Shigeyoshi Saito

Impact of surface coating and particle size on the uptake of small and ultrasmall superparamagnetic iron oxide nanoparticles by macrophages.

Purpose Magnetic resonance imaging (MRI) using contrast agents like superparamagnetic iron oxide (SPIO) is an extremely versatile technique to diagnose diseases and to monitor treatment. This study tested the relative importance of particle size and surface coating for the optimization of MRI contrast and labeling efficiency of macrophages migrating to remote inflammation sites. Materials and methods We tested four SPIO and ultrasmall superparamagnetic iron oxide (USPIO), alkali-treated dextran magnetite (ATDM) with particle sizes of 28 and 74 nm, and carboxymethyl dextran magnetite (CMDM) with particle sizes of 28 and 72 nm. Mouse macrophage RAW264 cells were incubated with SPIOs and USPIOs, and the labeling efficiency of the cells was determined by the percentage of Berlin blue-stained cells and by measuring T2 relaxation times with 11.7-T MRI. We used trypan blue staining to measure cell viability. Results Analysis of the properties of the nanoparticles revealed that ATDM-coated 74 nm particles have a lower T2 relaxation time than the others, translating into a higher ability of MRI negative contrast agent. Among the other three candidates, CMDM-coated particles showed the highest T2 relaxation time once internalized by macrophages. Regarding labeling efficiency, ATDM coating resulted in a cellular uptake higher than CMDM coating, independent of nanoparticle size. None of these particle formulations affected macrophage viability. Conclusion This study suggests that coating is more critical than size to optimize the SPIO labeling of macrophages. Among the formulations tested in this study, the best MRI contrast and labeling efficiency are expected with ATDM-coated 74 nm nanoparticles.

Control of the temperature rise in magnetic hyperthermia with use of an external static magnetic field

Our purpose in this study was to investigate the usefulness of a method for controlling the temperature rise in magnetic hyperthermia (MH) using an external static magnetic field (SMF), and to derive an empirical equation for describing the energy dissipation of magnetic nanoparticles (MNPs) in the presence of both the alternating magnetic field (AMF) and SMF through phantom experiments. We made a device that allows for MH in the presence of an SMF with a field-free point (FFP) using a Maxwell coil pair. We measured the temperature rise of MNPs under various conditions of AMF and SMF and various distances from the FFP (d), and calculated the specific absorption rate (SAR) from the initial slope of the temperature curve. The SAR values decreased with increasing strength of SMF (Hs) and d. The extent of their decrease with d increased with an increase of the gradient of SMF (Gs). The relationships between SAR and Hs and between SAR and d could be well approximated by Rosensweigs equation in which the amplitude of AMF (Hac) is replaced by √[Hac(2)]/√[Hac(2)+Hs(2)], except for the case when Gs was small. In conclusion, the use of an external SMF with an FFP will be effective for controlling the temperature rise in MH in order to reduce the risk of heating surrounding healthy tissues, and our empirical equation will be useful for estimating SAR in the presence of both the AMF and SMF and for designing an effective local heating system for MH.

Detection and Early Phase Assessment of Radiation-Induced Lung Injury in Mice Using Micro-CT

Radiation therapy is an important therapeutic modality for thoracic malignancies. However, radiation-induced pulmonary injuries such as radiation pneumonitis and fibrosis are major dose-limiting factors. Previous research shows that micro-computed tomography (micro-CT) can detect radiation-induced lung injuries a few months following irradiation, but studies to assess the early response of lung tissue are lacking. The aim of this study was to determine if micro-CT could be used to detect and assess early-phase radiation–induced lung injury in mice. Twenty-one animals were divided into three groups: normal (n = 7), one day after x-ray exposure (n = 7), and at four days after x-ray exposure (n = 7). The x-ray-exposed groups received a single dose of 20 Gy, to the whole lung. Histology showed enlargements of the air space (Lm: mean chord length) following irradiation. 40.5±3.8 µm and 60.0±6.9 µm were observed after one and four days, respectively, compared to 26.5±3.1 µm in normal mice. Three-dimensional micro-CT images were constructed and histograms of radiodensity - Hounsfield Units (HU) - were used to assess changes in mouse lungs. Radiation-induced lung injury was observed in irradiated mice, by the use of two parameters which were defined as shifts in peak HU between −200 to −800 HU (PeakHU) and increase in the number of pixels at −1000 HU (Number-1000). These parameters were correlated with histological changes. The results demonstrate that micro-CT can be used for the early detection and assessment of structural and histopathological changes resulting from radiation-induced lung injury in mice. Micro-CT has the advantage, over traditional histological techniques, of allowing longitudinal studies of lung disease progression and assessment of the entire lung, while reducing the number of animals required for such studies.

