementing an indentation testing protocol using the ultrasound transducer as an indenter, this method can also provide an indentation stiffness measurement of cartilage.The feasibility of the proposed method was examined using normal and proteoglycan-depleted cartilage specimens. It was found that the true ultrasound speed measured by the in situ calibration method was sensitive to the proteoglycan depletion (1735+/-35 m/s for normal, and 1598+/-28 m/s for proteoglycan-depleted cartilage), and that the measured cartilage thickness was consistently accurate regardless of the tissue condition. The measured indentation stiffness of articular cartilage was also sensitive to the tissue condition. Thus, this study demonstrates that the proposed ultrasonic indentation technique can be used to accurately identify the abnormality of articular cartilage in situ.     

Monitoring Radiofrequency Ablation Using Real-Time Ultrasound Nakagami Imaging Combined with Frequency and Temporal Compounding Techniques

Gas bubbles induced during the radiofrequency ablation (RFA) of tissues can affect the detection of ablation zones (necrosis zone or thermal lesion) during ultrasound elastography. To resolve this problem, our previous study proposed ultrasound Nakagami imaging for detecting thermal-induced bubble formation to evaluate ablation zones. To prepare for future applications, this study (i) created a novel algorithmic scheme based on the frequency and temporal compounding of Nakagami imaging for enhanced ablation zone visualization, (ii) integrated the proposed algorithm into a clinical scanner to develop a real-time Nakagami imaging system for monitoring RFA, and (iii) investigated the applicability of Nakagami imaging to various types of tissues. The performance of the real-time Nakagami imaging system in visualizing RFA-induced ablation zones was validated by measuring porcine liver (n = 18) and muscle tissues (n = 6). The experimental results showed that the proposed algorithm can operate on a standard clinical ultrasound scanner to monitor RFA in real time. The Nakagami imaging system effectively monitors RFA-induced ablation zones in liver tissues. However, because tissue properties differ, the system cannot visualize ablation zones in muscle fibers. In the future, real-time Nakagami imaging should be focused on the RFA of the liver and is suggested as an alternative monitoring tool when advanced elastography is unavailable or substantial bubbles exist in the ablation zone.     

In vivo quantitative microvasculature phenotype imaging of healthy and malignant tissues using a fiber-optic confocal laser microprobe

Real-time in vivo imaging of the microvasculature may help both earlier clinical detection of disease and the understanding of tumor-host interaction at various stages of progression. In vivo confocal and multiphoton microscopy is often hampered by bulky optics setup and has limited access to internal organs. A fiber-optic setup avoids these limitations and offers great user maneuverability. We report here the in vivo validation of a fiber-optic confocal fluorescence microprobe imaging system. In addition, we developed an automated fractal-based image analysis to characterize microvascular morphology based on vessel diameter distribution, density, volume fraction, and fractal dimension from real-time data. The system is optimized for use in the far-red and near-infrared region. The flexible 1.5-mm-diameter fiber-optic bundle and microprobe enable great user maneuverability, with a field of view of 423 x 423 microm and a tissue penetration of up to 15 microm. Lateral and axial resolutions are 3.5 and 15 microm. We show that it is possible to obtain high temporal and spatial resolution images of virtually any abdominal viscera in situ using a far-red blood pool imaging probe. Using an orthotopic model of pancreatic ductal adenocarcinoma, we characterized the tumor surface capillary and demonstrated that the imaging system and analysis can quantitatively differentiate between the normal and tumor surface capillary. This clinically approved fiber-optic system, together with the fractal-based image analysis, can potentially be applied to characterize other tumors in vivo and may be a valuable tool to facilitate their clinical evaluation.     

An improved microindentation technique to measure changes in properties of arterial intima during atherogenesis

A fully automated instrument was developed that measured the amount and time-course of indentation of arterial intima by a spherical tipped probe 25 micrometers in diameter loaded routinely by forces of 10 microN. The relationship of depth of indentation to time was non-exponential but reached an apparent asymptotic value in about 6 s. Routine indentations were recorded 10 s after loading, the relationship of indentation and force being essentially linear between 0-100 microN. The instrument is an order of magnitude more sensitive than one previously described (Gow and Vaishnav, 1975, J. Appl. Physiol. 38, 344-350) and produces indentations of 1-4 micrometers in the intima of rabbit thoracic aorta with a reproducibility of 1-2%. Preliminary usage of the new microindentor has shown increases in the compliance of the thoracic aorta intima in rabbits during cholesterol feeding.     

