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    Akira Ito

    ABSTRACT Skeletal muscular tissues were constructed using magnetic force-based tissue engineering (Mag-TE) techniques. Mouse myoblast C2C12 cells labeled with magnetite cationic liposomes (MCLs) were seeded into a well of 24-well... more
    ABSTRACT Skeletal muscular tissues were constructed using magnetic force-based tissue engineering (Mag-TE) techniques. Mouse myoblast C2C12 cells labeled with magnetite cationic liposomes (MCLs) were seeded into a well of 24-well ultra-low cell attachment culture plates. When a magnet was positioned underneath the well, cells accumulated evenly onto the culture surface and formed a multilayered cell sheet. Furthermore, because an angiogenic potential of transplants is considered to be important for the long-term maintenance of cell survival and tissue functions, a vascular endothelial growth factor (VEGF) gene-modified C2C12 (C2C12/VEGF) cell sheets were also fabricated by the Mag-TE technique. The secretion level of C2C12/VEGF sheets was 3.0 ng/day, indicating that VEGF gene-expressing cell sheets were successfully fabricated. Since the shape of artificial tissue constructs can be controlled by magnetic force, a cellular string-like assembly was formed by placing a linear-shaped magnetic field concentrator with a magnet. These cellular sheets and strings shrank and did not maintain their shapes for an additional in vitro culture period during myogenic differentiation. On the other hand, when a silicone plug was positioned at the center of well during the fabrication of cell sheets, the cell sheets shrank and formed a ring-like assembly around the plug. After 6-d cultivation of cell rings in differentiation medium, the C2C12 cells differentiated to form multinucleated myotubes. Thus, these procedures can provide a novel strategy for skeletal muscular tissue engineering.
    The purpose of this study was to analyze histologic, biochemical, and biomechanical differences between zonal, regional, and anatomic locations of porcine menisci. We evaluated six menisci removed from pigs. Medial and lateral menisci... more
    The purpose of this study was to analyze histologic, biochemical, and biomechanical differences between zonal, regional, and anatomic locations of porcine menisci. We evaluated six menisci removed from pigs. Medial and lateral menisci were divided into three regions: anterior, middle, and posterior. In each portion, the central zone (CZ) and peripheral zone (PZ) were examined histologically (hematoxylin & eosin, safranin O/Fast green, and picrosiriusred staining), using scanning electron microscopy, biochemically (hydroxyproline assay for collagen content and dimethylmethylene blue assay for glycosaminoglycan [GAG] content), and biomechanically (compression testing). Collagen content in the CZ was lower than that in the PZ. GAG content in the CZ was higher than that in the PZ. GAG content in the PZ of the posterior portion was significantly higher than that in the anterior and middle portions. Compression strength in the CZ was higher than that in the PZ. The differences in cellular...
    Moyamoya disease (MMD) is a chronic occlusive cerebrovascular disease with unknown etiology that is characterized by the development of abnormal vascular networks at the base of the brain. Recent genome-wide studies identified RNF213 as... more
    Moyamoya disease (MMD) is a chronic occlusive cerebrovascular disease with unknown etiology that is characterized by the development of abnormal vascular networks at the base of the brain. Recent genome-wide studies identified RNF213 as an important MMD susceptibility gene. However, the exact mechanism by which the RNF213 abnormality leads to MMD remains unknown. Thus, we sought to clarify the role of RNF213 in angiogenesis under ischemic conditions using conventional RNF213 knockout mice. We assessed the infarction volume, cerebral edema, and vascular density in the ischemic brain after transient middle cerebral artery occlusion (tMCAO). To further evaluate systemic angiogenesis following chronic ischemia, we investigated blood flow recovery using laser speckle flowmetry, the severity of ambulatory impairments, and vascular density in the hind-limb after permanent femoral artery ligation. Results were compared between homozygous RNF213 knockout mice (RNF213 -/-) and wild-type litte...
