Boyd M. Knosp

A Comparative Study of Eya1 and Eya4 Protein Function and Its Implication in Branchio-oto-renal Syndrome and DFNA10

Allele variants of EYA1 and EYA4, two members of the vertebrate Eya gene family, underlie two types of inherited human deafness, branchio-oto-renal (BOR) syndrome and DFNA10, respectively. To clarify how mutations in these two genes and their encoded proteins impact the normal biology of hearing, we completed a number of functional studies using the yeast-two-hybrid system. We verified that bait constructs of the homologous region (Eya1HR and Eya4HR) interact with Six1 prey constructs, although no interaction with Dach1 prey was demonstrable. To compare interaction affinities, we evaluated α-galactosidase activity after cotransformation of Eya1HR/Six1 and Eya4HR/Six1 and found that the latter interaction was weaker. By immunofluorescence staining, we showed Eya4HR localization to the cytoplasm. After coexpression of Six1, Eya4HR was translocated to the nucleus. Results with Eya1HR were similar. Translation of mutant constructs (Eya4HRR564X and Eya1HRR539X) could not be demonstrated. Using dual Eya-containing constructs (with two wild-type alleles or wild-type and mutant alleles), we confirmed no translation of the mutant allele, even if the mutation was nontruncating. These results are consistent with clinical data and implicate haploinsufficiency as the cause of BOR syndrome and DFNA10.

Three-dimensional relationships between tumor cells and microcirculation with double cyanine immunolabeling, laser scanning confocal microscopy, and computer-assisted reconstruction: an alternative to cast corrosion preparations.

The morphology of the microcirculation of uveal melanomas is a reliable market of tumor progression. Scanning electron microscopy of cast corrosion preparations can generate three-dimensional views of these vascular patterns, but this technique sacrifices the tumor parenchyma. Formalin-fixed wet tissue sections 100-150 microns thick from uveal melanomas were stained with the lectin Ulex europaeus agglutinin I (UEAI) and proliferating cell nuclear antigen (PCNA) to demonstrate simultaneously the tumor blood vessels and proliferating tumor cells. Indocarbocyanine (Cy3) was used as a fluorophore for UEAI and indodicarbocyanine (Cy5) was used for PCNA. Double labeled sections were examined with a laser scanning confocal microscope. Images of both stains were digitized at the same 5-microns intervals and each of the two images per interval was combined digitally to form one image. These combined images were visualized through voxel processing to study the relationship between melanoma cells expressing PCNA and various microcirculatory patterns. This technique produces images comparable to scanning electron microscopy of cast corrosion preparations while permitting simultaneous localization of melanoma cells expressing PCNA. The microcirculatory tree can be viewed from any perspective and the relationship between tumor cells and the tumor blood vessels can be studied concurrently in three dimensions. This technique is an alternative to cast corrosion preparations.

Ultrasound characterization of acute myocardial ischemia by quantitative texture analysis

In this study we tested the efficacy of quantitative texture analysis in the identification of acute myocardial ischemia using an ultrasound data acquisition system that digitizes and stores echocardiographic data in polar format. In nine closed-chest dogs, data were acquired before and after coronary occlusion using a 2.4 MHz echocardiographic system. Regions of interest were analyzed at end-diastole and end-systole from the ischemic area and normal area at the same depth of field. Ultrasound data were evaluated using previously reported quantitative gray level texture measures. After occlusion, texture changes indicative of ischemia were found in systolic images. The directional component of the data analysis was important; analysis in the azimuthal direction was more accurate than in the axial direction. Six texture measures exhibited significant changes in the ischemic region from control to occlusion when analyzing data in the azimuthal direction. One false positive result occurred (significant texture change in the normal region from control to occlusion) in the azimuthal direction. Several false positive alterations in the normal regions from control to occlusion were found when the texture was evaluated in the axial direction. For accurate assessment of ischemic changes, preocclusion image data were required. We conclude that quantitative echocardiographic texture analysis using polar format data can identify subtle changes in myocardial texture such as that due to acute ischemia, using data acquired through the chest wall.

Stereotactic Atlas of the Macaca mulatta Thalamus and Adjacent Basal Ganglia Nuclei

I. Introduction. II. Material and Methods. III. Stereotactic Technique with Visualization of the Intracerebral Landmarks. IV. Nuclear Outlines and Nomenclature. V. References. VI. Abbreviations. VII. Cytoarchitectonic Atlas Plates and Nuclear Maps. VIII. Instructions for Using the CD. IX. Addendum (Compact Disk). Index.

