Preclinical magnetic resonance imaging and spectroscopy in the fields of radiological technology, medical physics, and radiology.

Magnetic resonance imaging (MRI) is an indispensable diagnostic imaging technique used in the clinical setting. MRI is advantageous over X-ray and computed tomography (CT), because the contrast provided depends on differences in the density of various organ tissues. In addition to MRI systems in hospitals, more than 100 systems are used for research purposes in Japan in various fields, including basic scientific research, molecular and clinical investigations, and life science research, such as drug discovery, veterinary medicine, and food testing. For many years, additional preclinical imaging studies have been conducted in basic research in the fields of radiation technology, medical physics, and radiology. The preclinical MRI research includes studies using small-bore and whole-body MRI systems. In this review, we focus on the animal study using small-bore MRI systems as "preclinical MRI". The preclinical MRI can be used to elucidate the pathophysiology of diseases and for translational research. This review will provide an overview of previous preclinical MRI studies such as brain, heart, and liver disease assessments. Also, we provide an overview of the utility of preclinical MRI studies in radiological physics and technology.

In vivo MR imaging for tumor-associated initial neovascularization by supramolecular contrast agents.

Microvascular imaging is required to understand tumor angiogenesis development; however, an appropriate whole-body imaging method has not yet been established. Here, we successfully developed a supramolecular magnetic resonance (MR) contrast agent for long-term whole-tissue observation in a single individual. Fluorescein- and Gd-chelate-conjugated polyethylene glycols (PEGs) were synthesized, and their structures were optimized. Spectroscopic and pharmacokinetic analyses suggested that the fluorescein-conjugated linear and 8-arm PEGs with a molecular weight of approximately 10 kDa were suitable to form a supramolecular structure to visualize the microvessel structure and blood circulation. Microvascular formation was evaluated in a glioma cell transplantation model, and neovascularization around the glioma tissue at 5 days was observed, with the contrast agent leaking out into the cancer tissue. In contrast, after 12 days, microvessel structures were formed inside the glioma tissue, but the agents did not leak out. These imaging data for the first time proved that the microvessels formed inside cancer tissues at the early stage are very leaky, but that they form continuous microvessels after 12 days.

Interstitial-fluid shear stresses induced by vertically oscillating head motion lower blood pressure in hypertensive rats and humans.

The mechanisms by which physical exercise benefits brain functions are not fully understood. Here, we show that vertically oscillating head motions mimicking mechanical accelerations experienced during fast walking, light jogging or treadmill running at a moderate velocity reduce the blood pressure of rats and human adults with hypertension. In hypertensive rats, shear stresses of less than 1 Pa resulting from interstitial-fluid flow induced by such passive head motions reduced the expression of the angiotensin II type-1 receptor in astrocytes in the rostral ventrolateral medulla, and the resulting antihypertensive effects were abrogated by hydrogel introduction that inhibited interstitial-fluid movement in the medulla. Our findings suggest that oscillatory mechanical interventions could be used to elicit antihypertensive effects.

3D Printing of Collagen Scaffold with Enhanced Resolution in a Citrate-Modulated Gellan Gum Microgel Bath.

3D printing in a microgel-based supporting bath enables the construction of complex structures with soft and watery biomaterials but the low print resolution is usually an obstacle to its practical application in tissue engineering. Herein, high-resolution printing of a 3D collagen organ scaffold is realized by using an engineered Gellan gum (GG) microgel bath containing trisodium citrate (TSC). The introduction of TSC into the bath system not only mitigates the aggregation of GG microgels, leading to a more homogeneous bath morphology but also suppresses the diffusion of the collagen ink in the bath due to the dehydration effect of TSC, both of which contribute to the improvement of print resolution. 3D collagen organ structures such as hand, ear, and heart are successfully constructed with high shape fidelity in the developed bath. After printing, the GG and TSC can be easily removed by washing with water, and the obtained collagen product exhibits good cell affinity in a tissue scaffold application. This work offers an easy-to-operate strategy for developing a microgel bath for high-resolution printing of collagen, providing an alternative path to in vitro 3D organ construction.

