Fluence-to-dose conversion coefficients from monoenergetic neutrons below 20 MeV based on the VIP-man anatomical model

A new set of fluence-to-absorbed dose and fluence-to-effective dose conversion coefficients have been calculated for neutrons below 20 MeV using a whole-body anatomical model, VIP-Man, developed from the high-resolution transverse colour photographic images of the National Library of Medicine's Visible Human Project. Organ dose calculations were performed using the Monte Carlo code MCNP for 20 monoenergetic neutron beams between 1 x 10(-9) MeV and 20 MeV under six different irradiation geometries: anterior-posterior, posterior-anterior, right lateral, left lateral, rotational and isotropic. The absorbed dose for 24 major organs and effective dose results based on the realistic VIP-Man are presented and compared with those based on the simplified MIRD-based phantoms reported in the literature. Effective doses from VIP-Man are not significantly different from earlier results for neutrons in the energy range studied. There are, however, remarkable deviations in organ doses due to the anatomical differences between the image-based and the earlier mathematical models.     

Brightness-compensated 3-D optical flow algorithm for monitoring cochlear motion patterns

A method for three-dimensional motion analysis designed for live cell imaging by fluorescence confocal microscopy is described. The approach is based on optical flow computation and takes into account brightness variations in the image scene that are not due to motion, such as photobleaching or fluorescence variations that may reflect changes in cellular physiology. The 3-D optical flow algorithm allowed almost perfect motion estimation on noise-free artificial sequences, and performed with a relative error of <10% on noisy images typical of real experiments. The method was applied to a series of 3-D confocal image stacks from an in vitro preparation of the guinea pig cochlea. The complex motions caused by slow pressure changes in the cochlear compartments were quantified. At the surface of the hearing organ, the largest motion component was the transverse one (normal to the surface), but significant radial and longitudinal displacements were also present. The outer hair cell displayed larger radial motion at their basolateral membrane than at their apical surface. These movements reflect mechanical interactions between different cellular structures, which may be important for communicating sound-evoked vibrations to the sensory cells. A better understanding of these interactions is important for testing realistic models of cochlear mechanics.     

Dose-volume delivery guided proton therapy using beam on-line PET system

Proton therapy is one form of radiotherapy in which the irradiation can be concentrated on a tumor using a scanned or modulated Bragg peak. Therefore, it is very important to evaluate the proton-irradiated volume accurately. The proton-irradiated volume can be confirmed by detection of pair annihilation gamma rays from positron emitter nuclei generated by the target nuclear fragment reaction of irradiated proton nuclei and nuclei in the irradiation target using a positron emission tomography (PET) apparatus, and dose-volume delivery guided proton therapy (DGPT) can thereby be achieved using PET images. In the proton treatment room, a beam ON-LINE PET system (BOLPs) was constructed so that a PET apparatus of the planar-type with a high spatial resolution of about 2 mm was mounted with the field of view covering the isocenter of the beam irradiation system. The position and intensity of activity were measured using the BOLPs immediately after the proton irradiation of a gelatinous water target containing 16O nuclei at different proton irradiation energy levels. The change of the activity-distribution range against the change of the physical range was observed within 2 mm. The experiments of proton irradiation to a rabbit and the imaging of the activity were performed. In addition, the proton beam energy used to irradiate the rabbit was changed. When the beam condition was changed, the difference between the two images acquired from the measurement of the BOLPs was confirmed to clearly identify the proton-irradiated volume.     