    We studied 141 patients to evaluate the pathogenesis and clinical picture of high-risk unstable angina (UA), designated as impending myocardial infarction (IMI) in this study, or severe early post-infarction angina (PIA). IMI and PIA were... more
    We studied 141 patients to evaluate the pathogenesis and clinical picture of high-risk unstable angina (UA), designated as impending myocardial infarction (IMI) in this study, or severe early post-infarction angina (PIA). IMI and PIA were diagnosed when chest pain appeared at rest and lasted 15 min or more despite extensive pharmacological therapy during hospital stay among consecutive 510 patients with UA. All patients underwent coronary angiography urgently within 72 h after chest pain, and were divided into 2 subgroups according to ST segment shifts during chest pain. In IMI, 42 patients with ST depression had higher incidences of prior myocardial infarction (MI), worsening UA, multivessel disease and complex lesions such as eccentric irregular lesion or ulceration. On the contrary, in 44 with ST elevation, new onset UA, single vessel disease and coronary thrombus (CT) were dominant. In PIA, 32 patients with ST elevation revealed higher incidences in Q wave MI, ST elevation at the MI onset, single vessel disease and CT, compared to 23 with ST depression who showed a high proportion of complex lesions. Thus, it was evident that there was a common link between the pathogenesis of IMI and PIA. The therapeutic options were also different in the groups according to ST segment shift. We conclude that ST segment shifts during chest pain may be useful for determining the pathogenesis and clinical features of high-risk UA.
    Skeletal muscle regeneration requires migration, proliferation and fusion of myoblasts to form multinucleated myotubes. In our previous study, we showed that insulin-like growth factor (IGF)-I gene delivery stimulates the proliferation... more
    Skeletal muscle regeneration requires migration, proliferation and fusion of myoblasts to form multinucleated myotubes. In our previous study, we showed that insulin-like growth factor (IGF)-I gene delivery stimulates the proliferation and differentiation of mouse myoblast C2C12 cells and promotes the contractile force generated by tissue-engineered skeletal muscles. The aim of this study was to investigate the effects of the extracellular matrix on IGF-I gene-engineered C2C12 cells in vitro. Retroviral vectors for doxycycline (Dox)-inducible expression of the IGF-I gene were transduced into C2C12 cells. When cultured on a type IV collagen-coated surface, we observed significant increases in the migration speed and number of IGF-I gene-engineered C2C12 cells with Dox addition, designated as C2C12/IGF (+) cells. Co-culture of C2C12/IGF (+) cells and parental C2C12 cells, which had been cultured in differentiation medium for 3 days, greatly enhanced myotube formation. Moreover, type I...
    Hypofunction of the N-methyl-d-aspartate (NMDA) subtype of glutamate receptors may be involved in the pathophysiology of schizophrenia. Many studies have investigated peripheral NMDA receptor-related glutamatergic amino acid levels... more
    Hypofunction of the N-methyl-d-aspartate (NMDA) subtype of glutamate receptors may be involved in the pathophysiology of schizophrenia. Many studies have investigated peripheral NMDA receptor-related glutamatergic amino acid levels because of their potential as biological markers. Peripheral d-serine levels and the ratio of d-serine to total serine have been reported to be significantly lower in patients with schizophrenia than in controls. Peripheral d-serine levels and the d-/l-serine ratio have also been reported to significantly increase in patients with schizophrenia as their clinical symptoms improve from the time of admission to the time of discharge. In this study, we examined whether peripheral NMDA receptor-related glutamatergic amino acids levels were altered in patients with treatment-resistant schizophrenia compared to controls and whether these peripheral amino acids levels were altered by clozapine treatment. Twenty-two patients with treatment-resistant schizophrenia ...
    The computed tomography (CT) using the high-energy proton beam has the potential advantage of superior density resolution in the soft tissue analysis as compared to the conventional X-ray CT.We have developed the "first... more
    The computed tomography (CT) using the high-energy proton beam has the potential advantage of superior density resolution in the soft tissue analysis as compared to the conventional X-ray CT.We have developed the "first generation" type proton CT system using a 25-MeV proton beam to study the density distributions in small biomedical samples (<6 mm ϕ).The specimens were fixed in a cylindrical sample holder that was immersed in a water bath. The sample holder was scanned in front of the external semimicro beam of protons with a width of 0.06 mm at full width of half maximum (FWHM). The residual energy of the penetrated proton beam was analyzed with a solid state detector (SSD). The obtained integral density projection views were reconstructed into the density image by the filtered back projection method using the Shepp-Logan filter function.The spatial resolution of 0.1 mm and the density resolution of 0.1% were obtained by our system.The technique was applied to the eyes of small animals to study the changes in the density distributions of lenses, which develop with lens opacification in cataracts.
