Fumihiko Suwa

Oligodendrocyte Precursor Cells Support Blood-Brain Barrier Integrity via TGF-β Signaling

Trophic coupling between cerebral endothelium and their neighboring cells is required for the development and maintenance of blood-brain barrier (BBB) function. Here we report that oligodendrocyte precursor cells (OPCs) secrete soluble factor TGF-β1 to support BBB integrity. Firstly, we prepared conditioned media from OPC cultures and added them to cerebral endothelial cultures. Our pharmacological experiments showed that OPC-conditioned media increased expressions of tight-junction proteins and decreased in vitro BBB permeability by activating TGB-β-receptor-MEK/ERK signaling pathway. Secondly, our immuno-electron microscopic observation revealed that in neonatal mouse brains, OPCs attach to cerebral endothelial cells via basal lamina. And finally, we developed a novel transgenic mouse line that TGF-β1 is knocked down specifically in OPCs. Neonates of these OPC-specific TGF-β1 deficient mice (OPC-specific TGF-β1 partial KO mice: PdgfraCre/Tgfb1flox/wt mice or OPC-specific TGF-β1 total KO mice: PdgfraCre/Tgfb1flox/flox mice) exhibited cerebral hemorrhage and loss of BBB function. Taken together, our current study demonstrates that OPCs increase BBB tightness by upregulating tight junction proteins via TGF-β signaling. Although astrocytes and pericytes are well-known regulators of BBB maturation and maintenance, these findings indicate that OPCs also play a pivotal role in promoting BBB integrity.

A torque removal study on the primary stability of orthodontic titanium screw mini-implants in the cortical bone of dog femurs.

The aim of this study was to biomechanically evaluate the primary stability of pure titanium orthodontic mini-implants, inserted into pre-drilled cavities of differing diameters. Mini-implants (1.2 mm diameter) were placed into 1.0 mm and 1.2 mm diameter cavities prepared in the mid-region of the bilateral hind leg femurs of anesthetized beagles. Removal torque strengths were measured immediately, 1, 3, 6, 9 and 12 weeks post-insertion of the implant. For mini-implants placed into 1-mm cavities, removal torque values decrease over the first 6 weeks (p<0.01), after which values remained static. Average values obtained immediately, 1, 3 and 6 weeks post-insertion were 10.98, 8.83, 7.20 and 5.12 Ncm, respectively . Immediately post-insertion, removal torque values of mini-implants placed in a 1.2-mm cavity, were 11-fold lower than those placed in 1.0-mm cavities, which then demonstrated a significant increase in strength from 3 weeks (1.35 Ncm) to 6 weeks (5.17 Ncm) post-insertion (p<0.01). Measurements 6, 9 and 12 weeks post-insertion were similar to those in the 1.0-mm cavity. Initial stability of titanium mini-implants is considered necessary for immediate and early use in orthodontics, and an implant without this initial stability should be replaced or isolated until it develops the appropriate stability supported by osseointegration.

Reconstruction of a tissue‐engineered skin containing melanocytes

The objective of this study was to establish a new method for reconstruction of a tissue‐engineered skin containing melanocytes by employing tissue engineering. The keratinocytes, melanocytes and dermal fibroblasts were isolated and purified from human foreskin biopsies. Then the cells were used to construct a tissue‐engineered skin containing melanocytes. The localization of melanocytes in the tissue‐engineered skin was detected by DOPA staining, S‐100 immunohistochemical staining and transmission electron microscope (TEM). The results showed that the melanocytes could be detected in the basal layer of the constructed skin and the melanocytes showed dendritic morphology. Moreover the constructed skins were used to repair the athymic mice skin defects. Animal experiment results indicated that the skin equivalents could successfully repair full thickness skin defects in athymic mice and generated black skins by 6 weeks after grafting. Melanocytes located in the basal layer of the athymic mice skin could also be detected by using the S‐100 immunohistochemical staining. Our established method is useful to repair the full‐thickness skin defects.

In vitro construction of scaffold-free bilayered tissue-engineered skin containing capillary networks.