SAF values for internal photon emitters calculated for the RPI-P pregnant-female models using Monte Carlo methods

Estimates of radiation absorbed doses from radionuclides internally deposited in a pregnant woman and her fetus are very important due to elevated fetal radiosensitivity. This paper reports a set of specific absorbed fractions (SAFs) for use with the dosimetry schema developed by the Society of Nuclear Medicine's Medical Internal Radiation Dose (MIRD) Committee. The calculations were based on three newly constructed pregnant female anatomic models, called RPI-P3, RPI-P6, and RPI-P9, that represent adult females at 3-, 6-, and 9-month gestational periods, respectively. Advanced Boundary REPresentation (BREP) surface-geometry modeling methods were used to create anatomically realistic geometries and organ volumes that were carefully adjusted to agree with the latest ICRP reference values. A Monte Carlo user code, EGS4-VLSI, was used to simulate internal photon emitters ranging from 10 keV to 4 MeV. SAF values were calculated and compared with previous data derived from stylized models of simplified geometries and with a model of a 7.5-month pregnant female developed previously from partial-body CT images. The results show considerable differences between these models for low energy photons, but generally good agreement at higher energies. These differences are caused mainly by different organ shapes and positions. Other factors, such as the organ mass, the source-to-target-organ centroid distance, and the Monte Carlo code used in each study, played lesser roles in the observed differences in these. Since the SAF values reported in this study are based on models that are anatomically more realistic than previous models, these data are recommended for future applications as standard reference values in internal dosimetry involving pregnant females.     

Microdosimetry of astatine-211 single-cell irradiation: role of daughter polonium-211 diffusion

A microdosimetric analysis of previously published data on 211At-albumin, free 211At, and 211At-C215 irradiation of Colo-205 cells in a slowly rotating single-cell suspension is presented. A custom-built computer program based on the Monte Carlo method was used to simulate the irradiation and the energy deposition in individual cell nuclei. Separate simulations were made for the assumption that the 211Po atom stays in the position where it is created, and that it diffuses away. The mean event number at which 37% of all cells survived, n37, and the frequency mean specific energy per event, zF, were estimated. The Poisson distribution of events and simulated single and multievent distributions of specific energy were used to find the single-cell specific energy at which the probability of survival is reduced to 37%, z37. The calculated single-cell radiosensitivity values show that 211Po atoms, created on a cell surface by the decay of 211At atoms, will diffuse from the cell during its life-span. The increasing distance to the cell nucleus will drastically decrease the probability of the emitted alpha particle to hit the nucleus. This will result in fewer alpha-particle events in the cell nucleus. For dispersed cells, the diffusion of 211Po atoms will reduce the total dose from cell-bound 211At by a factor of 2.     

Multi-dimensional laser confocal microscopy on live cells in submicroliter volumes using glass capillaries

Imaging live cells using laser confocal microscopy requires the use of complex and rather cumbersome incubation chamber systems in order to maintain the correct physiological conditions. The volume of these chambers is in the range of a few hundred microliters. Here we present an easy and convenient alternative in the form of glass capillaries that accommodate volumes of 0.2-10 microliters. The capillaries can be loaded with both suspension and adherent cells. The loaded capillaries are taped on microscope slides and submerged into the immersion oil that covers the objective. The correct temperature is maintained using a thermostat-controlled objective heater. We demonstrate that using microlens enhanced -rotating Nipkow disc based confocal illumination, in combination with cold CCD cameras, maximum resolution multicolor time lapse fluorescence images can be obtained from live cells. The images obtained are free from disturbing optical distortions. Imaging in submicroliter volumes allows for fluorescence visualization of very rare cell types isolated using flow or affinity sorting or obtained by fine needle biopsies.     

A study of the electric field distribution in erythrocyte and rod shape cells from direct RF exposure

This paper shows the importance of using realistic cell shapes with the proper geometry and orientation to study the mechanisms of direct cellular effects from radiofrequency (RF) exposure. For this purpose, the electric field distribution within erythrocyte, rod and ellipsoidal cell models is calculated by using a finite element technique with adaptive meshing. The three cell models are exposed to linearly polarized electromagnetic plane waves of frequencies 900 and 2450 MHz. The results show that the amplification of the electric field within the membrane of the erythrocyte shape cell is more significant than that observed in other cell geometries. The results obtained show the dependence of the induced electric field distribution on frequency, electrical properties of membrane and cytoplasm and the orientation of the cell with respect to the applied field. The analysis of the transition of an erythrocyte shape to an ellipsoidal one shows that a uniformly shelled ellipsoid model is a rough approximation if a precise simulation of bioeffects in cells is desired.     

