Biomedical subjects

K Moriyama

Publications and source records attributed to K Moriyama.

At least 19 recordsLinked to original sources

PubMed2007

Identification of genes related to mechanical stress in human periodontal ligament cells using microarray analysis.

BACKGROUND AND OBJECTIVE: Differential expression of genes in human periodontal ligament (PDL) under mechanical stress, such as orthodontic force, is thought to be involved in the remodeling of PDL cells and periodontal tissues. However, little is known about the genes expressed in PDL cells under mechanical stress. MATERIAL AND METHODS: We employed microarray analysis to assess, in a comprehensive manner, the gene expression profiles in PDL cells compressed by a static force using an in vitro three-dimensional culture system. Six genes were selected and validated by quantitative real-time polymerase chain reaction analysis, consistent with the microarray data. RESULTS: The microarray data revealed that 108 of 30,000 genes tested were differentially expressed by mechanical force loading. Among them, 85 genes were up-regulated by mechanical stress, while 23 genes were down-regulated, judging by the thresholds of a two-fold increase/decrease compared with the controls. Thirty-two of the up-regulated and eight of the down-regulated genes, well-characterized in protein function, were involved in numerous biological processes including cell communication, cell signaling, cell cycle, stress response, and calcium release. However, several genes differentially expressed in our microarray data have not been well defined as stress-response molecules. CONCLUSION: Our microarray is the first to show the gene profile in PDL cells caused by mechanical stress; however, further studies to clarify the physiological function of these molecules in PDL cells are required.

Calcium↗

PubMed2004

Leukotrienes mediate 5-hydroxytryptamine-induced plasma extravasation in the rat knee joint via CysLT-type receptors.

OBJECTIVE AND DESIGN: The aim of this study was to investigate whether leukotrienes synthesized by 5-lipoxygenase (5-LO) and acting via leukotriene (LT) receptors contribute to 5-hydroxytryptamine (5-HT)-induced knee joint plasma extravasation (PE). MATERIALS AND METHODS: Knee joints of rats under anesthesia were perfused with 5-HT and synovial vascular Evans Blue dye leakage was measured spectrophotometrically. A series of 5-LO inhibitors and LT receptor antagonists were investigated for their ability to inhibit 5-HT-induced synovial PE. RESULTS: Inhibitors of 5-LO (NDGA and REV 5901) significantly attenuated 5-HT-induced plasma extravasation. MK 571, LY 171883, BAY u9773 (CysLT receptor antagonists) and REV 5901 (a CysLT receptor antagonist and a 5-LO inhibitor) were equally effective in inhibiting 5-HT-induced PE, indicating that leukotrienes mediate 5-HT-induced PE via CysLT receptors. In contrast, antagonists selective for LTB(4) receptors (BLT(1) and BLT(2) receptors) failed to reduce 5-HT-induced PE. CONCLUSIONS: These results demonstrate that leukotrienes, specifically cysteinyl-leukotrienes contribute to synovial plasma extravasation and suggest that leukotrienes act downstream of 5-HT in the inflammatory cascade.

Animals↗

PubMed2003

Recombinant monoclonal thyrotropin-stimulation blocking antibody (TSBAb) established from peripheral lymphocytes of a hypothyroid patient with primary myxedema.

Anti-TSH receptor antibodies (TRAbs) have been known to be involved in Graves' disease and primary hypothyroidism. We previously isolated and reconstituted immunoglobulin (Ig) genes of Epstein-Barr virus-transformed B cell clones producing monoclonal TRAbs obtained from Graves' patients. In the present study, we performed a similar experiment using a B cell clone, 32A-5, derived from a patient with primary hypothyroidism. The variable region genes of Ig heavy (H) and light (L) chains were isolated and sequenced from the 32A-5 clone. A significant number of somatic mutations were found in variable regions of H and L chain gene segments. Each pair of H and L chain cDNAs was ligated into an expression vector for IgG1 production and stably introduced into myeloma cells. The transfectants were injected ip into BALB/c mice to yield ample volume of the antibody for following applications. Interactions of recombinant 32A-5 with Graves' sera with varying thyroid-stimulating antibody (TSAb) activities were studied. The recombinant antibody tended to suppress TSAb activities in 10 of 15 Graves' sera, in which four were significantly inhibited. In summary, this is the first study to analyze human monoclonal TSH-stimulation blocking antibodies (TSBAb) at the molecular level. Use of human recombinant monoclonal TSBAb may be an analytical tool for molecular-basis etiology and an alternative therapeutic path for Graves' disease.