Many types of skin substitutes have been constructed using exogenous materials. Angiogenesis is an important factor for tissue-engineered skin constructs. In this study, we constructed a scaffold-free bilayered tissue-engineered skin containing a capillary network. First, we cocultured dermal fibroblasts with dermal microvascular endothelial cells at a ratio of 2 : 1. A fibrous sheet was formed by the interactions between the fibroblasts and the endothelial cells, and capillary-like structures were observed after 20 days of coculture. Epithelial cells were then seeded on the fibrous sheet to assemble the bilayered tissue. HE staining showed that tissue-engineered skin exhibited a stratified epidermis after 7 days. Immunostaining showed that the epithelium promoted the formation of capillary-like structures. Transmission electron microscopy (TEM) analysis showed that the capillary-like structures were typical microblood vessels. ELISA demonstrated that vascularization was promoted by significant upregulation of vascularization associated growth factors due to interactions among the 3 types of cells in the bilayer, as compared to cocultures of fibroblast and endothelial cells and monocultures.

Analysis of MRI findings in minimum invasive treatment for habitual temporomandibular joint dislocation by autologous blood injection around the temporomandibular joint capsule.

The aim of this study was to investigate magnetic resonance imaging (MRI) findings following autologous blood injection (ABI) for habitual temporomandibular joint (TMJ) dislocation. MRI was performed one hour and four and twelve weeks after ABI, revealing three types of significant findings. The first type was similar to hematoma and/or joint effusion in the articular capsule of the TMJ (type I). The second showed sporadic and diffuse T2 emphasis around the TMJ capsule (type II). The third involved a decreased range of condyle movement compared to before ABI (type III). Furthermore, we analyzed the three types of significant MRI findings. At one hour after ABI, type I was Grade 0 in 0 of 14 patients, Grade 1 in 8, Grade 2 in 2, and Grade 3 in 4. Type II was seen in 9 of the 14 cases and type III in 8. After twelve weeks, all cases of type I were Grade 0, no type II cases were evident, and type III was seen in 11 cases. Injecting autologous blood into surrounding TMJ tissues is an important factor in ABI. Minimally invasive treatment for habitual TMJ dislocation using ABI around the TMJ capsule appears to represent a very effective and safe treatment.

Comparative study of the ramification patterns of the subclavian branches as the subclavian artery passes in front or behind the scalenus anterior muscle

We investigated the ramification patterns of four subclavian branches (i.e., vertebral artery, internal thoracic artery, thyrocervical trunk, and costocervical trunk) as the subclavian artery passes in front or behind the scalenus anterior muscle. The investigation was carried out on 56 cadavers (112 cases) during student dissection practice sessions at Osaka Dental University. In 110 of the 112 cases, the subclavian artery passed behind the scalenus anterior muscle. The pattern of ramification of the subclavian branches in these cases was classified into six types (types A–F). In the remaining two cases (two cadavers), the subclavian artery passed in front of the scalenus anterior muscle. In both of these latter cases, the pattern of ramification of the subclavian branches differed from the six pattern types observed as the subclavian artery passed behind the scalenus anterior muscle: the first branch was the vertebral artery; the second, the costocervical trunk; the third, the thyrocervical trunk; the fourth, the internal thoracic artery. This same pattern of ramification was observed in three previously reported cases (two cadavers) in which the subclavian artery passed in front of the scalenus anterior muscle. Taken together, these observations indicate that the ramification pattern reported here and in a previous investigation for the subclavian artery passing in front of the scalenus anterior muscle is characteristic of this anatomical condition.

The Effect of Surface Roughness Difference on Bone Integration of Anodic Oxidized Ti Alloy Implants

At present, immediate loading of dental implants is of great interest. The key of this issue is how quickly implant can acquire firm fixation in the jaw bone, and how quickly the superstructure can be fabricated. In order to achieve these, it is necessary a large surface area of the implant and to promote early contact of the surrounding bone which can bear the load. In addition to this, anodic oxidization was carried out to prevent ion elution from titanium and titanium alloy in the body. The experiment was conducted with cylindrical Ti alloy test pieces of 4 different surface roughnesses (Ra=1.1μm, 1.2μm, 2.7μm, 4,7μm). These test pieces were treated by sandblasting and anodization, and implanted into the diaphysis of the femur of New Zealand white rabbits. Six weeks later, shear loading tests and histological examination were carried out. Strong interfacial bonding strength and active new bone formation were confirmed in the peripheral area of the test pieces having a surface roughness of Ra=2.7μm and 4,7μm. Judging from the stable fixation against the shear loading in the bone tissue seen in group C(Ra=2.7μm), it was

Transcriptional mRNA of bone morphogenetic proteins 2, 3, 4, and 5 in trigeminal nerve, benign and malignant peripheral nerve sheath tumors.

