metabolism of glycogen (glycogen, antibody, immunohistochemisry)
Book / Paper
Book:
1.
Otto Baba : 6, Ishiyaku Publishers,Inc., Tokyo, Feb. 2022.
(Keyword)
頭頸部の動脈 / 頭頸部の静脈
2.
Otto Baba and 落合 知子 : 医歯薬学系博物館事典, 雄山閣, Tokyo, May 2021.
3.
Otto Baba : 口腔解剖学 第2版, Ishiyaku Publishers,Inc., Tokyo, Feb. 2018.
(Keyword)
頭頸部の動脈 / 頭頸部の静脈
Academic Paper (Judged Full Paper):
1.
Yuchao Zhang, Yishan Sun, Jungang Shi, Peng Xu, Yingli Wang, Jianping Liu, Xinyu Gong, Yaru Wang, Yubin Tang, Haobo Liu, Xindi Zhou, Zhiqiao Lin, Otto Baba, Tsuyoshi Morita, Biao Yu and Lifeng Pan : Decoding the molecular mechanism of selective autophagy of glycogen mediated by autophagy receptor STBD1., Proceedings of the National Academy of Sciences of the United States of America, Vol.121, No.37, 2024.
(Summary)
Autophagy of glycogen (glycophagy) is crucial for the maintenance of cellular glucose homeostasis and physiology in mammals. STBD1 can serve as an autophagy receptor to mediate glycophagy by specifically recognizing glycogen and relevant key autophagic factors, but with poorly understood mechanisms. Here, we systematically characterize the interactions of STBD1 with glycogen and related saccharides, and determine the crystal structure of the STBD1 CBM20 domain with maltotetraose, uncovering a unique binding mode involving two different oligosaccharide-binding sites adopted by STBD1 CBM20 for recognizing glycogen. In addition, we demonstrate that the LC3-interacting region (LIR) motif of STBD1 can selectively bind to six mammalian ATG8 family members. We elucidate the detailed molecular mechanism underlying the selective interactions of STBD1 with ATG8 family proteins by solving the STBD1 LIR/GABARAPL1 complex structure. Importantly, our cell-based assays reveal that both the STBD1 LIR/GABARAPL1 interaction and the intact two oligosaccharide binding sites of STBD1 CBM20 are essential for the effective association of STBD1, GABARAPL1, and glycogen in cells. Finally, through mass spectrometry, biochemical, and structural modeling analyses, we unveil that STBD1 can directly bind to the Claw domain of RB1CC1 through its LIR, thereby recruiting the key autophagy initiation factor RB1CC1. In all, our findings provide mechanistic insights into the recognitions of glycogen, ATG8 family proteins, and RB1CC1 by STBD1 and shed light on the potential working mechanism of STBD1-mediated glycophagy.
(Keyword)
Autophagy / Humans / Glycogen / Autophagy-Related Protein 8 Family / Binding Sites / Adaptor Proteins, Signal Transducing / Protein Binding / Crystallography, X-Ray / Animals / Microtubule-Associated Proteins / Models, Molecular
Takafumi Kodama, Seiya Watanabe, Isana Kayanuma, Akira Sasaki, Daisuke Kurokawa, Otto Baba, Mitsuru Jimbo and Fumiya Furukawa : Gluconeogenesis during development of the grass puffer (Takifugu niphobles)., Comparative Biochemistry and Physiology. Part A: Molecular & Integrative Physiology, Vol.295, 2024.
(Summary)
During the development of teleost fish, the sole nutrient source is the egg yolk. The yolk consists mostly of proteins and lipids, with only trace amounts of carbohydrates such as glycogen and glucose. However, past evidence in some fishes showed transient increase in glucose during development, which may have supported the development of the embryos. Recently, we found in zebrafish that the yolk syncytial layer (YSL), an extraembryonic tissue surrounding the yolk, undergoes gluconeogenesis. However, in other teleost species, the knowledge on such gluconeogenic functions during early development is lacking. In this study, we used a marine fish, the grass puffer (Takifugu niphobles) and assessed possible gluconeogenic functions of their YSL, to understand the difference or shared features of gluconeogenesis between these species. A liquid chromatography (LC) / mass spectrometry (MS) analysis revealed that glucose and glycogen content significantly increased in the grass puffer during development. Subsequent real-time PCR results showed that most of the genes involved in gluconeogenesis increased in segmentation stages and/or during hatching. Among these genes, many were expressed in the YSL and liver, as shown by in situ hybridization analysis. In addition, glycogen immunostaining revealed that this carbohydrate source was accumulated in many tissues at segmentation stage but exclusively in the liver in hatched individuals. Taken together, these results suggest that developing grass puffer undergoes gluconeogenesis and glycogen synthesis during development, and that gluconeogenic activity is shared in YSL of zebrafish and grass puffer.
Tsuyoshi Morita, Shin Matsumoto and Otto Baba : Expression of secretory calcium-binding phosphoprotein (scpp) genes in medaka during the formation and replacement of pharyngeal teeth, BMC Oral Health, Vol.23, No.1, 744, 2023.
(Summary)
Analyses of tooth families and tooth-forming units in medaka with regard to tooth replacement cycles and the localization of odontogenic stem cell niches in the pharyngeal dentition clearly indicate that continuous tooth replacement is maintained. The secretory calcium-binding phosphoprotein (scpp) gene cluster is involved in the formation of mineralized tissues, such as dental and bone tissues, and the genes encoding multiple SCPPs are conserved in fish, amphibians, reptiles, and mammals. In the present study, we examined the expression patterns of several scpp genes in the pharyngeal teeth of medaka to elucidate their roles during tooth formation and replacement. Himedaka (Japanese medaka, Oryzias latipes) of both sexes (body length: 28 to 33 mm) were used in this study. Real-time quantitative reverse transcription-polymerase chain reaction (PCR) (qPCR) data were evaluated using one-way analysis of variance for multi-group comparisons, and the significance of differences was determined by Tukey's comparison test. The expression of scpp genes was examined using in situ hybridization (ISH) with a digoxigenin-labeled, single-stranded antisense probe. qPCR results showed that several scpp genes were strongly expressed in pharyngeal tissues. ISH analysis revealed specific expression of scpp1, scpp5, and sparc in tooth germ, and scpp5 was continually expressed in the odontoblasts of teeth attached to pedicles, but not in the osteoblasts of pedicles. In addition, many scpp genes were expressed in inner dental epithelium (ide), but not in odontoblasts, and scpp2 consistently showed epithelial-specific expression in the functional teeth. Taken together, these data indicate that specific expression of scpp2 and scpp5 may play a critical role in pharyngeal tooth formation in medaka. We characterized changes in the expression patterns of scpp genes in medaka during the formation and replacement of pharyngeal teeth.
(Keyword)
Humans / Animals / Oryzias / Calcium / Phosphoproteins / Odontogenesis / Bone and Bones / Mammals
C Heer E de, E C Zois, E Bridges, B der Vegt van, H Sheldon, A W Veldman, C M Zwager, T der Sluis van, S Haider, Tsuyoshi Morita, Otto Baba, P C Schröder, S Jong de, L A Harris and M Jalving : Glycogen synthase 1 targeting reveals a metabolic vulnerability in triple-negative breast cancer., Journal of Experimental & Clinical Cancer Research, Vol.42, No.1, 2023.
(Summary)
Our findings highlight GYS1 as potential therapeutic target in breast cancer, especially in TNBC and other highly proliferative subsets.
Katsuhiko Ono, Hitoshi Gotoh, Tadashi Nomura, Tsuyoshi Morita, Otto Baba, Mami Matsumoto, Sei Saitoh and Nobuhiko Ohno : Ultrastructural characteristics of oligodendrocyte precursor cells in the early postnatal mouse optic nerve observed by serial block-face scanning electron microscopy, PLoS ONE, Vol.17, No.12, 1-18, 2022.
(Summary)
Oligodendrocyte precursor cells (OPC) arise from restricted regions of the central nervous system (CNS) and differentiate into myelin-forming cells after migration, but their ultrastructural characteristics have not been fully elucidated. This study examined the three-dimensional ultrastructure of OPCs in comparison with other glial cells in the early postnatal optic nerve by serial block-face scanning electron microscopy. We examined 70 putative OPCs (pOPC) that were distinct from other glial cells according to established morphological criteria. The pOPCs were unipolar in shape with relatively few processes, and their Golgi apparatus were localized in the perinuclear region with a single cisterna. Astrocytes abundant in the optic nerve were distinct from pOPCs and had a greater number of processes and more complicated Golgi apparatus morphology. All pOPCs and astrocytes contained a pair of centrioles (basal bodies). Among them, 45% of pOPCs extended a short cilium, and 20% of pOPCs had centrioles accompanied by vesicles, whereas all astrocytes with basal bodies had cilia with invaginated ciliary pockets. These results suggest that the fine structures of pOPCs during the developing and immature stages may account for their distinct behavior. Additionally, the vesicular transport of the centrioles, along with a short cilium length, suggests active ciliogenesis in pOPCs.
E Christos Zois, M Anne Hendriks, Syed Haider, Elisabete Pires, Esther Bridges, Dimitra Kalamida, Dimitrios Voukantsis, Christoffer B Lagerholm, N Rudolf S Fehrmann, A Dunnen Wilfred F den, I Andrei Tarasov, Otto Baba, John Morris, M Francesca Buffa, O James S McCullagh, Mathilde Jalving and L Adrian Harris : Liver glycogen phosphorylase is upregulated in glioblastoma and provides a metabolic vulnerability to high dose radiation., Cell Death & Disease, Vol.13, No.6, 573, 2022.
(Summary)
Channelling of glucose via glycogen, known as the glycogen shunt, may play an important role in the metabolism of brain tumours, especially in hypoxic conditions. We aimed to dissect the role of glycogen degradation in glioblastoma (GBM) response to ionising radiation (IR). Knockdown of the glycogen phosphorylase liver isoform (PYGL), but not the brain isoform (PYGB), decreased clonogenic growth and survival of GBM cell lines and sensitised them to IR doses of 10-12 Gy. Two to five days after IR exposure of PYGL knockdown GBM cells, mitotic catastrophy and a giant multinucleated cell morphology with senescence-like phenotype developed. The basal levels of the lysosomal enzyme alpha-acid glucosidase (GAA), essential for autolysosomal glycogen degradation, and the lipidated forms of gamma-aminobutyric acid receptor-associated protein-like (GABARAPL1 and GABARAPL2) increased in shPYGL U87MG cells, suggesting a compensatory mechanism of glycogen degradation. In response to IR, dysregulation of autophagy was shown by accumulation of the p62 and the lipidated form of GABARAPL1 and GABARAPL2 in shPYGL U87MG cells. IR increased the mitochondrial mass and the colocalisation of mitochondria with lysosomes in shPYGL cells, thereby indicating reduced mitophagy. These changes coincided with increased phosphorylation of AMP-activated protein kinase and acetyl-CoA carboxylase 2, slower ATP generation in response to glucose loading and progressive loss of oxidative phosphorylation. The resulting metabolic deficiencies affected the availability of ATP required for mitosis, resulting in the mitotic catastrophy observed in shPYGL cells following IR. PYGL mRNA and protein levels were higher in human GBM than in normal human brain tissues and high PYGL mRNA expression in GBM correlated with poor patient survival. In conclusion, we show a major new role for glycogen metabolism in GBM cancer. Inhibition of glycogen degradation sensitises GBM cells to high-dose IR indicating that PYGL is a potential novel target for the treatment of GBMs.
Arief Waskitho, Yumiko Yamamoto, S Raman, Fumiya Kano, Huijiao Yan, R Raju, S Afroz, Tsuyoshi Morita, Daisuke Ikutame, Kazuo Okura, Masamitsu Ohshima, Akihito Yamamoto, Otto Baba and Yoshizo Matsuka : Peripherally Administered Botulinum Toxin Type A Localizes Bilaterally in Trigeminal Ganglia of Animal Model, Toxins, Vol.13, No.10, 704, 2021.
Huijiao Yan, Masamitsu Ohshima, Raju Raju, S Raman, Kazumitsu Sekine, Arief Waskitho, Miho Inoue, Masahisa Inoue, Otto Baba, Tsuyoshi Morita, Mayu Miyagi and Yoshizo Matsuka : Dentin-pulp complex tissue regeneration via three-dimensional cell sheet layering, Tissue Engineering. Part C, Methods, Vol.27, No.10, 559-570, 2021.
(Summary)
The dentin-pulp complex is a unique structure in teeth that contains both hard and soft tissues. Generally, deep caries and trauma cause damage to the dentin-pulp complex, and if left untreated, this damage will progress to irreversible pulpitis. The aim of this study was to fabricate a layered cell sheet composed of rat dental pulp (DP) cells and odontogenic differentiation of pulp (OD) cells and to investigate the ability to regenerate the dentin-pulp complex in a scaffold tooth. We fabricated two single cell sheets composed of DP cells (DP cell sheet) or OD cells (OD cell sheet) and a layered cell sheet made by layering both cells. The characteristics of the fabricated cell sheets were analyzed using light microscopy, scanning electron microscope (SEM), hematoxylin-eosin (HE) staining, and immunohistochemistry (IHC). Furthermore, the cell sheets were transplanted into the subrenal capsule of immunocompromised mice for 8 weeks. After this, the regenerative capacity to form dentin-like tissue was evaluated using micro-computed tomography (micro-CT), HE staining, and IHC. The findings of SEM and IHC confirmed that layered cell sheets fabricated by stacking OD cells and DP cells maintained their cytological characteristics. Micro-CT of layered cell sheet transplants revealed a mineralized capping of the access cavity in the crown area, similar to that of natural dentin. In contrast, the OD cell sheet group demonstrated the formation of irregular fragments of mineralized tissue in the pulp cavity, and the DP cell sheet did not develop any hard tissue. Moreover, bone volume/tissue volume (BV/TV) showed a significant increase in hard tissue formation in the layered cell sheet group compared with that in the single cell sheet group ( < 0.05). HE staining also showed a combination of soft and hard tissue formation in the layered cell sheet group. Furthermore, IHC confirmed that the dentin-like tissue generated from the layered cell sheet expressed characteristic markers of dentin but not bone equivalent to that of a natural tooth. In conclusion, this study demonstrates the feasibility of regenerating dentin-pulp complex using a bioengineered tissue designed to simulate the anatomical structure. Impact statement The dentin-pulp complex can be destroyed by deep caries and trauma, which may cause pulpitis and progress to irreversible pulpitis, apical periodontitis, and even tooth loss. Current treatments cannot maintain pulp health, and teeth can become brittle. We developed a three-dimensional (3D) layered cell sheet using dental pulp cells and odontogenic differentiation of pulp cells for dentin-pulp complex regeneration. Our layered cell sheet enables the regeneration of an organized 3D dentin-pulp-like structure comparable with that of natural teeth. This layered cell sheet technology may contribute to dentin-pulp complex regeneration and provide a novel method for complex tissue engineering.
Han Zhu, Zhang Weizhi, Ning Wanshan, Wang Chenwei, Deng Wankun, Li Zhidan, Shang Zehua, Shen Xiaofei, Liu Xiaohui, Otto Baba, Tsuyoshi Morita, Chen Lu, Xue Yu and Jia Da : Model-based analysis uncovers mutations altering autophagy selectivity in human cancer, Nature Communications, Vol.12, No.3258, 1-18, 2021.
(Summary)
Autophagy can selectively target protein aggregates, pathogens, and dysfunctional organelles for the lysosomal degradation. Aberrant regulation of autophagy promotes tumorigenesis, while it is far less clear whether and how tumor-specific alterations result in autophagic aberrance. To form a link between aberrant autophagy selectivity and human cancer, we establish a computational pipeline and prioritize 222 potential LIR (LC3-interacting region) motif-associated mutations (LAMs) in 148 proteins. We validate LAMs in multiple proteins including ATG4B, STBD1, EHMT2 and BRAF that impair their interactions with LC3 and autophagy activities. Using a combination of transcriptomic, metabolomic and additional experimental assays, we show that STBD1, a poorly-characterized protein, inhibits tumor growth via modulating glycogen autophagy, while a patient-derived W203C mutation on LIR abolishes its cancer inhibitory function. This work suggests that altered autophagy selectivity is a frequently-used mechanism by cancer cells to survive during various stresses, and provides a framework to discover additional autophagy-related pathways that influence carcinogenesis.
