Registration and Analysis of Developmental Image Sequences

Mapping images into the same anatomical coordinate system via image registration is a fundamental step when studying physiological processes, such as brain development. Standard registration methods are applicable when biological structures are mapped to the same anatomy and their appearance remains constant across the images or changes spatially uniformly. However, image sequences of animal or human development often do not follow these assumptions, and thus standard registration methods are unsuited for their analysis. In response, this dissertation tackles the problems of i) registering developmental image sequences with spatially non-uniform appearance change and ii) reconstructing a coherent 3D volume from serially sectioned images with non-matching anatomies between the sections. There are three major contributions presented in this dissertation. First, I develop a similarity metric that incorporates a time-dependent appearance model into the registration framework. The proposed metric allows for longitudinal image registration in the presence of spatially non-uniform appearance change over time—a common medical imaging problem for longitudinal magnetic resonance images of the neonatal brain. Next, a method is introduced for registering longitudinal developmental datasets with missing time points using an appearance atlas built from a population. The proposed method is applied to a longitudinal study of young macaque monkeys with incomplete image sequences. The final contribution is a template-free registration method to reconstruct images of serially sectioned biological samples into a coherent 3D volume. The method is applied to confocal fluorescence microscopy images of serially sectioned embryonic mouse brains.Doctor of Philosoph

Human Embryology

The study of human embryology has a very long history. Modern embryology owes its initial development to the key embryo collections that began in the 19th century. The first large collection was that of Carnegie, and this was followed later by the major 7 collections. The second role of the Carnegie collection was for researchers to establish a defined set of Carnegie stages based on embryo morphological features. Today, embryos are imaged three-dimensionally (3D) by a range of imaging modalities including, magnetic resonance microscopy (MRM), episcopic fluorescence image capture (EFIC), phase-contrast X-ray computed tomography (pCT), and optical projection tomography (OPT). Historically, embryo serial images were reconstructed using wax-plate and model techniques. The above new 3D imaging techniques now allow 3D computer reconstructions, analysis, and even 3D printing. This chapter will describe how the classical embryology collections and techniques have developed into today’s imaging and analysis techniques, giving new insights to human embryonic development

Canalization of the Vestibular Plate in the Absence of Urethral Fusion Characterizes Development of the Human Clitoris: The Single Zipper Hypothesis.

PurposeWe characterized the early gestation development of the female external genitalia using optical projection tomography to visualize anatomical structures at high resolution.Materials and methodsFirst and early second trimester human female fetal external genitalia were collected with consent after voluntary termination. Specimens labeled with anti-E-Cadherin antibody underwent analysis with optical projection tomography. Histological sections were immunostained for androgen receptor, 5α-reductase, Ki67 for proliferation and Caspase 3 for apoptosis.ResultsThree-dimensional reconstructions demonstrated proximal to distal canalization of the epithelial vestibular plate and formation of a vestibular groove, which remained open. Ki67 was observed throughout with greatest density in the dorsal vestibular plate at the level of the opening groove. Staining for Caspase 3 was minimal in all sections. Androgen receptor staining was seen throughout the mesenchyme and in the apical epithelium of the dorsal vestibular groove. Throughout the epithelium and epidermis 5α-reductase staining was observed.ConclusionsEarly development of the external genitalia in the female is analogous to that in the male, demonstrating a similar opening zipper driving canalization of the vestibular plate with localized epithelial proliferation in the absence of significant apoptosis. Thus we hypothesize that the mechanism underlying the opening zipper must be androgen independent and the absence of androgen driven urethral fusion characterizes the normal development of the human clitoris

FishNet: an online database of zebrafish anatomy

Background: Over the last two decades, zebrafish have been established as a genetically versatile model system for investigating many different aspects of vertebrate developmental biology. With the credentials of zebrafish as a developmental model now well recognized, the emerging new opportunity is the wider application of zebrafish biology to aspects of human disease modelling. This rapidly increasing use of zebrafish as a model for human disease has necessarily generated interest in the anatomy of later developmental phases such as the larval, juvenile, and adult stages, during which many of the key aspects of organ morphogenesis and maturation take place. Anatomical resources and references that encompass these stages are non-existent in zebrafish and there is therefore an urgent need to understand how different organ systems and anatomical structures develop throughout the life of the fish. Results: To overcome this deficit we have utilized the technique of optical projection tomography to produce three-dimensional (3D) models of larval fish. In order to view and display these models we have created FishNet http://www.fishnet.org.au, an interactive reference of zebrafish anatomy spanning the range of zebrafish development from 24 h until adulthood. Conclusion: FishNet contains more than 36 000 images of larval zebrafish, with more than 1 500 of these being annotated. The 3D models can be manipulated on screen or virtually sectioned. This resource represents the first complete embryo to adult atlas for any species in 3D

