Signal Improvement and Contrast Enhancement in Magnetic Resonance Imaging

This thesis reports advances in magnetic resonance imaging (MRI), with the ultimate goal of improving signal and contrast in biomedical applications. More specifically, novel MRI pulse sequences have been designed to characterize microstructure, enhance signal and contrast in tissue, and image functional processes. In this thesis, rat brain and red bone marrow images are acquired using iMQCs (intermolecular multiple quantum coherences) between spins that are 10 μm to 500 μm apart. As an important application, iMQCs images in different directions can be used for anisotropy mapping. We investigate tissue microstructure by analyzing anisotropy mapping. At the same time, we simulated images expected from rat brain without microstructure. We compare those with experimental results to prove that the dipolar field from the overall shape only has small contributions to the experimental iMQC signal. Besides magnitude of iMQCs, phase of iMQCs should be studied as well. The phase anisotropy maps built by our method can clearly show susceptibility information in kidneys. It may provide meaningful diagnostic information. To deeply study susceptibility, the modified-crazed sequence is developed. Combining phase data of modified-crazed images and phase data of iMQCs images is very promising to construct microstructure maps. Obviously, the phase image in all above techniques needs to be highly-contrasted and clear. To achieve the goal, algorithm tools from Susceptibility-Weighted Imaging (SWI) and Susceptibility Tensor Imaging (STI) stands out superb useful and creative in our system.

Application of a rank-based genetic association test to age-at-onset data from the Collaborative Study on the Genetics of Alcoholism study.

Association studies of quantitative traits have often relied on methods in which a normal distribution of the trait is assumed. However, quantitative phenotypes from complex human diseases are often censored, highly skewed, or contaminated with outlying values. We recently developed a rank-based association method that takes into account censoring and makes no distributional assumptions about the trait. In this study, we applied our new method to age-at-onset data on ALDX1 and ALDX2. Both traits are highly skewed (skewness > 1.9) and often censored. We performed a whole genome association study of age at onset of the ALDX1 trait using Illumina single-nucleotide polymorphisms. Only slightly more than 5% of markers were significant. However, we identified two regions on chromosomes 14 and 15, which each have at least four significant markers clustering together. These two regions may harbor genes that regulate age at onset of ALDX1 and ALDX2. Future fine mapping of these two regions with densely spaced markers is warranted.

A Plasmonic Gold Nanostar Theranostic Probe for In Vivo Tumor Imaging and Photothermal Therapy.

Nanomedicine has attracted increasing attention in recent years, because it offers great promise to provide personalized diagnostics and therapy with improved treatment efficacy and specificity. In this study, we developed a gold nanostar (GNS) probe for multi-modality theranostics including surface-enhanced Raman scattering (SERS) detection, x-ray computed tomography (CT), two-photon luminescence (TPL) imaging, and photothermal therapy (PTT). We performed radiolabeling, as well as CT and optical imaging, to investigate the GNS probe's biodistribution and intratumoral uptake at both macroscopic and microscopic scales. We also characterized the performance of the GNS nanoprobe for in vitro photothermal heating and in vivo photothermal ablation of primary sarcomas in mice. The results showed that 30-nm GNS have higher tumor uptake, as well as deeper penetration into tumor interstitial space compared to 60-nm GNS. In addition, we found that a higher injection dose of GNS can increase the percentage of tumor uptake. We also demonstrated the GNS probe's superior photothermal conversion efficiency with a highly concentrated heating effect due to a tip-enhanced plasmonic effect. In vivo photothermal therapy with a near-infrared (NIR) laser under the maximum permissible exposure (MPE) led to ablation of aggressive tumors containing GNS, but had no effect in the absence of GNS. This multifunctional GNS probe has the potential to be used for in vivo biosensing, preoperative CT imaging, intraoperative detection with optical methods (SERS and TPL), as well as image-guided photothermal therapy.

Quantitative genetics of CTCF binding reveal local sequence effects and different modes of X-chromosome association.

Associating genetic variation with quantitative measures of gene regulation offers a way to bridge the gap between genotype and complex phenotypes. In order to identify quantitative trait loci (QTLs) that influence the binding of a transcription factor in humans, we measured binding of the multifunctional transcription and chromatin factor CTCF in 51 HapMap cell lines. We identified thousands of QTLs in which genotype differences were associated with differences in CTCF binding strength, hundreds of them confirmed by directly observable allele-specific binding bias. The majority of QTLs were either within 1 kb of the CTCF binding motif, or in linkage disequilibrium with a variant within 1 kb of the motif. On the X chromosome we observed three classes of binding sites: a minority class bound only to the active copy of the X chromosome, the majority class bound to both the active and inactive X, and a small set of female-specific CTCF sites associated with two non-coding RNA genes. In sum, our data reveal extensive genetic effects on CTCF binding, both direct and indirect, and identify a diversity of patterns of CTCF binding on the X chromosome.