Evaluation of Cardiac Function in Young Mdx Mice Using MRI with Feature Tracking and Self-Gated Magnetic Resonance Cine Imaging.

This study aimed to evaluate cardiac function in a young mouse model of Duchenne muscular dystrophy (mdx) using cardiac magnetic resonance imaging (MRI) with feature tracking and self-gated magnetic resonance cine imaging. Cardiac function was evaluated in mdx and control mice (C57BL/6JJmsSlc mice) at 8 and 12 weeks of age. Preclinical 7-T MRI was used to capture short-axis, longitudinal two-chamber view and longitudinal four-chamber view cine images of mdx and control mice. Strain values were measured and evaluated from cine images acquired using the feature tracking method. The left ventricular ejection fraction was significantly less (p < 0.01 each) in the mdx group at both 8 (control, 56.6 ± 2.3% mdx, 47.2 ± 7.4%) and 12 weeks (control, 53.9 ± 3.3% mdx, 44.1 ± 2.7%). In the strain analysis, all strain value peaks were significantly less in mdx mice, except for the longitudinal strain of the four-chamber view at both 8 and 12 weeks of age. Strain analysis with feature tracking and self-gated magnetic resonance cine imaging is useful for assessing cardiac function in young mdx mice.

Application of Magnetic Resonance Strain Analysis Using Feature Tracking in a Myocardial Infarction Model.

This study validates the usefulness of myocardial strain analysis with cardiac cine magnetic resonance imaging (MRI) by evaluating the changes in the cardiac function and myocardial strain values longitudinally in a myocardial disease model. Six eight-week-old male Wistar rats were used as a model of myocardial infarction (MI). Cine images were taken in the short axis, two-chamber view longitudinal axis, and four-chamber view longitudinal axis directions in rats 3 and 9 days after MI and in control rats, with preclinical 7-T MRI. The control images and the images on days 3 and 9 were evaluated by measuring the ventricular ejection fraction (EF) and the strain values in the circumferential (CS), radial (RS), and longitudinal directions (LS). The CS decreased significantly 3 days after MI, but there was no difference between the images on days 3 and 9. The two-chamber view LS was -9.7 ± 2.1% at 3 days and -13.9 ± 1.4% at 9 days after MI. The four-chamber view LS was -9.9 ± 1.5% at 3 days and -11.9 ± 1.3% at 9 days after MI. Both the two- and four-chamber LS values were significantly decreased 3 days after MI. Myocardial strain analysis is, therefore, useful for assessing the pathophysiology of MI.

Quantitative Susceptibility Mapping and Amide Proton Transfer-Chemical Exchange Saturation Transfer for the Evaluation of Intracerebral Hemorrhage Model.

This study aimed to evaluate an intracerebral hemorrhage (ICH) model using quantitative susceptibility mapping (QSM) and chemical exchange saturation transfer (CEST) with preclinical 7T-magnetic resonance imaging (MRI) and determine the potential of amide proton transfer-CEST (APT-CEST) for use as a biomarker for the early detection of ICH. Six Wistar male rats underwent MRI, and another six underwent histopathological examinations on postoperative days 0, 3, and 7. The ICH model was created by injecting bacterial collagenase into the right hemisphere of the brain. QSM and APT-CEST MRI were performed using horizontal 7T-MRI. Histological studies were performed to observe ICH and detect iron deposition at the ICH site. T2-weighted images (T2WI) revealed signal changes associated with hemoglobin degeneration in red blood cells, indicating acute-phase hemorrhage on day 0, late-subacute-phase hemorrhage on day 3, and chronic-phase hemorrhage on day 7. The susceptibility alterations in each phase were detected using QSM. QSM and Berlin blue staining revealed hemosiderin deposition in the chronic phase. APT-CEST revealed high magnetization transfer ratios in the acute phase. Abundant mobile proteins and peptides were observed in early ICH, which were subsequently diluted. APT-CEST imaging may be a reliable noninvasive biomarker for the early diagnosis of ICH.