    Inhibitors of histone deacetylases (HDACs) are a promising class of anticancer agents that have an effect on gene regulation. The naturally occurring cyclic depsipeptide FK228 containing disulfide and Largazole possessing thioester... more
    Inhibitors of histone deacetylases (HDACs) are a promising class of anticancer agents that have an effect on gene regulation. The naturally occurring cyclic depsipeptide FK228 containing disulfide and Largazole possessing thioester functionalities act as pro-drugs and share the same HDAC inhibition mechanism in cell. Inspired from these facts, we have reported bicyclic tetrapeptide disulfide HDAC inhibitors resembling FK228 with potent activity and enhanced selectivity. In the present study, we report the design and synthesis of several mono and bicyclic tetrapeptide thioester HDAC inhibitors that share the inhibition mechanism similar to Largazole. Most of the compounds showed HDAC1 and HDAC4 inhibition and p21 promoting activity in nanomolar ranges. Among these the monocyclic peptides 1, 2 and bicyclic peptide, 4 are notable demanding more advanced research to be promising anticancer drug candidates.
    The induction of gastric tumor and intestinal metaplasia was examined in 8-week-old male JCL/SD rats exposed to localized X-irradiation of the gastric region. The animals were each given two 20 Gy fractions of X-rays, with a one-week... more
    The induction of gastric tumor and intestinal metaplasia was examined in 8-week-old male JCL/SD rats exposed to localized X-irradiation of the gastric region. The animals were each given two 20 Gy fractions of X-rays, with a one-week interval between fractions (total, 40 Gy). Nine atypical hyperplasias (20%) and 13 adenocarcinomas (28%) in the pyloric mucosa of the glandular stomach were found in 46 animals with X-irradiation. The incidence of intestinal metaplasia was 93% in the pyloric mucosa, 50% in the fundic mucosa and 96% in both the pyloric and fundic mucosa. Type B metaplasia (intestinal metaplasia without Paneth cells) was most common and type C (intestinal metaplasia with Paneth cells) was less frequent. No gastric tumor or intestinal metaplasia appeared in non-irradiated control rats. This study shows that local X-irradiation of the gastric region induced both gastric tumor and intestinal metaplasia independently.
    The naturally occurring cyclic depsipeptide, FK228 inhibits histone deacetylase (HDAC) enzymes after reductive cleavage of intra-molecular disulfide bond. One of the sulfhydryl groups produced in the reduction interacts with zinc atom... more
    The naturally occurring cyclic depsipeptide, FK228 inhibits histone deacetylase (HDAC) enzymes after reductive cleavage of intra-molecular disulfide bond. One of the sulfhydryl groups produced in the reduction interacts with zinc atom that involved in the catalytic mechanism of type 1 and 2 HDACs such as HDAC1, HDAC4, and HDAC6. In the present study, we describe the development of CHAP31, trapoxin B and HC-toxin based cyclic tetrapeptides with intra-molecular disulfide bond as HDAC inhibitors. The bicyclic tetrapeptides disulfide showed potent HDAC1 and HDAC4 inhibition and p21 promoting activity.
    Sumoylation is a post-translational modification that plays an important role in a wide range of cellular processes. Among the proteins involved in the sumoylation pathway, Ubc9 is the sole E2-conjugating enzyme required for sumoylation... more
    Sumoylation is a post-translational modification that plays an important role in a wide range of cellular processes. Among the proteins involved in the sumoylation pathway, Ubc9 is the sole E2-conjugating enzyme required for sumoylation and plays a central role by interacting with almost all of the partners required for sumoylation. Ubc9 has been implicated in a variety of human malignancies. In order to exploit the therapeutic potential of Ubc9, we have identified the potential site to target for rational drug design using molecular modeling approaches. The structural information derived was then used to prioritize hits from a small-molecule library for biological assay using a virtual screening protocol that involves shape matching with a known inhibitor inhibitors and docking of a small-molecule library utilizing computational approaches that incorporate both ligand and protein flexibility. Nineteen compounds were acquired from different chemical vendors and were tested for Ubc9 ...