Tsuyoshi Kano, Tsuyoshi Morita, Kaori Sumida, Hiromichi Yumoto and Otto Baba : Expression of fibroblast growth factor receptor1, -2c, and -3c transcripts in mouse molars after tooth eruption, Anatomical Science International, 2021.
Resmi Raju, Masamitsu Ohshima, Miho Inoue, Tsuyoshi Morita, Yan Huijiao, Arief Waskitho, Otto Baba, Masahisa Inoue and Yoshizo Matsuka : Three-dimensional periodontal tissue regeneration using a bone-ligament complex cell sheet, Scientific Reports, Vol.10, No.1, 1656, 2020.
(Summary)
Periodontal tissue is a distinctive tissue structure composed three-dimensionally of cementum, periodontal ligament (PDL) and alveolar bone. Severe periodontal diseases cause fundamental problems for oral function and general health, and conventional dental treatments are insufficient for healing to healthy periodontal tissue. Cell sheet technology has been used in many tissue regenerations, including periodontal tissue, to transplant appropriate stem/progenitor cells for tissue regeneration of a target site as a uniform tissue. However, it is still difficult to construct a three-dimensional structure of complex tissue composed of multiple types of cells, and the transplantation of a single cell sheet cannot sufficiently regenerate a large-scale tissue injury. Here, we fabricated a three-dimensional complex cell sheet composed of a bone-ligament structure by layering PDL cells and osteoblast-like cells on a temperature responsive culture dish. Following ectopic and orthotopic transplantation, only the complex cell sheet group was demonstrated to anatomically regenerate the bone-ligament structure along with the functional connection of PDL-like fibers to the tooth root and alveolar bone. This study represents successful three-dimensional tissue regeneration of a large-scale tissue injury using a bioengineered tissue designed to simulate the anatomical structure.
Mugen Koyama, Fumiya Furukawa, Yuka Koga, Shohei Funayama, Suehiro Furukawa, Otto Baba, Ching-Chun Lin, Pung-Pung Hwang, Shunsuke Moriyama and Sei-Ichi Okumura : Gluconeogenesis and glycogen metabolism during development of Pacific abalone, Haliotis discus hannai., American Journal of Physiology. Regulatory, Integrative and Comparative Physiology, Vol.318, No.3, R619-R633, 2020.
(Summary)
In lecithotrophic larvae, egg yolk nutrients are essential for development. Although yolk proteins and lipids are the major nutrient sources for most animal embryos and larvae, the contribution of carbohydrates to development has been less understood. In this study, we assessed glucose and glycogen metabolism in developing Pacific abalone, a marine gastropod mollusc caught and cultured in east Asia. We found that glucose and glycogen content gradually elevated in developing abalone larvae, and coincident expression increases of gluconeogenic genes and glycogen synthase suggested abalone larvae had activated gluconeogenesis and glycogenesis during this stage. At settling, however, glycogen sharply decreased, with concomitant increases in glucose content and expression of Pyg and G6pc, suggesting the settling larvae had enhanced glycogen conversion to glucose. A liquid chromatography-mass spectrometry (LC/MS)-based metabolomic approach that detected intermediates of these pathways further supported active metabolism of glycogen. Immunofluorescence staining and in situ hybridization suggested the digestive gland has an important role as glycogen storage tissue during settlement, while many other tissues also showed a capacity to metabolize glycogen. Finally, inhibition of glycolysis affected survival of the settling veliger larvae, revealing that glucose is, indeed, an important nutrient source in settling larvae. Our results suggest glucose and glycogen are required for proper energy balance in developing abalone and especially impact survival during settling.
Takuma Iwasa, S Afroz, Miho Inoue, Rieko Arakaki, Masamitsu Ohshima, R Raju, Arief Waskitho, Inoue Masahisa, Otto Baba and Yoshizo Matsuka : IL-10 and CXCL2 in trigeminal ganglia in neuropathic pain, Neuroscience Letters, Vol.703, 132-138, 2019.
Elizabeth Haythorne, Maria Rohm, Martijn de Bunt van, F Melissa Brereton, I Andrei Tarasov, S Thomas Blacker, Gregor Sachse, Mariana Dos Santos Silva, Raul Exposito Terron, Simon Davis, Otto Baba, Roman Fischer, R Michael Duchen, Patrik Rorsman, I James MacRae and M Frances Ashcroft : Diabetes causes marked inhibition of mitochondrial metabolism in pancreatic β-cells., Nature Communications, Vol.10, No.1, 2019.
(Summary)
Diabetes is a global health problem caused primarily by the inability of pancreatic β-cells to secrete adequate levels of insulin. The molecular mechanisms underlying the progressive failure of β-cells to respond to glucose in type-2 diabetes remain unresolved. Using a combination of transcriptomics and proteomics, we find significant dysregulation of major metabolic pathways in islets of diabetic βV59M mice, a non-obese, eulipidaemic diabetes model. Multiple genes/proteins involved in glycolysis/gluconeogenesis are upregulated, whereas those involved in oxidative phosphorylation are downregulated. In isolated islets, glucose-induced increases in NADH and ATP are impaired and both oxidative and glycolytic glucose metabolism are reduced. INS-1 β-cells cultured chronically at high glucose show similar changes in protein expression and reduced glucose-stimulated oxygen consumption: targeted metabolomics reveals impaired metabolism. These data indicate hyperglycaemia induces metabolic changes in β-cells that markedly reduce mitochondrial metabolism and ATP synthesis. We propose this underlies the progressive failure of β-cells in diabetes.
S Afroz, Rieko Arakaki, Takuma Iwasa, Masamitsu Ohshima, Maki Hosoki, Miho Inoue, Otto Baba, Yoshihiro Okayama and Yoshizo Matsuka : CGRP induces differential regulation of cytokines from satellite glial cells in trigeminal ganglia and orofacial nociception, International Journal of Molecular Sciences, Vol.20, No.3, 711, 2019.
(Summary)
Neuron-glia interactions contribute to pain initiation and sustainment. Intra-ganglionic (IG) secretion of calcitonin gene-related peptide (CGRP) in the trigeminal ganglion (TG) modulates pain transmission through neuron-glia signaling, contributing to various orofacial pain conditions. The present study aimed to investigate the role of satellite glial cells (SGC) in TG in causing cytokine-related orofacial nociception in response to IG administration of CGRP. For that purpose, CGRP alone (10 μL of 10 M), Minocycline (5 μL containing 10 μg) followed by CGRP with one hour gap (Min + CGRP) were administered directly inside the TG in independent experiments. Rats were evaluated for thermal hyperalgesia at 6 and 24 h post-injection using an operant orofacial pain assessment device (OPAD) at three temperatures (37, 45 and 10 °C). Quantitative real-time PCR was performed to evaluate the mRNA expression of IL-1β, IL-6, TNF-α, IL-1 receptor antagonist (IL-1RA), sodium channel 1.7 (NaV 1.7, for assessment of neuronal activation) and glial fibrillary acidic protein (GFAP, a marker of glial activation). The cytokines released in culture media from purified glial cells were evaluated using antibody cytokine array. IG CGRP caused heat hyperalgesia between 6-24 h (paired- test, < 0.05). Between 1 to 6 h the mRNA and protein expressions of GFAP was increased in parallel with an increase in the mRNA expression of pro-inflammatory cytokines IL-1β and anti-inflammatory cytokine IL-1RA and NaV1.7 (one-way ANOVA followed by Dunnett's post hoc test, < 0.05). To investigate whether glial inhibition is useful to prevent nociception symptoms, Minocycline (glial inhibitor) was administered IG 1 h before CGRP injection. Minocycline reversed CGRP-induced thermal nociception, glial activity, and down-regulated IL-1β and IL-6 cytokines significantly at 6 h (-test, < 0.05). Purified glial cells in culture showed an increase in release of 20 cytokines after stimulation with CGRP. Our findings demonstrate that SGCs in the sensory ganglia contribute to the occurrence of pain via cytokine expression and that glial inhibition can effectively control the development of nociception.
Furukawa Fumiya, Irachi Shotaro, Koyama Mugen, Otto Baba, Akimoto Hajime, Okumura Sei-ichi, Kagawa Hirohiko and Uchida Katsuhisa : Changes in glycogen concentration and gene expression levels of glycogen-metabolizing enzymes in muscle and liver of developing masu salmon., Comparative Biochemistry and Physiology. Part A: Molecular & Integrative Physiology, Vol.225, 72-84, 2018.
(Summary)
Glycogen, as an intracellular deposit of polysaccharide, takes important roles in energy balance of many animals. In fish, however, the role of glycogen during development is poorly understood. In the present study, we assessed changes in glycogen concentration and gene expression patterns of glycogen-metabolizing enzymes in developing masu salmon (Oncorhynchus masou masou), a salmonid species inhabiting west side of North Pacific Ocean. As we measured glycogen levels in the bodies and yolk sacs containing the liver separately, the glycogen concentration increased in both parts as the fish developed, whereas it transiently decreased in the yolk sac after hatching, implying glycogen synthesis and breakdown in these tissues. Immunofluorescence staining using anti-glycogen monoclonal antibody revealed localization of glycogen in the liver, muscle and yolk syncytial layer of the pre-hatching embryos and hatched larvae. In order to estimate glycogen metabolism in the fish, the genes encoding homologs of glycogen synthase (gys1 and gys2) and glycogen phosphorylase (pygma, pygmb and pygl) were cloned, and their expression patterns were assessed by quantitative PCR and in situ hybridization. In the fish, gys1 and gys2 were robustly expressed in the muscle and liver, respectively. Also, expression of pyg isoforms was found in muscle, liver and yolk syncytial layer during hatching. With changes in glycogen concentration and expression patterns of relevant genes, our results suggest, for the first time, possible involvement of glycogen in energy balance of fish embryos, especially during hatching.
(Keyword)
Glycogen synthase / Glycogen phosphorylase / energy metabolism / Fish development / Hatching
Kaori Sumida, Yoshinori Ando, Shinichiro Seki, Kikuji Yamashita, Akira Fujimura, Otto Baba and Seiichiro Kitamura : Anatomical status of the human palatopharyngeal sphincter and its functional implications., Surgical and Radiologic Anatomy : SRA, Vol.39, No.11, 1191-1201, 2017.
(Summary)
The PPS is a muscle distinct from the SCP. Its contraction produces Passavant's ridge and conceivably enhances the efficiency of velopharyngeal closure by pressing the salpingopharyngeal fold and the musculus uvulae ridge against the velum.
Shinichiro Seki, Kaori Sumida, Kikuji Yamashita, Otto Baba and Seiichiro Kitamura : Gross anatomical classification of the courses of the human lingual artery., Surgical and Radiologic Anatomy : SRA, Vol.39, No.2, 195-203, 2017.
(Summary)
The present study provides detailed information on the courses of lingual artery which will be of clinical importance especially in the super-selective arterial angiography.
(Keyword)
Gross anatomical classification / human lingual artery
Anthi Demetriadou, Julia Morales-Sanfrutos, Marianna Nearchou, Otto Baba, Kyriacos Kyriacou, W Edward Tate, Anthi Drousiotou and P Petros Petrou : Mouse Stbd1 is N-myristoylated and affects ER-mitochondria association and mitochondrial morphology., Journal of Cell Science, Vol.130, No.5, 903-915, 2017.
(Summary)
Starch binding domain-containing protein 1 (Stbd1) is a carbohydrate-binding protein that has been proposed to be a selective autophagy receptor for glycogen. Here, we show that mouse Stbd1 is a transmembrane endoplasmic reticulum (ER)-resident protein with the capacity to induce the formation of organized ER structures in HeLa cells. In addition to bulk ER, Stbd1 was found to localize to mitochondria-associated membranes (MAMs), which represent regions of close apposition between the ER and mitochondria. We demonstrate that N-myristoylation and binding of Stbd1 to glycogen act as major determinants of its subcellular targeting. Moreover, overexpression of non-myristoylated Stbd1 enhanced the association between ER and mitochondria, and further induced prominent mitochondrial fragmentation and clustering. Conversely, shRNA-mediated Stbd1 silencing resulted in an increase in the spacing between ER and mitochondria, and an altered morphology of the mitochondrial network, suggesting elevated fusion and interconnectivity of mitochondria. Our data unravel the molecular mechanism underlying Stbd1 subcellular targeting, support and expand its proposed function as a selective autophagy receptor for glycogen and uncover a new role for the protein in the physical association between ER and mitochondria.
Kikuji Yamashita, Dalkhsuren Shine-Od, Aldartsogt Dolgorsuren, Kaori Sumida, Tsuyoshi Morita and Otto Baba : Effects of Dietary Cure Containing AFIR (Antioxidant Far-Infrared Energy Radiating) Ceramics on Living Body of Hatchery Sea Bream (Pagrus Major), International Journal of Innovative Studies in Sciences and Engineering Technology, Vol.2, No.10, 7-16, 2016.
21.
Yuki Oe, Otto Baba, Hitoshi Ashida, C Kouichi Nakamura and Hajime Hirase : Glycogen distribution in the microwave-fixed mouse brain reveals heterogeneous astrocytic patterns., Glia, Vol.64, No.9, 1532-1545, 2016.
(Summary)
In the brain, glycogen metabolism has been implied in synaptic plasticity and learning, yet the distribution of this molecule has not been fully described. We investigated cerebral glycogen of the mouse by immunohistochemistry (IHC) using two monoclonal antibodies that have different affinities depending on the glycogen size. The use of focused microwave irradiation yielded well-defined glycogen immunoreactive signals compared with the conventional periodic acid-Schiff method. The IHC signals displayed a punctate distribution localized predominantly in astrocytic processes. Glycogen immunoreactivity (IR) was high in the hippocampus, striatum, cortex, and cerebellar molecular layer, whereas it was low in the white matter and most of the subcortical structures. Additionally, glycogen distribution in the hippocampal CA3-CA1 and striatum had a 'patchy' appearance with glycogen-rich and glycogen-poor astrocytes appearing in alternation. The glycogen patches were more evident with large-molecule glycogen in young adult mice but they were hardly observable in aged mice (1-2 years old). Our results reveal brain region-dependent glycogen accumulation and possibly metabolic heterogeneity of astrocytes. GLIA 2016;64:1532-1545.
Tsuyoshi Morita, Kaoru Fujikawa, Otto Baba and Shibata Shunichi : An in situ hybridization study of Hyaluronan synthase (Has) mRNA in developing mouse molar and incisor tooth germs, Gene Expression Patterns, Vol.21, No.1, 28-40, 2016.
(Summary)
Hyaluronan (HA) is a major constituent molecule in most extracellular matrices and is synthesized by Hyaluronan synthase (Has). In the present study, we examined expression patterns of Has1, -2, -3 mRNA in developing mouse molar and incisor tooth germs from embryonic day (E) 11.5 to postnatal day (P) 7, focusing on Hertwig's epithelial root sheath (HERS) and the apical bud in particular. Has1 mRNA expression was not detected in all tooth germs examined. Has2 mRNA was expressed in the surrounding mesenchyme from E12.0 to 18.0 in both molar and incisor tooth germs, but disappeared after birth. Meanwhile, Has3 mRNA was exclusively expressed within the enamel organ, especially in the inner enamel epithelium (IEE), stellate reticulum (SR), and stratum intermedium (SI) until the early bell stage at E16.0. Has3 mRNA disappeared as IEE differentiated into differentiating ameloblasts (dABs), but remained in SI until the root developmental stage of the molar tooth germ at P7. Has3 mRNA was also expressed in HERS until P7. In incisors, Has3 mRNA was expressed in the apical bud, especially in the transit-amplifying (TA) cell region from E16.0 to P7, and in the papillary layer (PL) adjacent to the mature enamel. These gene expression patterns suggested that Has3 is the main control factor for prenatal and postnatal HA synthesis of the tooth germ, and may in part regulate crown and root formation of the tooth germ, maintenance of stem cell niches in the apical bud as well as mineral transport in PL.
K Fujikawa, T Yokohama-Tamaki, T Morita, Otto Baba, C Qin and S Shibata : An in situ hybridization study of perlecan, DMP1, and MEPE in developing condylar cartilage of the fetal mouse mandible and limb bud cartilage., European Journal of Histochemistry, Vol.59, No.3, 2015.
