Three‐dimensional multiphoton/optical coherence tomography for diagnostic applications in dermatology

A preliminary clinical trial using state‐of‐the‐art multiphoton tomography (MPT) and optical coherence tomography (OCT) for three‐dimensional (3D) multimodal in vivo imaging of normal skin, nevi, scars and pathologic skin lesions has been conducted. MPT enabled visualization of sub‐cellular details with axial and transverse resolutions of <2 μm and <0.5 μm, respectively, from a volume of 0.35 × 0.35 × 0.2 mm³ at a frame rate of 0.14 Hz (512 × 512 pixels). State‐of‐the‐art OCT, operating at a center wavelength of 1300 nm, was capable of acquiring 3D images depicting the layered architecture of skin with axial and transverse resolutions ～8 μm and ～20 μm, respectively, from a volume of 7 × 3.5 × 1.5 mm³ at a frame rate of 46 Hz (1024 × 1024 pixels). This study demonstrates the clinical diagnostic potential of MPT/OCT for pre‐screening relatively large areas of skin using 3D OCT to identify suspicious regions at microscopic level and subsequently using high resolution MPT to obtain zoomed in, sub‐cellular level information of the respective regions (© 2013 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Application of 3-Dimensional Printing Technology to Construct an Eye Model for Fundus Viewing Study

Objective

To construct a life-sized eye model using the three-dimensional (3D) printing technology for fundus viewing study of the viewing system.

Methods

We devised our schematic model eye based on Navarro''s eye and redesigned some parameters because of the change of the corneal material and the implantation of intraocular lenses (IOLs). Optical performance of our schematic model eye was compared with Navarro''s schematic eye and other two reported physical model eyes using the ZEMAX optical design software. With computer aided design (CAD) software, we designed the 3D digital model of the main structure of the physical model eye, which was used for three-dimensional (3D) printing. Together with the main printed structure, polymethyl methacrylate(PMMA) aspherical cornea, variable iris, and IOLs were assembled to a physical eye model. Angle scale bars were glued from posterior to periphery of the retina. Then we fabricated other three physical models with different states of ammetropia. Optical parameters of these physical eye models were measured to verify the 3D printing accuracy.

Results

In on-axis calculations, our schematic model eye possessed similar size of spot diagram compared with Navarro''s and Bakaraju''s model eye, much smaller than Arianpour''s model eye. Moreover, the spherical aberration of our schematic eye was much less than other three model eyes. While in off- axis simulation, it possessed a bit higher coma and similar astigmatism, field curvature and distortion. The MTF curves showed that all the model eyes diminished in resolution with increasing field of view, and the diminished tendency of resolution of our physical eye model was similar to the Navarro''s eye. The measured parameters of our eye models with different status of ametropia were in line with the theoretical value.

Conclusions

The schematic eye model we designed can well simulate the optical performance of the human eye, and the fabricated physical one can be used as a tool in fundus range viewing research.     

High resolution MR eye protocol optimization: Comparison between 3D-CISS, 3D-PSIF and 3D-VIBE sequences

PurposeThe purpose of this study was to compare selected MRI pulse sequences and to evaluate their utility for depicting specific anatomic regions in the eye.MethodsA High-Resolution (HR) 0.08 × 0.08 × 0.60 mm³ MRI protocol was developed on a 1.5-T clinical system and applied in the left eye of an albino rabbit, utilizing a small field of view surface coil. The comprehensive MRI protocol consisted of two 3D (T2/T1)w sequences (3D-PSIF and 3D-CISS), and one 3D T1w sequence (3D-VIBE). The T1w 3D-VIBE sequence was acquired, before and after intravenous injection of 0.2 mmol/kgr gadolinium-DTPA. Signal-to-Noise Ratios (SNR) and Contrast-to-Noise Ratios (CNR) amongst specific eye anatomical areas were calculated for each sequence. The presence of artifacts was rated subjectively utilizing a 5 point scale.Results3D-PSIF and 3D-CISS provide better delineation and visualization of the eye as compared with 3D-VIBE sequences. 3D-CISS images present the highest SNR and revealed better discrimination of the ocular surrounding tissues; its basic drawback though is related to ghost artifacts appearing in the anterior chamber and resulting in the lowest image quality. In post-contrast imaging, the T1w 3D-VIBE sequence provided the best overall image quality. Moreover, 3D (T2/T1)w sequences can provide good anatomic depiction of the eye segments. Agreement between the two independent readers was good.ConclusionsOptimization of a comprehensive MR eye imaging protocol is achieved. A higher SNR, a better spatial resolution and a reduction of the total scan time were obtained, thus making clinical MRI systems more reliable in eye imaging.     

