Practical Applications of in Vivo and ex Vivo MRI in Toxicologic Pathology Using a Novel High-performance Compact MRI System

Magnetic resonance imaging (MRI) is widely used in preclinical research and drug development and is a powerful noninvasive method for assessment of phenotypes and therapeutic efficacy in murine models of disease. In vivo MRI provides an opportunity for longitudinal evaluation of tissue changes and phenotypic expression in experimental animal models. Ex vivo MRI of fixed samples permits a thorough examination of multiple digital slices while leaving the specimen intact for subsequent conventional hematoxylin and eosin (H&E) histology. With the advent of new compact MRI systems that are designed to operate in most conventional labs without the cost, complexity, and infrastructure needs of conventional MRI systems, the possibility of MRI becoming a practical modality is now viable. The purpose of this study was to investigate the capabilities of a new compact, high-performance MRI platform (M22; Aspect Imaging, Israel) as it relates to preclinical toxicology studies. This overview will provide examples of major organ system pathologies with an emphasis on how compact MRI can serve as an important adjunct to conventional pathology by nondestructively providing 3-dimensional (3-D) digital data sets, detailed morphological insights, and quantitative information. Comparative data using compact MRI for both in vivo and ex vivo are provided as well as validation using conventional H&E.

Keywords

MRI, ex vivo, in vivo, magnetic resonance histology (MRH), toxicology, pathology, preclinical imaging.

Introduction

Magnetic resonance imaging (MRI) is a powerful method for noninvasive in vivo assessment in preclinical research, as well as drug development and for phenotyping small animal models of disease. In preclinical studies, in vivo MRI can be used in longitudinal studies to monitor progression, regression, and therapeutic responses of diseases noninvasively. As such, there is no longer a need to sacrifice animals at interim time points and the same animal can be imaged and reimaged longitudinally thereby becoming its own control. In 1993,a novel approach of using MRI on fixed tissue specimens and perfusion-fixed laboratory animals to generate 3-dimensional (3-D) digital images of the samples was introduced and the term magnetic resonance histology (MRH) was coined (Johnson et al. 1993). This was followed by several neurotoxicology publications (Lester et al. 1999, 2000; Maronpot,Sills, and Johnson 2004; Morgan, Horsfield, and Steward 2004; Sills et al. 2004) that clearly demonstrated the value of MRH as a complementary adjunct to conventional histopathology. However, up until this time, widespread adoption of MRH has been hampered by the high purchase costs of conventional superconducting MRI systems, in addition to the significant siting and installation, operation, and ongoing maintenance costs of these MRI systems. In addition, there are significant safety concerns and complexities associated with operating superconducting MRI systems, which require dedicated technical staff with specific MR-based physics expertise to operate the instruments and their complicated sequences and imaging protocols.

A new compact, high-performance MRI platform (M22; Aspect Imaging, Israel) has been developed using a novel magnet design and set of associated software and application-based approaches that reduce the cost and complexity of conventional systems. This new MRI platform provides the opportunity for pathologists with no prior MR or imaging expertise to obtain diagnostic-quality in vivo MRI and ex vivo MRH images of experimental rodents, thereby greatly enhancing conventional histopathology in preclinical toxicology studies and in the development of rodent models of human disease (Geninatti-Crich et al. 2011; Schmid et al. 2013). Unlike superconducting MRI systems, this system is portable and self-shielded so it can be placed in most laboratories or research facilities and does not require a specially shielded room, cryogens or coolants, or dedicated electrical or plumbing supply. In addition, this system has dedicated software and hardware with preprogrammed protocols and sample handling systems to easily allow pathologists to perform high-throughput imaging of animals in vivo or of fixed samples ex vivo. The advantages of this new system are the ability for longitudinally monitoring disease (in vivo MRI) and rapid acquisition of multiple electronic slices of fixed tissue (ex vivo MRH), thereby providing 3-D digital morphologically detailed data of an entire target organ while leaving the specimen intact for follow-on conventional histopathology. This leads to a more comprehensive assessment of toxicological effects and disease progression in contrast to the limited number of 2-dimensional (2-D) tissue slices afforded by conventional histopathology.

The purpose of this article is to describe the utility of this new compact MRI platform in preclinical toxicologic pathology by providing examples of practical toxicology and experimental biology applications.

Methods

In Vivo Compact MRI

In vivo MRI was performed using the M2, a compact, high-performance MRI system, equipped with a set of application-specific radiofrequency (RF) coils: 60-mm rat whole body coil, 35-mm mouse whole body coil, in addition to 30-mm and 20-mm rat and mouse head coils, respectively (Figure 1). For in vivo imaging, animals were maintained in an anesthetized state with 2% isoflorane in O2 and placed on a specially designed heated bed where physiological signals were monitored throughout the experiment to ensure the animals’ wellbeing. The optimal RF coil was selected for scanning according to the animal model and body part of interest. All experiments were performed in accordance with the guidelines and approval of the Animal Care and Use Committees of the various organizations providing the animal models.

Ex Vivo MR-based Histology

High-resolution ex vivo MRH of fixed samples was performed on the same M2 compact MRI system (Figure 1) equipped with a 20-mm RF coil and a custom-designed automated feeding and scanning mechanism for high-throughput pathological imaging. In this configuration, the compact MRI system allows for high-throughput, automated scanning of up to 10 fixed samples without technician intervention. Briefly, fixed samples are placed into specially designed disposable capsules that are given a unique bar code. The operator assigns predefined MR protocols from a library of organ- and pathology-specific protocols. The appropriate protocol is associated with the bar-coded sample, according to the organ or pathology of interest. Up to 10 capsules, each with their own application-specific imaging protocol can then be loaded into the system, which automatically advances the sample into the MRI unit. The specified imaging protocol is executed, the capsule is ejected after imaging, and the next sample is then automatically advanced and its unique imaging protocol is executed. The process continues automatically until all 10 samples are imaged. Once loaded into capsules and given a bar code, no technician intervention or supervision is required throughout the imaging process. As such, samples can be left unattended to be scanned overnight, for example, so that the system can be used during the day for in vivo imaging or to run additional capsules during the day. Effectively, throughput can be maximized so that the system can be used 24 hr per day.

Samples were fixed using conventional immersion fixation techniques, transferred to phosphate-buffered saline or saline for 24 hr, and then into an MR transparent solution (Fluorinert FC, 3M, USA) to avoid tissue dehydration.