(Summary)
The main purpose of this in situ hybridization study was to investigate mRNA expression of three bone/cartilage matrix components (perlecan, DMP1, and MEPE) in developing primary (tibial) and secondary (condylar) cartilage. Perlecan mRNA expression was first detected in newly formed chondrocytes in tibial cartilage at E13.0, but this expression decreased in hypertrophic chondrocytes at E14.0. In contrast, at E15.0, perlecan mRNA was first detected in the newly formed chondrocytes of condylar cartilage; these chondrocytes had characteristics of hypertrophic chondrocytes, which confirmed the previous observation that progenitor cells of developing secondary cartilage rapidly differentiate into hypertrophic chondrocytes. DMP1 mRNA was detected in many chondrocytes within the lower hypertrophic cell zone in tibial cartilage at E14.0. In contrast, DMP1 mRNA expression was only transiently detected in a few chondrocytes of condylar cartilage at E15.0. Thus, DMP1 may be less important in the developing condylar cartilage than in the tibial cartilage. Another purpose of this study was to test the hypothesis that MEPE may be a useful marker molecule for cartilage. MEPE mRNA was not detected in any chondrocytes in either tibial or condylar cartilage; however, MEPE immunoreactivity was detected throughout the cartilage matrix. Western immunoblot analysis demonstrated that MEPE antibody recognized two bands, one of 67 kDa and another of 59 kDa, in cartilage-derived samples. Thus MEPE protein may gradually accumulate in the cartilage, even though mRNA expression levels were below the limits of detection of in situ hybridization. Ultimately, we could not designate MEPE as a marker molecule for cartilage, and would modify our original hypothesis.
Robert de Weerd van, Alvaro M Berbís, Marrion Sparrius, J Janneke Maaskant, Maikel Boot, J Nanne Paauw, Nadine Vries de, Louis Boon, Otto Baba, Javier F Cañada, Jeroen Geurtsen, Jesús Jiménez-Barbero and J Ben Appelmelk : A murine monoclonal antibody to glycogen: characterization of epitope-fine specificity by saturation transfer difference (STD) NMR spectroscopy and its use in mycobacterial capsular -glucan research., ChemBioChem, Vol.16, No.6, 977-989, 2015.
(Summary)
Mycobacterium tuberculosis, the causative agent of tuberculosis (TB), is a major pathogen responsible for 1.5 million deaths annually. This bacterium is characterized by a highly unusual and impermeable cell envelope, which plays a key role in mycobacterial survival and virulence. Although many studies have focused on the composition and functioning of the mycobacterial cell envelope, the capsular -glucan has received relatively minor attention. Here we show that a murine monoclonal antibody (Mab) directed against glycogen cross-reacts with mycobacterial -glucans, polymers of (1-4)-linked glucose residues with (1-6)-branch points. We identified the Mab epitope specificity by saturation transfer difference NMR and show that the (1-4)-linked glucose residues are important in glucan-Mab interaction. The minimal epitope is formed by (linear) maltotriose. Notably, a Mycobacterium mutant lacking the branching enzyme GlgB does not react with the Mab; this suggests that the (1-6)-branches form part of the epitope. These seemingly conflicting data can be explained by the fact that in the mutant the linear form of the -glucan (amylose) is insoluble. This Mab was subsequently used to develop several techniques helpful in capsular -glucan research. By using a capsular glucan-screening methodology based on this Mab we were able to identify several unknown genes involved in capsular -glucan biogenesis. Additionally, we developed two methods for the detection of capsular -glucan levels. This study therefore opens new ways to study capsular -glucan and to identify possible targets for further research.
E Dorte Kristensen, H Peter Albers, Clara Prats, Otto Baba, B Jesper Birk and P Jørgen F Wojtaszewski : Human muscle fibre type-specific regulation of AMPK and downstream targets by exercise., The Journal of Physiology, Vol.593, No.8, 2053-2069, 2015.
(Summary)
AMP-activated protein kinase (AMPK) is a regulator of energy homeostasis during exercise. Studies suggest muscle fibre type-specific AMPK expression. However, fibre type-specific regulation of AMPK and downstream targets during exercise has not been demonstrated. We hypothesized that AMPK subunits are expressed in a fibre type-dependent manner and that fibre type-specific activation of AMPK and downstream targets is dependent on exercise intensity. Pools of type I and II fibres were prepared from biopsies of vastus lateralis muscle from healthy men before and after two exercise trials: (1) continuous cycling (CON) for 30 min at 69 ± 1% peak rate of O2 consumption (V O2 peak ) or (2) interval cycling (INT) for 30 min with 6 × 1.5 min high-intensity bouts peaking at 95 ± 2% V O2 peak . In type I vs. II fibres a higher β1 AMPK (+215%) and lower γ3 AMPK expression (-71%) was found. α1 , α2 , β2 and γ1 AMPK expression was similar between fibre types. In type I vs. II fibres phosphoregulation after CON was similar (AMPK(Thr172) , ACC(Ser221) , TBC1D1(Ser231) and GS(2+2a) ) or lower (TBC1D4(Ser704) ). Following INT, phosphoregulation in type I vs. II fibres was lower (AMPK(Thr172) , TBC1D1(Ser231) , TBC1D4(Ser704) and ACC(Ser221) ) or higher (GS(2+2a) ). Exercise-induced glycogen degradation in type I vs. II fibres was similar (CON) or lower (INT). In conclusion, a differentiated response to exercise of metabolic signalling/effector proteins in human type I and II fibres was evident during interval exercise. This could be important for exercise type-specific adaptations, i.e. insulin sensitivity and mitochondrial density, and highlights the potential for new discoveries when investigating fibre type-specific signalling.
Yosuke Takezawa, Otto Baba, Shinichi Kohsaka and Kazuyuki Nakajima : Accumulation of glycogen in axotomized adult rat facial motoneurons., Journal of Neuroscience Research, Vol.93, No.6, 913-921, 2015.
(Summary)
This study biochemically determined glycogen content in the axotomized facial nucleus of adult rats up to 35 days postinsult. The amounts of glycogen in the transected facial nucleus were significantly increased at 5 days postinsult, peaked at 7 days postinsult, and declined to the control levels at 21-35 days postinsult. Immunohistochemical analysis with antiglycogen antibody revealed that the quantity of glycogen granules in the axotomized facial nucleus was greater than that in the control nucleus at 7 days postinjury. Dual staining methods with antiglycogen antibody and a motoneuron marker clarified that the glycogen was localized mainly in motoneurons. Immunoblotting and quantification analysis revealed that the ratio of inactive glycogen synthase (GS) to total GS was significantly decreased in the injured nucleus at about 1-3 days postinsult and significantly increased from 7 to 14 days postinsult, suggesting that glycogen is actively synthesized in the early period postinjury but suppressed after 7 days postinsult. The enhanced glycogen at about 5-7 days postinsult is suggested to be responsible for the decrease in inactive GS levels, and the decrease of glycogen after 7 days postinsult is considered to be caused by increased inactive GS levels and possibly the increase in active glycogen phosphorylase.
(Keyword)
Animals / Axotomy / Disease Models, Animal / Facial Nucleus / Glial Fibrillary Acidic Protein / glucose / Glycogen / Glycogen Synthase / Male / Motor Neurons / Rats / Rats, Wistar / Time Factors
H Peter Albers, T Andreas J Pedersen, B Jesper Birk, E Dorte Kristensen, F Birgitte Vind, Otto Baba, Jane Nøhr, Kurt Højlund and P Jørgen F Wojtaszewski : Human muscle fiber type-specific insulin signaling: impact of obesity and type 2 diabetes., Diabetes, Vol.64, No.2, 485-497, 2014.
(Summary)
Skeletal muscle is a heterogeneous tissue composed of different fiber types. Studies suggest that insulin-mediated glucose metabolism is different between muscle fiber types. We hypothesized that differences are due to fiber type-specific expression/regulation of insulin signaling elements and/or metabolic enzymes. Pools of type I and II fibers were prepared from biopsies of the vastus lateralis muscles from lean, obese, and type 2 diabetic subjects before and after a hyperinsulinemic-euglycemic clamp. Type I fibers compared with type II fibers have higher protein levels of the insulin receptor, GLUT4, hexokinase II, glycogen synthase (GS), and pyruvate dehydrogenase-E1 (PDH-E1) and a lower protein content of Akt2, TBC1 domain family member 4 (TBC1D4), and TBC1D1. In type I fibers compared with type II fibers, the phosphorylation response to insulin was similar (TBC1D4, TBC1D1, and GS) or decreased (Akt and PDH-E1). Phosphorylation responses to insulin adjusted for protein level were not different between fiber types. Independently of fiber type, insulin signaling was similar (TBC1D1, GS, and PDH-E1) or decreased (Akt and TBC1D4) in muscle from patients with type 2 diabetes compared with lean and obese subjects. We conclude that human type I muscle fibers compared with type II fibers have a higher glucose-handling capacity but a similar sensitivity for phosphoregulation by insulin.
Dawud Abduweli, Otto Baba, J Makoto Tabata, Kazunori Higuchi, Hiroshi Mitani and Yoshiro Takano : Tooth replacement and putative odontogenic stem cell niches in pharyngeal dentition of medaka (Oryzias latipes)., Microscopy, Vol.63, No.2, 141-153, 2014.
(Summary)
The small-sized teleost fish medaka, Oryzias latipes, has as many as 1000 pharyngeal teeth undergoing continuous replacement. In this study, we sought to identify the tooth-forming units and determine its replacement cycles, and further localize odontogenic stem cell niches in the pharyngeal dentition of medaka to gain insights into the mechanisms whereby continuous tooth replacement is maintained. Three-dimensional reconstruction of pharyngeal epithelium and sequential fluorochrome labeling of pharyngeal bones and teeth indicated that the individual functional teeth and their successional teeth were organized in families, each comprising up to five generations of teeth and successional tooth germs, and that the replacement cycle of functional teeth was approximately 4 weeks. BrdU label/chase experiments confirmed the existence of clusters of label-retaining epithelial cells at the posterior end of each tooth family where the expression of pluripotency marker Sox2 was confirmed by in situ hybridization. Label-retaining cells were also identified in the mesoderm immediately adjacent to the posterior end of each tooth family. These data suggest the importance of existence of slow-cycling dental epithelial cells and Sox2 expressions at the posterior end of each tooth family to maintain continuous tooth formation and replacement in the pharyngeal dentition of medaka.
Otto Baba, S Masato Ota, Tatsuo Terashima, J Makoto Tabata and Yoshiro Takano : Expression of transcripts for fibroblast growth factor 18 and its possible receptors during postnatal dentin formation in rat molars., Odontology, Vol.103, No.2, 136-142, 2013.
(Summary)
Fibroblast growth factors (FGFs) regulate the proliferation and differentiation of various cells via their respective receptors (FGFRs). During the early stages of tooth development in fetal mice, FGFs and FGFRs have been shown to be expressed in dental epithelia and mesenchymal cells at the initial stages of odontogenesis and to regulate cell proliferation and differentiation. However, little is known about the expression patterns of FGFs in the advanced stages of tooth development. In the present study, we focused on FGF18 expression in the rat mandibular first molar (M1) during the postnatal crown and root formation stages. FGF18 signals by RT-PCR using cDNAs from M1 were very weak at postnatal day 5 and were significantly up-regulated at days 7, 9 and 15. Transcripts were undetectable by in situ hybridization (ISH) but could be detected by in situ RT-PCR in the differentiated odontoblasts and cells of the sub-odontoblastic layer in both crown and root portions of M1 at day 15. The transcripts of FGFR2c and FGFR3, possible candidate receptors of FGF18, were detected by RT-PCR and ISH in differentiated odontoblasts throughout postnatal development. These results suggest the continual involvement of FGF18 signaling in the regulation of odontoblasts during root formation where it may contribute to dentin matrix formation and/or mineralization.
S Shibata, Y Sakamoto, Otto Baba, C Qin, G Murakami and H B Cho : An immunohistochemical study of matrix proteins in the craniofacial cartilage in midterm human fetuses., European Journal of Histochemistry, Vol.57, No.4, e39, 2013.
(Summary)
Immunohistochemical localization of collagen types I, II, and X, aggrecan, versican, dentin matrix protein (DMP)-1, martix extracellular phosphoprotein (MEPE) were performed for Meckel's cartilage, cranial base cartilage, and mandibular condylar cartilage in human midterm fetuses; staining patterns within the condylar cartilage were compared to those within other cartilaginous structures. Mandibular condylar cartilage contained aggrecan; it also had more type I collagen and a thicker hypertrophic cell layer than the other two types of cartilage; these three characteristics are similar to those of the secondary cartilage of rodents. MEPE immunoreactivity was first evident in the cartilage matrix of all types of cartilage in the human fetuses and in Meckel's cartilage of mice and rats. MEPE immunoreactivity was enhanced in the deep layer of the hypertrophic cell layer and in the cartilaginous core of the bone trabeculae in the primary spongiosa. These results indicated that MEPE is a component of cartilage matrix and may be involved in cartilage mineralization. DMP-1 immunoreactivity first became evident in human bone lacunae walls and canaliculi; this pattern of expression was comparable to the pattern seen in rodents. In addition, chondroid bone was evident in the mandibular (glenoid) fossa of the temporal bone, and it had aggrecan, collagen types I and X, MEPE, and DMP-1 immunoreactivity; these findings indicated that chondroid bone in this region has phenotypic expression indicative of both hypertrophic chondrocytes and osteocytes.
Clara Prats, Alba Gomez-Cabello, Pernille Nordby, L Jesper Andersen, W Jørn Helge, Flemming Dela, Otto Baba and Thorkil Ploug : An optimized histochemical method to assess skeletal muscle glycogen and lipid stores reveals two metabolically distinct populations of type I muscle fibers., PLoS ONE, Vol.8, No.10, e77774, 2013.
(Summary)
Skeletal muscle energy metabolism has been a research focus of physiologists for more than a century. Yet, how the use of intramuscular carbohydrate and lipid energy stores are coordinated during different types of exercise remains a subject of debate. Controversy arises from contradicting data from numerous studies, which used different methodological approaches. Here we review the "pros and cons" of previously used histochemical methods and describe an optimized method to ensure the preservation and specificity of detection of both intramyocellular carbohydrate and lipid stores. For optimal preservation of muscle energy stores, air drying cryosections or cycles of freezing-thawing need to be avoided. Furthermore, optimization of the imaging settings in order to specifically image intracellular lipid droplets stained with oil red O or Bodipy-493/503 is shown. When co-staining lipid droplets with associated proteins, Bodipy-493/503 should be the dye of choice, since oil red O creates precipitates on the lipid droplets blocking the light. In order to increase the specificity of glycogen stain, an antibody against glycogen is used. The resulting method reveals the existence of two metabolically distinct myosin heavy chain I expressing fibers: I-1 fibers have a smaller crossectional area, a higher density of lipid droplets, and a tendency to lower glycogen content compared to I-2 fibers. Type I-2 fibers have similar lipid content than IIA. Exhaustive exercise lead to glycogen depletion in type IIA and IIX fibers, a reduction in lipid droplets density in both type I-1 and I-2 fibers, and a decrease in the size of lipid droplets exclusively in type I-1 fibers.
Yan Liu, Li Zeng, Keli Ma, Otto Baba, Pen Zheng, Yang Liu and Yin Wang : Laforin-malin complex degrades polyglucosan bodies in concert with glycogen debranching enzyme and brain isoform glycogen phosphorylase., Molecular Neurobiology, Vol.49, No.2, 645-657, 2013.
(Summary)
In Lafora disease (LD), the deficiency of either EPM2A or NHLRC1, the genes encoding the phosphatase laforin and E3 ligase, respectively, causes massive accumulation of less-branched glycogen inclusions, known as Lafora bodies, also called polyglucosan bodies (PBs), in several types of cells including neurons. The biochemical mechanism underlying the PB accumulation, however, remains undefined. We recently demonstrated that laforin is a phosphatase of muscle glycogen synthase (GS1) in PBs, and that laforin recruits malin, together reducing PBs. We show here that accomplishment of PB degradation requires a protein assembly consisting of at least four key enzymes: laforin and malin in a complex, and the glycogenolytic enzymes, glycogen debranching enzyme 1 (AGL1) and brain isoform glycogen phosphorylase (GPBB). Once GS1-synthesized polyglucosan accumulates into PBs, laforin recruits malin to the PBs where laforin dephosphorylates, and malin degrades the GS1 in concert with GPBB and AGL1, resulting in a breakdown of polyglucosan. Without fountional laforin-malin complex assembled on PBs, GPBB and AGL1 together are unable to efficiently breakdown polyglucosan. All these events take place on PBs and in cytoplasm. Deficiency of each of the four enzymes causes PB accumulation in the cytoplasm of affected cells. Demonstration of the molecular mechanisms underlying PB degradation lays a substantial biochemical foundation that may lead to understanding how PB metabolizes and why mutations of either EPM2A or NHLRC1 in humans cause LD. Mutations in AGL1 or GPBB may cause diseases related to PB accumulation.
