Ritsuo Takagi

Identification of potential biomarkers of lymph node metastasis in oral squamous cell carcinoma by cDNA microarray analysis

We surveyed the expression of 557 cancer‐related genes in 15 cases of well‐differentiated OSCC by cDNA microarray analysis. To identify potential biomarkers for lymph node metastasis, all microarray data were compared by the Mann‐Whitney test and the significance analysis of microarrays between OSCCs with and those without lymph node metastasis. The tissues of OSCCs with lymph node metastasis exhibited increased expression levels of MMP‐1, MMP‐3, uPA, integrin‐α3, paxillin, tenascin C and IL‐6 transcripts. All of these genes were included in common clusters on the Cluster/TreeView analysis, implying that functional gene groups of proteolytic enzymes and integrin‐related molecules are involved in cervical lymph node metastasis. The results of RTQ‐PCR for differentially expressed genes were in accord with those of cDNA microarray analyses, suggesting that the data obtained by microarray gene expression analyses were valid. Consistent with cooperative expression patterns, immunohistochemical analyses demonstrated that products of MMP‐1, MMP‐3 and uPA were colocalized to components of the neoplastic stroma, particularly mononuclear inflammatory cells with well‐developed eosinophilic cytoplasm. Our results suggest that expression levels of molecules involved in tissue remodeling and cell–ECM adhesion, especially MMP‐1 and integrin‐α3, can provide an accurate biomarker system for predicting the risk of cervical lymph node metastasis in OSCC.

Curcumin prevents diabetic cardiomyopathy in streptozotocin-induced diabetic rats: Possible involvement of PKC–MAPK signaling pathway

The development of diabetic cardiomyopathy is accompanied with a high membrane-bound protein kinase C (PKC) levels. Curcumin is a naturally occurring compound which is known to inhibit PKC activity. However, the effects of curcumin on ameliorating diabetic cardiomyopathy are still undefined. We evaluated whether curcumin treatment is associated with the modulation of PKC-α and -β₂-mitogen-activated protein kinase (MAPK) pathway in experimental diabetic cardiomyopathy. Diabetes was induced in male Sprague-Dawley rats by streptozotocin (STZ). Curcumin (100mg/kg/day) was started three weeks after STZ injection and was given for 8 weeks. We demonstrate that curcumin significantly prevented diabetes-induced translocation of PKC-α and -β2 to membranous fraction and diabetes-induced increased phosphorylation of p38MAPK and extracellular regulated-signal kinase (ERK)1/2 in left ventricular tissues of diabetic rats. Curcumin treatment also markedly decreased NAD(P)H oxidase subunits (p67phox, p22phox, gp91phox), growth factors (transforming growth factor-β, osteopontin) and myocyte enhancer factor-2 protein expression as well as inhibited NF-κB activity at nuclear level. Furthermore, curcumin decreased the mRNA expression of transcriptional coactivator p300 and atrial natriuretic peptide, decreased accumulation of ECM protein and reversed the increment of superoxide production in left ventricular tissues, as evidenced by dihydroethidium staining. It is also significantly lowered plasma glucose and attenuated oxidative stress, as determined by lipid peroxidation and activity of anti-oxidant enzyme, and as a result attenuated cardiomyocyte hypertrophy, myocardial fibrosis and left ventricular dysfunction. Taken together, it is suggested that curcumin by inhibiting PKC-α and -β₂-MAPK pathway may be useful as an adjuvant therapy for the prevention of diabetic cardiomyopathy.

The Association of Temporomandibular Joint Pain With Abnormal Bone Marrow in the Mandibular Condyle

PURPOSE This study investigated the association between temporomandibular joint pain and bone marrow alterations in the mandibular condyle seen on magnetic resonance (MR) images. PATIENTS AND METHODS The study was based on 112 temporomandibular joints in 112 patients with disc displacement without reduction. Thirty-four patients with abnormal bone marrow on MR images were compared with a control group of 78 patients with normal bone marrow. The analysis was based on proton density and T2-weighted MR images in the oblique sagittal and coronal planes. The degree of pain was correlated to the status of the bone marrow using statistical methods. RESULTS The degree of pain in joints with abnormal bone marrow was higher than in joints with normal bone marrow signal on MR images (P = .0045). CONCLUSION Because the stage of internal derangement was similar in both groups, more intensive pain appears to be associated with bone marrow alterations.

Thermal Generation of Spin Current in an Antiferromagnet.

