Makoto Tsuchimochi

Intraoperative gamma cameras for radioguided surgery: Technical characteristics, performance parameters, and clinical applications

Several small gamma cameras (SGCs) intended for surgical use are now in development or currently being marketed. In this review, we discuss the characteristics, performance, and clinical use of SGCs which are hand-held and small enough to be easily managed by surgeons during their procedures. We expect that SGCs have the potential to be used more widely in radioguided surgery. As advancing molecular imaging technologies will broaden clinical indications, SGCs will likely be used and integrated with other imaging modalities into numerous types of radioguided surgery in the near future.

Evaluation of the Efficacy of a Small CdTe γ-Camera for Sentinel Lymph Node Biopsy

We previously reported the basic performance of a prototype small cadmium telluride (CdTe) γ-camera (SSGC) intended for use in radioguided surgeries. In this study, we sought to confirm the favorable previous results and to extend the preliminary findings to examine the efficacy of the SSGC in an animal study and a clinical setting for sentinel lymph node biopsy. Methods: The prototype SSGC (1,024 pixels; field of view, 44.8 × 44.8 mm), equipped with a parallel-hole collimator, was used in both animal and clinical studies. 99mTc-phytate (18.5 MBq) was injected into the tongues and legs of 6 rabbits. In the clinical study, 74 MBq of 99mTc-phytate was injected into peritumoral regions in 8 patients with oral cancer. The detection of hot nodes by the SSGC was compared with that by a conventional scintillation γ-camera (CGC). Results: The SSGC detected 29 hot nodes in images of 6 rabbits after injection. The number of hot nodes was the same as the number seen in CGC studies, but the CGC required a longer acquisition time to produce comparable images. There were no differences between the SSGC and the CGC in terms of activity ratios and hot node-to-background ratios. The biodistribution of 99mTc-phytate in removed tissues was evaluated by contact radiography, and radioactivity was assayed with a γ-well counter. The mean ± SD radioactivity in specimens was 0.15% ± 0.15%, with a range of 0.01%–0.62%. In the clinical study, the SSGC detected 30 hot nodes with a 5- to 60-s acquisition time at 4 h after injection. The SSGC documented all hot nodes depicted by the CGC at 4 h after injection. Conclusion: The SSGC showed significant potential for the detection of sentinel lymph nodes in lymphoscintigraphy. The results of the studies suggested that the SSGC facilitates the exploration of hot nodes in sentinel lymph node biopsy.

Dual-modality imaging with 99mTc and fluorescent indocyanine green using surface-modified silica nanoparticles for biopsy of the sentinel lymph node: an animal study

BackgroundWe propose a new approach to facilitate sentinel node biopsy examination by multimodality imaging in which radioactive and near-infrared (NIR) fluorescent nanoparticles depict deeply situated sentinel nodes and fluorescent nodes with anatomical resolution in the surgical field. For this purpose, we developed polyamidoamine (PAMAM)-coated silica nanoparticles loaded with technetium-99m (99mTc) and indocyanine green (ICG).MethodsWe conducted animal studies to test the feasibility and utility of this dual-modality imaging probe. The mean diameter of the PAMAM-coated silica nanoparticles was 30 to 50 nm, as evaluated from the images of transmission electron microscopy and scanning electron microscopy. The combined labeling with 99mTc and ICG was verified by thin-layer chromatography before each experiment. A volume of 0.1 ml of the nanoparticle solution (7.4 MBq, except for one rat that was injected with 3.7 MBq, and 1 μg of an ICG derivative [ICG-sulfo-OSu]) was injected submucosally into the tongue of six male Wistar rats.ResultsScintigraphic images showed increased accumulation of 99mTc in the neck of four of the six rats. Nineteen lymph nodes were identified in the dissected neck of the six rats, and a contact radiographic study showed three nodes with a marked increase in uptake and three nodes with a weak uptake. NIR fluorescence imaging provided real-time clear fluorescent images of the lymph nodes in the neck with anatomical resolution. Six lymph nodes showed weak (+) to strong (+++) fluorescence, whereas other lymph nodes showed no fluorescence. Nodes showing increased radioactivity coincided with the fluorescent nodes. The radioactivity of 15 excised lymph nodes from the four rats was assayed using a gamma well counter. Comparisons of the levels of radioactivity revealed a large difference between the high-fluorescence-intensity group (four lymph nodes; mean, 0.109% ± 0.067%) and the low- or no-fluorescence-intensity group (11 lymph nodes; mean, 0.001% ± 0.000%, p < 0.05). Transmission electron microscopy revealed that small black granules were localized to and dispersed within the cytoplasm of macrophages in the lymph nodes.ConclusionAlthough further studies are needed to determine the appropriate dose of the dual-imaging nanoparticle probe for effective sensitivity and safety, the results of this animal study revealed a novel method for improved node detection by a dual-modality approach for sentinel lymph node biopsy.