High-Resolution CT for Small-Animal Imaging Research

Preclinical imaging has a critical role in phenotyping, in drug discovery, and in providing a basic understanding of mechanisms of disease. Translating imaging methods from humans to small animals is not an easy task. The purpose of this work is to review high-resolution computed tomography (CT) also known as micro-CT for small-animal imaging. We present the principles, the technologies, the image quality parameters, and the types of applications. We show that micro-CT can be used to provide not only morphological but also functional information such as cardiac function or vascular permeability. Another way in which micro-CT can be used in the study of both function and anatomy is by combining it with other imaging modalities, such as positron emission tomography or single-photon emission tomography. Compared to other modalities, micro-CT imaging is usually regarded as being able to provide higher throughput at lower cost and higher resolution. The limitations are usually associated with the relatively poor contrast mechanisms and the radiation damage, although the use of novel nanoparticle-based contrast agents and careful design of studies can address these limitations.

Live Imaging of Host-Parasite Interactions in a Zebrafish Infection Model Reveals Cryptococcal Determinants of Virulence and Central Nervous System Invasion.

UNLABELLED: The human fungal pathogen Cryptococcus neoformans is capable of infecting a broad range of hosts, from invertebrates like amoebas and nematodes to standard vertebrate models such as mice and rabbits. Here we have taken advantage of a zebrafish model to investigate host-pathogen interactions of Cryptococcus with the zebrafish innate immune system, which shares a highly conserved framework with that of mammals. Through live-imaging observations and genetic knockdown, we establish that macrophages are the primary immune cells responsible for responding to and containing acute cryptococcal infections. By interrogating survival and cryptococcal burden following infection with a panel of Cryptococcus mutants, we find that virulence factors initially identified as important in causing disease in mice are also necessary for pathogenesis in zebrafish larvae. Live imaging of the cranial blood vessels of infected larvae reveals that C. neoformans is able to penetrate the zebrafish brain following intravenous infection. By studying a C. neoformans FNX1 gene mutant, we find that blood-brain barrier invasion is dependent on a known cryptococcal invasion-promoting pathway previously identified in a murine model of central nervous system invasion. The zebrafish-C. neoformans platform provides a visually and genetically accessible vertebrate model system for cryptococcal pathogenesis with many of the advantages of small invertebrates. This model is well suited for higher-throughput screening of mutants, mechanistic dissection of cryptococcal pathogenesis in live animals, and use in the evaluation of therapeutic agents. IMPORTANCE: Cryptococcus neoformans is an important opportunistic pathogen that is estimated to be responsible for more than 600,000 deaths worldwide annually. Existing mammalian models of cryptococcal pathogenesis are costly, and the analysis of important pathogenic processes such as meningitis is laborious and remains a challenge to visualize. Conversely, although invertebrate models of cryptococcal infection allow high-throughput assays, they fail to replicate the anatomical complexity found in vertebrates and, specifically, cryptococcal stages of disease. Here we have utilized larval zebrafish as a platform that overcomes many of these limitations. We demonstrate that the pathogenesis of C. neoformans infection in zebrafish involves factors identical to those in mammalian and invertebrate infections. We then utilize the live-imaging capacity of zebrafish larvae to follow the progression of cryptococcal infection in real time and establish a relevant model of the critical central nervous system infection phase of disease in a nonmammalian model.

Investigation and Development of a Fully 3D Tilt Capable Hybrid SPECT - CT System for Dedicated Breast Imaging

X-ray mammography has been the gold standard for breast imaging for decades, despite the significant limitations posed by the two dimensional (2D) image acquisitions. Difficulty in diagnosing lesions close to the chest wall and axilla, high amount of structural overlap and patient discomfort due to compression are only some of these limitations. To overcome these drawbacks, three dimensional (3D) breast imaging modalities have been developed including dual modality single photon emission computed tomography (SPECT) and computed tomography (CT) systems. This thesis focuses on the development and integration of the next generation of such a device for dedicated breast imaging. The goals of this dissertation work are to: [1] understand and characterize any effects of fully 3-D trajectories on reconstructed image scatter correction, absorbed dose and Hounsifeld Unit accuracy, and [2] design, develop and implement the fully flexible, third generation hybrid SPECT-CT system capable of traversing complex 3D orbits about a pendant breast volume, without interference from the other. Such a system would overcome artifacts resulting from incompletely sampled divergent cone beam imaging schemes and allow imaging closer to the chest wall, which other systems currently under research and development elsewhere cannot achieve.