3-d reconstruction and morphological analysis of normal rectum

There is no documentation of the 3-D dimensions of normal rectum. We present a method for the architectural and volumetric analysis of the rectum which makes use of a computerized 3-D reconstruction and CT scans of cross-sections of the rectum. The technique is simple, fast, yet potentially reliable. An attempt has been made to calculate the volume, area and average diameter of non-distended rectums (normal). Once we can standardize the technique, then we can study dimensions of the rectum in adult males and females. This thesis constitutes a unified presentation of the essential aspects of the method used in this study. We plan to study the dimensions of distended rectums (normal) measured in single column Barium Enema X-rays in two views. Once the technique is standardized, it can be applied to study the dimensions of diseased rectums

Three-Dimensional Myoarchitecture of the Lower Esophageal Sphincter and Esophageal Hiatus Using Optical Sectioning Microscopy.

Studies to date have failed to reveal the anatomical counterpart of the lower esophageal sphincter (LES). We assessed the LES and esophageal hiatus morphology using a block containing the human LES and crural diaphragm, serially sectioned at 50 μm intervals and imaged at 8.2 μm/pixel resolution. A 3D reconstruction of the tissue block was reconstructed in which each of the 652 cross sectional images were also segmented to identify the boundaries of longitudinal (LM) and circular muscle (CM) layers. The CM fascicles on the ventral surface of LES are arranged in a helical/spiral fashion. On the other hand, the CM fascicles from the two sides cross midline on dorsal surface and continue as sling/oblique muscle on the stomach. Some of the LM fascicles of the esophagus leave the esophagus to enter into the crural diaphragm and the remainder terminate into the sling fibers of the stomach. The muscle fascicles of the right crus of diaphragm which form the esophageal hiatus are arranged like a "noose" around the esophagus. We propose that circumferential squeeze of the LES and crural diaphragm is generated by a unique myo-architectural design, each of which forms a "noose" around the esophagus

Energy-Filtering Transmission Electron Microscopy of Biological Specimens

By energy-filtering transmission electron microscopy (EFTEM) electrons can be separated by their energy losses. An electron-energy filter, added to the microscope column allows the measurement of the energy distribution of transmitted electrons that have lost energy (\u3c 2,000 eV, with an energy resolution of ~ 1 eV). These filtered electrons, recorded either as a spectrum or as an image, are composed of two parts superimposed on top of each other: (a) the unspecific energy-loss population (= the continuum) and (b) the specific element-related energy-loss population (= the edges). At the edges, electron data in spectra and images are mathematically processed, to obtain the desired element-related net-intensity values or images. These data are related to the total transmitted electron intensity, from the zero-and low-loss spectral region giving the relative spectral-or image intensity ratios (SR*x, IR*x), which can be related to the element concentration. The acquisition of the zero-loss and low-loss data is hampered by the restricted dynamic range of the TV camera. By improvements through the introduction of calibrated attenuation filters in the optical path to the TV-camera, more reliable values for SR*x and IR*x can be acquired. By addition of Bio-standards adjacent to the tissue, a known and unknown concentration of the element present in the same ultrathin section and the bias in the concentration estimation, can be obtained. Some practical examples are given for the estimation of the iron concentration in siderosomes, boron in melanosomes and calcium in calcium oxalate monohydrate crystals

Image registration of ex-vivo MRI to sparsely sectioned histology of hippocampal and neocortical temporal lobe specimens.

Intractable or drug-resistant epilepsy occurs in up to 30% of epilepsy patients, with many of these patients undergoing surgical excision of the affected brain region to achieve seizure control. Recent magnetic resonance imaging (MRI) sequences and analysis techniques have the potential to detect abnormalities not identified with diagnostic MRI protocols. Prospective studies involving pre-operative imaging and collection of surgically-resected tissue provide a unique opportunity for verification and tuning of these image analysis techniques, since direct comparison can be made against histopathology, and can lead to better prediction of surgical outcomes and potentially less invasive procedures. To carry out MRI and histology comparison, spatial correspondence between the MR images and the histology images must be found. Towards this goal, a novel pipeline is presented here for bringing ex-vivo MRI of surgically-resected temporal lobe specimens and digital histology into spatial correspondence. The sparsely-sectioned histology images represent a challenge for 3D reconstruction which we address with a combined 3D and 2D registration algorithm that alternates between slice-based and volume-based registration with the ex-vivo MRI. We evaluated our registration method on specimens resected from patients undergoing anterior temporal lobectomy (N=7) and found our method to have a mean target registration error of 0.76±0.66 and 0.98±0.60 mm for hippocampal and neocortical specimens respectively. This work allows for the spatially-local comparison of histology with post-operative MRI and paves the way for eventual correlation with pre-operative MRI image analysis techniques