Resting-State Temporal Synchronization Networks Emerge from Connectivity Topology and Heterogeneity

Spatial patterns of coherent activity across different brain areas have been identified during the resting-state fluctuations of the brain. However, recent studies indicate that resting-state activity is not stationary, but shows complex temporal dynamics. We were interested in the spatiotemporal dynamics of the phase interactions among resting-state fMRI BOLD signals from human subjects. We found that the global phase synchrony of the BOLD signals evolves on a characteristic ultra-slow (<0.01Hz) time scale, and that its temporal variations reflect the transient formation and dissolution of multiple communities of synchronized brain regions. Synchronized communities reoccurred intermittently in time and across scanning sessions. We found that the synchronization communities relate to previously defined functional networks known to be engaged in sensory-motor or cognitive function, called resting-state networks (RSNs), including the default mode network, the somato-motor network, the visual network, the auditory network, the cognitive control networks, the self-referential network, and combinations of these and other RSNs. We studied the mechanism originating the observed spatiotemporal synchronization dynamics by using a network model of phase oscillators connected through the brain’s anatomical connectivity estimated using diffusion imaging human data. The model consistently approximates the temporal and spatial synchronization patterns of the empirical data, and reveals that multiple clusters that transiently synchronize and desynchronize emerge from the complex topology of anatomical connections, provided that oscillators are heterogeneous.     

Three‐dimensional interventional photoacoustic imaging for biopsy needle guidance with a linear array transducer

Needle placement is important for many clinical interventions, such as tissue biopsy, regional anesthesia and drug delivery. It is essential to visualize the spatial position of the needle and the target tissue during the interventions using appropriate imaging techniques. Based on the contrast of optical absorption, photoacoustic imaging is well suited for the guidance of interventional procedures. However, conventional photoacoustic imaging typically provides two‐dimensional (2D) slices of the region of interest and could only visualize the needle and the target when they are within the imaging plane of the probe at the same time. This requires great alignment skill and effort. To ease this problem, we developed a 3D interventional photoacoustic imaging technique by fast scanning a linear array ultrasound probe and stitching acquired image slices. in vivo sentinel lymph node biopsy experiment shows that the technique could precisely locate a needle and a sentinel lymph node in a tissue volume while a perfusion experiment demonstrates that the technique could visualize the 3D distribution of injected methylene blue dye underneath the skin at high temporal and spatial resolution. The proposed technique provides a practical way for photoacoustic image‐guided interventions.      

High-Resolution Harmonics Ultrasound Imaging for Non-Invasive Characterization of Wound Healing in a Pre-Clinical Swine Model

This work represents the first study employing non-invasive high-resolution harmonic ultrasound imaging to longitudinally characterize skin wound healing. Burn wounds (day 0-42), on the dorsum of a domestic Yorkshire white pig were studied non-invasively using tandem digital planimetry, laser speckle imaging and dual mode (B and Doppler) ultrasound imaging. Wound depth, as measured by B-mode imaging, progressively increased until day 21 and decreased thereafter. Initially, blood flow at the wound edge increased up to day 14 and subsequently regressed to baseline levels by day 21, when the wound was more than 90% closed. Coinciding with regression of blood flow at the wound edge, there was an increase in blood flow in the wound bed. This was observed to regress by day 42. Such changes in wound angiogenesis were corroborated histologically. Gated Doppler imaging quantitated the pulse pressure of the primary feeder artery supplying the wound site. This pulse pressure markedly increased with a bimodal pattern following wounding connecting it to the induction of wound angiogenesis. Finally, ultrasound elastography measured tissue stiffness and visualized growth of new tissue over time. These studies have elegantly captured the physiological sequence of events during the process of wound healing, much of which is anticipated based on certain dynamics in play, to provide the framework for future studies on molecular mechanisms driving these processes. We conclude that the tandem use of non-invasive imaging technologies has the power to provide unprecedented insight into the dynamics of the healing skin tissue.     