Analytic microdosimetry for radioimmunotherapeutic alpha emitters.

Analytic microdosimetry using Fourier transform techniques has been applied to internal alpha emitters. These techniques need revision and simplification for use with short-lived radionuclides such as those which may be useful for radioimmunotherapy. Analytic methods may have advantages over Monte Carlo methods in some cases (e.g., where time is important). Applications to eight different source geometries show the usefulness of these techniques. Comparisons of some of the results of Monte Carlo calculations prove its accuracy. For a uniform source of 5.867-MeV alphas spread throughout the volume outside a cell surface, the two methods agree well. Results are within 1% both for the average specific energy and for the number of hits. Analytic microdosimetry provides an alternate method to use for the critical evaluation of models that seek to predict the relation between alpha energy deposition and cell survival data. Similarly, it may be helpful to point the way toward the rational interpretation of general biological results for antibodies labeled with alpha emitters.     

Three-dimensional reconstruction of confocal laser microscopy images to study the behaviour of osteoblastic cells grown on biomaterials.

The adhesion, spreading and cytoskeletal organization of osteoblastic cells seeded onto titanium and titania/hydroxyapatite composite coating (TiO2/HA) were studied using images acquired by confocal laser scanning microscopy. The fluorescence staining technique was employed to visualize actin cytoskeletal organization of cells, 2-D images were exhaustive when the cells were seeded at low density (in the first 24 h of incubation), but they were less clear when the cells proliferated and appeared stacked. Since the shareware software were not satisfactory, a new 3-D image reconstruction was developed using ordinary software and a model was obtained directly from the optical section set, in order to achieve a more realistic and faithful vision of morphological structures and to evaluate the behaviour of bone cells grown on materials. The results showed that the cells grown on titanium conform to the irregular substrate surfaces maximizing the contact between the cell membrane and the substrate and proliferate disposing close to each other. On the contrary, the osteoblasts seeded onto TiO2/HA coating develop clusters where the cells aggregated extending processes in order to establish intercellular connections. Cell aggregation is an early and critical event leading to cell differentiation and mineralization process and could be a first signal of the tendency of TiO2/HA coating to stimulate cell differentiation.     

A Study of volumetric variations of basal nuclei in the normal human brain by magnetic resonance imaging

Knowledge of the effects of healthy aging on brain structures is necessary to identify abnormal changes due to diseases. Many studies have demonstrated age‐related volume changes in the brain using MRI. 60 healthy individuals who had normal MRI aged from 20 years to 80 years were examined and classified into three groups: Group I: 21 persons; nine males and 12 females aging between 20‐39 years old. Group II: 22 persons; 11 males and 11 females aging between 40‐59 years old. Group III: 17 persons; eight males and nine females aging between 60‐80 years old. Volumetric analysis was done to evaluate the effect of age, gender and hemispheric difference in the caudate and putamen by the slicer 4.3.3.1 software using 3D T1‐weighted images. Data were analyzed by student's unpaired t test, ANOVA and regression analysis. The volumes of the measured and corrected caudate nuclei and putamen significantly decreased with aging in males. There was a statistically insignificant relation between the age and the volume of the measured caudate nuclei and putamen in females but there was a statistically significant relation between the age and the corrected caudate nuclei and putamen. There was no significant difference on the caudate and putamen volumes between males and females. There was no significant difference between the right and left caudate nuclei volumes. There was a leftward asymmetry in the putamen volumes. The results can be considered as a base to track individual changes with time (aging and CNS diseases). Clin. Anat. 30:175–182, 2017. © 2017 Wiley Periodicals, Inc.     