The longitudinal spin Seebeck effect has been investigated for a uniaxial antiferromagnetic insulator Cr(2)O(3), characterized by a spin-flop transition under magnetic field along the c axis. We have found that a temperature gradient applied normal to the Cr(2)O(3)/Pt interface induces inverse spin Hall voltage of spin-current origin in Pt, whose magnitude turns out to be always proportional to magnetization in Cr(2)O(3). The possible contribution of the anomalous Nernst effect is confirmed to be negligibly small. The above results establish that an antiferromagnetic spin wave can be an effective carrier of spin current.

FGF23 is mainly synthesized by osteocytes in the regularly distributed osteocytic lacunar canalicular system established after physiological bone remodeling.

This study aimed to evaluate whether the immunolocalization of fibroblast growth factor (FGF) 23 and dentin matrix protein 1 (DMP1) is associated with the spatial regularity of the osteocyte lacunar canalicular system(s) (OLCS). Femora of 12-weeks-old male ICR mice were fixed with 4% paraformaldehyde, decalcified with a 10% EDTA solution and then embedded in paraffin. We have devised a triple staining procedure that combines silver impregnation, alkaline phosphatase (ALPase) immunohistochemistry and tartrate-resistant acid phosphatase (TRAPase) enzyme histochemistry on a single paraffin section. This procedure permitted the visualization of ALPase-positive plump osteoblasts and several TRAPase-positive osteoclasts on those bone matrices featuring irregularly arranged OLCS, and of ALPase-positive bone lining cells on the bone matrix displaying the well-arranged OLCS. As observations proceeded from the metaphysis toward the diaphysis, the endosteal cortical bone displayed narrower bands of calcein labeling, accompanied by increased regularity of the OLCS. This implies that the speed of bone deposition during bone remodeling would affect the regularity of the OLCS. While DMP1 was evenly localized in all regions of the cortical bones, FGF23 was more abundantly localized in osteocytes of cortical bones with regularly arranged OLCS. In cortical bones, the endosteal area featuring regular OLCS exhibited more intense FGF23 immunoreaction when compared to the periosteal region, which tended to display irregular OLCS. In summary, FGF23 appears to be synthesized principally by osteocytes in the regularly distributed OLCS that have been established after bone remodeling.

Clinical study on prognostic factors for autotransplantation of teeth with complete root formation

Autotransplantation is often performed to replace a missing tooth, but tooth autotransplantation has been reported in fewer teeth with complete root formation than those with incomplete root formation. The aim of this prospective study was to evaluate the factors that affect the prognosis of autotransplantation of teeth with complete root formation. 109 patients with 117 transplants were studied. Of the 117 transplants investigated, 14 (12%) failed during the observation period. The overall 1-year survival rate was 96%; the 5-year survival rate was 84%. The major causes of failure were unsuccessful initial healing and replacement root resorption with periodontal inflammation. Factors significantly associated with unsuccessful transplantation, in single factor analysis, were age 40 years or more, molar tooth as donor, probing pocket depth to 4mm or more, history of root canal treatment, multi-rooted teeth and fixation with sutures. Pocket depth of 4mm or more and history of root canal treatment appeared to increase the risk of unsuccessful transplantation in multivariate analysis. It is suggested that the pocket depth of the donor tooth and history of root canal treatment are related to the healing of paratransplantal tissue and root resorption.

A clinical study of alveolar bone tissue engineering with cultured autogenous periosteal cells: Coordinated activation of bone formation and resorption

In ongoing clinical research into the use of cultured autogenous periosteal cells (CAPCs) in alveolar bone regeneration, CAPCs were grafted into 33 sites (15 for alveolar ridge augmentation and 18 for maxillary sinus lift) in 25 cases. CAPCs were cultured for 6weeks, mixed with particulate autogenous bone and platelet-rich plasma, and then grafted into the sites. Clinical outcomes were determined from high-resolution three-dimensional computed tomography (3D-CT) images and histological findings. No serious adverse events were attributable to the use of grafted CAPCs. Bone regeneration was satisfactory even in cases of advanced atrophy of the alveolar process. Bone biopsy after bone grafting with CAPCs revealed prominent recruitment of osteoblasts and osteoclasts accompanied by angiogenesis around the regenerated bone. 3D-CT imaging suggested that remodeling of the grafted autogenous cortical bone particles was faster in bone grafting with CAPCs than in conventional bone grafting. The use of CAPCs offers cell-based bone regeneration therapy, affording complex bone regeneration across a wide area, and thus expanding the indications for dental implants. Also, it enables the content of particulate autogenous bone in the graft material to be reduced to as low as 40%, making the procedure less invasive, or enabling larger amounts of graft materials to be prepared. It may also be possible to dispense with the use of autogenous bone altogether in the future. The results suggest that CAPC grafting induces bone remodeling, thereby enhancing osseointegration and consequently reducing postoperative waiting time after dental implant placement.