Animals↗

PubMed2002

Drug-induced neutropenia associated with anti-neutrophil cytoplasmic antibodies (ANCA): possible involvement of complement in granulocyte cytotoxicity.

Although antineutrophil antibodies are thought to be involved in drug-induced neutropenia, neither the precise mechanisms nor the particular antigens on the neutrophil surface have yet been clarified. Recently, we examined a patient with Graves' disease who developed antineutrophil cytoplasmic antibodies (ANCA) after propylthiouracil treatment and exhibited neutropenia. Because several target antigens of ANCA are expressed on the surface of neutrophils, it was suggested that ANCA might contribute to neutropenia. The patient's serum bound specifically to neutrophils and HL-60 cells differentiated into granulocytes, and lysed the HL-60 cells via a complement-mediated mechanism. Furthermore, two representative ANCA antigens, proteinase 3 and myeloperoxidase, significantly inhibited both the binding and cytotoxicity of the serum. Finally, tumour necrosis factor-alpha, which is known to up-regulate cell surface expression of several ANCA antigens, enhanced both the binding and cytotoxicity of the serum. These findings suggest that ANCA induced by propylthiouracil contributed to leucopenia through a complement-mediated mechanism.

Antibodies, Antineutrophil Cytoplasmic↗

PubMed2002

The phytochemical lindleyin, isolated from Rhei rhizoma, mediates hormonal effects through estrogen receptors.

Some plant compounds or herb mixtures are popular alternatives to conventional therapies and contain organic compounds that bind to some nuclear receptors, such as the estrogen receptor (ER), to exert various biological effects. We studied the effect of various herbal extracts on ERalpha and ERbeta isoforms. One herbal extract, Rhei rhizoma (rhubarb), acts as an agonist to both ERalpha and ERbeta. The phytochemical lindleyin, a major component of rhubarb, might contribute to this estrogenic activity through ERalpha and ERbeta. 4-Hydroxytamoxifen, an ER antagonist, completely reversed the estrogenic activity of lindleyin. Lindleyin binds to ERalpha in vitro, as demonstrated using a fluorescent polarization assay. The in vivo effect of rhubarb extract was studied using a vitellogenin assay system in the freshwater fish, Japanese medaka (Oryzias latipes). There were marked increases in serum vitellogenin levels in male medaka exposed to rhubarb extract. We conclude that lindleyin, a component of some herbal medicines, is a novel phytoestrogen and might trigger many of the biological responses evoked by the physiological estrogens.

Cells, Cultured↗

PubMed2001

Novel mutations in ABCA1 gene in Japanese patients with Tangier disease and familial high density lipoprotein deficiency with coronary heart disease.

Mutations in the ATP-binding cassette transporter 1 (ABCA1) gene have been recently identified as the molecular defect in Tangier disease (TD) and familial high density lipoprotein deficiency (FHA). We here report novel mutations in the ABCA1 gene in two sisters from a Japanese family with TD who have been described previously (S. Ohtaki, H. Nakagawa, N. Kida, H. Nakamura, K. Tsuda, S. Yokoyama, T. Yamamura, S. Tajima, A. Yamamoto, Atherosclerosis 49 (1983)) and a family with FHA. Both probands of TD and FHA developed coronary heart disease. Sequence analysis of the ABCA1 gene from the patients with TD revealed a homozygous G to A transition at nucleotide 3805 of the cDNA resulting in the substitution of Asp 1229 with Asn in exon 27, and a C to T at nucleotide 6181 resulting in the substitution of Arg 2021 with Trp in exon 47. Sequence analysis of the ABCA1 gene from the FHA patient revealed a homozygous 4 bp CGCC deletion from nucleotide 3787 to 3790 resulting in premature termination by frameshift at codon 1224. These mutations were confirmed by restriction digestion analysis, and were not found in 141 control subjects. Our findings indicate that mutations in the ABCA1 gene are associated with TD as well as FHA.