Ultrasound Characterization of Acoustic Properties of Acute Intracardiac Thrombi: Studies in a New Experimental Model

Echocardiographic differentiation among intracavitary thrombus, cavity noise, and adjacent myocardium can be difficult. As an initial step toward quantitative thrombus characterization with ultrasound, 11 dogs were studied with an in vivo intracardiac thrombus model to delineate the acoustic properties of acute thrombi. The apical coronary arteries were ligated, and subsequently injections of 5% sodium rescinoleate and 1000 units of thrombin at the endocardium-blood interface created left ventricular mural thrombi. Echocardiographic images were obtained in long- and short-axis views with a digital acquisition system, and a statistical analysis of echo intensities was performed in regions of interest in the thrombus, surrounding ventricular cavity and adjacent myocardium. Statistical measurements used to evaluate echo intensities in each region of interest included mean gray level, standard deviation, skewness, and kurtosis. The results showed that thrombus could be distinguished from myocardium (by mean gray level and standard deviation) only in short-axis views, where regions of interest could be placed at similar depths of field. Mean gray level, standard deviation, and skewness all distinguished thrombus from intracavitary blood regardless of the region of interest placement. The phase of the cardiac cycle at which data were acquired did not alter the results. We conclude that acute intracardiac thrombi can be distinguished from surrounding blood and myocardium with ultrasound tissue characterization techniques that may have the potential for clinical application.

Dose anisotropy around an Au-198 seed source

For interstitial implants with radioactive Au-198 seeds, the dosimetric calculations usually ignore the finite source size and employ a point source approximation, resulting in a computed isotropic dose distribution. However, the measured radiation fluence from Au-198 seeds is reported to be anisotropic, suggesting some amount of dose anisotropy in tissue. We have measured this dose anisotropy around Au-198 seeds (2.5 X 0.8 mm2) using Kodak X-OMAT XV-2 films placed in contact with individual seeds in a phantom. Autoradiographs obtained for various exposure times were digitized and studied with an image analysis computer network. The networks overall spatial resolution was about 0.01 cm. The optical isodensity contours around the seed were obtained. The useful range of optical density (OD) for the system was found to be 1 to 2 OD units. Within this range, the shape of an optical isodensity contour would be identical to an isodose contour. Proximal to the source, the contours were elliptical in shape, elongated along the seed axis. However, further away, the elongation was in the direction normal to the seed axis. This was in agreement with the reported data on the radiation fluence around Au-198 seeds. It was concluded that measurable dose anisotropy existed around Au-198 seeds. However, it was too small to be of any clinical significance.

A Multi-agent System Architecture for End-User Level Grid Monitoring Using Geographic Information Systems (MAGGIS): Architecture and Implementation

This paper illustrates a Multi-Agent system architecture for end-user level Grid monitoring using Geographical Information Systems (MAGGIS). The purpose of this research is to investigate MAGGIS architecture and implementation issues, and to verify the following two hypotheses: 1.) multi-agent systems provide an effective and scalable architecture to synthesize various Grid information providers for monitoring Grid resources; and 2.) geographic information systems (GIS) provide an ideal solution to organizing and managing the geographic aspect of Grid resource information as well as to providing an effective user interface for monitoring Grid status. The MAGGIS framework is implemented in a Grid portal environment based on the open Grid service architecture. It is observed that the MAGGIS not only helps end-users monitor the status of Grid resources, but also provides quick and comprehensive information for resource scheduling and management on behalf of user applications.

Distributed computation for diffusion problem in a P2P-enhanced computing system

Basic exploration of diffusion equation solvers in distributed computing systems has been a very important issue for computational fluid dynamics (CFD). This paper presents a fundamental study of a distributed computing solution for diffusive phenomena in a P2P enhanced Grid system. We propose a simple distributed system with the architecture of multi-clients and multi-servers (MCMS). Multithreading is implemented on both the client and server nodes, enabling them to communicate. This paper focuses on architecture, model implementation, multithreading, network communication, domain decomposition/composition, problem convergence, fault-tolerance, and dynamic load balancing. A discussion of the results from conducted numerical experiments is useful for performing future intensive CFD computations on Grids.

Nuclear Outlines and Nomenclature

The identification of thalamic nuclei was based on the cytoarchitecture and the available data on the topography of their subcortical connections. The outlines of the nuclei outside the ventral tier, for example the midline nuclei, were based solely on the cytoarchitecture as described by Olszewski (1952) and also by Ilinsky and KultasIlinsky (1987). The ventral tier nuclei include the regions that receive motor-related and somatosensory afferents. The topographic relationships of the subcortical afferents in the ventral tier are more obvious in the sagittal section plane as the major afferent-receiving zones are arranged back to front from the anterior to the posterior pole of the thalamus with a limited co-presence in the coronal plane (Asanuma et al., 1983a,b; Ilinsky and Kultas-Ilinsky, 1987; Ilinsky et al., 1993). At the same time the adjacent afferent zones interdigitize profusely. This interdigitation is difficult to detect in the coronal plane but is quite obvious in the sagittal plane (Ilinsky et al., 1993). In the process of the atlas preparation the initial outlines of the ventral tier nuclei were very detailed and closely followed these extremely wavy boundaries. However, this amount of the detail became an impediment in 3D computer reconstructions and, therefore, the original outlines were edited to smooth out the contours. Nonetheless, the degree of the interdigitation retained in the atlas is still significant and is expressed by intrusions of the colors of the adjacent nuclei into one other. This is especially impressive in the coronal section plane.

Stereotactic Technique with Visualization of the Intracerebral Landmarks

Details of the stereotactic technique on experimental animals (cats and monkeys) with the use of ventriculographic landmarks and the intracerebral coordinate system have been described in detail earlier (Ilinsky et al., 1978; 1980; Ilinsky and KultasIlinsky, 1982). Below is a brief summary of the method that consists of three stages.