Validation of a freehand 3D ultrasound system for morphological measures of the medial gastrocnemius muscle

Muscle volume and length are important parameters for examining the force-generating capabilities of muscle and their evaluation is necessary in studies that investigate muscle morphology and mechanical changes due to age, function, pathology, surgery and training. In this study, we assessed the validity and reliability of in vivo muscle volume and muscle belly length measurement using a multiple sweeps freehand 3D ultrasound (3DUS). The medial gastrocnemius of 10 subjects was scanned at three ankle joint angles (15°, 0° and ?15° dorsiflexion) three times using the freehand 3DUS and once on the following day using magnetic resonance imaging (MRI). All freehand 3DUS and MRI images were segmented, volumes rendered and volumes and muscle belly lengths measured. The freehand 3DUS overestimated muscle volume by 1.9±9.1 mL, 1.1±3.8% difference and underestimated muscle belly length by 3.0±5.4 mm, 1.3±2.2% difference. The intra-class correlation coefficients (ICC) for repeated freehand 3DUS system measures of muscle volume and muscle belly length were greater than 0.99 and 0.98, respectively. The ICCs for the segmentation process reliability for the freehand 3DUS system and MRI for muscle volume were both greater than 0.99 and muscle belly length were 0.97 and 0.99, respectively. Freehand 3DUS is a valid and reliable method for the measurement of human muscle volume and muscle belly length in vivo. It could be used as an alternative to MRI for measuring in vivo muscle morphology and thus allowing the determination of PCSA and estimation of the force-generating capacity of individual muscles within the setting of a biomechanics laboratory.     

Imaging cortical dynamics at high spatial and temporal resolution with novel blue voltage-sensitive dyes

Conventional imaging techniques have provided high-resolution imaging either in the spatial domain or in the temporal domain. Optical imaging utilizing voltage-sensitive dyes has long had the unrealized potential to achieve high resolution in both domains simultaneously, providing subcolumnar spatial detail with millisecond precision. Here, we present a series of developments in voltage-sensitive dyes and instrumentation that make functional imaging of cortical dynamics practical, in both anesthetized and awake behaving preparations, greatly facilitating exploration of the cortex. We illustrate this advance by analyzing the millisecond-by-millisecond emergence of orientation maps in cat visual cortex.     

Mechano-acoustic determination of Young's modulus of articular cartilage

The compressive stiffness of an elastic material is traditionally characterized by its Young's modulus. Young's modulus of articular cartilage can be directly measured using unconfined compression geometry by assuming the cartilage to be homogeneous and isotropic. In isotropic materials, Young's modulus can also be determined acoustically by the measurement of sound speed and density of the material. In the present study, acoustic and mechanical techniques, feasible for in vivo measurements, were investigated to quantify the static and dynamic compressive stiffness of bovine articular cartilage in situ. Ultrasound reflection from the cartilage surface, as well as the dynamic modulus were determined with the recently developed ultrasound indentation instrument and compared with the reference mechanical and ultrasound speed measurements in unconfined compression (n=72). In addition, the applicability of manual creep measurements with the ultrasound indentation instrument was evaluated both experimentally and numerically. Our experimental results indicated that the sound speed could predict 47% and 53% of the variation in the Young's modulus and dynamic modulus of cartilage, respectively. The dynamic modulus, as determined manually with the ultrasound indentation instrument, showed significant linear correlations with the reference Young's modulus (r(2)=0.445, p<0.01, n=70) and dynamic modulus (r(2)=0.779, p<0.01, n=70) of the cartilage. Numerical analyses indicated that the creep measurements, conducted manually with the ultrasound indentation instrument, were sensitive to changes in Young's modulus and permeability of the tissue, and were significantly influenced by the tissue thickness. We conclude that acoustic parameters, i.e. ultrasound speed and reflection, are indicative to the intrinsic mechanical properties of the articular cartilage. Ultrasound indentation instrument, when further developed, provides an applicable tool for the in vivo detection of cartilage mechano-acoustic properties. These techniques could promote the diagnostics of osteoarthrosis.     