Comparative Study of Experimental Ocular Melanoma Using Two Implantation Techniques of B16-F10 Melanocytes

Objective: The aim of this work was to evaluate the behaviour of B16-F10 melanoma cell cultures implanted in the anterior chamber of the eye of New Zealand white rabbits by studying the clinical-pathological and ultrastructural characteristics of the lesions. Methods: One group (A) (consisting of 30 rabbits) was transclerally inoculated (1 mm from sclero-corneal limbus) with 4×10⁶ melanocytes and another group (B) (also 30 animals) was inoculated once per week for 3 consecutive weeks with 5×10⁶ cells (total 15×10⁶); 30 animals acted as the control group (C). All the lesions were processed for optic and electronic microscopy. Results: Tumoral growth in group A was 43% (13/30) and in group B 80% (24/30). All lesions were pigmented and none perforated the eyeball. Microscopically, they were a mixture of epithelioid and fusiform cells disposed around the blood vessels. Ultrastructurally, the presence of melanosomes in different stages of maturation and aberrant melanosomes were characteristic. Conclusion: We suggest that the transcleral inoculation of 15×10⁶ B16-F10 melanocytes into the anterior chamber of the eye of New Zealand white rabbits may be a valid and reproducible method for obtaining an experimental ocular melanoma model.     

Estimating narwhal (Monodon monoceros) age using tooth layers and aspartic acid racemization of eye lens nuclei

Counting growth-layer groups (GLGs) in teeth is one of the most precise and widely accepted methods for aging marine mammals. Male narwhals have a large erupted tusk that can be used for aging, but this tusk is often difficult or expensive to obtain from hunters and most females do not display the tusk; thus, alternative methods for narwhal aging are needed. In this study, we aged narwhals by counting annual GLGs in embedded tusks and by measuring the change in the ratio of D- and L-enantiomers of aspartic acid in the eye lens nucleus that occurs as the animal ages (the aspartic acid racemization [AAR] technique). Absolute age estimates were estimated for seven tusks aged ≤15 yr. Estimated age was a significant predictor of aspartic acid D/L ratios with a racemization rate (K_asp) of 9.72 × 10⁻⁴/year ± 2.28 × 10⁻⁴ and a (D/L)₀ of 3.46 × 10⁻² ± 1.78 × 10⁻³ (r² = 0.74). Results from our study, which included more younger GLG-aged animals than previously evaluated, confirms AAR can be used to generate age estimates for narwhals.     

High Resolution 3D Imaging of Ex-Vivo Biological Samples by Micro CT

Non-destructive volume visualization can be achieved only by tomographic techniques, of which the most efficient is the x-ray micro computerized tomography (μCT).High resolution μCT is a very versatile yet accurate (1-2 microns of resolution) technique for 3D examination of ex-vivo biological samples^{1, 2}. As opposed to electron tomography, the μCT allows the examination of up to 4 cm thick samples. This technique requires only few hours of measurement as compared to weeks in histology. In addition, μCT does not rely on 2D stereologic models, thus it may complement and in some cases can even replace histological methods^{3, 4}, which are both time consuming and destructive. Sample conditioning and positioning in μCT is straightforward and does not require high vacuum or low temperatures, which may adversely affect the structure. The sample is positioned and rotated 180° or 360°between a microfocused x-ray source and a detector, which includes a scintillator and an accurate CCD camera, For each angle a 2D image is taken, and then the entire volume is reconstructed using one of the different available algorithms^5-7. The 3D resolution increases with the decrease of the rotation step. The present video protocol shows the main steps in preparation, immobilization and positioning of the sample followed by imaging at high resolution.     