Real-time reflectance confocal microscopy: comparison of two-dimensional images and three-dimensional image stacks for detection of cervical precancer

Confocal microscopy can provide real-time, 2-D and 3-D images of the cellular morphology and tissue architecture features that pathologists use to detect precancerous lesions without the need for tissue removal, sectioning, and staining. The utility of 3-D confocal image stacks of epithelial tissue for detecting dysplasia has not yet been explored. We aim to extract morphometry and tissue architecture information from 2-D confocal reflectance images and 3-D image stacks from fresh, unstained cervical biopsies and compare their potential for detecting dysplasia. Nine biopsies are obtained from eight patients; confocal images are acquired pre- and postacetic acid at multiple epithelial depths in 1.5 mum-intervals. Postacetic acid images are processed to segment cell nuclei; after segmentation, 2-D images taken at 50 mum below the tissue surface, and the entire 3-D image stacks are processed to extract morphological and architectural features. Data are analyzed to determine which features gave the best separation between normal and high-grade cervical precancer. Most significant differences are obtained from parameters extracted from the 3-D image stacks. However, in all cases where the 2-D features were multiplicatively scaled by the depth of acquisition divided by the epithelial thickness or scaled by the scattering coefficient, the significance level is equal to or greater than the comparable feature extracted from the 3-D image stacks. A linear discriminant function previously developed to separate 19 samples of normal tissue and high-grade cervical precancer based on the nuclear-to-cytoplasm (N/C) ratio and epithelial scattering coefficient is prospectively applied to the nine biopsies examined to determine the accuracy with which it could separate normal tissue from cervical intra epithelial neoplasia (CIN) 23. For the entire data set of 28 biopsies, a sensitivity and specificity of 100% is produced using this discriminant function; the scattering coefficient provides more discriminative capacity than the N/C ratio. The success of the scaled 2-D image features has important implications for using confocal microscopy to detect precancer in the clinic. Acquisition of the epithelial thickness or scattering coefficient requires less time than 3-D image sets and little additional effort is required to gain the added information compared to 2-D images alone.     

K-edge digital subtraction imaging with dichromatic x-ray sources: SNR and dose studies

The aim of the present work is to analytically evaluate the signal to noise ratio (SNR) and the delivered dose in K-edge digital subtraction imaging (KES) using two types of x-ray sources: a monochromatic x-ray source (available at synchrotron radiation facilities and considered as gold standard) and a quasi-monochromatic compact source. The energy separation DeltaE between the two monochromatic beams is 1 keV and 4 keV for the two sources, respectively. The evaluation has been performed for both radiography and computed tomography. Different geometries have been studied to mimic clinical situations. In mammography, a pathology perfused by a contrast agent has been modelled; in angiography, a vessel superimposed to a ventricle or a stand-alone artery stenosis has been studied. The SNR and the skin dose have been calculated as a function of the detail diameter, the contrast agent (iodine and gadolinium), and its concentration in the tissues. Results show that for DeltaE = 4 keV a slightly higher delivered dose is required to obtain the same SNR with respect to DeltaE < 1 keV. A similar study has been performed for KES-CT. Computer simulations of CT images performed with Snark software are shown to validate the analytical calculations.     

A Computational Approach to Understand Phenotypic Structure and Constitutive Mechanics Relationships of Single Cells

The goal of this study is to construct a representative 3D finite element model (FEM) of individual cells based on their sub-cellular structures that predicts cell mechanical behavior. The FEM simulations replicate atomic force microscopy (AFM) nanoindentation experiments on live vascular smooth muscle cells. Individual cells are characterized mechanically with AFM and then imaged in 3D using a spinning disc confocal microscope. Using these images, geometries for the FEM are automatically generated via image segmentation and linear programming algorithms. The geometries consist of independent structures representing the nucleus, actin stress fiber network, and cytoplasm. These are imported into commercial software for mesh refinement and analysis. The FEM presented here is capable of predicting AFM results well for 500 nm indentations. The FEM results are relatively insensitive to both the exact number and diameter of fibers used. Despite the localized nature of AFM nanoindentation, the model predicts that stresses are distributed in an anisotropic manner throughout the cell body via the actin stress fibers. This pattern of stress distribution is likely a result of the geometric arrangement of the actin network.     