A local bone anabolic effect of rhFGF2-impregnated gelatin hydrogel by promoting cell proliferation and coordinating osteoblastic differentiation.

UNLABELLED The bone anabolic effect of rhFGF2 is attributed to activation of proliferation and differentiation of osteoblasts. Concomitant up-regulation of Runx2 and Bmp2 implies a coordinative function of FGF/FGFR signaling on osteoblast differentiation. INTRODUCTION Duration and tissue concentration of growth factor exposure are important in tissue regeneration. This study analyzed the availability of rhFGF2 using a sustained release gelatin hydrogel system. To examine biological aspects of the bone anabolic effect, we carried out morphological and cell proliferation assays together with gene expression analyses of osteoblast related genes induced by rhFGF2 using localizing and quantifying procedures in vivo. MATERIALS AND METHODS Bone formation induced by implantation of gelatin hydrogel impregnated with 20 microg rhFGF2 (rhFGF2(+)) onto mice maxillae was analyzed by micro computed tomography, proliferating cell nuclear antigen (PCNA) immunohistochemistry, in situ hybridization and quantitative real time polymerase chain reaction combined with laser microdissection (LMD-QPCR). RESULTS The bony maxilla was augmented to 1.58 times its original volume (p=0.002) by the implantation of rhFGF2(+) gelatin hydrogel. An increased number of PCNA-positive nuclei were observed among differentiated osteoblasts as well as undifferentiated mesenchymal cells. Fgfr1, Fgfr2 and Runx2 were shown to be co-expressed mainly in differentiated osteoblasts but also in osteoblast marker-negative spindle-shaped cells that were scattered within the outer layer of hyperplastic periosteum. LMD-QPCR revealed up-regulation of Bmp2 expression accompanied by increased transcription of Fgfr1, Fgfr2 and Runx2 by rhFGF2 controlled release. CONCLUSIONS rhFGF2 sustained release results in bone formation on the maxilla by positively regulating the expansion and differentiation of osteoblastic cells. It is suggested that FGF/FGFR signaling coordinates a bone anabolic effect by simultaneously activating RUNX2 and BMP2 pathways. The gelatin hydrogel system, which enables a sustained slow rate of release of rhFGF2 in tissue has advantages of optimizing bone regeneration.

Diagnostic value of integrin α3, β4, and β5 gene expression levels for the clinical outcome of tongue squamous cell carcinoma†

The objective of the current study was to identify biomarkers that reflect the clinical course of squamous cell carcinoma of the tongue (TSCC).

Intermittent PTH Administration Stimulates Pre‐Osteoblastic Proliferation Without Leading to Enhanced Bone Formation in Osteoclast‐Less c‐fos−/− Mice

This study aimed to investigate the behavior and ultrastructure of osteoblastic cells after intermittent PTH treatment and attempted to elucidate the role of osteoclasts on the mediation of PTH‐driven bone anabolism. After administering PTH intermittently to wildtype and c‐fos−/− mice, immunohistochemical, histomorphometrical, ultrastructural, and statistical examinations were performed. Structural and kinetic parameters related to bone formation were increased in PTH‐treated wildtype mice, whereas in the osteoclast‐deficient c‐fos−/− mice, there were no significant differences between groups. In wildtype and knockout mice, PTH administration led to significant increases in the number of cells double‐positive for alkaline phosphatase and BrdU, suggesting active pre‐osteoblastic proliferation. Ultrastructural examinations showed two major pre‐osteoblastic subtypes: one rich in endoplasmic reticulum (ER), the hypER cell, and other with fewer and dispersed ER, the misER cell. The latter constituted the most abundant preosteoblastic phenotype after PTH administration in the wildtype mice. In c‐fos−/− mice, misER cells were present on the bone surfaces but did not seem to be actively producing bone matrix. Several misER cells were shown to be positive for EphB4 and were eventually seen rather close to osteoclasts in the PTH‐administered wildtype mice. We concluded that the absence of osteoclasts in c‐fos−/− mice might hinder PTH‐driven bone anabolism and that osteoclastic presence may be necessary for full osteoblastic differentiation and enhanced bone formation seen after intermittent PTH administration.