    The present study was designed to examine the effect of the intestinal carcinogen 1,2-dimethylhydrazine (DMH) on grafted colorectal mucosa implanted into the glandular stomach of rats. Four groups were studied: Group 1 received the... more
    The present study was designed to examine the effect of the intestinal carcinogen 1,2-dimethylhydrazine (DMH) on grafted colorectal mucosa implanted into the glandular stomach of rats. Four groups were studied: Group 1 received the operation and DMH, Group 2 received the operation alone, Group 3 received DMH alone and Group 4 (controls) received only a sham operation. For Groups 1 and 2, about 8-mm diameter segments of colorectal tissue obtained from various sites in the large intestine of 8-week-old male F344 rats were isologously implanted into the fundic region of the stomachs of age-matched rats. DMH was injected at a dose of 20 mg/kg body weight i.m. per week for 20 weeks beginning 4 weeks after the operation. The animals were then observed for 8 months after the initial DMH treatment. In Group 1, adenocarcinomas developed in 41 of 60 successful implants (68%). Furthermore, poorly differentiated type tumors were observed in the grafts obtained from the rectum. This finding was contrary to that for intrinsic rectal tumors, all of which were well differentiated. The histochemical staining of mucin in the tissues from different sites of the large intestine revealed that sulfomucin, which normally exists essentially only in the intrinsic descending colon or rectum, was present in the grafts from the proximal ascending or ascending colon. No gastric tumors were observed in the control rats, which received either DMH or sham operations alone. Tumors in the intrinsic large intestine were observed only in rats that received DMH. These results indicate that colorectal mucosa implanted into the glandular stomach, like the intrinsic large intestine, is still sensitive to tumorigenesis caused by DMH.
    ... Bull. Amer. Me-teor. Soc., 65, 114–125. Kobayashi, C., K. Takano, S. Kusunoki, M. Sugi,and A. Kitoh, 2000: Seasonal predictability in win-ter over eastern Asia using the JMA global model. Quart. J. Roy. Meteor. Soc., 126, 2111– 2123.... more
    ... Bull. Amer. Me-teor. Soc., 65, 114–125. Kobayashi, C., K. Takano, S. Kusunoki, M. Sugi,and A. Kitoh, 2000: Seasonal predictability in win-ter over eastern Asia using the JMA global model. Quart. J. Roy. Meteor. Soc., 126, 2111– 2123. ...
    Cytochrome b5 is an integral membrane protein located on the outer surface of the endoplasmic reticulum (ER). This cytochrome is considered to be synthesized on free ribosomes and to be inserted post-translationally into the ER membrane,... more
    Cytochrome b5 is an integral membrane protein located on the outer surface of the endoplasmic reticulum (ER). This cytochrome is considered to be synthesized on free ribosomes and to be inserted post-translationally into the ER membrane, without participation of a signal recognition particle. To elucidate the signal responsible for targeting of cytochrome b5 to the ER membrane in vivo, DNAs encoding various derivatives of the cytochrome were constructed and introduced into cultured mammalian COS cells, and the subcellular distributions of the derivatives expressed in the cells were then analyzed. The deletion of more than 11 amino acid residues at the carboxy-terminal end of cytochrome b5 abolished the targeting and anchoring of the cytochrome to the ER membrane. Fusion proteins consisting of the carboxy-terminal 10 amino acid residues of cytochrome b5 and passenger proteins with the hydrophobic portion could be localized in the ER membrane. Thus, the last 10 amino acid residues of cytochrome b5 carry information necessary for the cytochrome to be targeted to the ER membrane.