Evaluation by Bone Scintigraphy of Osteogenic Activity of Commercial Bioceramics (Porous β-TCP and HAp Particles) Subcutaneously Implanted in Rats:

Osteogenic potential of biomaterials used in bone regenerative therapy has been mainly examined in an animal-implantation study. We have here evaluated the applicability of bone scintigraphy in imaging ectopic bone formation, especially its initial phase, by β-tricalcium phosphate (β-TCP) particles that were implanted in rat dorsal subcutaneous tissues. In implanted osteogenic osteosarcoma cells used as a positive control, osteoid formation was found by histological examination and bone scintigraphy using 99mTc- hydroxymethyl diphosphonate (HMDP) at 2 and 3 weeks post-implantation, respectively, while the microfocuscomputed tomography (μCT) system required further mineralization, which occurred at 4 weeks. Implantation of β-TCP particles alone induced only faint biomineralization inside the particles, which could be microscopically detected by calcein chelation at 2 weeks post-implantation, but not by other histological examinations (e.g., HE staining) or μCT. However, the bone scintigraphy successfully detected this microscopic change at 1 week. Implanted hydroxyapatite (HAp) particles alone used as a negative control did not induce mineralization at microscopic levels, and therefore nothing was detected by either calcein chelation or bone scintigraphy. In conclusion, the bone scintigraphic methodology, although exhibiting less quantitation and resolution, would be applicable as a non-invasive, highly sensitive methodology in detecting the initial, microscopic changes associated with mineralization.

Quantification of disc displacement in internal derangement of the temporomandibular joint using magnetic resonance imaging

Many measures have been developed to determine the extent of disc displacement in internal derangements of the temporomandibular joint (TMJ) using magnetic resonance imaging. The purpose of this study was to develop a quantitative method of analyzing disc position and to evaluate the positions of the disc in internal derangements of the TMJ (group 1, with reduction; group 2, without reduction). Magnetic resonance images of 150 TMJs in 20 healthy volunteers and 55 patients with internal derangements were evaluated. The anatomical points of interest of the TMJ, including the anterior (DA) and posterior (DP) points of the disc, were marked on parasagittal magnetic resonance images of the TMJ disc taken in both the closed- and the open-mouth positions. All points were recorded using an x-y coordinate system, with reference to a referral line. In the closed-mouth position, the DP in patients in group 1 was situated in a more-anterior direction than the DP in volunteers. The DP in group 2 was located further anterior and inferior than the DP in group 1. However, the position of the DA did not differ between group 1 and group 2. In the open-mouth position, the DP was displaced anteroinferiorly to a greater extent in group 2 than in group 1 (one-way ANOVA, followed by Scheffe’s test; P < 0.0001). The distance between the disc points in the closed- and open-mouth positions was also evaluated. Comparison of the disc point position in the closed- and open-mouth positions in symptomatic and asymptomatic displaced TMJ discs revealed no significant difference. In conclusion, most of our results quantitatively support previously reported findings in imaging, surgical, and histopathological studies of TMJ internal derangement. We suggest that our measure of disc position of the TMJ would be useful to assess the status and response to treatment of internal derangements of the TMJ.