In vivo determination of young's modulus for the human cornea

The present paper describes a calculation technique to determine a value of Young's modulus for the cornea when the intraocular pressure, the shape of the cornea and the thickness variation along the cornea are known fromin vivo measurements. Twenty-eight persons were examined in both eyes and a mean value of Young's modulus for the cornea was calculated to 2.45×10⁴±0.57×10⁴ N/m² (S.D.).     

Exploring TOF capabilities of PET detector blocks based on large monolithic crystals and analog SiPMs

Monolithic scintillators are more frequently used in PET instrumentation due to their advantages in terms of accurate position estimation of the impinging gamma rays both planar and depth of interaction, their increased efficiency, and expected timing capabilities. Such timing performance has been studied when those blocks are coupled to digital photosensors showing an excellent timing resolution.In this work we study the timing behaviour of detectors composed by monolithic crystals and analog SiPMs read out by an ASIC. The scintillation light spreads across the crystal towards the photosensors, resulting in a high number of SiPMs and ASIC channels fired. This has been studied in relation with the Coincidence Timing Resolution (CTR). We have used LYSO monolithic blocks with dimensions of 50 × 50 × 15 mm³ coupled to SiPM arrays (8 × 8 elements with 6 × 6 mm² area) which compose detectors suitable for clinical applications.While a CTR as good as 186 ps FWHM was achieved for a pair of 3 × 3 × 5 mm³ LYSO crystals, when using the monolithic block and the SiPM arrays, a raw CTR over 1 ns was observed. An optimal timestamp assignment was studied as well as compensation methods for the time-skew and time-walk errors. This work describes all steps followed to improve the CTR. Eventually, an average detector time resolution of 497 ps FWHM was measured for the whole thick monolithic block. This improves to 380 ps FWHM for a central volume of interest near the photosensors. The timing dependency with the photon depth of interaction and planar position are also included.     

In-vivo dose measurements with MOSFET dosimeters during MV portal imaging

BackgroundThe purpose of this study was to investigate the feasibility of MOSFET dosimeter in measuring eye dose during 2D MV portal imaging for setup verification in radiotherapy.Materials and methodsThe in-vivo dose measurements were performed by placing the dosimeters over the eyes of 30 brain patients during the acquisition of portal images in linear accelerator by delivering 1 MU with the field sizes of 10 × 10 cm² and 15 × 15 cm².ResultsThe mean doses received by the left and right eyes of 10 out of 30 patients when both eyes were completely inside the anterior portal field were found to be 2.56 ± 0.2 cGy and 2.75 ± 0.2, respectively. Similarly, for next 10 patients out of the same 30 patients the mean doses to left and right eyes when both eyes were completely out of the anterior portal fields were found to be 0.13 ± 0.02 cGy and 0.17 ± 0.02 cGy, respectively. The mean doses to ipsilateral and contralateral eye for the last 10 patients when one eye was inside the anterior portal field were found to be 3.28 ± 0.2 cGy and 0.36 ± 0.1 cGy, respectively.ConclusionThe promising results obtained during 2D MV portal imaging using MOSFET have shown that this dosimeter is well suitable for assessing low doses during imaging thereby enabling to optimize the imaging procedure using the dosimetric data obtained. In addition, the documentation of the dose received by the patient during imaging procedure is possible with the help of an in-built software in conjunction with the MOSFET reader module.     