Or Kakhlon, Hava Glickstein, Naomi Feinstein, Yan Liu, Otto Baba, Tatsuo Terashima, Orhan Hasan Akman, Salvatore Dimauro and Alexander Lossos : Polyglucosan neurotoxicity caused by glycogen branching enzyme deficiency can be reversed by inhibition of glycogen synthase., Journal of Neurochemistry, Vol.127, No.1, 101-113, 2013.
(Summary)
Uncontrolled elongation of glycogen chains, not adequately balanced by their branching, leads to the formation of an insoluble, presumably neurotoxic, form of glycogen called polyglucosan. To test the suspected pathogenicity of polyglucosans in neurological glycogenoses, we have modeled the typical glycogenosis Adult Polyglucosan Body Disease (APBD) by suppressing glycogen branching enzyme 1 (GBE1, EC 2.4.1.18) expression using lentiviruses harboring short hairpin RNA (shRNA). GBE1 suppression in embryonic cortical neurons led to polyglucosan accumulation and associated apoptosis, which were reversible by rapamycin or starvation treatments. Further analysis revealed that rapamycin and starvation led to phosphorylation and inactivation of glycogen synthase (GS, EC 2.4.1.11), dephosphorylated and activated in the GBE1-suppressed neurons. These protective effects of rapamycin and starvation were reversed by overexpression of phosphorylation site mutant GS only if its glycogen binding site was intact. While rapamycin and starvation induce autophagy, autophagic maturation was not required for their corrective effects, which prevailed even if autophagic flux was inhibited by vinblastine. Furthermore, polyglucosans were not observed in any compartment along the autophagic pathway. Our data suggest that glycogen branching enzyme repression in glycogenoses can cause pathogenic polyglucosan buildup, which might be corrected by GS inhibition.
Yin Wang, Keli Ma, Peixiang Wang, Otto Baba, Helen Zhang, M Jack Parent, Pan Zheng, Yang Liu, A Berge Minassian and Yan Liu : Laforin prevents stress-induced polyglucosan body formation and Lafora disease progression in neurons., Molecular Neurobiology, Vol.48, No.1, 49-61, 2013.
(Summary)
Glycogen, the largest cytosolic macromolecule, is soluble because of intricate construction generating perfect hydrophilic-surfaced spheres. Little is known about neuronal glycogen function and metabolism, though progress is accruing through the neurodegenerative epilepsy Lafora disease (LD) proteins laforin and malin. Neurons in LD exhibit Lafora bodies (LBs), large accumulations of malconstructed insoluble glycogen (polyglucosans). We demonstrated that the laforin-malin complex reduces LBs and protects neuronal cells against endoplasmic reticulum stress-induced apoptosis. We now show that stress induces polyglucosan formation in normal neurons in culture and in the brain. This is mediated by increased glucose-6-phosphate allosterically hyperactivating muscle glycogen synthase (GS1) and is followed by activation of the glycogen digesting enzyme glycogen phosphorylase. In the absence of laforin, stress-induced polyglucosans are undigested and accumulate into massive LBs, and in laforin-deficient mice, stress drastically accelerates LB accumulation and LD. The mechanism through which laforin-malin mediates polyglucosan degradation remains unclear but involves GS1 dephosphorylation by laforin. Our work uncovers the presence of rapid polyglucosan metabolism as part of the normal physiology of neuroprotection. We propose that deficiency in the degradative phase of this metabolism, leading to LB accumulation and resultant seizure predisposition and neurodegeneration, underlies LD.
Nahar Kamrun, Akemi Tetsumura, Yoshikazu Nomura, Satoshi Yamaguchi, Otto Baba, Shin Nakamura, Hiroshi Watanabe and Tohru Kurabayashi : Visualization of the superior and inferior borders of the mandibular canal: a comparative study using digital panoramic radiographs and cross-sectional computed tomography images., Oral Surgery, Oral Medicine, Oral Pathology and Oral Radiology, Vol.115, No.4, 550-557, 2013.
(Summary)
The visibility of the superior border was very poor on panoramic images. The use of cross-sectional CT images remarkably improved this poor visualization.
Li Zeng, Yin Wang, Otto Baba, Pan Zheng, Yang Liu and Yan Liu : Laforin is required for the functional activation of malin in endoplasmic reticulum stress resistance in neuronal cells., The FEBS Journal, Vol.279, No.14, 2467-2478, 2012.
(Summary)
Mutations in either EPM2A, the gene encoding a dual-specificity phosphatase named laforin, or NHLRC1, the gene encoding an E3 ubiquitin ligase named malin, cause Lafora disease in humans. Lafora disease is a fatal neurological disorder characterized by progressive myoclonus epilepsy, severe neurological deterioration and accumulation of poorly branched glycogen inclusions, called Lafora bodies or polyglucosan bodies, within the cell cytoplasm. The molecular mechanism underlying the neuropathogenesis of Lafora disease remains unknown. Here, we present data demonstrating that in the cells expressing low levels of laforin protein, overexpressed malin and its Lafora disease-causing missense mutants are stably polyubiquitinated. Malin and malin mutants form ubiquitin-positive aggregates in or around the nuclei of the cells in which they are expressed. Neither wild-type malin nor its mutants elicit endoplasmic reticulum stress, although the mutants exaggerate the response to endoplasmic reticulum stress. Overexpressed laforin impairs the polyubiquitination of malin while it recruits malin to polyglucosan bodies. The recruitment and activities of laforin and malin are both required for the polyglucosan body disruption. Consistently, targeted deletion of laforin in brain cells from Epm2a knockout mice increases polyubiquitinated proteins. Knockdown of Epm2a or Nhlrc1 in neuronal Neuro2a cells shows that they cooperate to allow cells to resist ER stress and apoptosis. These results reveal that a functional laforin-malin complex plays a critical role in disrupting Lafora bodies and relieving ER stress, implying that a causative pathogenic mechanism underlies their deficiency in Lafora disease.
Hiroko Ida-Yonemochi, Mitsushiro Nakatomi, Hidemitsu Harada, Hiroki Takata, Otto Baba and Hayato Ohshima : Glucose uptake mediated by glucose transporter 1 is essential for early tooth morphogenesis and size determination of murine molars., Developmental Biology, Vol.363, No.1, 52-61, 2011.
(Summary)
Glucose is an essential source of energy for body metabolism and is transported into cells by glucose transporters (GLUTs). Well-characterized class I GLUT is subdivided into GLUTs1-4, which are selectively expressed depending on tissue glucose requirements. However, there is no available data on the role of GLUTs during tooth development. This study aims to clarify the functional significance of class I GLUT during murine tooth development using immunohistochemistry and an in vitro organ culture experiment with an inhibitor of GLUTs1/2, phloretin, and Glut1 and Glut2 short interfering RNA (siRNA). An intense GLUT1-immunoreaction was localized in the enamel organ of bud-stage molar tooth germs, where the active cell proliferation occurred. By the bell stage, the expression of GLUT1 in the dental epithelium was dramatically decreased in intensity, and subsequently began to appear in the stratum intermedium at the late bell stage. On the other hand, GLUT2-immunoreactivity was weakly observed in the whole tooth germs throughout all stages. The inhibition of GLUTs1/2 by phloretin in the bud-stage tooth germs induced the disturbance of primary enamel knot formation, resulting in the developmental arrest of the explants and the squamous metaplasia of dental epithelial cells. Furthermore, the inhibition of GLUTs1/2 in cap-to-bell-stage tooth germs reduced tooth size in a dose dependent manner. These findings suggest that the expression of GLUT1 and GLUT2 in the dental epithelial and mesenchymal cells seems to be precisely and spatiotemporally controlled, and the glucose uptake mediated by GLUT1 plays a crucial role in the early tooth morphogenesis and tooth size determination.
(Keyword)
Animals / Biological Transport / Cell Line / Dose-Response Relationship, Drug / Enamel Organ / Epithelium / Female / Gene Expression Regulation, Developmental / glucose / Glucose Transporter Type 1 / Glucose Transporter Type 2 / immunohistochemistry / In Situ Hybridization / Male / Mice / Mice, Inbred ICR / Molar / Odontogenesis / Phloretin / Pregnancy / RNA Interference / Reverse Transcriptase Polymerase Chain Reaction / Time Factors / Tissue Culture Techniques / Tooth Germ
Keijiro Munakata, Kayoko Ookata, Hiroyuki Doi, Otto Baba, Tatsuo Terashima, Shigehisa Hirose and Akira Kato : Histological demonstration of glucose transporters, fructose-1,6-bisphosphatase, and glycogen in gas gland cells of the swimbladder: is a metabolic futile cycle operating?, Biochemical and Biophysical Research Communications, Vol.417, No.1, 564-569, 2011.
(Summary)
Luminal surface of the swimbladder is covered by gas gland epithelial cells and is responsible for inflating the swimbladder by generating O(2) from Root-effect hemoglobin that releases O(2) under acidic conditions. Acidification of blood is achieved by lactic acid secreted from gas gland cells, which are poor in mitochondria but rich in the glycolytic activity. The acidic conditions are locally maintained by a countercurrent capillary system called rete mirabile. To understand the regulation of anaerobic metabolism of glucose in the gas gland cells, we analyzed the glucose transporter expressed there and the fate of ATP generated by glycolysis. The latter is important because the ATP should be immediately consumed otherwise it strongly inhibits the glycolysis rendering the cells unable to produce lactic acid anymore. Expression analyses of glucose transporter (glut) genes in the swimbladder of fugu (Takifugu rubripes) by RT-PCR and in situ hybridization demonstrated that glut1a and glut6 are expressed in gas gland cells. Immunohistochemical analyses of metabolic enzymes demonstrated that a gluconeogenesis enzyme fructose-1,6-bisphosphatase (Fbp1) and a glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (Gapdh) are highly expressed in gas gland cells. The simultaneous catalyses of glycolysis and gluconeogenesis reactions suggest the presence of a futile cycle in gas gland cells to maintain the levels of ATP low and to generate heat that helps reduce the solubility of O(2).
Natsuko Sogabe, Rieko Maruyama, Otto Baba, Takayuki Hosoi and Masae Goseki-Sone : Effects of long-term vitamin K(1) (phylloquinone) or vitamin K(2) (menaquinone-4) supplementation on body composition and serum parameters in rats., Bone, Vol.48, No.5, 1036-1042, 2011.
(Summary)
Vitamin K is a cofactor for -glutamyl carboxylase, which is an essential enzyme for the -carboxylation of vitamin K-dependent proteins such as osteocalcin and matrix Gla protein. Although it has been suggested that vitamin K plays an important role in the improvement of bone metabolism, the relationship between dietary vitamin K intake and bone metabolism has not been thoroughly investigated. Moreover, vitamin K is thought to have other actions beyond influencing the -carboxylation status. In the present study, we examined the effects of the long-term addition of phylloquinone (PK) or menaquinone-4 (MK-4) to a control diet on bone mineral density, bone strength, body composition, and serum parameters in rats. A total of 23 female Sprague-Dawley strain rats (6 weeks old) were divided into three groups: basic control diet group, PK diet (PK: 600mg/kg diet) group, and MK diet (MK-4: 600mg/kg diet) group. Three months after starting the experimental diet, the addition of PK to the basic control diet significantly increased the bone mineral density (BMD) of the femur (p<0.05). In the MK group, there was no significant difference in the BMD of the femur. However, two types of bone strength parameter: the minimum cross-sectional moment of inertia and the polar moment of inertia, were significantly higher in the MK group than in the control (p<0.05, respectively). Furthermore, the femoral bone parameters (the width, dry weight and ash weight, and cortical, cancellous, trabecular, and total bone mineral contents) in the MK group were increased significantly compared with the control. Interestingly, the addition of PK or MK-4 significantly decreased the total fat accumulation (p<0.01 and p<0.05, respectively), and serum triglycerides were reduced by 48% in the PK group and 29% in the MK group compared with the control. There were no significant differences in the levels of serum calcium, phosphorus, alkaline phosphatase, growth hormone, insulin-like growth hormone-1, insulin-like growth hormone binding protein-3, and cross-linked N-teleopeptide of type I collagen among the three groups. This is the first study to demonstrate the effect of the long-term addition of PK or MK-4 to the control diet on body composition and serum parameters in an in vivo system using rats. Further studies on the mechanism of vitamin K supplementation in the regulation of bone metabolism would provide valuable data on the prevention of lifestyle-related disorders, including osteoporosis.
(Keyword)
Adipose Tissue / Animals / Blood Glucose / Body Composition / Bone Density / cholesterol / Cytokines / Diet / Dietary Supplements / Female / Femur / Hormones / Organ Size / Rats / Rats, Sprague-Dawley / Serum / Time Factors / Tomography, X-Ray Computed / Triglycerides / Vitamin K 1 / Vitamin K 2
Masud Ahmad, Hachiro Iseki, Dawud Abduweli, Otto Baba, J Makoto Tabata and Yoshiro Takano : Ultrastructural and histochemical evaluation of appositional mineralization of circumpulpal dentin at the crown- and root-analog portions of rat incisors., Journal of Electron Microscopy, Vol.60, No.1, 79-87, 2010.
(Summary)
Mineralization of circumpulpal dentin has been interpreted in such a way that predentin matrix is abruptly converted to almost fully mineralized dentin at the mineralization front. A group of investigators pointed out the existence of intermediary layer along the mineralization front of rat incisor dentin and claimed that dentin mineralization is a rather transient process. Owing to a paucity of information, however, the entity of transient mineralization of dentin has remained elusive. Here we confirmed the existence of a lightly mineralized layer (LL) along the mineralization front of rat incisor dentin, recognizable by both light and electron microscopy, in routinely processed specimens. LL less than 3 µm thick was shown to be located along the mineralization front of crown-analog dentin and tapered out toward the root analog of the incisor. Electron microscopy revealed that mineral deposition first occurred in the non-collagenous matrix of LL and that mineralization of collagen fibers took place sometime later at the conventional mineralization front. Microscopic appearance of the mineral phase of LL varied considerably depending on the histological processing of ultrathin sections, thus explaining the inconsistent interpretation of dentin mineralization in previous studies. These data suggest that mineralization of circumpulpal dentin in rat incisors proceeds in a stepwise or a transient manner, initiated by crystal deposition in the non-collagenous matrix followed by massive mineral deposition in collagen fibers at the mineralization front. The thickness of LL where only the non-collagenous matrix is mineralized may vary in relation to differences in the local non-collagenous matrix and also the rate of collagen mineralization in the respective portions of circumpulpal dentin.
Hiroto Nakayama, Kazuo Takakuda, N Hiroko Matsumoto, Atsushi Miyata, Otto Baba, J Makoto Tabata, Tatsuo Ushiki, Tsuyoshi Oda, D Marc McKee and Yoshiro Takano : Effects of altered bone remodeling and retention of cement lines on bone quality in osteopetrotic aged c-Src-deficient mice., Calcified Tissue International, Vol.86, No.2, 172-183, 2010.
(Summary)
Cement lines represent mineralized, extracellular matrix interfacial boundaries at which bone resorption by osteoclasts is followed by bone deposition by osteoblasts. To determine the contribution of cement lines to bone quality, the osteopetrotic c-Src mouse model-where cement lines accumulate and persist as a result of defective osteoclastic resorption-was used to investigate age-related changes in structural and mechanical properties of bone having long-lasting cement lines. Cement lines of osteopetrotic bones in c-Src knockout mice progressively mineralized with age up to the level that the entire matrix of cement lines was lost by EDTA decalcification. While it was anticipated that suppressed and abnormal remodeling, together with the accumulation of cement line interfaces, would lead to defective bone quality with advancing age of the mutant mice, unexpectedly, three-point bending tests of the long bones of 1-year-old c-Src-deficient mice indicated significantly elevated strength relative to age-matched wild-type bones despite the presence of numerous de novo microcracks. Among these microcracks in the c-Src bones, there was no sign of preferential propagation or arrest of microcracks along the cement lines in either fractured or nonfractured bones of old c-Src mice. These data indicate that cement lines are not the site of a potential internal failure of bone strength in aged c-Src osteopetrotic mice and that abundant and long-lasting cement lines in these osteopetrotic bones appear to have no negative impacts on the mechanical properties of this low-turnover bone despite their progressive hypermineralization (and thus potential brittleness) with age.