Magnetic resonance imaging findings of nodular fasciitis in the mental region.

Nodular fasciitis (NF) is a benign reactive lesion of the soft tissues related to the fascia and characterized by fibroblastic proliferation. The most common site is the upper extremities (46%), followed by the head and neck region (20%). In the orofacial region, the lesion typically develops within the subcutaneous structures overlying the angle and inferior border of the mandible and the zygoma. Magnetic resonance imaging (MRI) findings of NF in the orofacial region are almost unreported in the literature. In the present case report, we describe MRI findings of mental NF in a 19-year-old woman. MRI revealed a well-defined, round soft-tissue mass lying on the mentum. On T1-weighted MRI, the lesion was isointense to skeletal muscle; it was hyperintense to skeletal muscle on T2-weighted MRI, and was enhanced by Gd-diethylenetriamine pentaacetic acid (DTPA). Histologic examination revealed abundant myxoid degeneration dispersed in the lesion. The T2-weighted higher heterogeneous signal intensity was likely due to abundant myxoid degeneration or the cellular component of the lesion. A strong bright signal intensity belt appeared in the periphery of the lesion on Gd-DTPA enhancement. This rim enhancement appearred to represent small arterioles and venules that were visible in the peripheral area on histologic examination.

Dual-Labeled Near-Infrared/99mTc Imaging Probes Using PAMAM-Coated Silica Nanoparticles for the Imaging of HER2-Expressing Cancer Cells

We sought to develop dual-modality imaging probes using functionalized silica nanoparticles to target human epidermal growth factor receptor 2 (HER2)-overexpressing breast cancer cells and achieve efficient target imaging of HER2-expressing tumors. Polyamidoamine-based functionalized silica nanoparticles (PCSNs) for multimodal imaging were synthesized with near-infrared (NIR) fluorescence (indocyanine green (ICG)) and technetium-99m (99mTc) radioactivity. Anti-HER2 antibodies were bound to the labeled PCSNs. These dual-imaging probes were tested to image HER2-overexpressing breast carcinoma cells. In vivo imaging was also examined in breast tumor xenograft models in mice. SK-BR3 (HER2 positive) cells were imaged with stronger NIR fluorescent signals than that in MDA-MB231 (HER2 negative) cells. The increased radioactivity of the SK-BR3 cells was also confirmed by phosphor imaging. NIR images showed strong fluorescent signals in the SK-BR3 tumor model compared to muscle tissues and the MDA-MB231 tumor model. Automatic well counting results showed increased radioactivity in the SK-BR3 xenograft tumors. We developed functionalized silica nanoparticles loaded with 99mTc and ICG for the targeting and imaging of HER2-expressing cells. The dual-imaging probes efficiently imaged HER2-overexpressing cells. Although further studies are needed to produce efficient isotope labeling, the results suggest that the multifunctional silica nanoparticles are a promising vehicle for imaging specific components of the cell membrane in a dual-modality manner.

Immunohistochemical localization of parathyroid hormone-related protein (PTHrP) and serum PTHrP in normocalcemic patients with oral squamous cell carcinoma.