Fetal Gyrification in Cynomolgus Monkeys: A Concept of Developmental Stages of Gyrification

Our article summarizes a series of studies about fetal gyrification and its relation to cerebral growth in cynomolgus monkeys. Based on the cerebral growth (i.e., brain weight, cerebral volume, and frontooccipital length of the cerebral hemisphere) and the developmental pattern of gyrification in each sulcus of cynomolgus monkeys, we divided the gyrification process into four stages: Stage 1. Demarcation of cerebral lobes and limbic gyri; Stage 2. Demarcation of neocortical gyri; Stage 3. Emergence of secondary and tertiary sulci; and Stage 4. Growth of sulcal length and depth. Each stage of those gyrification processes was influenced by different developmental events, such as the emergence of corticocortical long‐associative fiber tracts, cortical maturations, and subcortical white‐matter development. This is the first report to systematically propose gyrification processes closely related to the order of phyologenetical development of the cerebral cortex in primates. Anat Rec, 2012.

Molecular imaging of mesothelioma by detection of manganese-superoxide dismutase activity using manganese-enhanced magnetic resonance imaging

Malignant mesothelioma (MM) is a fatal malignancy with a rapidly increasing incidence in industrialized countries because of the widespread use of asbestos in the past centuries. Early diagnosis of MM is critical for a better prognosis, but this is often difficult because of the lack of disease‐specific diagnostic imaging. Here, we report that manganese‐enhanced magnetic resonance imaging (MEMRI) represents a promising approach for a more selective mesothelioma imaging by monitoring a high‐level expression of manganese‐superoxide dismutase (Mn‐SOD), which is observed in many MM. We found that most human MM cells overexpressed Mn‐SOD protein compared with human mesothelial cells and that NCI‐H226 human MM cells highly expressed Mn‐SOD and augmented Mn accumulation when loaded with manganese chloride (MnCl2). The cells showed marked T1‐signal enhancement on in vitro MRI after incubation with MnCl2 because of the T1 shortening effect of Mn2+. H226 subcutaneous tumor was preferentially enhanced compared with a lung adenocarcinoma cell tumor and another human MM cell tumor in MnCl2‐enhanced T1‐weighted MR image (T1WI), correlating with their respective Mn‐SOD expression levels. Moreover, in a more clinically relevant setting, H226 xenografted pleural tumor was markedly enhanced and readily detected by MEMRI using manganese dipyridoxyl diphosphate (MnDPDP), a clinically used contrast agent, as well as MnCl2. Therefore, we propose that MEMRI can be a potentially powerful method for noninvasive detection of MM, with high spatial resolution and marked signal enhancement, by targeting Mn‐SOD.

Dynamic contrast-enhanced MRI of the liver in Mrp2-deficient rats using the hepatobiliary contrast agent Gd-EOB-DTPA.

ObjectivesThe objective of this study was to compare the hepatic uptake and biliary excretion of gadolinium-ethoxybenzyl-diethylenetriamine pentaacetic acid (Gd-EOB-DTPA) in control and multidrug resistance–associated protein 2 (Mrp2)–deficient rats by noninvasive dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) and the impact of Mrp2 deficiency on organic anion-transporting polypeptide 1 (Oatp1) transporters and liver vascularization by immunohistochemistry. Materials and MethodsTwenty rats were used in the normal control (n = 10) and Mrp2-deficient rat groups (n = 10). Dynamic contrast-enhanced magnetic resonance imaging studies were performed using Gd-EOB-DTPA (0.025 mmol Gd/kg; 0.1 mL/kg body weight) as the contrast agent. The percentages of relative enhancement were calculated at each time point after Gd-EOB-DTPA injection. In addition, relative enhancement maps were generated through pixel-by-pixel calculations before the injection and at 5, 10, 20, 30, and 40 minutes after the injection. After the DCE-MRI study, blood was sampled from all rats and 6 blood sample parameters, serum aspartate aminotransferase, alanine aminotransferase, total bilirubin, direct bilirubin, indirect bilirubin, and total bile acids, were measured. Rat livers were processed for histologic diagnosis to clarify contrast agent uptake/efflux by examining Oatp1, Mrp2, and platelet endothelial cell adhesion molecule immunohistochemical staining. ResultsThe relative enhancement of the Mrp2-deficient, Eisai hyperbilirubinuria rats (EHBRs) (48.6% [3.4%]) was significantly lower than that of the control rats (64.0% [3.2%]; P < 0.001) 5 minutes after the Gd-EOB-DTPA injection. Thereafter, the relative enhancement observed in the EHBRs (10 minutes, 59.6% [5.4%]; 20 minutes, 67.8% [4.1%]; 30 minutes, 69.1% [4.2%]; 40 minutes, 71.0% [4.2%]; P < 0.0001) was significantly higher than that in the control rats at the same time points after the Gd-EOB-DTPA injection. The aspartate aminotransferase and alanine aminotransferase values were not significantly different between the 2 groups. However, total bilirubin, direct bilirubin, indirect bilirubin, and total bile acids levels in EHBRs were significantly higher than those in the control rats. The percentages of the Mrp2-positive cells in the control rats were higher compared with the EHBRs (control, 0.3% [0.1%]; EHBR, 14.1% [3.6%]; P < 0.01). However, the percentages of the Oatp1-positive cells were not different between the 2 groups. Moreover, the percentages of the platelet endothelial cell adhesion molecule–positive cells in the blood vessels of the control rat livers were higher compared with the EHBRs (control, 17.5% [3.3%]; EHBR, 9.5% [3.9%]; P < 0.01). ConclusionsThe utility of noninvasive DCE-MRI with Gd-EOB-DTPA as a tool for the assessment of Mrp2-deficient hyperbilirubinuria rats was demonstrated. We also clarified that the lower vascular density in the EHBRs may cause delayed uptake of the contrast agent compared with the control rats. In addition, the lower Mrp2 transporter expression may cause the lower efflux of the contrast agent from the Mrp2-deficient rats compared with the control rats.