Longitudinal Evaluation Using Preclinical 7T-Magnetic Resonance Imaging/Spectroscopy on Prenatally Dose-Dependent Alcohol-Exposed Rats.

Prenatal alcohol exposure causes many detrimental alcohol-induced defects in children, collectively known as fetal alcohol spectrum disorders (FASD). This study aimed to evaluate a rat model of FASD, in which alcohol was administered at progressively increasing doses during late pregnancy, using preclinical magnetic resonance (MR) imaging (MRI) and MR spectroscopy (MRS). Wistar rats were orally administered 2.5 mL/day of ethanol (25% concentration) on gestational day 15, and postnatal fetuses were used as FASD models. Four groups were used: a control group (non-treatment group) and three groups of FASD model rats that received one, two, or four doses of ethanol, respectively, during the embryonic period. Body weight was measured every other week until eight weeks of age. MRI and MRS were performed at 4 and 8 weeks of age. The volume of each brain region was measured using acquired T2-weighted images. At 4 weeks of age, body weight and cortex volume were significantly lower in the three FASD model groups (2.5 × 1: 304 ± 6 mm3, p < 0.05; 2.5 × 2: 302 ± 8 mm3, p < 0.01; 2.5 × 4: 305 ± 6 mm3, p < 0.05) than they were in the non-treatment group (non-treatment: 313 ± 6 mm3). The FASD model group that received four doses of alcohol (2.5 × 4: 0.72 ± 0.09, p < 0.05) had lower Taurine/Cr values than the non-treatment group did (non-treatment: 0.91 ± 0.15), an effect that continued at 8 weeks of age (non-treatment: 0.63 ± 0.09; 2.5 × 4: 0.52 ± 0.09, p < 0.05). This study is the first to assess brain metabolites and volume over time using MRI and MRS. Decreases in brain volume and taurine levels were observed at 4 and 8 weeks of age, suggesting that the effects of alcohol persisted beyond adulthood.

XCR1+ conventional dendritic cell-induced CD4+ T helper 1 cell activation exacerbates cardiac remodeling after ischemic myocardial injury.

Cardiac remodeling has no established therapies targeting inflammation. CD4+ T-cell subsets have been reported to play significant roles in healing process after ischemic myocardial injury, but their detailed mechanisms of activation remain unknown. To explore immune reactions during cardiac remodeling, we applied a non-surgical model of coronary heart disease (CHD) induced by a high-fat diet (HFD-CHD) in SR-BI-/-/ApoeR61h/h mice. Flow cytometry analyses throughout the period of progressive cardiac dysfunction revealed that CD4+ T Helper 1 (Th1) cells were predominantly activated in T-cell subsets. Probucol was reported to attenuate cardiac dysfunction after coronary artery ligation model (ligation-MI) in rats. To determine whether probucol suppress cardiac remodeling after HFD-CHD, we treated SR-BI-/-/ApoeR61h/h mice with probucol. We found treatment with probucol in HFD-CHD mice reduced cardiac dysfunction, with attenuated activation of Th1 cells. RNA-seq analyses revealed that probucol suppressed the expression of CXCR3, a Th1-related chemokine receptor, in the heart. XCR1+ cDC1 cells, which highly expresses the CXCR3 ligands CXCL9 and CXCL10, were predominantly activated after HFD-CHD. XCR1+ cDC1 lineage skewing of pre-DC progenitors was observed in bone marrow, with subsequent systemic expansion of XCR1+ cDC1 cells after HFD-CHD. Activation of CXCR3+ Th1 cell and XCR1+ cDC1 cells was also observed in ligation-MI. Notably, post-MI depletion of XCR1+ cDC1 cells suppressed CXCR3+ Th1 cell activation and prevented cardiac dysfunction. In patient autopsy samples, CXCR3+ Th1 and XCR1+ cDC1 cells infiltrated the infarcted area. In this study, we identified a critical role of XCR1+ cDC1-activated CXCR3+ Th1 cells in ischemic cardiac remodeling.

