Remote Non-invasive Stereoscopic Imaging of Blood Vessels: First In-vivo Results of a New Multispectral Contrast Enhancement Technology

We describe a contactless optical technique selectively enhancing superficial blood vessels below variously pigmented intact human skin by combining images in different spectral bands. Two CMOS-cameras, with apochromatic lenses and dual-band LED-arrays, simultaneously streamed Left (L) and Right (R) image data to a dual-processor PC. Both cameras captured color images within the visible range (VIS, 400–780 nm) and grey-scale images within the near infrared range (NIR, 910–920 nm) by sequentially switching between LED-array emission bands. Image-size-settings of 1280 × 1024 for VIS & 640 × 512 for NIR produced 12 cycles/s (1 cycle = 1 VIS L&R-pair + 1 NIR L&R-pair). Decreasing image-size-settings (640 × 512 for VIS and 320 × 256 for NIR) increased camera-speed to 25 cycles/s. Contrasts from below the tissue surface were algorithmically distinguished from surface shadows, reflections, etc. Thus blood vessels were selectively enhanced and back-projected into the stereoscopic VIS-color-image using either a 3D-display or conventional shutter glasses. As a first usability reconnaissance we applied this custom-built mobile stereoscopic camera for several clinical settings: • blood withdrawal; • vein inspection in dark skin; • vein detection through iodide; • varicose vein and nevi pigmentosum inspection. Our technique improves blood vessel visualization compared to the naked eye, and supports depth perception

The Relation Between Histological, Tumor-Biological and Clinical Parameters in Deep and Superficial Leiomyosarcoma and Leiomyoma

Purpose: Leiomyosarcomas (LMS) of deep and superficial tissues were examined to identify prognostic markers explaining their different biological behaviour and to define differences between cutaneous and subcutaneous LMS. LMS and leiomyomas (LM) of the skin were compared to and consistent differences that could aid in the (sometimes difficult) diagnosis

Lipid-rich Plaques Detected by Near-infrared Spectroscopy Are More Frequently Exposed to High Shear Stress

High wall shear stress (WSS) and near-infrared spectroscopy (NIRS) detected lipid-rich plaque (LRP) are both known to be associated with plaque destabilization and future adverse cardiovascular events. However, knowledge of spatial co-localization of LRP and high WSS is lacking. This study investigated the co-localization of LRP based on NIRS and high WSS. Fifty-three patients presenting acute coronary syndrome underwent NIRS-intravascular-ultrasound (NIRS-IVUS) imaging of a non-culprit coronary artery. WSS was obtained using WSS profiling in 3D-reconstructions of the coronary arteries based on fusion of IVUS-segmented lumen and CT-derived 3D-centerline. Thirty-eight vessels were available for final analysis and divided into 0.5 mm/45° sectors. LRP sectors, as identified by NIRS, were more often colocalized with high WSS than sectors without LRP. Moreover, there was a dose-dependent relationship between lipid content and high WSS exposure. This study is a first step in understanding the evolution of LRPs to vulnerable plaques. [Figure not available: see fulltext.

Real-time volumetric lipid imaging in vivo by intravascular photoacoustics at 20 frames per second

Lipid deposition can be assessed with combined intravascular photoacoustic/ultrasound (IVPA/US) imaging. To date, the clinical translation of IVPA/US imaging has been stalled by a low imaging speed and catheter complexity. In this paper, we demonstrate imaging of lipid targets in swine coronary arteries in vivo, at a clinically useful frame rate of 20 s−1. We confirmed image contrast for atherosclerotic plaque in human samples ex vivo. The system is on a mobile platform and provides real-time data visualization during acquisition. We achieved an IVPA signal-to-noise ratio of 20 dB. These data show that clinical translation of IVPA is possible in principle

Combined optical coherence tomography and intravascular ultrasound radio frequency data analysis for plaque characterization. Classification accuracy of human coronary plaques in vitro