Low frequency cMUT technology: Application to measurement of brain movement and assessment of bone quality

Following recent advances in medical ultrasound imaging methods almost all human tissues can currently be examined. There are, however, two exceptions: the human skeleton and the brain, because bone tissue is a strongly attenuating and defocusing medium, rendering classical pulse-echo imaging methods inappropriate. Specific imaging approaches within low frequency bands, i.e. 200 kHz–2 MHz, have therefore recently been developed and the results are very promising: (1) the technique for the bone is axial transmission measurement, which consists of using elastic guided modes to characterize all elastic constants of the medium; (2) for brain exploration, it has been demonstrated that brain movement can be measured (i.e. brain pulsatility) with elastography techniques. However, there are certain limitations in the fabrication of low frequency probes with classical technology, which involve finding an alternative to the traditional PZT. Capacitive Micromachined Ultrasonic Transducers (cMUTs) can overcome these limitations and greatly improve these new imaging modalities. The study presented here represents technological development with several goals: (1) the design and fabrication of two different low frequency linear arrays for bone and brain exploration, respectively the testing of axial transmission measurements with a cMUT probe and; (2) comparison with a PZT probe; (3) the development of an imaging method based on the elastography of brain pulsatility, its implementation in a commercial ultrasound scanner and clinical trials for the validation. The results obtained with cMUT and PZT probes are compared.     

Non‐invasive sentinel lymph node mapping and needle guidance using clinical handheld photoacoustic imaging system in small animal

Translating photoacoustic imaging (PAI) into clinical setup is a challenge. Handheld clinical real‐time PAI systems are not common. In this work, we report an integrated photoacoustic (PA) and clinical ultrasound imaging system by combining light delivery with the ultrasound probe for sentinel lymph node imaging and needle guidance in small animal. The open access clinical ultrasound platform allows seamless integration of PAI resulting in the development of handheld real‐time PAI probe. Both methylene blue and indocyanine green were used for mapping the sentinel lymph node using 675 and 690 nm wavelength illuminations, respectively. Additionally, needle guidance with combined ultrasound and PAI was demonstrated using this imaging system. Up to 1.5 cm imaging depth was observed with a 10 Hz laser at an imaging frame rate of 5 frames per second, which is sufficient for future translation into human sentinel lymph node imaging and needle guidance for fine needle aspiration biopsy.      

A Novel Application of Musculoskeletal Ultrasound Imaging

Ultrasound is an attractive modality for imaging muscle and tendon motion during dynamic tasks and can provide a complementary methodological approach for biomechanical studies in a clinical or laboratory setting. Towards this goal, methods for quantification of muscle kinematics from ultrasound imagery are being developed based on image processing. The temporal resolution of these methods is typically not sufficient for highly dynamic tasks, such as drop-landing. We propose a new approach that utilizes a Doppler method for quantifying muscle kinematics. We have developed a novel vector tissue Doppler imaging (vTDI) technique that can be used to measure musculoskeletal contraction velocity, strain and strain rate with sub-millisecond temporal resolution during dynamic activities using ultrasound. The goal of this preliminary study was to investigate the repeatability and potential applicability of the vTDI technique in measuring musculoskeletal velocities during a drop-landing task, in healthy subjects. The vTDI measurements can be performed concurrently with other biomechanical techniques, such as 3D motion capture for joint kinematics and kinetics, electromyography for timing of muscle activation and force plates for ground reaction force. Integration of these complementary techniques could lead to a better understanding of dynamic muscle function and dysfunction underlying the pathogenesis and pathophysiology of musculoskeletal disorders.     

Ultrasound enhanced delivery of molecular imaging and therapeutic agents in Alzheimer's disease mouse models

Alzheimer's disease is a neurodegenerative disorder typified by the accumulation of a small protein, beta-amyloid, which aggregates and is the primary component of amyloid plaques. Many new therapeutic and diagnostic agents for reducing amyloid plaques have limited efficacy in vivo because of poor transport across the blood-brain barrier. Here we demonstrate that low-intensity focused ultrasound with a microbubble contrast agent may be used to transiently disrupt the blood-brain barrier, allowing non-invasive, localized delivery of imaging fluorophores and immunotherapeutics directly to amyloid plaques. We administered intravenous Trypan blue, an amyloid staining red fluorophore, and anti-amyloid antibodies, concurrently with focused ultrasound therapy in plaque-bearing, transgenic mouse models of Alzheimer's disease with amyloid pathology. MRI guidance permitted selective treatment and monitoring of plaque-heavy anatomical regions, such as the hippocampus. Treated brain regions exhibited 16.5+/-5.4-fold increase in Trypan blue fluorescence and 2.7+/-1.2-fold increase in anti-amyloid antibodies that localized to amyloid plaques. Ultrasound-enhanced delivery was consistently reproduced in two different transgenic strains (APPswe:PSEN1dE9, PDAPP), across a large age range (9-26 months), with and without MR guidance, and with little or no tissue damage. Ultrasound-mediated, transient blood-brain barrier disruption allows the delivery of both therapeutic and molecular imaging agents in Alzheimer's mouse models, which should aid pre-clinical drug screening and imaging probe development. Furthermore, this technique may be used to deliver a wide variety of small and large molecules to the brain for imaging and therapy in other neurodegenerative diseases.     