Evaluation of the efficacy of creatine chemical exchange saturation transfer imaging in assessing testicular maturity.

Purpose: Microscopic testicular sperm extraction is the most effective treatment for NOA, but the sperm retrieval rate is low and depends on testicular maturity. However, there are limited useful tests to assess testicular maturity. Chemical exchange saturation transfer (CEST) imaging is a new magnetic resonance imaging (MRI) technique that can image the distribution of trace substances in vivo. We focused on the potential role of creatine (Cr) in testes and hypothesized that Cr-CEST could indicate intratesticular spermatogenesis. Methods: We performed Cr-CEST by using 7T MRI on wild-type C57B6/J mice and several types of male infertility models such as Sertoli-cell only (SCO) (Kitw/Kitwv), maturation arrest (MA) (Zfp541 knockout mouse and Kctd19 knockout mouse), and teratozoospermia (Tbc1d21 knockout mouse). After performing Cr-CEST, histological analysis was performed. Results: The SCO and MA models showed decreased CEST signal intensity (p < 0.05), while no reduction was observed in the teratozoospermia model (p = 1.0). CEST signal intensity increased as the spermatogenesis stage progressed from the SCO model to the MA and teratozoospermia models. Furthermore, CEST signal intensity was reduced in 4-week-old wild-type mice with immature testes (p < 0.05). Conclusions: This study suggests that Cr-CEST evaluates intratesticular spermatogenesis noninvasively and provides a new therapeutic strategy for treating male infertility.

Creatine Chemical Exchange Saturation Transfer (Cr-CEST) Imaging Can Evaluate Cisplatin-induced Testicular Damage.

PURPOSE: This study aimed to investigate the ability of creatine-chemical exchange saturation transfer (Cr-CEST) technique assessed through 7-T MRI to evaluate cisplatin-induced testicular damage. METHODS: We used 8-10 weeks C57BL/6 mice (n = 10) that were divided into a control group (n = 5) and a cisplatin-treated group (n = 5). The cisplatin group received cisplatin at a dose of 15 mg/kg, via intraperitoneal injection, while the control group received saline. MR images of mouse testes were acquired under anesthesia 18 days after the injection using a horizontal 7-T scanner. The pulse sequence consisted of rapid acquisition with a relaxation enhancement (RARE) with magnetization transfer. The Z-spectra were collected using a 2000-ms saturation pulse at a B1 amplitude of 1.2 µT, with frequencies varying from -4.8 to +4.8 parts per million (ppm). Maps of magnetization transfer ratio with asymmetric analysis (MTRasym) were reconstructed at a Cr metabolite concentration of 1.8 ppm. RESULTS: The Cr-CEST effect was significantly reduced in the cisplatin-treated group compared to the control group (MTRasym of control mice vs. cisplatin-treated mice: 6.9 [6-7.5] vs. 5.2 [4-5.5], P = 0.008). Correlation analysis revealed a strong correlation between the Cr-CEST effect and the pathological score (ρ = 0.93, P < 0.001). CONCLUSION: Cr-CEST MRI can be useful for the evaluation of cisplatin-induced testicular damage in mice.

Quantitative behavioral evaluation of a non-human primate stroke model using a new monitoring system.