(Keyword)
aging patterrn / Animals / Bone Remodeling / Bone Resorption / Bone and Bones / Calcification, Physiologic / Disease Models, Animal / Disease Progression / extracellular matrix / Fractures, Bone / Mice / knockout mice / Osteoclasts / Osteopetrosis / Protein-Tyrosine Kinases / Tensile Strength / Weight-Bearing / src-Family Kinases
S A D Atukorala, Keiji Inohaya, Otto Baba, J Makoto Tabata, K R A R Ratnayake, Dawud Abduweli, Shohei Kasugai, Hiroshi Mitani and Yoshiro Takano : Scale and tooth phenotypes in medaka with a mutated ectodysplasin-A receptor: implications for the evolutionary origin of oral and pharyngeal teeth., Archives of Histology and Cytology, Vol.73, No.3, 139-148, 2010.
(Summary)
Ectodermal contribution to the induction of pharyngeal teeth that form in the endodermal territory of the oropharyngeal cavity in some teleost fishes has been a matter of considerable debate. To determine the role of ectodermal cell signaling in scale and tooth formation and thereby to gain insights in evolutionary origin of teeth, we analyzed scales and teeth in rs-3 medaka mutants characterized by reduced scale numbers due to aberrant splicing of the ectodysplasin-A receptor (edar). Current data show that, in addition to a loss of scales (83% reduction), a drastic loss of teeth occurred in both oral (43.5% reduction) and pharyngeal (73.5% reduction) dentitions in rs-3. The remaining scales of rs-3 were irregular in shape and nearly 3 times larger in size relative to those of the wild-type. In contrast, there was no abnormality in size and shape in the remaining teeth of rs-3. In wild-type medaka embryos, there was a direct contact between the surface ectoderm and rostral endoderm in pharyngeal regions before the onset of pharyngeal tooth formation. However, there was no sign of ectodermal cell migration in the pharyngeal endoderm and hence no direct evidence of any ectodermal contribution to pharyngeal odontogenesis. These data suggest differential roles for Eda-Edar signaling in the induction and growth of scales and teeth and support the intrinsic odontogenic competence of the rostral endoderm in medaka.
Jeroen Geurtsen, Sunita Chedammi, Joram Mesters, Marlène Cot, N Nicole Driessen, Tounkang Sambou, Ryo Kakutani, Roy Ummels, Janneke Maaskant, Hiroki Takata, Otto Baba, Tatsuo Terashima, Nicolai Bovin, E Christina M J Vandenbroucke-Grauls, Jérôme Nigou, Germain Puzo, Anne Lemassu, Mamadou Daffé and J Ben Appelmelk : Identification of mycobacterial alpha-glucan as a novel ligand for DC-SIGN: involvement of mycobacterial capsular polysaccharides in host immune modulation., The Journal of Immunology, Vol.183, No.8, 5221-5231, 2009.
(Summary)
Mycobacterium tuberculosis possesses a variety of immunomodulatory factors that influence the host immune response. When the bacillus encounters its target cell, the outermost components of its cell envelope are the first to interact. Mycobacteria, including M. tuberculosis, are surrounded by a loosely attached capsule that is mainly composed of proteins and polysaccharides. Although the chemical composition of the capsule is relatively well studied, its biological function is only poorly understood. The aim of this study was to further assess the functional role of the mycobacterial capsule by identifying host receptors that recognize its constituents. We focused on alpha-glucan, which is the dominant capsular polysaccharide. Here we demonstrate that M. tuberculosis alpha-glucan is a novel ligand for the C-type lectin DC-SIGN (dendritic cell-specific ICAM-3-grabbing nonintegrin). By using related glycogen structures, we show that recognition of alpha-glucans by DC-SIGN is a general feature and that the interaction is mediated by internal glucosyl residues. As for mannose-capped lipoarabinomannan, an abundant mycobacterial cell wall-associated glycolipid, binding of alpha-glucan to DC-SIGN stimulated the production of immunosuppressive IL-10 by LPS-activated monocyte-derived dendritic cells. By using specific inhibitors, we show that this IL-10 induction was DC-SIGN-dependent and also required acetylation of NF-kappaB. Finally, we demonstrate that purified M. tuberculosis alpha-glucan, in contrast to what has been reported for fungal alpha-glucan, was unable to activate TLR2.
Y Yagi, N Suda, Y Yamakoshi, Otto Baba and Keiji Moriyama : In vivo application of amelogenin suppresses root resorption., Journal of Dental Research, Vol.88, No.2, 176-181, 2009.
(Summary)
Amelogenin is recognized as an enamel protein associated with enamel formation. Besides this well-known function, remarkable root resorption has been seen in amelogenin-null mutant mice. Moreover, in vitro culture studies showed that amelogenin suppressed osteoclast differentiation. These studies raised the hypothesis that amelogenin can inhibit root resorption by reducing odontoclast number. To examine this hypothesis, we applied porcine amelogenins in a rat root resorption model, in which maxillary first molars were replanted after being air-dried. Compared with untreated and carrier-treated tooth roots, the application dramatically reduced the odontoclast number on root surfaces and inhibited cementum and root dentin resorption. Amelogenin significantly reduced the number of human odontoclastic cells in culture. It also inhibited RANKL expression in mouse bone marrow cell cultures. All these findings support our hypothesis that amelogenin application suppresses root resorption by inhibiting odontoclast number, and suggest that this is mediated by the regulation of RANKL expression.
Takuya Notani, J Makoto Tabata, Hachiro Iseki, Otto Baba and Yoshiro Takano : Introduction of a three-dimensional and layered (TDL) culture, a novel primary co-culture method for ameloblasts and pulp-derived cells., Archives of Histology and Cytology, Vol.72, No.3, 187-198, 2009.
(Summary)
The enamel organ engaged in enamel matrix formation in tooth germs comprises four different cell types: the ameloblasts, the cells of the stratum intermedium, stellate reticulum, and the outer enamel epithelium, each characterized by distinct structural features. In ordinary primary cultures of tooth-derived cells, these cells generally become flat in profile and hardly regain their original profiles comparable to those in vivo, even under conditions that can induce the expression of functional markers from these cells. To overcome this limitation inherent to the cell culture of tooth-derived cells, we introduced a novel co-culture method, a "three-dimensional and layered (TDL) culture", a three-dimensional (3D) culture of dental pulp-derived cells dispersed in type I collagen gel combined with a layered culture of enamel epithelial cells seeded on top of the gel to establish thereby a culture condition where the functional tooth-derived cells regain their original structures and spatial arrangements. We subjected the TDL gels thus prepared to floating cultures and found that, in the layered epithelial cells, those facing the 3D gel became cuboidal/short columnar in shape, showed cell polarity and well-developed intercellular junctions, had PAS positive material in their cytoplasm, and expressed a distinct immunoreactivity for cyotokeratin 14 and amelogenins. Pulpal cells in the gel displayed a strong ALP activity throughout the 3D gel. The current observations have clearly shown that the structural and functional features reminiscent of early secretory ameloblasts could be restored in the enamel organ-derived cells in a TDL culture.
Kazunori Higuchi, Peerapong Santiwong, Hitoshi Tamaki, Tatsuo Terashima, Hiroto Nakayama, Takuya Notani, Hachiro Iseki, Otto Baba and Yoshiro Takano : Development and terminal differentiation of pulp and periodontal nerve elements in subcutaneous transplants of molar tooth germs and incisors of the rat., European Journal of Oral Sciences, Vol.116, No.4, 324-333, 2008.
(Summary)
Ectopic tooth transplants are known to receive rich innervation of local neurons, but the precise location and structural features of neurites in the pulp and periodontal ligament (PDL) of such transplants are unclear. In this experiment, the molar tooth germs of rat embryos and incisors of young rats were subcutaneously transplanted into the dorsal regions of rats and processed, at various time intervals, for immunohistochemical demonstration of neural elements. Teeth with periodontal tissue elements developed in most of the molar transplants in 6 or 8 wk and received rich innervation, including some autonomic fibres, in the pulp. Nerve elements were also confirmed to be present in the PDL of these transplants, including specialized nerve ending-like structures reminiscent of the periodontal Ruffini endings. Mechanoreceptor-like structures were also induced in the regenerated PDL of similarly transplanted incisors, although the success rate was low. We conclude that rich and highly ordered innervation of the pulp, and occasional development of mechanoreceptors in the regenerated PDL of ectopic dental transplants, imply a high probability of successful induction of teeth with both nociceptive and mechanical sensations in the ectopic tooth and/or tooth germ transplant systems, although differentiation of mechanoreceptor-like nerve endings occurred in only a few rare cases.
Giannantonio Panza, Jan Stöhr, Eva Birkmann, Detlev Riesner, Dieter Willbold, Otto Baba, Tatsuo Terashima and Christian Dumpitak : Aggregation and amyloid fibril formation of the prion protein is accelerated in the presence of glycogen., Rejuvenation Research, Vol.11, No.2, 365-369, 2008.
(Summary)
Prion diseases like Creutzfeldt-Jakob disease in humans or scrapie in sheep and goats are infectious neurodegenerative diseases. Their infectious agent, called prion, is composed mainly of aggregated and misfolded prion protein and non-proteinaceous components. An example of such a common non-proteinaceous secondary component of natural prions is the polysaccharide scaffold. We studied the influence of such a polysaccharide on the conformational transition of PrP applying an in vitro conversion system. Here we report that glycogen supports and accelerates PrP amorphous aggregation similar to seeded aggregation and leads to co-aggregates. Furthermore, PrP fibril formation was highly accelerated in the presence of glycogen.
Shunichi Shibata, Otto Baba, Tsuyoshi Oda, Tamaki Yokohama-Tamaki, Chunlin Qin, T William Butler, Yasunori Sakakura and Yoshiro Takano : An immunohistochemical and ultrastructural study of the pericellular matrix of uneroded hypertrophic chondrocytes in the mandibular condyle of aged c-src-deficient mice., Archives of Oral Biology, Vol.53, No.3, 220-230, 2007.
(Summary)
These results indicate that long-term survived, uneroded hypertrophic chondrocytes, at least in a part, acquire osteocytic characteristics.
(Keyword)
Aggrecans / aging / Animals / Biomarkers / Chondrocytes / Collagen Type I / Collagen Type II / Collagen Type X / extracellular matrix / Extracellular Matrix Proteins / Glycoproteins / Hypertrophy / immunohistochemistry / Mandibular Condyle / Mice / Mice, Mutant Strains / transmission electron microscopy / Phosphoproteins / Proto-Oncogene Proteins pp60(c-src) / Versicans
Carmen M Louzao, Begoña Espiña, R Mercedes Vieytes, V Felix Vega, A Juan Rubiolo, Otto Baba, Tatsuo Terashima and M Luis Botana : "Fluorescent glycogen" formation with sensibility for in vivo and in vitro detection., Glycoconjugate Journal, Vol.25, No.6, 503-510, 2007.
(Summary)
There are presently many methods of detecting complex carbohydrates, and particularly glycogen. However most of them require radioisotopes or destruction of the tissue and hydrolysis of glycogen to glucose. Here we present a new method based on the incorporation of 2-NBDG (2-{N-[7-nitrobenz-2-oxa-1, 3-diazol 4-yl] amino}-2-deoxyglucose), a D-glucose fluorescent derivative, into glycogen. Two kinds of approaches were carried out by using Clone 9 rat hepatocytes as a cellular model; (1) Incubation of cell lysates with 2-NBDG, carbohydrate precipitation in filters and measurement of fluorescence in a microplate reader (2) Incubation of living hepatocytes with 2-NBDG and recording of fluorescence images by confocal microscopy. 2-NBDG labeled glycogen in both approaches. We confirmed this fact by comparison to the labeling obtained with a specific monoclonal anti-glycogen antibody. Also drugs that trigger glycogen synthesis or degradation induced an increase or decrease of fluorescence, respectively. This is a simple but efficient method of detecting glycogen with 2-NBDG. It could be used to record changes in glycogen stores in living cells and cell-free systems and opens the prospect of understanding the role of this important energy reserve under various physiological and pathophysiological conditions.
Chia-Hsi Joshua Chang, Su-Mei Wu, Yung-Che Tseng, Yi-Chun Lee, Otto Baba and Pung-Pung Hwang : Regulation of glycogen metabolism in gills and liver of the euryhaline tilapia (Oreochromis mossambicus) during acclimation to seawater., The Journal of Experimental Biology, Vol.210, No.Pt 19, 3494-3504, 2007.
(Summary)
Glucose, which plays a central role in providing energy for metabolism, is primarily stored as glycogen. The synthesis and degradation of glycogen are mainly initialized by glycogen synthase (GS) and glycogen phosphorylase (GP), respectively. The present study aimed to examine the glycogen metabolism in fish liver and gills during acute exposure to seawater. In tilapia (Oreochromis mossambicus) gill, GP, GS and glycogen were immunocytochemically colocalized in a specific group of glycogen-rich (GR) cells, which are adjacent to the gill's main ionocytes, mitochondrion-rich (MR) cells. Na+/K+-ATPase activity in the gills, protein expression and/or activity of GP and GS and the glycogen content of the gills and liver were examined in tilapia after their acute transfer from freshwater (FW) to 25 per thousand seawater (SW). Gill Na+/K+-ATPase activity rapidly increased immediately after SW transfer. Glycogen content in both the gills and liver were significantly depleted after SW transfer, but the depletion occurred earlier in gills than in the liver. Gill GP activity and protein expression were upregulated 1-3 h post-transfer and eventually recovered to the normal level as determined in the control group. At the same time, GS protein expression was downregulated. Similar changes in liver GP and GS protein expression were also observed but they occurred later at 6-12 h post-transfer. In conclusion, GR cells are initially stimulated to provide prompt energy for neighboring MR cells that trigger ion-secretion mechanisms. Several hours later, the liver begins to degrade its glycogen stores for the subsequent energy supply.
Atsushi Miyata, Otto Baba, Tsuyoshi Oda, Isao Ishikawa and Yoshiro Takano : Diverse effects of c-src deficiency on molar tooth development and eruption in mice., Archives of Histology and Cytology, Vol.70, No.1, 63-78, 2007.
(Summary)
C-src deficiency is characterized by osteopetrosis due to impaired bone resorption by hypofunctional osteoclasts and the resultant failure of tooth eruption. In preliminary observations, we frequently encountered erupted molars in c-src deficient mice unlike in other osteopetrotic animals. Here we examine the effects of c-src deficiency on the development of molar teeth with an emphasis on the spatial relation of growing teeth with the surrounding bones. In c-src deficient mice, the magnitude of tooth impaction differed considerably among the types of molars; all maxillary 1st molars were totally impacted deep in the alveolar sockets, whereas most mandibular 1st molars fully erupted into oral cavity. Distribution of osteoclasts in the alveolar bone was identical among all types of molars, and electron microscopy revealed signs of bone resorbing activity in these osteoclasts despite the absence of a ruffled border. From early development, the alveolar space was much narrower in the upper molar tooth germs than in the lower ones in both wild type and homozygous animals, and particularly so in the upper 1st molars. Current observations thus indicate a significant contribution of "hypofunctional osteoclasts" in c-src deficient mice in molar tooth development except for the upper 1st molars, which appear to require highly functional osteoclasts to gain sufficient space for them to grow normally. Taken together, these findings on the seemingly tooth-type specific effects of c-src deficiency on the development and eruption of molar teeth in c-src deficient mice can be attributed to the given differential spatial relation of the respective tooth germs with the surrounding bones in the presence of hypofunctional osteoclasts.
Yung-Che Tseng, Chang-Jen Huang, Chia-Hsi Joshua Chang, Wen-Yuan Teng, Otto Baba, Ming-Ji Fann and Pung-Pung Hwang : Glycogen phosphorylase in glycogen-rich cells is involved in the energy supply for ion regulation in fish gill epithelia., American Journal of Physiology. Regulatory, Integrative and Comparative Physiology, Vol.293, No.1, R482-91, 2007.