ATP Binding Cassette Transporter 1↗

PubMed2001

Degradation and fracture of Ni-Ti superelastic wire in an oral cavity.

Superelastic Ni-Ti wire is widely used in orthodontic clinics, but delayed fracture in the oral cavity has been observed. Because hydrogen embrittlement is known to cause damage to Ti alloy systems, orthodontic wires were charged with hydrogen using an electro-chemical system in saline. Tensile tests were carried out, and fracture surfaces were observed after hydrogen charging. The strength of the Co-Cr alloy and stainless steel used in orthodontic treatment, was not affected by the hydrogen charging. However, Ni-Ti wire showed significant decreases in strength. The critical stress of martensite transformation was increased with increasing hydrogen charging, and the alloy was embrittled. The fractured surface of the alloys with severe hydrogen charging exhibited dimple patterns similar to those in the alloys from patients. In view of the galvanic current in the mouth, the fracture of the Ni-Ti alloys might be attributed to the degradation of the mechanical properties due to hydrogen absorption.

Absorption↗

PubMed2001

Characterization of the secretable ectodomain of thyrotropin receptor produced by the recombinant baculovirus system.

Thyrotropin receptor (TSHR) is a member of the glycoprotein hormone receptor family and an autoantigen of Graves' disease. Various attempts have been made to obtain a large amount of soluble ectodomain of TSHR in insect or mammalian cells, but most of them failed to secrete the overexpressed ectodomain. In the present study, we observed that about one-third of the ectodomain protein (sTSHR-gp), in which the signal peptide of TSHR was replaced by the baculovirus-encoded glycoprotein 67-signal peptide, was secreted into the culture medium and the remainder stayed within cells in the recombinant baculovirus system. Microsequencing the N-terminal of the purified protein confirmed that the baculovirus signal peptide was cleaved at the expected site. Carbohydrate studies using several glycosidases and lectins revealed that the secreted form of the ectodomain had biantennary carbohydrate, whereas the non-secreted form had high-mannose. Moreover, the secreted form of sTSHR-gp exhibited high-affinity ligand binding, whereas the non-secreted form did not show any significant ligand binding. Regarding the interactions of TSHR ectodomains with anti-TSHR antibodies, both the secreted and non-secreted forms of sTSHR-gp, almost completely neutralized the stimulatory and inhibitory anti-TSHR antibody activities. In conclusion, we succeeded in secreting the ectodomain of TSHR into culture medium, which was capable of binding to TSH and neutralizing anti-TSHR antibody activities.

Amino Acid Sequence↗

PubMed2001

Cloning and expression of the chick p63 gene.

We have isolated a chick cDNA for p63, a member of the p53 transcription factor family. This cDNA encodes a protein of 582 amino acids for an alpha isoform in the C-terminal region, while lacking the N-terminal transactivation domain. The chick p63 gene is first expressed in the prospective cutaneous ectoderm at stage 6 and later in the developing epithelia. The p63 expression is intense in specialized epithelial structures, such as apical ectodermal ridge of the limb bud, epithelia of branchial arches and feather buds. Furthermore, we have found that the transcripts are detected in the interdigital epithelium, intersomite epithelium, and epaxial dermamyotome.

Amino Acid Sequence↗

PubMed2001

MC3T3-E1-conditioned medium-induced mineralization by clonal rat dental pulp cells.