Histochemical Probing of Potato Periderm with Neutral Red: A Sensitive Cytofluorochrome for the Hydrophobic Domain of Suberin

A technique is described which uses the lipid fluorochrome neutral red as a cytochemical probe to detect the hydrophobic domain of the lignosuberin matrix in native and wound periderm of potato tuber. Toluidine blue O is used as a counterstain to quench autofluorescence. The neutral red technique appears to be specific for the hydrophobic/lipid domain of suberin and is significantly more sensitive than Sudan III and IV. The fluorochrome was extensively used on paraffin-embedded tissue with excellent results but also worked on freehand sections of fresh periderm tissue. In tuber tissue undergoing wound-healing, the pattern of suberin fluorescence obtained with the neutral red probe was identical in specificity to the color pattern obtained with Sudan III/IV, but somewhat different than that observed when berberine was used. Results obtained with the neutral red probe and berberine probe visually demonstrated that during ligno-suberin biosynthesis, the depositions of hydrophobic/lipid and phenolic/lignin-like components in potato tuber periderm were separate processes. The deposition of these components does not necessarily require their simultaneous presence because the fluorescence from these probes showed that the components were not consistently present together on the cell walls.     

Imaging modalities to assess structural birth defects in mutant mouse models

Assessment of structural birth defects (SBDs) in animal models usually entails conducting detailed necropsy for anatomical defects followed by histological analysis for tissue defects. Recent advances in new imaging technologies have provided the means for rapid phenotyping of SBDs, such as using ultra‐high frequency ultrasound biomicroscopy, optical coherence tomography, micro‐CT, and micro‐MRI. These imaging modalities allow the detailed assessment of organ/tissue structure, and with ultrasound biomicroscopy, structure and function of the cardiovascular system also can be assessed noninvasively, allowing the longitudinal tracking of the fetus in utero. In this review, we briefly discuss the application of these state‐of‐the‐art imaging technologies for phenotyping of SBDs in rodent embryos and fetuses, showing how these imaging modalities may be used for the detection of a wide variety of SBDs. Birth Defects Research (Part C) 90:176–184, 2010. © 2010 Wiley‐Liss, Inc.     

Fluid load support during localized indentation of cartilage with a spherical probe

Interstitial fluid pressurization, a consequence of a biphasic tissue structure, is essential to the load bearing and lubrication properties of articular cartilage. Focal tissue degradation may interfere with this protective mechanism, eventually leading to gross degeneration and osteoarthritis. Our long-term goal is to determine whether local contacts can be used as a means to probe local tissue integrity and functionality. In the present work, Hertzian rate-controlled microindentation was used as a model of the more complicated sliding system to directly determine the effects of contact radius and deformation rate on interstitial load support. During localized contact between a steel spherical probe and bovine articular cartilage, the equilibrium and non-equilibrium responses were well-fit by the Hertz model (R(2)>0.998) with a mean equilibrium contact modulus of 0.93 MPa. The effective contact modulus and fluid load fraction were independent of indentation depth, contact radius, and normal force; both increased monotonically with indentation rate. At 21 μm/s indentation rate, the cartilage was effectively stiffened by 6-fold with the fluid pressure supporting 85% of the contact force. The results motivated a simple analytical model that directly links the tribomechanical response (including fluid load support) and the Peclet number to measurable material properties and controllable experimental variables. This paper demonstrates that tribological contacts can be used to probe local functional properties. Such measurements can add important insights into the roles of focal tissue damage and impaired local functionality in the pathogenesis of osteoarthritis.