(Summary)
The molecular and cellular mechanisms behind glycogen metabolism and the energy metabolite translocation between mammal neurons and astrocytes have been well studied. A similar mechanism is proposed for rapid mobilization of local energy stores to support energy-dependent transepithelial ion transport in gills of the Mozambique tilapia (Oreochromis mossambicus). A novel gill glycogen phosphorylase isoform (tGPGG), which catalyzes the initial degradation of glycogen, was identified in branchial epithelial cells of O. mossambicus. Double in situ hybridization and immunocytochemistry demonstrated that tGPGG mRNA and glycogen were colocalized in glycogen-rich cells (GRCs), which surround ionocytes (labeled with a Na(+)-K(+)-ATPase antiserum) in gill epithelia. Concanavalin-A (a marker for the apical membrane) labeling indicated that GRCs and mitochondria-rich cells share the same apical opening. Quantitative real-time PCR analyses showed that tGPGG mRNA expression levels specifically responded to environmental salinity changes. Indeed, the glycogen content, glycogen phosphorylase (GP) protein level and total activity, and the density of tGPGG-expressing cells (i.e., GRCs) in fish acclimated to seawater (SW) were significantly higher than those in freshwater controls. Short-term acclimation to SW caused an evident depletion in the glycogen content of GRCs. Taken altogether, tGPGG expression in GRCs is stimulated by hyperosmotic challenge, and this may catalyze initial glycogen degradation to provide the adjacent ionocytes with energy to carry out iono- and osmoregulatory functions.
Otto Baba, Atsushi Miyata, Tatsuhiko Abe, Shunichi Shibata, Yukiko Nakano, Tatsuo Terashima, Tsuyoshi Oda, Akira Kudo and Yoshiro Takano : Formation of acellular cementum-like layers, with and without extrinsic fiber insertion, along inert bone surfaces of aging c-Src gene knockout mice., European Journal of Oral Sciences, Vol.114, No.6, 524-534, 2006.
(Summary)
To investigate the long-term effects of c-src deficiency on skeletal and dental tissues, we examined the lower jaws and long bones of c-src gene knockout (c-src KO) mice by histological and histochemical methods. Numerous multinucleated osteoclasts were distributed throughout the mandible in 5-wk-old c-src KO mice, but by 14 wk they had almost completely disappeared from the alveolar bone, leaving tartrate-resistant acid phosphatase (TRAP)-positive layers along the bone surface. Deposition of osteopontin-positive mineralized tissue, reminiscent of acellular afibrillar cementum (AAC), was confirmed along the TRAP-positive bone surface at 14 wk. The layer progressively thickened up to 21 months. A comparable mineralized layer was noted along the trabeculae of long bones as thickened cement lines. In the periostin-rich areas of jaw bones, but not in the long bones, portions of AAC-like mineralized layers were often replaced with and/or covered by acellular extrinsic fiber cementum (AEFC)-like tissue. These data suggest that the deposition of AAC-like mineralized tissue is a general phenomenon that may occur along inert or slowly remodeling bone surfaces under conditions characterized by reduced bone-resorbing activity, whereas the induction of AEFC-like tissue seems to be associated with the expression of certain molecules that are particularly abundant in the microenvironment of the periodontal ligament.
Yoshiyuki Nemoto, Kazunori Higuchi, Otto Baba, Akira Kudo and Yoshiro Takano : Multinucleate osteoclasts in medaka as evidence of active bone remodeling., Bone, Vol.40, No.2, 399-408, 2006.
(Summary)
Putative sites of bone resorption in the acellular bony skeleton of the medaka fish (Oryzias latipes) were investigated primarily by RNA in situ hybridization and histological analysis. Numerous cells that displayed intense enzymatic activity of tartrate-resistant acid phosphatase (TRAP), the main marker of osteoclasts, were distributed in the pharyngeal region of this fish. Moreover, these cells expressed cathepsin K, an osteoclast-specific gene, as well as the genes for TRAP and vacuolar-type proton ATPase (V-ATPase). Some of the TRAP-positive cells displayed all of the morphological characteristics equivalent to those of mammalian- and bird-type osteoclasts. These cells were associated primarily with the shedding teeth and their supporting bones (pedicles), where alkaline phosphatase (ALPase)-positive osteoblasts were also located, implying progressive bone remodeling associated with tooth replacement in these regions. In contrast, the inner aspects of the neural and hemal arches of the vertebral column, which were the only sites of bone resorption other than the tooth-bearing bones, showed sporadically aligned flat mononuclear TRAP-positive cells without a ruffled border, indicating a different mode of bone remodeling in these regions. These results suggest the feasibility of medaka as a model animal for the investigation of bone-related abnormalities and their genetic backgrounds.
Yoshiro Takano, Yukiko Nakano, Yoko Yamamoto-Shuda, Otto Baba and Tatsuo Terashima : Proteolysis on maturing enamel surface, as shown by gel-coating methods., European Journal of Oral Sciences, Vol.114 Suppl 1, 2006.
(Summary)
Degradation of enamel matrix proteins, and their removal during early maturation, is critical for the growth of large enamel crystals in the subsequent processes of enamel maturation. In this study, we sought to demonstrate, using in vivo zymography, the exact sites of proteolysis in maturing enamel and its relationship to the overlying ameloblasts. The maturing enamel surfaces of rat and bovine incisors were exposed and painted either with pre-exposed autoradiographic emulsion or with densely fluorescein-conjugated (DQ) gelatin. After a few hours, photographic development of the emulsion revealed alternate black and white banding patterns over the maturing enamel surface. DQ gelatin also revealed similar banding patterns of fluorescent and non-fluorescent regions. White, powdery areas of emulsion and fluorescent bands of DQ gelatin both corresponded to the areas of ruffle-ended ameloblasts, at least up to the mid stages of enamel maturation, implicating a predominant contribution of ruffle-ended ameloblasts in the degradation of enamel matrix proteins. Powdery white bands in autoradiographic emulsion shifted from the areas of ruffle-ended to smooth-ended ameloblasts in late maturation in both bovine and rat incisors and were not influenced by proteinase inhibitors or heat inactivation, implicating non-enzymatic interactions. DQ gelatin, in fact, did not generate any fluorescence in such smooth-ended ameloblast regions.
S C Young, S-W Kim, C Qin, Otto Baba, T W Butler, R R Taylor, D J Bartlett, P J Vacanti and C P Yelick : Developmental analysis and computer modelling of bioengineered teeth., Archives of Oral Biology, Vol.50, No.2, 259-265, 2005.
(Summary)
Here we present the developmental progression of bioengineered pig teeth from 1 to 25 weeks of development. We demonstrate that 2-25 week implants contained embryonic tooth bud- and cap-stage tooth structures consisting of dental epithelium expressing the sonic hedgehog gene and condensed dental mesenchyme. Implants harvested at 18-25 weeks also contained tooth bud-like structures, as well as mature tooth structures containing enamel, dentin and pulp tissues. Immunohistochemical analyses confirmed the expression of dentin- and enamel-specific proteins in differentiated bioengineered tooth tissues. Three-dimensional computer modelling further demonstrated a spatial organization of enamel, dentin and pulp tissues resembling that of natural teeth. We conclude that bioengineered teeth commonly exhibit morphological stages characteristic of naturally forming teeth. Furthermore, the presence of immature tooth buds at all times assayed and increased numbers of bioengineered tooth structures over time suggests that porcine dental progenitor cells maintain the ability to form teeth for at least 25 weeks.
Katrina MacAulay, S Anne Blair, Eric Hajduch, Tatsuo Terashima, Otto Baba, Calum Sutherland and S Harinder Hundal : Constitutive activation of GSK3 down-regulates glycogen synthase abundance and glycogen deposition in rat skeletal muscle cells., The Journal of Biological Chemistry, Vol.280, No.10, 9509-9518, 2005.
(Summary)
The effects of inhibition or constitutive activation of glycogen synthase kinase-3 (GSK3) on glycogen synthase (GS) activity, abundance, and glycogen deposition in L6 rat skeletal muscle cells were investigated. GS protein expression increased approximately 5-fold during differentiation of L6 cells (comparing cells at the end of day 5 with those at the beginning of day 3). However, exposure of undifferentiated myoblasts (day 3) to 50 microM SB-415286, a GSK3 inhibitor, led to a significant elevation in GS protein that was not accompanied by changes in the abundance of GLUT4, another late differentiation marker. In contrast, stable expression of a constitutively active form of GSK3beta (GSK3S9A) led to a significant reduction (approximately 80%) in GS protein that was antagonized by SB-415286. Inhibition of GSK3 or expression of the constitutively active GSK3S9A did not result in any detectable changes in GS mRNA abundance. However, the increase in GS protein in undifferentiated myoblasts or that seen following incubation of cells expressing GSK3S9A with GSK3 inhibitors was blocked by cycloheximide suggesting that GSK3 influences GS abundance possibly via control of mRNA translation. Consistent with the reduction in GS protein, cells expressing GSK3S9A were severely glycogen depleted as judged using a specific glycogen-staining antibody. Inhibiting GSK3 in wild-type or GSK3S9A-expressing cells using SB-415286 resulted in an attendant activation of GS, but not that of glucose transport. However, GS activation alone was insufficient for stimulating glycogen deposition. Only when muscle cells were incubated simultaneously with insulin and SB-415286 or with lithium (which stimulates GS and glucose transport) was an increase in glycogen accretion observed. Our findings suggest that GSK3 activity is an important determinant of GS protein expression and that while glycogen deposition in muscle cells is inherently dependent upon the activity/expression of GS, glucose transport is a key rate-determining step in this process.
Y Kitahara, N Suda, T Terashima, Otto Baba, K Mekaapiruk, E V Hammond, Y Takano and K Ohyama : Accelerated bone formation and increased osteoblast number contribute to the abnormal tooth germ development in parathyroid hormone-related protein knockout mice., Bone, Vol.35, No.5, 1100-1106, 2004.
(Summary)
Our previous study showed that tooth germs at late embryonic stage [later than embryonic day 17.5 (E17.5)] and neonatal homozygous parathyroid hormone-related protein (PTHrP)-knockout mice are compressed or penetrated by the surrounding alveolar bone tissue. In vivo and in vitro studies have shown that the development of the tooth germ proper is not disturbed, but insufficient alveolar bone resorption, due to the decreased number and hypofunction of osteoclasts, is the main cause of this abnormality. In addition to the insufficient alveolar bone resorption, progressive bone formation toward tooth germs was observed in homozygous mice, suggesting that accelerated bone formation also contributes to this abnormality. To further investigate this, homozygous mice at E14.0 and E15.5, when alveolar bone is forming, were used for histochemical and bone histomorphometric analyses. In contrast to the late embryonic stage, the alveolar bone did not yet compress developing tooth germs in homozygous mice on E14.0, but a larger amount of bone tissue was seen compared to wild-type littermates. Histomorphometric analysis of bone at E14.0 revealed that the osteoblast numbers and surfaces in the mandibles and in the bone collar of femora of homozygous mice were significantly higher than those of wild-type mice. However, unlike our previous study showing the osteoclast surface on E18.5 in homozygous mice to be significantly lower than that of wild-type mice, this study at E14.0 showed no significant difference between the two genotypes. To evaluate the amount of calcification around tooth germs, 3D images of mandibles were reconstructed from the calcein-labeled sections of the wild-type and mutant mice. Labeling was performed at E14.0, and the mice were sacrificed 1 h after the calcein injection to minimize the effect of bone resorption. Comparison of the 3D images revealed that the labeled surface was larger around developing tooth germs in homozygous mouse than in wild-type mouse. On day E15.5, osteoblasts approached the enamel organ of homozygous mice but this was not observed in wild-type mice. In this study, we report a systemic increase in osteoblast number and accelerated bone formation in homozygous PTHrP-knockout mice, both of which contribute to the abnormal tooth development.
Otto Baba, Chunlin Qin, C Jan Brunn, N James Wygant, W Bradley McIntyre and T William Butler : Colocalization of dentin matrix protein 1 and dentin sialoprotein at late stages of rat molar development., Matrix Biology, Vol.23, No.6, 371-379, 2004.
(Summary)
Dentin matrix protein 1 (DMP1) and dentin sialophosphoprotein (DSPP) are acidic proteins found in the extracellular matrices of bones and teeth. Recent data from gene knockouts, along with those of gene mutations, indicate that these two phosphoproteins are critical for bone and tooth development and/or maintenance. However, the precise functions of the two proteins have not been elucidated. In order to gain insights into their functions in tooth formation, we performed systematic, comparative investigations on the immunolocalization of DMP1 and dentin sialoprotein (DSP, a cleaved fragment of DSPP), using the rat first molar at different developmental stages as a model. Immunohistochemistry (IHC) was performed with specific, monoclonal antibodies against the COOH-terminal fragments of DMP1 and against DSP. In 1-day- and 1-week-old rats, weak immunoreactions for DMP1 were observed in dentinal tubules while stronger reactions for DSP were seen in the tubules and predentin. In rats older than 2 weeks, immunoreactions for DMP1 were found in dentinal tubules, predentin and odontoblasts. In 5-week- and 8-week-old rats, strong immunoreactions for DMP1 were widely distributed in odontoblasts and predentin. The distribution pattern of DSP was strikingly similar to that of DMP1 after 2 weeks and the localization of each was distinctly different from that of bone sialoprotein (BSP). The unique colocalization of DMP1 and DSPP in tooth development suggests that the two proteins play complementary and/or synergistic roles in formation and maintenance of healthy teeth.
(Keyword)
Animals / Antibodies, Monoclonal / Bone and Bones / Extracellular Matrix Proteins / Gene Expression Regulation, Developmental / immunohistochemistry / In Situ Hybridization / Integrin-Binding Sialoprotein / Molar / Odontoblasts / Phosphoproteins / Protein Precursors / Protein Structure, Tertiary / Rats / Rats, Sprague-Dawley / Sialoglycoproteins / Time Factors
Otto Baba, Chunlin Qin, C Jan Brunn, E Jarrod Jones, N James Wygant, W Bradley McIntyre and T William Butler : Detection of dentin sialoprotein in rat periodontium., European Journal of Oral Sciences, Vol.112, No.2, 163-170, 2004.
(Summary)
Cloning and sequencing of the cDNA indicates that dentin sialophosphoprotein (DSPP) is a precursor of both dentin sialoprotein (DSP) and dentin phosphoprotein (DPP). Dentin sialophosphoprotein must be proteolytically processed to form these two extracellular matrix (ECM) proteins. Numerous studies led us to conclude that DSP (and DSPP) are exclusively expressed by odontoblasts and preameloblasts. However, recent observations suggest a wider distribution. To test this hypothesis, we conducted systematic studies on rat first molar during root formation with immunohistochemical techniques using specific anti-DSP polyclonal and monoclonal antibodies. We also performed in situ hybridization, using high-stringency RNA probes to detect DSP transcripts. Immunohistochemical studies demonstrated that DSP is not only localized in odontoblasts, dentin ECM and preameloblasts, but also in alveolar bone, cellular cementum, osteocytes, cementocytes, and their matrices. The results of in situ hybridization were consistent with those from immunohistochemistry, showing the expression of DSP transcripts in osteoblasts of alveolar bone, fibroblasts in periodontal ligament and cementoblasts in cellular cementum. Together, these observations suggest that DSP is involved in formation of the periodontium as well as tooth structures.
Chunlin Qin, C Jan Brunn, Otto Baba, N James Wygant, W Bradley McIntyre and T William Butler : Dentin sialoprotein isoforms: detection and characterization of a high molecular weight dentin sialoprotein., European Journal of Oral Sciences, Vol.111, No.3, 235-242, 2003.
(Summary)
Dentin sialoprotein (DSP) is a glycoprotein accounting for 5-8% of the dentin non-collagenous proteins. The cDNA sequence predicts that rat DSP has 13 potential casein kinase phosphorylation sites and six potential N-linked glycosylation sites. However, its total phosphorylation level, as well as the nature and locations of the carbohydrate moieties, are unknown. Our findings in the present study show that rat DSP has 6.2 phosphates per molecule and that the majority of carbohydrates are attached to the protein through N-linked glycosylations. During our separation of dentin non-collagenous proteins with ion-exchange chromatography, we observed high molecular weight components eluting late in the salt gradient that were recognized by anti-DSP antibodies. We have purified these high molecular weight components using a monoclonal anti-DSP antibody affinity column. Data from amino acid analysis, phosphate level measurements and Edman degradation of tryptic peptides unequivocally proved that the very acidic, high molecular weight components are isoforms of DSP (designated HMW-DSP). Deglycosylation analysis indicates that the slower migration rate of HMW-DSP on SDS-PAGE results from its higher level of carbohydrate modifications.
R Emma Hudson, A David Pan, John James, M John Lucocq, A Simon Hawley, A Kevin Green, Otto Baba, Tatsuo Terashima and Grahame D Hardie : A novel domain in AMP-activated protein kinase causes glycogen storage bodies similar to those seen in hereditary cardiac arrhythmias., Current Biology, Vol.13, No.10, 861-866, 2003.