The dependence of x-ray scatter radiation on object shape, size, material composition and the CT acquisition trajectory, was investigated with a well-established beam stop array (BSA) scatter correction method. While the 2D scatter to primary ratio (SPR) was the main metric used to characterize total system scatter, a new metric called ‘normalized scatter contribution’ was developed to compare the results of scatter correction on 3D reconstructed volumes. Scatter estimation studies were undertaken with a sinusoidal saddle (±15° polar tilt) orbit and a traditional circular (AZOR) orbit. Clinical studies to acquire data for scatter correction were used to evaluate the 2D SPR on a small set of patients scanned with the AZOR orbit. Clinical SPR results showed clear dependence of scatter on breast composition and glandular tissue distribution, otherwise consistent with the overall phantom-based size and density measurements. Additionally, SPR dependence was also observed on the acquisition trajectory where 2D scatter increased with an increase in the polar tilt angle of the system.

The dose delivered by any imaging system is of primary importance from the patient’s point of view, and therefore trajectory related differences in the dose distribution in a target volume were evaluated. Monte Carlo simulations as well as physical measurements using radiochromic film were undertaken using saddle and AZOR orbits. Results illustrated that both orbits deliver comparable dose to the target volume, and only slightly differ in distribution within the volume. Simulations and measurements showed similar results, and all measured dose values were within the standard screening mammography-specific, 6 mGy dose limit, which is used as a benchmark for dose comparisons.

Hounsfield Units (HU) are used clinically in differentiating tissue types in a reconstructed CT image, and therefore the HU accuracy of a system is very important, especially when using non-traditional trajectories. Uniform phantoms filled with various uniform density fluids were used to investigate differences in HU accuracy between saddle and AZOR orbits. Results illustrate the considerably better performance of the saddle orbit, especially close to the chest and nipple region of what would clinically be a pedant breast volume. The AZOR orbit causes shading artifacts near the nipple, due to insufficient sampling, rendering a major portion of the scanned phantom unusable, whereas the saddle orbit performs exceptionally well and provides a tighter distribution of HU values in reconstructed volumes.

Finally, the third generation, fully-suspended SPECT-CT system was designed in and developed in our lab. A novel mechanical method using a linear motor was developed for tilting the CT system. A new x-ray source and a custom made 40 x 30 cm2 detector were integrated on to this system. The SPECT system was nested, in the center of the gantry, orthogonal to the CT source-detector pair. The SPECT system tilts on a goniometer, and the newly developed CT tilting mechanism allows ±15° maximum polar tilting of the CT system. The entire gantry is mounted on a rotation stage, allowing complex arbitrary trajectories for each system, without interference from the other, while having a common field of view. This hybrid system shows potential to be used clinically as a diagnostic tool for dedicated breast imaging.

A LabVIEW Platform for Preclinical Imaging Using Digital Subtraction Angiography and Micro-CT.

CT and digital subtraction angiography (DSA) are ubiquitous in the clinic. Their preclinical equivalents are valuable imaging methods for studying disease models and treatment. We have developed a dual source/detector X-ray imaging system that we have used for both micro-CT and DSA studies in rodents. The control of such a complex imaging system requires substantial software development for which we use the graphical language LabVIEW (National Instruments, Austin, TX, USA). This paper focuses on a LabVIEW platform that we have developed to enable anatomical and functional imaging with micro-CT and DSA. Our LabVIEW applications integrate and control all the elements of our system including a dual source/detector X-ray system, a mechanical ventilator, a physiological monitor, and a power microinjector for the vascular delivery of X-ray contrast agents. Various applications allow cardiac- and respiratory-gated acquisitions for both DSA and micro-CT studies. Our results illustrate the application of DSA for cardiopulmonary studies and vascular imaging of the liver and coronary arteries. We also show how DSA can be used for functional imaging of the kidney. Finally, the power of 4D micro-CT imaging using both prospective and retrospective gating is shown for cardiac imaging.

Correction for Eddy Current-Induced Echo-Shifting Effect in Partial-Fourier Diffusion Tensor Imaging.