Cancer cells produce parathyroid hormone-related protein (PTHrP) in the early phase of malignancy development, before hypercalcemia occurs. The relationship between PTHrP and the clinicopathologic features of oral squamous cell carcinoma is poorly understood. We studied 60 patients (43 men, 17 women; mean age, 64.8 ± 11.2 years) with primary oral squamous cell carcinoma, from whom pretreatment biopsy specimens were obtained. We examined the relationship among immunohistochemical PTHrP expression, serum PTHrP levels, clinical characteristics of the tumor, and histopathologic aspects of the tumor. The mean calcium concentration for the 60 patients was 9.1 ± 0.4 mg/dl. No patients had laboratory evidence of hypercalcemia before treatment. Six patients had serum levels of C-terminal (C)-PTHrP higher than the normal level of 55.3 pmol/l. There were no significant differences in serum C-PTHrP levels according to TNM stages. Abundant positive immunoreactivity for anti-PTHrP (1-34) antibody was recognized diffusely in the whole cytoplasm of many tumor cells. Anti-PTHrP (38-64) antibody staining tended to localize as small granules in the cytoplasm, especially close to the nuclear periphery. There was no correlation between the serum C-PTHrP concentration and the intensity of either immunostain. The intensity of PTHrP was proportionally related to the degree of differentiation or extent of keratinization (P < 0.05) and the histologic malignancy grade of the tumor (P < 0.05), when using antibody against PTHrP (1-34), but not when using antibody against PTHrP (38-64). Serum C-PTHrP levels did not correlate with the intensity of cellular PTHrP expression and characteristics of the tumor at the initial patient visit. The fragment that includes PTHrP (1-34) may be involved in the differentiation of oral squamous cell carcinoma. The differences between immunoreactivities may have been due to differing tissue malignancies and the use of different antibodies. The results suggest the need for caution when interpreting immunoreactivities of PTHrP in malignancies.

Non-invasive, quantitative assessment of the morphology of γ-irradiated human mesenchymal stem cells and periosteal cells using digital holographic microscopy

Abstract Purpose: To assure the quality of cells to be used in cell therapy, we examined the applicability of digital holographic microscopy (DHM) for non-invasive, quantitative assessment of changes in cell morphology. Materials and methods: Mesenchymal stem cells derived from adipose tissue (MSC-AT) and bone marrow (MSC-BM), in addition to human alveolar periosteal cells (PC) as a reference, were γ-ray irradiated (1 and 4 Gy), and their morphological changes were quantified without fixation using holographic microscopy. After detachment and fixation with ethanol, cell number and surface antigen expression were determined using an automated cell counter kit and flow-cytometry, respectively. Results: Among various indexes, only indexes related to cell size were significantly changed after γ-irradiation. Both BMC-AT and BMC-BM were enlarged and more sensitive to a low dose of γ-irradiation than PC. In contrast to PC, proteins related to DNA damage repair (γ-H2AX, p21waf1, p53 and Rb) were not substantially upregulated or sustained for a week in either MSC-AT or MSC-BM. Conclusion: Instead of DNA damage markers, we suggest that cell morphological parameters (e.g. cell volume) that are monitored by DHM could be a useful and more stable marker of MSC quality.

Quantitative single-cell motility analysis of platelet-rich plasma-treated endothelial cells in vitro.

Platelet‐rich plasma (PRP) has been widely applied in regenerative therapy due to its high concentration of growth factors. Previous in vitro and in vivo studies have provided evidence supporting the angiogenic activity of PRP. To more directly demonstrate how PRP acts on endothelial cells, we examined the PRP‐induced changes in the motility of human umbilical vein endothelial cells by examining the involvement of VEGF. Time‐lapse quantitative imaging demonstrated that in the initial phase (∼2 h) of treatment, PRP substantially stimulated cell migration in a wound‐healing assay. However, this effect of PRP was not sustained at significant levels beyond the initial phase. The average net distance of cell migration at 10 h was 0.45 ± 0.16 mm and 0.82 ± 0.23 mm in control and PRP‐stimulated cells, respectively. This effect was also demonstrated with recombinant human VEGF and was significantly attenuated by a neutralizing anti‐VEGF antibody. Immunofluorescent examination of paxillin and actin fibers demonstrated that PRP concomitantly up‐regulated focal adhesion and cytoskeletal formation. Western blotting analysis of phosphorylated VEGFR2 demonstrated that PRP mainly stimulated the phosphorylation of immature VEGFR2 in a dose‐ and time‐dependent manner, an action that was completely blocked by the neutralizing antibody. Taken together, these data suggest that PRP acts directly on endothelial cells via the activation of VEGFR2 to transiently up‐regulate their motility. Thus, the possibility that PRP desensitizes target endothelial cells for a relatively long period of time after short‐term activation should be considered when the controlled release system of PRP components is designed.