(Summary)
The AMP-activated protein kinase (AMPK) is an alphabetagamma heterotrimer that is activated by low cellular energy status and affects a switch away from energy-requiring processes and toward catabolism. While it is primarily regulated by AMP and ATP, high muscle glycogen has also been shown to repress its activation. Mutations in the gamma2 and gamma3 subunit isoforms lead to arrhythmias associated with abnormal glycogen storage in human heart and elevated glycogen in pig muscle, respectively. A putative glycogen binding domain (GBD) has now been identified in the beta subunits. Coexpression of truncated beta subunits lacking the GBD with alpha and gamma subunits yielded complexes that were active and normally regulated. However, coexpression of alpha and gamma with full-length beta caused accumulation of AMPK in large cytoplasmic inclusions that could be counterstained with anti-glycogen or anti-glycogen synthase antibodies. These inclusions were not affected by mutations that increased or abolished the kinase activity and were not observed by using truncated beta subunits lacking the GBD. Our results suggest that the GBD binds glycogen and can lead to abnormal glycogen-containing inclusions when the kinase is overexpressed. These may be related to the abnormal glycogen storage bodies seen in heart disease patients with gamma2 mutations.
Tatsuo TERASHIMA, Otto Baba, Yoshiro TAKANO, Kotaro ISHIBASHI and Shinichiro OIDA : Analysis of Enamel Formation in Mutant Rats with Hypoplastic Type of Amelogenesis Imperfecta, Dentistry in Japan, Vol.39, 8-14, 2003.
K Sena, Y Morotome, Otto Baba, T Terashima, Y Takano and I Ishikawa : Gene expression of growth differentiation factors in the developing periodontium of rat molars., Journal of Dental Research, Vol.82, No.3, 166-171, 2003.
(Summary)
Growth and differentiation factors (GDF) 5, 6, and 7 are known to play roles in tendon and ligament formation, and are therefore probably involved in the formation of periodontal ligament. In this study, we sought to determine temporal and spatial expression of GDF-5, -6, and -7 mRNA in developing periodontal tissue of rat molars using in situ hybridization. GDF gene expression in the periodontal ligament was first detected in cells associated with the initial process of periodontal ligament fiber bundle formation. Gene signals were also detected in cells located along the alveolar bone and cementum surfaces, the insertion sites of periodontal ligaments, during the course of root formation. GDF expression in these cells were down-regulated after completion of root formation. Our results appeared to suggest the involvement of GDF-5, -6, and -7 in the formation of the dental attachment apparatus.
Otto Baba, Nobuyuki Takahashi, Tatsuo Terashima, Wu Li, K Pamela DenBesten and Yoshiro Takano : Expression of alternatively spliced RNA transcripts of amelogenin gene exons 8 and 9 and its end products in the rat incisor., The Journal of Histochemistry and Cytochemistry, Vol.50, No.9, 1229-1236, 2002.
(Summary)
In addition to seven known exons of the amelogenin gene, recent studies have identified two exons downstream of amelogenin exon 7 in genomic DNA of mouse and rat. Here the spatial and temporal expression of mRNAs and of the translated proteins derived from alternative splicing of the amelogenin gene ending with exon 8 and exon 9 were examined by in situ hybridization (ISH) and immunohistochemistry (IHC). RNA signals for exons 8 and 9 were expressed in the ameloblast layer extending from early presecretory to postsecretory transitional stages of amelogenesis. IHC of amelogenin proteins that include sequences encoded by these exons demonstrated identical localization of these proteins in the ameloblast layer corresponding to RNA signals identified by ISH. There was intense immunostaining of the enamel matrix secreted by these cells. Western blotting analysis of rat enamel proteins revealed three distinct protein bands with sequences encoded by the new exons. These data confirmed the existence of the transcripts of alternatively spliced mRNAs coding for exons 8 and 9 of the amelogenin gene in rat tooth germs and suggest that the translated proteins contribute to the heterogeneity of amelogenins and have some significant roles in enamel formation and mineralization.
N Suda, Otto Baba, N Udagawa, T Terashima, Y Kitahara, Y Takano, T Kuroda, V P Senior, F Beck and E V Hammond : Parathyroid hormone-related protein is required for normal intramembranous bone development., Journal of Bone and Mineral Research, Vol.16, No.12, 2182-2191, 2001.
(Summary)
It is well established that parathyroid hormone-related protein (PTHrP) regulates chondrocytic differentiation and endochondral bone formation. Besides its effect on cartilage, PTHrP and its major receptor (type I PTH/PTHrP receptor) have been found in osteoblasts, suggesting an important role of PTHrP during the process of intramembranous bone formation. To clarify this issue, we examined intramembranous ossification in homozygous PTHrP-knockout mice histologically. We also analyzed phenotypic markers of osteoblasts and osteoclasts in vitro and in vivo. A well-organized branching and anastomosing pattern was seen in the wild-type mice. In contrast, marked disorganization of the branching pattern of bone trabeculae and irregularly aligned osteoblasts were recognized in the mandible and in the bone collar of the femur of neonatal homozygous mutant mice. In situ hybridization showed that most of the osteoblasts along the bone surfaces of the wild-type mice and some of the irregularly aligned osteoblastic cells in the homozygous mice expressed osteocalcin. Alkaline phosphatase (ALP) activity and expression of osteopontin messenger RNA (mRNA) in primary osteoblastic cells did not show significant differences between cultures derived from the mixture of heterozygous mutant and wild-type mice (+/? mice) and those from homozygous mutant mice. However, both mRNA and protein levels of osteocalcin in the osteoblastic cells of homozygous mutant mice were lower than those of +/? mice, and exogenous PTHrP treatment corrected this suppression. Immunohistochemical localization of characteristic markers of osteoclasts and ruffled border formation did not differ between genotypes. Cocultures of calvarial osteoblastic cells and spleen cells of homozygous mutant mice generated an equivalent number of tartrate-resistant acid phosphatase-positive (TRAP+) mononuclear and multinucleated cells and of pit formation to that of +/? mice, suggesting that osteoclast differentiation is not impaired in the homozygous mutant mice. These results suggest that PTHrP is required not only for the regulation of cartilage formation but also for the normal intramembranous bone development.
(Keyword)
Adenosine Triphosphatases / Animals / Bone Development / Carbonic Anhydrase II / Cathepsin K / Cathepsins / Cells, Cultured / Female / gene expression / Male / Mandible / Mice / Mice, Inbred C57BL / knockout mice / Osteoblasts / Osteocalcin / Osteoclasts / osteopontin / Parathyroid Hormone-Related Protein / Proteins / Sialoglycoproteins
M García-Rocha, A Roca, N La Iglesia De, Otto Baba, M J Fernández-Novell, C J Ferrer and J J Guinovart : Intracellular distribution of glycogen synthase and glycogen in primary cultured rat hepatocytes., The Biochemical Journal, Vol.357, No.Pt 1, 17-24, 2001.
(Summary)
Changes in the intracellular distribution of liver glycogen synthase (GS) might constitute a new regulatory mechanism for the activity of this enzyme at cellular level. Our previous studies indicated that incubation of isolated hepatocytes with glucose activated GS and resulted in its translocation from a homogeneous cytosolic distribution to the cell periphery. These studies also suggested a relationship with insoluble elements of the cytoskeleton, in particular actin. Here we show the translocation of GS in a different experimental model that allows the analysis of this phenomenon in long-term studies. We describe the reversibility of translocation of GS and its effect on glycogen distribution. Incubation of cultured rat hepatocytes with glucose activated GS and triggered its translocation to the hepatocyte periphery. The relative amount of the enzyme concentrated near the plasma membrane increased with time up to 8 h of incubation with glucose, when the glycogen stores reached their maximal value. The lithium-induced covalent activation of GS was not sufficient to cause its translocation to the cell periphery. The intracellular distribution of GS closely resembled that of glycogen. Our results showed an interaction between GS and an insoluble element of the hepatocyte matrix. Although no co-localization between actin filaments and GS was observed in any condition, disruption of actin cytoskeleton resulted in a significantly lower percentage of cells in which the enzyme translocated to the cell periphery in response to glucose. This observation suggests that the microfilament network has a role in the translocation of GS.
Y Takano, H Sakai, Otto Baba and T Terashima : Differential involvement of matrix vesicles during the initial and appositional mineralization processes in bone, dentin, and cementum., Bone, Vol.26, No.4, 333-339, 2000.
(Summary)
The distribution of matrix vesicles and its role in biological mineralization were examined in bone and dental hard tissues of the rat after daily administrations of 1-hydroxyethylidene-1, 1-bisphosphonate (HEBP), a potent inhibitor of mineralization, for 7 or 14 days. Newly formed, nonmineralized matrices of the HEBP-affected bone and mesodermal dental hard tissues other than circumpulpal dentin contained numerous mineral-filled matrix vesicles (MV), randomly distributed throughout the collagenous matrix. The distribution density of the mineral-filled MV in the HEBP-affected matrices of calvaria, metaphyseal trabecular bone, alveolar bone, and cellular cementum ranged from 60 to 70 per 100 microm(2), and no statistically significant differences were noted among the values. In the HEBP-affected dentin, however, MV were located only in the nonmineralized matrix of mantle dentin and totally absent in the circumpulpal dentin layers. Instead, the HEBP-affected circumpulpal dentin contained a dense meshwork of noncollagenous matrix enriched with calcium and phosphorus. Comparable meshwork structures were undetectable in nonmineralized matrices of the other hard tissues affected by HEBP. These observations suggest that a certain population of MV (60-70 per 100 microm(2)) is involved in the process of appositional mineralization in most of the mesodermal hard tissues, in addition to their well-known role in initial mineral induction in these tissues. Circumpulpal dentin appears to be an exception, where MV are not required for the appositional mineralization process. Exclusive localization of dentin phosphoproteins in circumpulpal dentin layers must take place to facilitate appositional mineralization at the calcification front, in the absence of MV.
Y Takano, H Sakai, Otto Baba, Y Sakamoto, T Terashima, K Ohya and N Kurosaki : Demonstration of putative Ca-binding domains in dentin matrix of rat incisors after daily injections of 1-hydroxyethylidene-1,1-bisphosphonate (HEBP)., European Journal of Oral Sciences, Vol.106 Suppl 1, 274-281, 1998.
(Summary)
In order to clarify the initial process of dentin mineralization, the inhibitory effect of 1-hydroxyethylidene-1,1-bisphosphonate (HEBP) on dentin mineralization was investigated. Rats (100 g) were subcutaneously injected with HEBP (8 mg P/kg) for 7 or 14 d, and the incisors were processed for Ca histochemistry and/or electron microscopy. HEBP-treated incisors demonstrated ladder-like alternate rows of mineralized and non-mineralized dentin at the apical end. GBHA revealed moderate Ca reactions in the non-mineralized circumpulpal dentin matrix where electron microscopy revealed rich distribution of fine mesh-like electron-dense material. Non-mineralized mantle dentin matrix was negative for Ca but contained numerous matrix vesicles (MVs) filled with crystalline and/or amorphous mineral deposits. Mineralization of circumpulpal dentin occurred independently of MV-rich mantle dentin layer in affected specimens. Our data provide histochemical evidence of possible Ca-binding property of the circumpulpal dentin matrix and its absence in the mantle dentin where MV-mediated mineralization occurs. In the mantle dentin, HEBP does not interfere with crystal growth in MVs but inhibits its outgrowth after membrane rupture. It is proposed that circumpulpal dentin matrix has a potential to mineralize independently of MV-mediated mineralization of mantle dentin, although MVs determine the initial site and timing of dentin mineralization.
T Iimura, Otto Baba, Y Maruoka, K Takeda, S Sasaki, H Shimokawa and S Oida : Homeobox gene expression during bone formation induced by BMP., Annals of the New York Academy of Sciences, Vol.785, 274-277, 1996.
Otto Baba : Production of monoclonal antibody that recognizes glycogen and its application for immunohistochemistry, The Journal of the Stomatological Society, Japan, Vol.60, No.2, 264-287, 1993.
(Keyword)
antibody / glycogen / immunohistochemistry
72.
S Shibata, Otto Baba, M Niikura, S Suzuki, Y Yamashita and T Ichijo : An ultrastructural study of mitotic chondrocytes in the proliferative zone of the rat tibial growth plate., Annals of Anatomy, Vol.175, No.1, 41-45, 1993.
(Summary)
Mitotic chondrocytes containing fairly prominent cell organelles were observed in the proliferative zone of the rat tibial growth plate by electron microscopy. During the later stage of mitosis, the endoplasmic reticulum was mostly vacuolated, and the Golgi stacks were replaced by spherical and cylindrical vacuoles and small vesicles. The cytoplasmic division began at early telophase. However, the cytoplasmic cleavage furrow was so narrow that dividing chondrocytes looked as if they were partitioned by a slit. This is probably one reason for the appearance of the binuclear cells that are occasionally observed in the tibial growth plate.
S Shibata, Otto Baba, Y Sakamoto, M Ohsako, Y Yamashita and T Ichijo : An ultrastructural study of the mitotic preosteoblasts in the primary spongiosa of the rat mandibular condyle., Bone, Vol.14, No.1, 35-40, 1993.
(Summary)
In this study, we observed mitotic preosteoblasts that have the structural features of osteoblasts in the primary spongiosa of the rat mandibular condyle. The rough endoplasmic reticulum and the Golgi apparatus showed remarkable disorganization during mitosis. The Golgi saccules were replaced by groups of large vacuoles and small vesicles. The cisternae of the rough endoplasmic reticulum also were vacuolized. Since this disorganization occurred in conjunction with the formation of the mitotic spindle, it is probably related to the changes of the microtubular cytoskeleton. Further, secretory granules were arrayed along the mitotic spindle microtubules at the metaphase, and concentrated around the midbody at the telophase. These findings indicate a close relationship exists between secretory granules and microtubules.
S Shibata, Otto Baba, M Ohsako, S Suzuki, Y Yamashita and T Ichijo : Ultrastructural observation on matrix fibers in the condylar cartilage of the adult rat mandible., The Bulletin of Tokyo Medical and Dental University, Vol.38, No.4, 53-61, 1991.
(Summary)
In this study, ultrastructural and immunohistochemical studies were performed on the adult rat (15 weeks old) mandibular condyle, with particular attention to the matrix fibers in the condylar cartilage. The fibrous zone had thick collagen fibrils which formed fibril bundles. These collagen fibrils consisted mainly of Type I collagen. From the proliferative zone to the mature zone, the density of the collagen fibrils became higher. In the hypertrophic zone, thick collagen fibrils were formed around the chondrocytes. Immunohistochemical study indicated that these collagen fibrils consisted mainly of Type I collagen. Therefore, it was confirmed that the hypertrophic chondrocytes in this tissue had one of the osteoblastic phenotypes.
S Shibata, Otto Baba, M Ohsako, S Shikano, T Terashima, Y Yamashita and T Ichijo : Histological observation of large light cells that seem to be surviving hypertrophic chondrocytes in the rat mandibular condyle., Archives of Oral Biology, Vol.36, No.7, 541-544, 1991.
(Summary)
Large light cells (more than 20 microns dia), including some with mitotic figures, appeared to be surviving hypertrophic chondrocytes. Thus at least a few hypertrophic chondrocytes in the rat mandibular condyle may survive, be released into the primary spongiosa, and divide.
Tatsuo Terashima, Otto Baba and Shinichiro Oida : 4. Expression of enamel protein hy the methods for in situ hybridiza tion and immunohistochemistry in the incisor of the amelogenesis imperfecta rat(ami)(8th Annual Meeting of the Society of Hard Tissue Biology July 24, 1999 in Asahikawa), Journal of Hard Tissue Biology, Vol.9, No.1, 26, 2000.
Kaori Sumida, Tsuyoshi Morita and Otto Baba : Considering the Morphological and Functional Implications of the Human Palatopharyngeus, Journal of Oral Health and Biosciences, Vol.34, No.2, 19-25, Mar. 2022.
Otto Baba : 知れば安心，もっと楽しい! 解剖学的口腔の観かた∼下顎骨を知る 第6回下顎骨の成長・歯の喪失による形態変化, The Journal of Dental Hygiene, Vol.40, No.4, 346-350, Mar. 2020.
3.
Otto Baba : 知れば安心，もっと楽しい! 解剖学的口腔の観かた∼下顎骨を知る 第5回下顎骨に付着する筋肉と咀嚼・嚥下運動, The Journal of Dental Hygiene, Vol.40, No.3, 236-239, Feb. 2020.
4.