Assessment of intraorbital structure volume using a numerical segmentation image technique (NSI): the fatty tissue and the eyeball

INTRODUCTION: Measurement of the degree of exophthalmos is one of the main methods used in the assessment of pathological processes that occur in the orbital space and is widely used. However, this only provides initial information about the volume relations between the intraorbital structures. The aims of our work were as follows: to draw up a new computer application, namely the numerical segmentation image (NSI) technique, for the automatic calculation of the volume of the intraorbital structures on the basis of magnetic resonance imaging (MRI) images, to determine its usefulness in the segmentation of fatty tissue and the eyeball and to estimate their volume in relation to the degree of exophthalmos. MATERIAL AND METHODS: A total of 45 patients (90 orbits) were included in the study. All the patients underwent MRI examination of the orbits by a 1.5 T scanner using a head coil. The degree of exophthalmos was determined clinically and radiologically in relation to the interzygomatic line. Quantitative assessment of the eyeball and fatty tissue was made using an NSI application. RESULTS: The influence of fatty tissue volume on the degree of exophthalmos was determined as being statistically significant (r = 0.367, p = 0.000374) but was smaller in comparison with the relationship between total eye muscle volume and degree of exophthalmos; eyeball volume was found to have the least influence (r = 0.344, p = 0.000374). Two eyeballs of significantly smaller volume were found in the group of 90 orbits analysed. CONCLUSIONS: The NSI technique is a clinically useful application, providing objective data calculated individually for each orbit. A credible protocol for estimating the degree of exophthalmos on the basis of the NSI technique should include the eye muscle volume, fatty tissue volume and, in cases where eyeball pathologies coexist, the eyeball volume as well.     

In vivo study of rat cortical hemodynamics using a stereotaxic‐apparatus‐compatible photoacoustic microscope

Brain imaging is an important technique in cognitive neuroscience. In this article, we designed a stereotaxic‐apparatus‐compatible photoacoustic microscope for the studies of rat cortical hemodynamics. Compared with existing optical resolution photoacoustic microscopy (ORPAM) systems, the probe owns feature of fast, light and miniature. In this microscope, we integrated a miniaturized ultrasound transducer with a center frequency of 10 MHz to detect photoacoustic signals and a 2‐dimensional (2D) microelectromechanical system (MEMS) scanner to achieve raster scanning of the optical focus. Based on phantom evaluation, this imaging probe has a high lateral resolution of 3.8 μm and an effective imaging domain of 2 × 2 mm². Different from conventional ORPAMs, combining with standard stereotaxic apparatus enables broad studies of rodent brains without any motion artifact. To show its capability, we successfully captured red blood cell flow in the capillary, monitored the vascular changes during bleeding and blood infusion and visualized cortical hemodynamics induced by middle cerebral artery occlusion.      

A compact high‐speed full‐field optical coherence microscope for high‐resolution in vivo skin imaging

A compact high‐speed full‐field optical coherence microscope has been developed for high‐resolution in vivo imaging of biological tissues. The interferometer, in the Linnik configuration, has a size of 11 × 11 × 5 cm³ and a weight of 210 g. Full‐field illumination with low‐coherence light is achieved with a high‐brightness broadband light‐emitting diode. High‐speed full‐field detection is achieved by using part of the image sensor of a high‐dynamic range CMOS camera. En face tomographic images are acquired at a rate of 50 Hz, with an integration time of 0.9 ms. The image spatial resolution is 0.9 μm × 1.2 μm (axial × transverse), over a field of view of 245 × 245 μm². Images of human skin, revealing in‐depth cellular‐level structures, were obtained in vivo and in real‐time without the need for stabilization of the subject. The system can image larger fields, up to 1 × 1 mm², but at a reduced depth.      

Axial eye length after retinal detachment surgery

Changes in the eye axial diameter were studied to assess the eye globe impact of conventional operation for retinal detachment. The study included 69 eyes in 69 patients operated on for rhegmatogenous retinal detachment. There were 46.4% of men and 53.6% of women, mean age 52.7 (+/- 15.21) years. Results of preoperative and postoperative ultrasonographic measurement of axial diameter are presented. The mean preoperative and postoperative eye axial diameter was 23.69 (+/- 1.84) mm and 24.43 (+/- 1.91) mm, respectively. Postoperative results showed the axial eye length to increase by a mean of 0.74 (+/- 0.44) mm, yielding a statistically significant difference from the preoperative measurement (p < 0.001). The mean myopia induced by this eyeball elongation was 1.77 D. The encircling band with and without segmental buckling used in surgical repair of retinal detachment creates circular and segmental indentation of the eyeball, thus increasing its axial length. The myopia induced by elongation of the eyeball results in considerable myopia, which requires appropriate correction in the early postoperative period to achieve favorable vision rehabilitation.     