Background: Recently, the common marmoset (Callithrix jacchus) has attracted significant interest as a non-human primate stroke model. Functional impairment in non-human primate stroke models should be evaluated quantitatively and successively after stroke, but conventional observational assessments of behavior cannot fully fit this purpose. In this paper, we report a behavioral analysis using MarmoDetector, a three-dimensional motion analysis, in an ischemic stroke model using photosensitive dye, along with an observational behavioral assessment and imaging examination. Methods: Ischemic stroke was induced in the left hemisphere of three marmosets. Cerebral infarction was induced by intravenous injection of rose bengal and irradiation with green light. The following day, the success of the procedure was confirmed by magnetic resonance imaging (MRI). The distance traveled, speed, activity time, and jumps/climbs were observed for 28 days after stroke using MarmoDetector. We also assessed the marmosets' specific movements and postural abnormalities using conventional neurological scores. Results: Magnetic resonance imaging diffusion-weighted and T2-weighted images showed hyperintense signals, indicating cerebral infarction in all three marmosets. MarmoDetector data showed that the both indices immediately after stroke onset and gradually improved over weeks. Neurological scores were the worst immediately after stroke and did not recover to pre-infarction levels during the observation period (28 days). A significant correlation was observed between MarmoDetector data and conventional neurological scores. Conclusion: In this study, we showed that MarmoDetector can quantitatively evaluate behavioral changes in the acute to subacute phases stroke models. This technique can be practical for research on the pathophysiology of ischemic stroke and for the development of new therapeutic methods.

Magnetic Resonance Imaging and Spectroscopy Analysis in a Pelizaeus-Merzbacher Disease Rat Model.

Pelizaeus−Merzbacher disease (PMD) is an X-linked recessive disorder of the central nervous system. We performed 7 Tesla magnetic resonance imaging of the brain in Tama rats, a rodent PMD model, and control rats, as well as evaluated the diagnostic values. In the white matter of the Tama rats, the T2 values were prolonged, which is similar to that observed in patients with PMD (60.7 ± 1.8 ms vs. 51.6 ± 1.3 ms, p < 0.0001). The apparent diffusion coefficient values in the white matter of the Tama rats were higher than those of the control rats (0.68 ± 0.03 × 10−3 mm2/s vs. 0.64 ± 0.03 × 10−3 mm2/s, p < 0.05). In proton magnetic resonance spectroscopy, the N-acetylaspartate (6.97 ± 0.12 mM vs. 5.98 ± 0.25 mM, p < 0.01) and N-acetylaspartate + N-acetylaspartylglutamate values of the Tama rats were higher (8.22 ± 0.17 mM vs. 7.14 ± 0.35 mM, p < 0.01) than those of the control rats. The glycerophosphocholine + phosphocholine values of the Tama rats were lower than those of the control rats (1.04 ± 0.09 mM vs. 1.45 ± 0.04 mM, p < 0.001). By using Luxol fast blue staining, we confirmed dysmyelination in the Tama rats. These results are similar to those of patients with PMD and other PMD animal models.

Metabolites Alterations and Liver Injury in Hepatic Encephalopathy Models Evaluated by Use of 7T-MRI.

This study is to observe a thioacetamide (TAA) administered Hepatic encephalopathy (HE) model rats at three and ten days after TAA administration using liver MRI and brain MR Spectroscopy (MRS) by use of 7T-MRI. Forty-two Wistar rats (control group, n = 14) were intraperitoneally administered at 300 mg/kg (low-dose group, n = 14) or 400 mg/kg (high-dose group, n = 14) doses of TAA for induced of HE. At three days after TAA administration, glutamine (Gln) measured by MRS in high-dose and low-dose TAA groups showed significant increases in comparison to those of the control group (p < 0.05). Other metabolites measured by MRS showed no significant changes. Liver T1ρ and T2 relaxation times significantly increased three days after TAA injection compared to pre-injection. There was a correlation between Gln levels in the brain and the relaxation time of the liver. Furthermore, Gln levels and relaxation time changed depending on the TAA dose. The Gln concentration in the brain increased with the deterioration of liver function, as inferred from the prolonged relaxation time of the liver. The prolonged relaxation time of the liver corresponded with the level of Gln in the brain. Gln concentration for the alterations of brain metabolites and T1ρ relaxation time for the assessment of liver damage are useful markers for inter-organ association analysis in the HE model.