Otto Baba : 知れば安心，もっと楽しい! 解剖学的口腔の観かた∼下顎骨を知る 第4回口腔壁を構成する下顎に付着する筋, The Journal of Dental Hygiene, Vol.40, No.2, 122-125, Jan. 2020.
5.
Otto Baba : 知れば安心，もっと楽しい! 解剖学的口腔の観かた∼下顎骨を知る 第3回下顎骨の基本構造③(下顎孔と下顎管と関節突起について), The Journal of Dental Hygiene, Vol.40, No.1, 12-15, Dec. 2019.
6.
Otto Baba : 知れば安心，もっと楽しい! 解剖学的口腔の観かた∼下顎骨を知る 第2回 下顎骨の基本構造②(下顎枝について), The Journal of Dental Hygiene, Vol.39, No.12, 1240-1242, Nov. 2019.
7.
Otto Baba : 知れば安心，もっと楽しい! 解剖学的口腔の観かた∼下顎骨を知る 第1回 下顎骨の基本構造①(概要と下顎体について), The Journal of Dental Hygiene, Vol.39, No.11, 1127-1129, Oct. 2019.
8.
Otto Baba : The New Insight into Radicular Dentin Formation, Journal of Oral Health and Biosciences, Vol.29, No.1, 1-6, Jun. 2016.
(Summary)
Dentin is mineralized connective tissue, which consists of two parts, coronal dentincovered by enamel and radicular dentin by cementum. Although both parts are formed by odontoblasts, biophysical and biochemical properties are indeed different, which evoke the specific system for radicular dentin formation. One of the critical factor of radicular dentin formation is Hertwig's epithelial root sheath (HERS), which leads proliferation and differentiation of odontoblasts under the control of TGF-/BMP signaling, and the ablation or over-expression of constituent molecules showed the specific malformation of radicular dentin. Wnt/-catenin signaling is the other candidate of dentin formation which may regulate differently on crown and root formation. Fibroblast growth factor (FGF) 18 is possible downstream molecule of this signaling. Interestingly, the expression of FGF18 transcripts was detected during root formation, but not crown formation in rat mandibular molar.While possible receptors, FGFR2 and FGFR3, were continuously observed in both crown and root formation, those suggested that coronal and radicular dentin might be formed under the continuous and/or reciprocal control of different FGFs. The clarification of these systems may open new insight into the tissue regeneration and/or engineering of tooth and periodontium.
Otto Baba, Qin Chunlin and T William Butler : Dentin matrix protein 1(DMP1)およびDentin sialoprotein(DSP)のラット臼歯成長過程におけるco-localization, Journal of Oral Biosciences, Vol.46, No.5, 377, Sep. 2004.
C Qin, Otto Baba and T W Butler : Post-translational modifications of sibling proteins and their roles in osteogenesis and dentinogenesis., Critical Reviews in Oral Biology and Medicine, Vol.15, No.3, 126-136, Jun. 2004.
(Summary)
The extracellular matrix (ECM) of bone and dentin contains several non-collagenous proteins. One category of non-collagenous protein is termed the SIBLING (Small Integrin-Binding LIgand, N-linked Glycoprotein) family, that includes osteopontin (OPN), bone sialoprotein (BSP), dentin matrix protein 1 (DMP1), dentin sialophosphoprotein (DSPP), and matrix extracellular phosphoglycoprotein (MEPE). These polyanionic SIBLING proteins are believed to play key biological roles in the mineralization of bone and dentin. Although the specific mechanisms involved in controlling bone and dentin formation are still unknown, it is clear that some functions of the SIBLING family members are dependent on the nature and extent of post-translational modifications (PTMs), such as phosphorylation, glycosylation, and proteolytic processing, since these PTMs would have significant effects on their structure. OPN and BSP are present in the ECM of bone and dentin as full-length forms, whereas amino acid sequencing indicates that DMP1 and DSPP exist as proteolytically processed fragments that result from scission of X-Asp bonds. We hypothesized that the processing of DMP1 and DSPP is catalyzed by the PHEX enzyme, since this protein, an endopeptidase that is predominantly expressed in bone and tooth, has a strong preference for cleavage at the NH2-terminus of aspartyl residue. We envision that the proteolytic processing of DMP1 and DSPP may be an activation process that plays a significant, crucial role in osteogenesis and dentinogenesis, and that a failure in this processing would cause defective mineralization in bone and dentin, as observed in X-linked hypophosphatemic rickets.
S Afroz, Rieko Arakaki, Takuma Iwasa, Masamitsu Ohshima, Maki Hosoki, Miho Inoue, Otto Baba, Yoshihiro Okayama and Yoshizo Matsuka : CGRP induced glial-cytokine expression, NFkB signaling and orofacial nociception, International Association for Dental Research, Washington, D.C., Mar. 2020.
2.
Otto Baba : Regulation of Odontoblasts in Various Phases of Dentin Formation, DENTISPHERE 4 (International Scientific Meeting 5th Asean Plus & Tokushima Joint International Conference), Surabaya, Indonesia, Nov. 2019.
3.
R Raju, Masamitsu Ohshima, Tsuyoshi Morita, Huijiao Yan, Miho Inoue, Otto Baba, Masahisa Inoue and Yoshizo Matsuka : A double-layered complex cell sheet can regenerate the periodontal tissue in a mice model, Dentisphere, Surabayai, Nov. 2019.
4.
S Afroz, Rieko Arakaki, Takuma Iwasa, Masamitsu Ohshima, Otto Baba, Y Okayama and Yoshizo Matsuka : Orofacial pain due to glial expressed cytokines in trigeminal ganglion, Dentisphere, Surabayai, Nov. 2019.
5.
Arief Waskitho, Yumiko Yamamoto, Tsuyoshi Morita, Huijiao Yan, R Raju, Masamitsu Ohshima, Junhel Dalanon, Otto Baba and Yoshizo Matsuka : Bilateral effects of unilateral administration of botulinum toxin A in chemotherapy induced neuropathy, Dentisphere, Surabayai, Nov. 2019.
6.
Otto Baba : Symposium: Perspectives on the Impact of William Butler's Mentoring, 97th General session & exhibition of the IADR, Vancouver, Jun. 2019.
7.
S Afroz, Rieko Arakaki, Takuma Iwasa, Masamitsu Ohshima, Maki Hosoki, Miho Inoue, Otto Baba, Yoshihiro Okayama and Yoshizo Matsuka : Orofacial nociception due to glial expressed cytokines in trigeminal ganglia, International Association for Dental Research, Vancouver, Jun. 2019.
8.
Kazumitsu Sekine, Yoshihito Naitou, Tetsuo Ichikawa, Otto Baba and Kenichi Hamada : Development of structural and chemical enforcement of neointimal growth as the blood contacting surface for the vascular prosthesis, 45th European Society for Artificial Organs 2018 Congress, Madrid, Sep. 2018.
9.
Y Kumei, J Zeredo, R Watahiki, K Kagiyama, K Fukasawa and Otto Baba : Common marmosets in Martian, Lunar, and zero gravity conditions: posture, behavior and motion sickness., The Joint Conference of the 7th International Symposium on Physical Sciences in Space & 25th European Low Gravity Research Association Biennial Symposium and General Assembly, Oct. 2017.
10.
R Watahiki, J Zeredo, K Kagiyama, H Hashimoto, N Ishioka, Y Inatomi, Otto Baba, S Aou, K Natsume and Y Kumei : Postural and behavioral of primates (Common Marmoset) to Martian, Lunar, and Zero gravity conditions., American Society for Gravitational and Space Research Annual Meeting, Oct. 2016.
Proceeding of Domestic Conference:
1.
Mikiya Satoh, Kaho Yumoto, Tsuyoshi Morita, Kaori Sumida and Otto Baba : Involvement of sox genes during the formation of medaka pharyngeal tooth and pedicle, 第64回歯科基礎医学会学術大会, Sep. 2022.
2.
Yui Ohno, Ryutaroh Tohgoh, Ayako Yamane, Tsuyoshi Morita, Kaori Sumida and Otto Baba : マウス象牙質の形成過程におけるFgf18の発現, 四国歯学会第60回例会, Jun. 2022.
3.
Waskitho Arief, Yumiko Yamamoto, Raman Swarna Lakshmi, Fumiya Kano, Huijiao Yan, Kazuo Okura, Daisuke Ikutame, Masamitsu Ohshima, Otto Baba, Akihito Yamamoto and Yoshizo Matsuka : Bilateral botulinum toxin type A effect on orofacial neuropathic pain of animal model, 徳島大学脳科学クラスターミニリトリート, Feb. 2022.
4.
Masahito Kawano, Mikiya Satoh, Kaho Yumoto, Kaori Sumida, Tsuyoshi Morita, 神尾 強司 and Otto Baba : 口蓋垂裂の一例の肉眼解剖学的および組織学的観察, 四国歯学会 第57回例会, Mar. 2021.
5.
Kazumitsu Sekine, Otto Baba and Kenichi Hamada : 表面修飾を施した多孔質チタンインプラントの組織封鎖性評価, 第58回日本人工臓器学会大会, Nov. 2020.
6.
Kazumitsu Sekine, Otto Baba and Kenichi Hamada : 骨代替インプラントを目的とした封鎖性の高いチタンスキャフォールドの開発, 第59回日本生体医工学会大会, May 2020.
7.
Kaori Sumida, Seiichiro Kitamura, Tsuyoshi Morita, 神尾 強司, 城 翠, 古田 琴, 濱 義之, 田畑 純 and Otto Baba : コモンリスザルの軟口蓋・咽頭の筋構築に関する肉眼解剖学的研究, 第125回日本解剖学会総会・全国学術集会 講演プログラム・抄録集 山口 2020 誌上開催, 162, Mar. 2020.
8.
Yui Miyagawa, Masaki Gotoh, Yukari Hatada, Tsuyoshi Morita, Tsuyoshi Kano, Kaori Sumida and Otto Baba : マダイ稚魚の歯の形成過程に関する組織学的研究, The 125th Annual Meeting of The Japanese Association of Anatomists, Mar. 2020.
9.
Tsuyoshi Morita, Tsuyoshi Kano, Kaori Sumida and Otto Baba : 歯・歯周組織の形成機構の解析および再生への応用の検討, 四国歯学会第56回例会, Feb. 2020.
10.
Shin Matsumoto, Soichiroh Kinjoh, Tsuyoshi Morita, Kaori Sumida, Tsuyoshi Kano and Otto Baba : メダカ咽頭歯における歯および歯周組織の形成遺伝子の発現, 四国歯学会第56回例会, 34, Feb. 2020.
11.
Takuma Iwasa, Afroz Shaista, Miho Inoue, Rieko Arakaki, Masamitsu Ohshima, Raju Resmi, Arief Waskitho, 井上 正久, Otto Baba and Yoshizo Matsuka : 痛みに対するサイトカイン療法の開発に向けて, 徳島県歯科医学大会, Feb. 2020.
12.
Tsuyoshi Kano, Tsuyoshi Morita, Kaori Sumida, Hiromichi Yumoto and Otto Baba : Gene expression of dentin formation related factors in dental pulp of mouse molar, after tooth eruption, 第61回歯科基礎医学会学術大会, Oct. 2019.
(Keyword)
pulp / 象牙質 / in situ hybridization
13.
Waskitho Arief, Yamamoto Yumiko, Tsuyoshi Morita, Yan Huijiao, Raju Resmi, Masamitsu Ohshima, Dalanon Junhel, Otto Baba and Yoshizo Matsuka : Orofacial neuropathic pain rat models induced by chemotherapy drugs, Tokushima University Bioscience Retreat, Sep. 2019.
14.
Raju Resmi, Masamitsu Ohshima, Miho Inoue, Tsuyoshi Morita, Yan Huijiao, Otto Baba, Inoue Masahisa and Yoshizo Matsuka : Development of a multilayered complex cell sheet to regenerate periodontal tissue, Tokushima University Bioscience Retreat, Sep. 2019.
15.
Raju Resmi, Masamitsu Ohshima, Miho Inoue, Tsuyoshi Morita, Yan Huijiao, Otto Baba, 井上 正久 and Yoshizo Matsuka : 骨芽細胞と歯根膜細胞との立体形成複合細胞シートによる歯周組織再生の確立, 硬組織再生生物学会, Aug. 2019.
16.
Kazumitsu Sekine, Otto Baba and Kenichi Hamada : 細胞賦活化処理による多孔質性状チタン足場材の生体内癒合性の向上を狙う, 第58回日本生体医工学会大会, Jun. 2019.
17.
Kaori Sumida, Seiichiro Kitamura, Tsuyoshi Morita, 神尾 強司, 田畑 純 and Otto Baba : ヒト，ウマ，イヌの茎突咽頭筋の走行・停止の比較からヒト茎突咽頭筋の機能を考える, 第124回日本解剖学会総会全国学術集会, Mar. 2019.
18.
Shin Matsumoto, Soichiroh Kinjoh, Sohshi Fujita, Tsuyoshi Morita, Tsuyoshi Kano, Kaori Sumida, Asuna Sugimoto, Tsutomu Iwamoto and Otto Baba : メダカ咽頭歯における分泌性カルシウム結合リンタンパク質(SCPP)遺伝子の発現, 第124回日本解剖学会総会・全国学術集会, Mar. 2019.
(Keyword)
SCPP / メダカ / 咽頭歯 / in situ hybridization
19.
Kazumitsu Sekine, Otto Baba and Kenichi Hamada : 細胞賦活性を施した多孔質チタン足場材に関する生体内癒合性評価, 第56回日本人工臓器学会大会, Nov. 2018.
20.
金城 聡一郎, Shin Matsumoto, 藤田 創詩, Kaori Sumida, Tsuyoshi Morita, 神尾 強司 and Otto Baba : 馬蹄腎の一例についての肉眼解剖学的および組織学的観察, 四国歯学会第53回例会・第37回総会, Jul. 2018.
21.
Kazumitsu Sekine, Otto Baba and Kenichi Hamada : Evaluation of Titanium surface modification for hard tissue prostheses, 第57回日本生体医工学会大会, Jun. 2018.
22.
Kaori Sumida, Seiichiro Kitamura, 神尾 強司, Tsuyoshi Morita, Kikuji Yamashita and Otto Baba : イヌの軟口蓋の筋構築に関する肉眼解剖学的研究, 日本解剖学会総会・第123回全国学術集会,, Mar. 2018.
23.
Kaori Sumida, Seiichiro Kitamura, Tsuyoshi Morita, Kikuji Yamashita and Otto Baba : Gross anatomical study of the human stylopharyngeus: origin, course,and insertion., The 122nd annual meeting of the Japanese association of anatomists, Mar. 2017.
Et cetera, Workshop:
1.
Raju Resmi, Masamitsu Ohshima, Tsuyoshi Morita, Yan Huijiao, Miho Inoue, Otto Baba, Inoue Masahisa and Yoshizo Matsuka : Three-dimensional periodontal tissue regeneration using a bone-ligament complex cell sheet, 発生・再生・遺伝クラスター ミニリトリート, Feb. 2020.
2.
Waskitho Arief, Yumiko Yamamoto, Tsuyoshi Morita, Yan Huijiao, Raju Resmi, Masamitsu Ohshima, Dalanon Junhel, Laksmi Swarna, Chavan P., Otto Baba and Yoshizo Matsuka : Botulinum toxin A administration in chemotherapy-induced neuropathic pain rat model, 徳島大学脳科学クラスターミニリトリート, Feb. 2020.
Common marmoset as a primate model for gerontology in dentistry (Project/Area Number: 16K15852 )
The common marmoset, a new-world monkey as a human-alternative model for studies on pediatric dentistry (Project/Area Number: 26463107 )
New paradigm of enamel maturation: Mechanisms of non-endocytotic resorption and degradation of enamel matrix proteins (Project/Area Number: 24390408 )
The study on the cells forming periodontium by observing the expression of dentin proteins as the markers of differentiation (Project/Area Number: 18591996 )
Establishment of organ culture system for tooth regeneration using pharyngeal teeth of medaka and its use for the exploration of dental stem cells (Project/Area Number: 18390485 )
The induction of periodontal tissues by dentin-derived matrix (Project/Area Number: 14370577 )
Structural analysis of the role of enamel matrix proteins in regeneration of biological hard tissues (Project/Area Number: 12557151 )
Biochemical and histological analysis of proteoglycans systhsized by cartilaginous cell line, ATDC5. (Project/Area Number: 07671963 )
Immunohistochemical and Enzyme-histochemical studies on the cellular networks of antigen-presenting cells in the dental and periodontal tissues. (Project/Area Number: 07457426 )