Optimal ultraviolet wavelength for in vivo photoacoustic imaging of cell nuclei

In order to image noninvasively cell nuclei in vivo without staining, we have developed ultraviolet photoacoustic microscopy (UV-PAM), in which ultraviolet light excites nucleic acids in cell nuclei to produce photoacoustic waves. Equipped with a tunable laser system, the UV-PAM was applied to in vivo imaging of cell nuclei in small animals. We found that 250 nm was the optimal wavelength for in vivo photoacoustic imaging of cell nuclei. The optimal wavelength enables UV-PAM to image cell nuclei using as little as 2 nJ laser pulse energy. Besides the optimal wavelength, application of a wavelength between 245 and 275 nm can produce in vivo images of cell nuclei with specific, positive, and high optical contrast.     

Cell detection in phase-contrast images used for alpha-particle track-etch dosimetry: a semi-automated approach

A novel alpha-particle irradiator has recently been developed that provides the ability to characterize cell response. The irradiator is comprised of a collimated, planar alpha-particle source which, from below, irradiates cells cultured on a track-etch material. Cells are imaged using phase-contrast microscopy before and following irradiation to obtain geometric information and survival rates; these can be used with data from alpha-particle track images to assess cell response. A key step in this process is determining cell location within the pre-irradiation images. Although this can be done completely by a human observer, the number of images requiring analysis makes the process time-consuming and tedious. To reduce the potential human error and decrease user interaction time, a semi-automated, computer-aided method of cell detection has been developed. The method employs a two-level adaptive thresholding technique to obtain size and position information about potential cell cytoplasms and nuclei. Proximity and geometry-based thresholds are then used to mark structures as cells. False-positive detections from the automated algorithm are due mostly to imperfections in the track-etch background, camera effects and cellular residue. To correct for these, a human observer reviews all detected structures, discarding false positives. When analysing two randomly selected cell dish image databases, the semi-automated method detected 92-94% of all cells and 94-97% of cells with a well-defined cytoplasm and nucleus while reducing human workload by 32-83%.     

Distinct expression patterns of ER alpha and ER beta in normal human mammary gland

AIM: Two oestrogen receptors (ERs) have been identified to date-the "classic" ER alpha and the more recently described ER beta. Although much is known about ER alpha at the mRNA and protein levels, our knowledge of the expression and distribution of ER beta protein is much more limited. The aim of this study was to compare the cellular distribution of ER alpha and ER beta in normal human mammary gland. METHODS: Formalin fixed, paraffin wax embedded material was obtained from reduction mammoplasty specimens, normal tissue adjacent to breast tumour, or fibroadenoma. Sections were immunohistochemically stained for ER alpha, ER beta, and the progesterone receptor. The staining pattern for each antibody was evaluated and compared. RESULTS: ER alpha was restricted to the cell nuclei of epithelial cells lining ducts and lobules. Although ER beta was also seen in these cells, additional strong staining was detected specifically in the cell nuclei of myoepithelial cells. Occasional staining was seen in surrounding stromal and endothelial cell nuclei and in lymphocytes. CONCLUSIONS: ER subtypes have distinct distribution patterns in the normal mammary gland. The widespread distribution of ER beta suggests that it may be the dominant ER in the mammary gland where it may be acting as a natural suppressor.     

Morphogenetic domains in the yolk syncytial layer of axiating zebrafish embryos.