In most diffusion tensor imaging (DTI) studies, images are acquired with either a partial-Fourier or a parallel partial-Fourier echo-planar imaging (EPI) sequence, in order to shorten the echo time and increase the signal-to-noise ratio (SNR). However, eddy currents induced by the diffusion-sensitizing gradients can often lead to a shift of the echo in k-space, resulting in three distinct types of artifacts in partial-Fourier DTI. Here, we present an improved DTI acquisition and reconstruction scheme, capable of generating high-quality and high-SNR DTI data without eddy current-induced artifacts. This new scheme consists of three components, respectively, addressing the three distinct types of artifacts. First, a k-space energy-anchored DTI sequence is designed to recover eddy current-induced signal loss (i.e., Type 1 artifact). Second, a multischeme partial-Fourier reconstruction is used to eliminate artificial signal elevation (i.e., Type 2 artifact) associated with the conventional partial-Fourier reconstruction. Third, a signal intensity correction is applied to remove artificial signal modulations due to eddy current-induced erroneous T2(∗) -weighting (i.e., Type 3 artifact). These systematic improvements will greatly increase the consistency and accuracy of DTI measurements, expanding the utility of DTI in translational applications where quantitative robustness is much needed.

Radiolabeled inhibitors as probes for imaging mutant IDH1 expression in gliomas: Synthesis and preliminary evaluation of labeled butyl-phenyl sulfonamide analogs.

INTRODUCTION: Malignant gliomas frequently harbor mutations in the isocitrate dehydrogenase 1 (IDH1) gene. Studies suggest that IDH mutation contributes to tumor pathogenesis through mechanisms that are mediated by the neomorphic metabolite of the mutant IDH1 enzyme, 2-hydroxyglutarate (2-HG). The aim of this work was to synthesize and evaluate radiolabeled compounds that bind to the mutant IDH1 enzyme with the goal of enabling noninvasive imaging of mutant IDH1 expression in gliomas by positron emission tomography (PET). METHODS: A small library of nonradioactive analogs were designed and synthesized based on the chemical structure of reported butyl-phenyl sulfonamide inhibitors of mutant IDH1. Enzyme inhibition assays were conducted using purified mutant IDH1 enzyme, IDH1-R132H, to determine the IC50 and the maximal inhibitory efficiency of the synthesized compounds. Selected compounds, 1 and 4, were labeled with radioiodine ((125)I) and/or (18)F using bromo- and phenol precursors, respectively. In vivo behavior of the labeled inhibitors was studied by conducting tissue distribution studies with [(125)I]1 in normal mice. Cell uptake studies were conducted using an isogenic astrocytoma cell line that carried a native IDH1-R132H mutation to evaluate the potential uptake of the labeled inhibitors in IDH1-mutated tumor cells. RESULTS: Enzyme inhibition assays showed good inhibitory potency for compounds that have iodine or a fluoroethoxy substituent at the ortho position of the phenyl ring in compounds 1 and 4 with IC50 values of 1.7 μM and 2.3 μM, respectively. Compounds 1 and 4 inhibited mutant IDH1 activity and decreased the production of 2-HG in an IDH1-mutated astrocytoma cell line. Radiolabeling of 1 and 4 was achieved with an average radiochemical yield of 56.6 ± 20.1% for [(125)I]1 (n = 4) and 67.5 ± 6.6% for [(18)F]4 (n = 3). [(125)I]1 exhibited favorable biodistribution characteristics in normal mice, with rapid clearance from the blood and elimination via the hepatobiliary system by 4 h after injection. The uptake of [(125)I]1 in tumor cells positive for IDH1-R132H was significantly higher compared to isogenic WT-IDH1 controls, with a maximal uptake ratio of 1.67 at 3 h post injection. Co-incubation of the labeled inhibitors with the corresponding nonradioactive analogs, and decreasing the normal concentrations of FBS (10%) in the incubation media substantially increased the uptake of the labeled inhibitors in both the IDH1-mutant and WT-IDH1 tumor cell lines, suggesting significant non-specific binding of the synthesized labeled butyl-phenyl sulfonamide inhibitors. CONCLUSIONS: These data demonstrate the feasibility of developing radiolabeled probes for the mutant IDH1 enzyme based on enzyme inhibitors. Further optimization of the labeled inhibitors by modifying the chemical structure to decrease the lipophilicity and to increase potency may yield compounds with improved characteristics as probes for imaging mutant IDH1 expression in tumors.