Toshi-ichiro Tanabe

Pathologic and enzyme histochemical studies on bone formation induced by bone morphogenetic protein in mouse muscle tissue.

Bone morphogenetic protein (BMP) irreversibly induced the differentiation of mesenchymal-type cells into osteoprogenitor cells for endochondral ossification. During the process of BMP-induced differentiation in mice, 4 cell type (chondroblasts, osteoblasts, chondroclasts, and osteoclasts) were examined for phosphatase and succinate dehydrogenase using a wide range of buffers (4.0 less than or equal to pH less than or equal to 9.2). During the chondroid tissue-forming stage (1 week), chondroblast-like or osteoblast-like cells expressed phosphatase activity at 6.8 less than or equal to pH less than or equal to 9.2; chondroclast-like or osteoclast-like cells expressed phosphatase activity at 4.0 less than or equal to pH less than or equal to 5.8. However, mature chondrocytes found in hyaline cartilage expressed phosphatase activity between 6.6 less than or equal to pH less than or equal to 7.6 (2 weeks). During the process of endochondral ossification, alkaline phosphatase activity decreased in osteoblast-like cells with traces of acid phosphatase activity still detectable. Chondroclastic and osteoclastic giant cells were characterized by intense succinate dehydrogenase activity.

Mechanisms of the immunosuppressive effects of mouse adipose tissue-derived mesenchymal stromal cells on mouse alloreactively stimulated spleen cells

The mechanisms of immunomodulation by mesenchymal stromal cells remain poorly understood. In this study, the effects of mouse adipose tissue-derived mesenchymal stromal cells (ASCs) on mouse spleen cells alloreactively stimulated by anti-CD3 and anti-CD28 antibody-coated (anti-CD3/CD28) beads were observed. Production of interferon-γ by the anti-CD3/CD28 bead-stimulated spleen cells was significantly suppressed in co-culture with ASCs. However, an augmented intensity of CD69 on the stimulated spleen cells was not suppressed in the presence of ASCs. The immunosuppressive effects of ASCs were partially mediated by one or more soluble factors (26% suppression). However, the ASCs require cell-cell contact in order to maximally exert suppression (88%). The suppressive effect of ASCs mediated by direct cell contact was partially reversed following knockdown of β2 microglobulin, a component of the major histocompatibility complex (MHC) class I molecule, with siRNA. The results of the study demonstrated that ASCs have significant immune modulatory effects on alloreactively stimulated spleen cells. The effects of ASCs on spleen cells are dependent on soluble factor(s) and cell contact, which is mediated by the MHC class I complex on ASCs.

Immunohistochemical and Enzyme Histochemical Studies in Bone Formation Induced by Bone Morphogenetic Protein (BMP) in Mouse Muscle Tissue

It has been a well-known phenomenon that bone morphogenetic protein (BMP) heterotopically induces bone formation by causing the differentiation of osteoprogenitor cells from mesenchymal cells [1,2]. Two hypotheses of the cell origination and mechanism of bone formation by BMP have been proposed: EMP irreversibly induces the differentiation of perivascular mesenchymal cells into osteoprogenitor cells [3,4]; BMP is capable of stimulating the differentiation of skeletal muscle into cartilage [5–7]. The biological roles of bone formation induced by BMP still remain unclear. In the presnt study, the process of chondro-osteogenesis and cell differentiation for endochondral ossification by BMP implanted into mouse skeletal muscle tissue, were evaluated in terms of the immunohistochemical method for laminin, fibronectin, glycosaminoglycan, and S-100 protein, and the enzyme histochemical method of phosphatase activity with wide range of buffers (pH 4.0 to pH 9.2).

Laminin and fibronectin in bone formation induced by bone morphogenetic protein (BMP) in mouse muscle tissue

The morphological features at the light and electron microscopic level and distribution of fibronectin and laminin using immunohistochemical methods were evaluated in chondro-osteogneic tissue induced by bone morphogenetic protein (BMP) in mouse skeletal muscle. Satellite cells were seen at the muscle fibers adjacent to the site of BMP implantation 3 days after BMP-implantation. At the ultrastructural level, the skeletal muscle had a continuous basal lamina, satellite cells were partially enveloped by basal lamina-like structure, and fibroblast-like cells with abundant mitochondria and endoplasmic reticulum were adjacent to the myofibrils. An enhanced immunoreactivity for laminin and fibronectin was observed at the basement membrane of muscle fibers and extracellular matrix of neighboring connective tissue on day 3. On the 7th day, the enhanced immunoreactivity of laminin was persistent on the muscle fibers adjacent to the BMP and myoblast-like satellite cells, however, fibronectin showed decreased reaction. On the 14th day specimen, the newly formed osteo-chondroid matrix with numerous chondroid cells and osteoblasts showed positive immunoreaction for laminin and fibronectin. During the process of resorption of the induced osteoid tissue, no immunoreactivity for laminin and fibronectin was observed. An enhanced immunoreactivity of these glycoproteins during the early stages of BMP induced chondro-ossification suggests that they play an important role in the differentiation of myo-fibroblasts and organization and integration of ground substances of connective tissue and chondro-osseous matrix.

Guided Bone Regeneration, Platelet Rich Plasma and Low-Intensity Ultrasound Irradiation for Dental Implants

Because a concept of an osteointegrated dental implant system was established, prognosis of a dental implant at treatment improved and the treatment of restoration for missing teeth was changed. However, a dental implant treatment into atrophic jaw bone requires bone augmentation, obviously. At this time, we studied for the purpose of establishing the evidence of each method for clinical application of these bone augmentation method, such as guided bone regeneration (GBR), and autogenous bone block graft (BBG). In addition, we pursued the basic study of the evidence about the bone formation with platelet rich plasma (PRP) which recognized the availability in clinic. Furthermore, we present the results of basic studies which we tested for the purpose of applying a low-intensity pulse ultrasound (LIPU) irradiation applied to a fracture treatment in orthopedics area to intra-oral area, specially the condition after implant placement. In the results of comparison with GBR site and BBG, the differences of labeling bands were observed with a fluorescence microscopy. There was much labeling bands on GBR sections in comparison with BBG. This meaning that the bone remodeling around implants at GBR site was more active than BBG site. And the new bone formation by PRP was identified on soft X-ray graphically at first week after PRP applied mandible bone defect (experimental side). At same region of first week specimen, we confirmed positive reactions of platelet derived growth factor