The yolk syncytial layer (YSL) of the teleostean yolk cell is known to play important roles in the induction of cellular mesendoderm, as well as the patterning of dorsal tissues. To determine how this extraembryonic endodermal compartment is subdivided and morphologically transformed during early development, we have examined collective movements of vitally stained YSL nuclei in axiating zebrafish embryos by using four-dimensional confocal microscopy. During blastulation, gastrulation, and early segmentation, zebrafish YSL nuclei display several highly patterned movements, which are organized into spatially distinct morphogenetic domains along the anterior-posterior and dorsal-ventral axes. During the late blastula period, with the onset of epiboly, nuclei throughout the YSL initiate longitudinal movements that are directed along the animal-vegetal axis. As epiboly progresses, nuclei progressively recede from the advancing margin of the epibolic YSL. However, a small group of nuclei is retained at the YSL margin to form a constricting blastoporal ring. During mid-gastrulation, YSL nuclei undergo convergent-extension behavior toward the dorsal midline, with a subset of nuclei forming an axial domain that underlies the notochord. These highly patterned movements of YSL nuclei share remarkable similarities to the morphogenetic movements of deep cells in the overlying zebrafish blastoderm. The macroscopic shape changes of the zebrafish yolk cell, as well as the morphogenetic movements of its YSL nuclei, are homologous to several morphogenetic behaviors that are regionally expressed within the vegetal endodermal cell mass of gastrulating Xenopus embryos. In contrast to the cellular endoderm of Xenopus, the dynamics of zebrafish YSL show that a syncytial endodermal germ layer can express a temporal sequence of morphogenetic domains without undergoing progressive steps of cell fate restriction.     

The relationship between the number of nuclei of an osteoclast and its resorptive capability in vitro

Summary This study examined the relationship between the number of nuclei in an osteoclast and its resorptive efficiency, as demonstrated by the size of the pit it can make in a mineralized tissue in 24 h in vitro. Osteoclasts released mechanically from prehatch chick long bones were cultured on dentine slices or on plastic dishes for periods of 6 or 24 h. The frequency distribution of the multinucleate tartrate-resistant acid phosphatase (TRAP)-positive cells with different numbers of nuclei was determined: the mean number of nuclei per cell was 6.92, with a mode of 4. 47% had 5 or fewer nuclei and only 11% more than 10 nuclei. The pits associated with 292 osteoclasts with known numbers of nuclei were measured using a confocal laser light microscope (Lasertec) and dedicated image analysis system, and depths, plan areas and volumes determined. There was a positive correlation between the number of nuclei per osteoclast and the volume of the pit made, but a trend for the volume resorbed per nucleus to decrease with increase in the number of nuclei per osteoclast.     

Mechanistic study of apoptosis induced by high-fluence low-power laser irradiation using fluorescence imaging techniques

Low-power laser irradiation (LPLI) can cause cell proliferation, differentiation, or death; however, the cellular mechanisms of these effects of LPLI, at high or low fluences, are not well known. To investigate the mechanism of high-fluence LPLI-induced apoptosis, both human lung adenocarcinoma cells (ASTC-a-1) and African green monkey SV40-transformed kidney fibroblast cells (COS-7) were irradiated with a He-Ne laser for 10 min under a fluence of 120 J/cm(2) and 80 J/cm(2), respectively. The dynamics of reactive oxygen species (ROS) generation was determined by measuring changes in fluorescence resulting from oxidation of intracellular dichlorodihydrofluorescein diacetate (H(2)DCFDA) to (DCF). The changes of mitochondrial membrane potential, DeltaPsim, were studied by measuring the reduction of cellular fluorescence of Rhodamine 123 dyes using confocal laser scanning microscopy. The activation of caspase-3 in cells transfected by [SCAT3] reporters was observed using fluorescence resonance energy transfer (FRET) imaging. The activity of caspase-8 during high-fluence LPLI-induced apoptosis was studied by monitoring the cellular distribution of [Bid-CFP] reporters using fluorescence imaging. The following temporal sequence of cellular events was observed during apoptosis induced by high-fluence LPLI (120 J/cm(2), ASTC-a-1 cells): (1) immediate generation of mitochondrial ROS following laser irradiation, reaching a maximum level 60 min after irradiation; (2) onset of DeltaPsim decrease 15 min after laser irradiation, reaching a minimum level 50 min after irradiation; and (3) activation of caspase-3 between 30 min and 180 min after laser irradiation. Our results also show that the high-fluence LPLI does not activate caspase-8, indicating that the induced apoptosis was initiated directly from mitochondrial ROS generation and DeltaPsim decrease, independent of the caspase-8 activation.