High resolution scintimammography helps in differentiating benign from malignant findings in scintigraphic hot spots

Several efforts have been focusing on the development of detectors devoted to high solution ^99mTc sestamibi scintimammography to improve sensitivity for non palpable lesions. To this aim new high resolution scintillation gamma camera was developed under the “Integiated Mammographic Imaging” project. The gamma camera, made by CAEN and Pol.Hi.Tech, has an overall dimension of 112×120×75mm3. It consists of an array of 1 in. PSPMTs Hamamatsu H8520-C12 closely packed, a NaI(T1) scintillation array (1.8×1.8×6mm³ pixel) and a general purpose collimator. By this gamma camera a clinical experience on a few patients with breast cancer suspicion was performed. In this paper we show how high resolution approach allows to better categorize the lesions on the basis of the morphology of the spatial distribution of the radiotracer in the breast tissue. By comparing conventional and high resolution images of a young patient (29 y.o.) with breast cancer suspicion, it appears clear how the Anger, camera images showed a defined hot spot, highly suggestive of malignant lesion; on the contrary, the high resolution scan shown a large and inhomogeneous uptake area with the absence of clear and focal character of the uptake, to be considered as a probably non malignant lesions. This resuh was confirmed by byoptical findings that diagnosed the echographic findings as a benign inflammatory lesion.     

Comparative Effect of the I3.1 Probiotic Formula in Two Animal Models of Colitis

Use of probiotic therapy is an active area of investigation to treat intestinal disorders. The clinical benefits of the I3.1 probiotic formula (Lactobacillus plantarum (CECT7484, CECT7485) and P. acidilactici (CECT7483)) were demonstrated in irritable bowel syndrome (IBS) patients in a randomized, double-blind, placebo-controlled clinical trial. The aim of this study was to evaluate the therapeutic effects of I3.1 in two experimental models of colitis, a dextran sulfate sodium (DSS)-induced colitis model and an interleukin (IL)-10-deficient mice model. Colitis was induced in 32 8-week-old Balb/c mice by administering 3% (w/v) DSS in drinking water for 5 days. Probiotics were administered orally (I3.1 or VSL#3, 1 × 10⁹ CFU daily) for 10 days before the administration of DSS. Also, probiotics (I3.1 or VSL#3, 1 × 10⁹ CFU daily) were administered orally to 36 6-week-old C57B6J IL-10(?/?) mice for 10 weeks. Body weight was recorded daily. Colon samples were harvested for histological examination and cytokine measurements. Body weight after DSS administration did not change in the I3.1 group, whereas the VSL#3 group had weight loss. Also, I3.1 normalized IL-6 to levels similar to that of healthy controls and significantly increased the reparative histologic score. In the IL-10-deficient model, both VSL#3 and I3.1 reduced the severity of colitis compared to untreated controls, and I3.1 significantly reduced the levels of IFN-γ compared to the other two groups. In conclusion, I3.1 displays a protective effect on two murine models of experimental colitis. Results suggest that the mechanism of action could be different from VSL#3.     

Acoustic resolution photoacoustic microscopy based on microelectromechanical systems scanner

Photoacoustic microscopy (PAM) can be classified as optical resolution (OR)‐PAM and acoustic resolution (AR)‐PAM depending on the type of resolution achieved. Using microelectromechanical systems (MEMS) scanner, high‐speed OR‐PAM system was developed earlier. Depth of imaging limits the use of OR‐PAM technology for many preclinical and clinical imaging applications. Here, we demonstrate the use of a high‐speed MEMS scanner for AR‐PAM imaging. Lateral resolution of 84 μm and an axial resolution of 27 μm with ~2.7 mm imaging depth was achieved using a 50 MHz transducer‐based AR‐PAM system. Use of a higher frequency transducer at 75 MHz has further improved the resolution characteristics of the system with a reduction in imaging depth and a lateral resolution of 53 μm and an axial resolution of 18 μm with ~1.8 mm imaging depth was achieved. Using the two‐axis MEMS scanner a 2 × 2 .5 mm² area was imaged in 3 seconds. The capability of achieving acoustic resolution images using the MEMS scanner makes it beneficial for the development of high‐speed miniaturized systems for deeper tissue imaging.      