Bone morphogenetic proteins (BMPs) have been shown to play an important role in cell growth and differentiation. BMPs, a rapidly expanding family closely related to transforming growth factor-beta (TGF-beta) superfamily, have been proven recently to possess a regulatory role and neurotrophic capacity in neurogenesis. The aim of the present study is to reveal the relationship among BMPs, peripheral nerve and neoplastic lesions of nerve sheath tumors. The mRNA transcriptions of BMP 2, 3, 4 and 5 in 12 cases of schwannoma, four cases of malignant schwannoma and three cases of trigeminal neuralgia were detected using an in situ hybridization technique. Our results demonstrated that the myelin sheaths of schwann cell from the peripheral neuroectomy of trigeminal neuralgia were positively expressing mRNA of BMP-2, 3, 4 and 5. However, the nerve fibers of trigeminal nerve showed only BMP-2 positive staining. All of the neoplastic lesions of nerve sheath showed a consistent but variant expression of BMP-2, 3, 4, and 5. Except for the BMP-4 mRNA, the expression signals of BMP-2, 3 and 5 mRNA in malignant schwannoma were relatively lower than in benign lesions. On the basis of the findings, we concluded that selected members of BMPs existed in the peripheral nerves and might contribute to the health maintenance, proliferation, regeneration and neoplastic transformation of the peripheral nerve system. Moreover, the effects of BMP-2, 3, 4 and 5 on peripheral nerve system and its neoplastic transformation might be widespread, diverse and antagonistic.

Injury of Microvasculization on Differential Positions in the Rats Tongue Dorsum after Nd: YAG Constant power Laser Irradiation

The purpose of this study was to examine the depth into penetration of the tissue and the circulatory structure of the microvasculature after irradiation the Nd: YAG laser on the lingual dorsum of a 9-week-old male wistar rat. The Nd: YAG laser was irradiated against differential position on the lingual dorsum of a rat under the 3Watt for 1.0 second.Wounds of tongue dorsum were observed macroscopically, microscopically and also examined by SEM.The following results were obtained;Microscopical findings: The penetration of left tongue dorsum anterior group was observed still the musculi longitudinalis superior. The penetration of right margo linguae group was observed still the musculi longitudinalis superior. The penetration of tongue dorsum posterior group was observed still the musculi tranversus linguae superior.SEM findings: left tongue dorsum anterior group disappeared to the circulatory structure of the microvasculature. Right margo linguae group disappeared to the circulatory structure of the microvas culature. Right tongue dorsum posterior group disappeared to the lamina propria arteriolar and venular networks, arterioles and venulea were exposed.Irradiation power of the Nd: YAG laser need to change by differential position on the lingual dorsum of the rat.

2313 Age-related changes of tanycytes in the median eminence of the male rat

SHUJI HIDAKA, EM1 ITATEYAMA, SEIYA KONDO, SEIICHI CHIBA, TETSUYA KAKUYA, MAMORU KUROKAWA, HIRONOBU YOSHIMATSU, TOSHIIE SAKATA Since the cloning of the obese (ob) gene and ob receptor (diabetes, db) genes, evidence has been accumulating that the ob system is a key regulatory pathway associated with mammalian body weight homeostasis. We investigated that hypothalamic histamine content of the Zucker fa/fa rat and db/db mouse, in both of which abnormalities of leptin receptor had been detected, was significantly lower than that in lean littermate. Intracerebroventricular (ICV) administration of 1 pg leptin increased hypothalamic histamine turnover rate in male Wister King A rat. The pretreatment of a-fluoromethylhistidine, a specific suicide inhibitor of a histamine synthesizing decarboxylase enzyme (HDC), attenuated the anorexic effects of leptin ICV administration. In conclusion, our present observations suggest that hypothalamic histamine is one of the important target of leptin action in the brain.