    We previously reported the production of human erythropoietin (hEpo) using genetically manipulated (GM) chickens. The recombinant hEpo was produced in the serum and egg white of the GM chickens, and the oligosaccharide chain structures of... more
    We previously reported the production of human erythropoietin (hEpo) using genetically manipulated (GM) chickens. The recombinant hEpo was produced in the serum and egg white of the GM chickens, and the oligosaccharide chain structures of the serum-derived hEpo were more favorable than those of the egg white-derived hEpo. In the present study, a retroviral vector encoding an expression cassette for
    This report is a proceeding of 22 lectures presented at the study meeting on the Investigation and Control of Boundary-Layer Transition held four times from September 1987 to February 1989. It contains reports on the analysis of... more
    This report is a proceeding of 22 lectures presented at the study meeting on the Investigation and Control of Boundary-Layer Transition held four times from September 1987 to February 1989. It contains reports on the analysis of transition mechanism, three-dimensional boundary-layer transition, transition control, and other topics. This group consists of researchers participating in applications research for fundamental studies on boundary layer transition, with the aim of exchanging their opinions and discussing them thoroughly. Recently, lectures and discussions have been ranging over more subjects including not only transition problems but also analysis of turbulence flows and control methods to reduce viscous resistance.
    Infraciliature and morphogenesis of three rumen ophryoscolecid ciliates, Diplodinium polygonale Dogiel, 1925, Diplodinium leche Imai et al., 1992, and Diplodinium nanum Imai, 1988, are described from pyridinated silver... more
    Infraciliature and morphogenesis of three rumen ophryoscolecid ciliates, Diplodinium polygonale Dogiel, 1925, Diplodinium leche Imai et al., 1992, and Diplodinium nanum Imai, 1988, are described from pyridinated silver carbonate-impregnated specimens. These three species have two polybrachykineties in the buccal area and a polybrachykinety in the dorsal ciliary zone. The vestibular polybrachykinety (VP) of D. polygonale and D. leche arises from the dorsal extremity of the adoral polybrachykinety (AP) as in Entodinium species, extending toward the left in D. polygonale and toward the left posterior in D. leche. The VP of D. nanum arises from the inner side of the AP, separate from its dorsal extremity, as in other Diplodinium species and extends toward the left posterior. These series of the polybrachykinety arrangements in D. polygonale, D. leche, and D. nanum can be regarded as transitional forms in the evolution of an Entodinium-like ancestor to Diplodinium. Morphogenesis of these three Diplodinium species is not different from that of other Diplodinium species.
    The tumorigenicity of 9-3',4',5',6'-tetrachloro-o-carboxy phenyl-6-hydroxy-2,4,5,7-tetraiodo-3-isoxanthone sodium (CAS: 632-68-8) [also called Food Red 105 (FR 105) or Rose... more
    The tumorigenicity of 9-3',4',5',6'-tetrachloro-o-carboxy phenyl-6-hydroxy-2,4,5,7-tetraiodo-3-isoxanthone sodium (CAS: 632-68-8) [also called Food Red 105 (FR 105) or Rose Bengal B], which is widely used for food or cosmetic coloring in Japan, was examined in (C57BL/6N X C3H/N)F1 mice. Animals were divided into 6 groups with 50 mice in each group, and they were given solutions of 0, 0.125, and 0.5% FR 105 in the drinking water starting at 6 weeks of age and ending at 101 weeks. The mean relative thyroid weights in both sexes of mice given 0.125 and 0.5% FR 105 were significantly increased compared to those of controls. Enlarged thyroid glands were composed exclusively of colloid goiters characterized by distended follicles lined with flattened follicular cells. The male mice given 0.5% FR 105 had follicular adenomas in the thyroid gland at an incidence of 22.9%, which was significantly higher than the incidence in controls (P less than .005). Radioactive 131I uptake in colloid goiters was markedly decreased compared to that in intact thyroid glands. The results indicate that FR 105 can induce colloid goiters and thyroid adenomas in a dose-dependent manner.