This study was performed to characterize coronary plaque types by optical coherence tomography (OCT) and intravascular ultrasound (IVUS) radiofrequency (RF) data analysis, and to investigate the possibility of error reduction by combining these techniques. Intracoronary imaging methods have greatly enhanced the diagnostic capabilities for the detection of high-risk atherosclerotic plaques. IVUS RF data analysis and OCT are two techniques focusing on plaque morphology and composition. Regions of interest were selected and imaged with OCT and IVUS in 50 sections, from 14 human coronary arteries, sectioned post-mortem from 14 hearts of patients dying of non-cardiovascular causes. Plaques were classified based on IVUS RF data analysis (VH-IVUSTM), OCT and the combination of those. Histology was the benchmark. Imaging with both modalities and coregistered histology was successful in 36 sections. OCT correctly classified 24; VH-IVUS 25, and VH-IVUS/OCT combined, 27 out of 36 cross-sections. Systematic misclassifications in OCT were intimal thickening classified as fibroatheroma in 8 cross-sections. Misclassifications in VH-IVUS were mainly fibroatheroma as intimal thickening in 5 cross-sections. Typical image artifacts were found to affect the interpretation of OCT data, misclassifying intimal thickening as fibroatheroma or thin-cap fibroatheroma. Adding VH-IVUS to OCT reduced the error rate in this study

PTEN Is Associated With Worse Local Control in Early Stage Supraglottic Laryngeal Cancer Treated With Radiotherapy

Objectives: The aim of this study was to establish the prognostic value of the epidermal growth factor receptor (EGFR) and phosphatase and tensin homolog deleted on chromosome 10 (PTEN) expression on local control in patients with early stage supraglottic laryngeal squamous cell carcinoma (LSCC) treated with radiotherapy only. Study design: Retrospective cohort study. Methods: Immunohistochemical staining for EGFR and PTEN was performed on pretreatment biopsies of a selected well-defined homogeneous group of 52 patients with T1-T2 supraglottic LSCC treated with radiotherapy between 1990 and 2008. Kaplan-Meier analysis and univariate and multivariate Cox Regression analyses were performed to correlate clinical data and expression levels of EGFR and PTEN with local control. Results: Kaplan-Meier survival analysis and Cox Regression analysis showed a significant association between PTEN expression and local control (hazard ratio [HR] = 3.26, 95% confidence interval [CI] = 1.14-9.33, P = .027) and between lymph node status and local control (HR = 3.60, 95% CI = 1.26-10.31, P = .017). Both were independent prognostic factors in a multivariate analysis (HR = 3.28, 95% CI = 1.14-9.39, P = .027 and HR = 3.62, 95% CI = 1.26-10.37, P = .017, respectively). There was no significant association between EGFR expression and local control (HR = 1.32, 95% CI = 1.17-10.14, P = .79). Conclusion: This study showed an association between both high PTEN expression and the presence of lymph node metastasis and deteriorated local control in early stage supraglottic LSCC treated with radiotherapy. Level of Evidence: NA

Cortactin expression assessment improves patient selection for a watchful waiting strategy in pT1cN0-staged oral squamous cell carcinomas with a tumor infiltration depth below 4 mm

BACKGROUND: In this feasibility study we aimed to evaluate the value of previously reported molecular tumor biomarkers associated with lymph node metastasis in oral squamous cell carcinoma (OSCC) to optimize neck strategy selection criteria. METHODS: The association between expression of cortactin, cyclin D1, FADD, RAB25, and S100A9 and sentinel lymph node status was evaluated in a series of 87 (cT1‐2N0) patients with OSCC treated with primary resection and SLNB procedure. RESULTS: Tumor infiltration depth and tumor pattern of invasion were independent prognostic markers for SLN status, while none of the tumor makers showed a better prognostic value to replace SLNB as neck staging technique in the total cohort. However, in the subgroup of patients with pT1N0 OSCC, cortactin expression (OR 16.0, 95%CI 2.0–127.9) was associated with SLN classification. CONCLUSIONS: Expression of cortactin is a promising immunohistochemical tumor marker to identify patients at low risk that may not benefit from SLNB or END

Focal areas of increased lipid concentration on the coating of microbubbles during short tone-burst ultrasound insonification