Ontogenetic pattern of gyrification in fetuses of cynomolgus monkeys

The ontogenetic pattern of gyrification and its relationship with cerebral cortical volume were examined in cynomolgus monkey fetuses. T(1)-weighted coronal magnetic resonance (MR) images at 7 T were acquired from the fixed cerebra of three male fetuses, each at embryonic days (EDs) 70 to 150, and the gyrification index (GI) of each slice was estimated. The mean GI was low (1.1-1.2) during EDs 70 to 90, and then increased dramatically on ED 100. The developmental profiles of the rostrocaudal GI distribution revealed that cortical convolution was more frequent in the parietooccipital region than in other regions during EDs 100 to 150, forming an adult-like pattern by ED 150. The mean GI was closely correlated with the volume of cortical gray matter (r=0.9877), and also with the volume of white matter/intermediate zone (r=0.8961). These findings suggest that cortical convolution is correlated with either the maturation of cortical gray matter or the development of white matter bundles. The characteristic GI distribution pattern of catarrhines was formed by ED 150 in correlation with the progressive sulcal infolding in the parietooccipital region of the cerebrum.

Increased N-acetylaspartate in model mouse of Pelizaeus-Merzbacher disease.

To evaluate the N‐acetylaspartate (NAA) and N‐acetylaspartylglutamate (NAAG) biochemical pathways in the brain of myelin synthesis‐deficient (msd) mouse, a model of Pelizaeus‐Merzbacher disease (PMD).

Spatial frequency-based analysis of mean red blood cell speed in single microvessels: investigation of microvascular perfusion in rat cerebral cortex.

Background Our previous study has shown that prenatal exposure to X-ray irradiation causes cerebral hypo-perfusion during the postnatal development of central nervous system (CNS). However, the source of the hypo-perfusion and its impact on the CNS development remains unclear. The present study developed an automatic analysis method to determine the mean red blood cell (RBC) speed through single microvessels imaged with two-photon microscopy in the cerebral cortex of rats prenatally exposed to X-ray irradiation (1.5 Gy). Methodology/Principal Findings We obtained a mean RBC speed (0.9±0.6 mm/sec) that ranged from 0.2 to 4.4 mm/sec from 121 vessels in the radiation-exposed rats, which was about 40% lower than that of normal rats that were not exposed. These results were then compared with the conventional method for monitoring microvascular perfusion using the arteriovenous transit time (AVTT) determined by tracking fluorescent markers. A significant increase in the AVTT was observed in the exposed rats (1.9±0.6 sec) as compared to the age-matched non-exposed rats (1.2±0.3 sec). The results indicate that parenchyma capillary blood velocity in the exposed rats was approximately 37% lower than in non-exposed rats. Conclusions/Significance The algorithm presented is simple and robust relative to monitoring individual RBC speeds, which is superior in terms of noise tolerance and computation time. The demonstrative results show that the method developed in this study for determining the mean RBC speed in the spatial frequency domain was consistent with the conventional transit time method.

Manganese-Enhanced MRI Reveals Early-Phase Radiation-Induced Cell Alterations In Vivo

For tumor radiotherapy, the in vivo detection of early cellular responses is important for predicting therapeutic efficacy. Mn(2+) is used as a positive contrast agent in manganese-enhanced MRI (MEMRI) and is expected to behave as a mimic of Ca(2+) in many biologic systems. We conducted in vitro and in vivo MRI experiments with Mn(2+) to investigate whether MEMRI can be used to detect cell alterations as an early-phase tumor response after radiotherapy. Colon-26 cells or a subcutaneously grafted colon-26 tumor model were irradiated with 20 Gy of X-rays. One day after irradiation, a significant augmentation of G2-M-phase cells, indicating a cell-cycle arrest, was observed in the irradiated cells in comparison with the control cells, although both early and late apoptotic alterations were rarely observed. The MEMRI signal in radiation-exposed tumor cells (R1: 0.77 ± 0.01 s(-1)) was significantly lower than that in control cells (R1: 0.82 ± 0.01 s(-1)) in vitro. MEMRI signal reduction was also observed in the in vivo tumor model 24 hours after irradiation (R1 of radiation: 0.97 ± 0.02 s(-1), control: 1.10 ± 0.02 s(-1)), along with cell-cycle and proliferation alterations identified with immunostaining (cyclin D1 and Ki-67). Therefore, MEMRI after tumor radiotherapy was successfully used to detect cell alterations as an early-phase cellular response in vitro and in vivo.