    Surgical resection is the accepted standard of care for patients with non-small cell lung cancer (NSCLC). Several imaging modalities play central roles in the detection and staging of the disease. The aim of this review is to evaluate the... more
    Surgical resection is the accepted standard of care for patients with non-small cell lung cancer (NSCLC). Several imaging modalities play central roles in the detection and staging of the disease. The aim of this review is to evaluate the utility of computed tomography (CT), magnetic resonance imaging (MRI), positron emission tomography (PET) and PET/CT for NSCLC staging. Radiographic staging refers to the use of CT as a non-invasive diagnostic technique. However, while the vast majority of patients undergo only CT, CT is a notoriously inaccurate means of tumor and nodal staging in many situations. PET/CT clearly improves the staging, particularly nodal staging, compared to CT or PET alone. In addition, as a result of the increased soft-tissue contrast, MRI is superior to CT for distinguishing between tissue characteristics. Endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA), which is a minimally invasive technique, also has pathological diagnostic potential. Extensive research and the resultant improvements in the understanding of genetics, histology, molecular biology and oncology are transforming our understanding of lung cancer, and it is clear that imaging modalities such as CT, MRI, PET and PET/CT will have an important role in its preoperative management. However, thoracic surgeons should also be aware of the limitations of these techniques.
    We report herein a case of primary malignant fibrous histiocytoma (MFH) of the ileum. A 71-year-old Japanese man was admitted to our hospital with symptoms of abdominal pain and anorexia. Computed tomography, magnetic resonance imaging, a... more
    We report herein a case of primary malignant fibrous histiocytoma (MFH) of the ileum. A 71-year-old Japanese man was admitted to our hospital with symptoms of abdominal pain and anorexia. Computed tomography, magnetic resonance imaging, a follow-through study of the small intestine, and angiography all demonstrated a tumor of the ileum suggestive of a primary malignancy. A partial resection of the ileum was performed. It was histopathologically and immunohistochemically diagnosed to be a storiform-type primary MFH of the ileum with peritoneal dissemination. There have been a total of 25 cases of primary small bowel MFH documented in the Japanese or Western literature including our case. The malignant potential of such tumors is high, and the prognosis tends to be very poor. Unfortunately, we could not conclude whether the poor outcome was due to a delayed diagnosis or instead to its biological malignant behavior, since the number of such reported cases is still too small to make any definitive conclusions.
    The tumorigenicity of hydrogen peroxide (HP) was examined by administering 0.4% HP solution in drinking water to various strains of mice. In C57BL mice, gastric lesions in the forestomach occurred in over 67% of the mice treated with HP... more
    The tumorigenicity of hydrogen peroxide (HP) was examined by administering 0.4% HP solution in drinking water to various strains of mice. In C57BL mice, gastric lesions in the forestomach occurred in over 67% of the mice treated with HP for 120 days and duodenal lesions were noted in over 80% of the mice that received HP for 60 days. Gastric lesions were found only in the glandular stomach and duodenal lesions were restricted to the peri-pyloric and proximal part of the duodenum. The lesions in both organs were often multiple. The gastric lesions were composed of erosion and hyperplasia and those in the duodenum were exclusively hyperplasia. In mice given 0.4% and 0.1% HP for 420 to 740 days, 5 and 1% of them, respectively, had duodenal cancer by histological criteria though they did not show any distant metastasis. In the control group, no cancer was noted in the same observation period. The incidence and average numbers of lesions in both the stomach and duodenum after 150 to 210 days of HP intake decreased or fell to zero if HP administration was interrupted for 10 to 30 days. In DBA and BALB mice given 0.4% HP for 90 to 210 days, the incidences of gastric lesions reached 30 and 10% at maximum, respectively, and the incidences of duodenal lesions were 60 to 100% and 40 to 69%, respectively.
    Adrenal myelolipomas are rare benign tumors composed of mature adipose tissue and hematopoietic elements that resemble bone marrow. They are usually asymptomatic, and most cases are incidentally found at radiological examination or... more
    Adrenal myelolipomas are rare benign tumors composed of mature adipose tissue and hematopoietic elements that resemble bone marrow. They are usually asymptomatic, and most cases are incidentally found at radiological examination or autopsy. Symptoms such as abdominal pain and increasing girth occur only when the tumor grows large. We report the case of a giant adrenal myelolipoma in a 51-year old man who presented with a huge abdominal mass and abdominal pain. The resected tumor weighed 6,000 g and could represent the largest such tumor ever documented in the literature. We discuss the diagnosis and treatment of this unusual tumor.

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