Acoustic behavior of lipid-coated microbubbles has been widely studied, which has led to several numerical microbubble dynamics models that incorporate lipid coating behavior, such as buckling and rupture. In this study we investigated the relationship between micro-bubble acoustic and lipid coating behavior on a nanosecond scale by using fluorescently labeled lipids. It is hypothesized that a local increased concentration of lipids, appearing as a focal area of increased fluorescence intensity (hot spot) in the fluorescence image, is related to buckling and folding of the lipid layer thereby highly influencing the microbubble acoustic behavior. To test this hypothesis, the lipid microbubble coating was fluorescently labeled. The vibration of the microbubble (n= 177; 2.3-10.3 μm in diameter) upon insonification at an ultrasound frequency of 0.5 or 1 MHz at 25 or 50 kPa acoustic pressure was recorded with the UPMC Cam, an ultra-high-speed fluorescence camera, operated at ∼4-5 million frames per second. During short tone-burst excitation, hot spots on the microbubble coating occurred at relative vibration amplitudes > 0.3 irrespective of frequency and acoustic pressure. Around resonance, the majority of the microbubbles formed hot spots. When the microbubble also deflated acoustically, hot spot formation was likely irreversible. Although compression-only behavior (defined as substantially more microbubble compression than expansion) and subharmonic responses were observed in those microbubbles that formed hot spots, both phenomena were also found in microbubbles that did not form hot spots during insonification. In conclusion, this study reveals hot spot formation of the lipid monolayer in the microbubble's compression phase. However, our experimental results show that there is no direct relationship between hot spot formation of the lipid coating and microbubble acoustic behaviors such as compression-only and the generation of a subharmonic response. Hence, our hypothesis that hot spots are related to acoustic buckling could not be verified

High-Resolution Imaging of Intracellular Calcium Fluctuations Caused by Oscillating Microbubbles

Ultrasound insonification of microbubbles can locally enhance drug delivery, but the microbubble–cell interaction remains poorly understood. Because intracellular calcium (Cai 2+) is a key cellular regulator, unraveling the Cai 2+ fluctuations caused by an oscillating microbubble provides crucial insight into the underlying bio-effects. Therefore, we developed an optical imaging system at nanometer and nanosecond resolution that can resolve Cai 2+ fluctuations and microbubble oscillations. Using this system, we clearly distinguished three Cai 2+ uptake profiles upon sonoporation of endothelial cells, which strongly correlated with the microbubble oscillation amplitude, severity of sonoporation and opening of cell–cell contacts. We found a narrow operating range for viable drug delivery without lethal cell damage. Moreover, adjacent cells were affected by a calcium wave propagating at 15 μm/s. With the unique optical system, we unraveled the microbubble oscillation behavior required for drug delivery and Cai 2+ fluctuations, providing new insight into the microbubble–cell interaction to aid clinical translation

Opening of endothelial cell–cell contacts due to sonoporation

Ultrasound insonification of microbubbles can locally increase vascular permeability to enhance drug delivery. To control and optimize the therapeutic potential, we need to better understand the underlying biological mechanisms of the drug delivery pathways. The aim of this in vitro study was to elucidate the microbubble-endothelial cell interaction using the Brandaris 128 ultra-high-speed camera (up to 25 Mfps) coupled to a custom-built Nikon confocal microscope, to visualize both microbubble oscillation and the cellular response. Sonoporation and opening of cell-cell contacts by single αVβ3-targeted microbubbles (n = 152) was monitored up to 4 min after ultrasound insonification (2 MHz, 100–400 kPa, 10 cycles). Sonoporation occurred when microbubble excursion amplitudes exceeded 0.7 μm. Quantification of the influx of the fluorescent model drug propidium iodide upon sonoporation showed that the size of the created pore increased for larger microbubble excursion amplitudes. Microbubble-mediated opening of cell-cell contacts occurred as a cellular response upon sonoporation and did not correlate with the microbubble excursion amplitude itself. The initial integrity of the cell-cell contacts affected the susceptibly to drug delivery, since cell-cell contacts opened more often when cells were only partially attached to their neighbors (48%) than when fully attached (14%). The drug delivery outcomes were independent of nonlinear microbubble behavior, microbubble location, and cell size. In conclusion, by studying the microbubble–cell interaction at nanosecond and nanometer resolution the relationship between drug delivery pathways and their underlying mechanisms was further unraveled. These novel insights will aid the development of safe and efficient microbubble-mediated drug delivery