A novel automatic 3D stitching algorithm for optical coherence tomography angiography and its application in dermatology

Optical coherence tomography (OCT) and OCT angiography (OCTA) techniques offer numerous advantages in clinical skin applications but the field of view (FOV) of current commercial systems are relatively limited to cover the entire skin lesion. The typical method to expand the FOV is to apply wide field objective lens. However, lateral resolution is often sacrificed when scanning with these lenses. To overcome this drawback, we developed an automated 3D stitching method for creating high-resolution skin structure and vascular volumes with large field of view, which was realized by montaging multiple adjacent OCT and OCTA volumes. The proposed stitching method is demonstrated by montaging 3 × 3 OCT and OCTA volumes (nine OCT/OCTA volumes as one data set with each volume covers 2.5 cm × 2.5 cm area) of healthy thin and thick skin from six volunteers. The proposed stitching protocol achieves high flexibility and repeatable for all the participants. Moreover, according to evaluation of structural similarity index and feature similarity index, our proposed stitched result has a superior similarity to single scanning protocol in large-scaled. We had also verified its improved performance through assessing metrics of vessel contrast-noise-ratio (CNR) from 2.07 ± 0.44 (single large-scaled scanning protocol) to 3.05 ± 0.51 (proposed 3 × 3 sub-volume stitching method).     

Modelling the visual world of a velvet worm

In many animal phyla, eyes are small and provide only low-resolution vision for general orientation in the environment. Because these primitive eyes rarely have a defined image plane, traditional visual-optics principles cannot be applied. To assess the functional capacity of such eyes we have developed modelling principles based on ray tracing in 3D reconstructions of eye morphology, where refraction on the way to the photoreceptors and absorption in the photopigment are calculated incrementally for ray bundles from all angles within the visual field. From the ray tracing, we calculate the complete angular acceptance function of each photoreceptor in the eye, revealing the visual acuity for all parts of the visual field. We then use this information to generate visual filters that can be applied to high resolution images or videos to convert them to accurate representations of the spatial information seen by the animal. The method is here applied to the 0.1 mm eyes of the velvet worm Euperipatoides rowelli (Onychophora). These eyes of these terrestrial invertebrates consist of a curved cornea covering an irregular but optically homogeneous lens directly joining a retina packed with photoreceptive rhabdoms. 3D reconstruction from histological sections revealed an asymmetric eye, where the retina is deeper in the forward-pointing direction. The calculated visual acuity also reveals performance differences across the visual field, with a maximum acuity of about 0.11 cycles/deg in the forward direction despite laterally pointing eyes. The results agree with previous behavioural measurements of visual acuity, and suggest that velvet worm vision is adequate for orientation and positioning within the habitat.     

The scaling of eye size with body mass in birds

We developed a simple method that uses skulls to estimate the diameter, and hence the mass, of birds'' eyes. Allometric analysis demonstrated that, within five orders (parrots, pigeons, petrels, raptors and owls) and across 104 families of flying birds, eye mass is proportional to (body mass)^0.68 over a range of body masses (6 g to 11.3 kg). As expected from their habits and visual ecology, raptors and owls have enlarged eyes, with masses 1.4 and 2.2 times greater than average birds of the same weight. Taking existing relationships for flight speed on body mass, we find that resolution increases close to (flight speed)^1.333. Consequently, large birds resolve objects at a longer time to contact than small birds. Eye radius and skull size co-vary in strict proportion, suggesting common physiological, aerodynamic and mechanical constraints. Because eye mass scales close to brain mass, metabolic rate and information processing could also be limiting, but the precise factors determining the scaling of eye to body have not been identified.