Dental pulp is thought to participate in supplementary mineralization, such as reparative dentin and pulp stones, but no direct proof of this has been reported. To study this process at a molecular level, we investigated the matrix mineralization of dental pulp using a clonal cell line (RPC-C2A) derived from rat incisor dental pulp. Mineralized nodules in extracellular matrix were formed by RPC-C2A cells cultured in the presence of conditioned medium (CM) from confluent osteoblastic MC3T3-E1 cells. These nodules were stained by the von Kossa method and with alizarin red S and quantified by the measurement of acid-soluble calcium deposition. This CM was most effective when collected 3-6 days after confluency and added at 50% to the culture medium. The CM-treated RPC-C2A cells showed high alkaline phosphatase activity, a high mRNA level of osteocalcin and decreases in the mRNA levels of osteopontin and osteonectin, but undetectable levels of mRNA of dentin sialophosphoprotein by Northern blot analyses. A pan-specific anti-transforming growth factor (TGF)-beta antibody and a soluble form of receptor for bone morphogenetic protein (BMP)-2/-4 did not neutralize the CM-induced mineralization. These results suggest that some soluble factor(s) other than TGF-beta or BMP-2/-4 in the CM from MC3T3-E1 cells cause differentiation of RPC-C2A cells to osteoblast-like cells.

Alkaline Phosphatase↗

PubMed2001

Apoptosis of periodontal ligament cells induced by mechanical stress during tooth movement.

The mechanical force generated during tooth movement creates compressed and cell-free areas in the periodontal membrane. The way in which periodontal ligament cells disappear at the compressed area during tooth movement remains unclear. In the present study we examined whether periodontal ligament cells undergo apoptosis by mechanical stress during tooth movement using the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick-end-labelling method (TUNEL). TUNEL-positive stainings of periodontal ligament cells began to appear at the compressed areas 12 h after tooth movement, and the number of those cells reached maximum at 24 h after tooth movement. Thereafter TUNEL-positive cells disappeared at 48 h, and direct and undermining bone resorption began at the same area 72 h after tooth movement. These results showed that compressed periodontal ligament cells were eliminated by apoptosis in the early phase of tooth movement.

Alveolar Bone Loss↗

PubMed2001

Phosphorylation of cofilin by LIM-kinase is necessary for semaphorin 3A-induced growth cone collapse.

Semaphorin 3A is a chemorepulsive axonal guidance molecule that depolymerizes the actin cytoskeleton and collapses growth cones of dorsal root ganglia neurons. Here we investigate the role of LIM-kinase 1, which phosphorylates an actin-depolymerizing protein, cofilin, in semaphorin 3A-induced growth cone collapse. Semaphorin 3A induced phosphorylation and dephosphorylation of cofilin at growth cones sequentially. A synthetic cell-permeable peptide containing a cofilin phosphorylation site inhibited LIM-kinase in vitro and in vivo, and essentially suppressed semaphorin 3A-induced growth cone collapse. A dominant-negative LIM kinase, which could not be activated by PAK or ROCK, suppressed the collapsing activity of semaphorin 3A. Phosphorylation of cofilin by LIM-kinase may be a critical signaling event in growth cone collapse by semaphorin 3A.

Actin Depolymerizing Factors↗

PubMed2001

Development of artificial seed crystal for crystallization of calcium phosphate.

An artifical seed crystal material consisting of calcium silicate hydrate (5CaO x 6SiO2 x 5H2O : tobermorite crystals) applicable for phosphorus removal by crystallization was developed. Card-house shaped tobermorite crystals were developed on the seed material where orthophosphate crystallized as a calcium phosphate. The seed material can be manufactured by mixing siliceous and calcareous raw materials, pelletizing and subsequent autoclaving. Laboratory experiments were conducted to apply the new developed seed crystal material in the phosphorus recovery from sludge sidestreams of a wastewater treatment plant. In this crystallization process, the performance the carbon dioxide degassingprocess usually carried out when applying crystallization was not necessary, the hydroxyapatite was able to crystallize at a pH of 8.0 to 8.5 without precipitation of calcium carbonates. In the treatment of a sidestream with orthophosphate concentrations of 50 mgl(-1) and COD concentrations between 200 to 400 mgl(-1), phosphorus removal efficiencies ranging from 75 to 85% were observed. The seed crystal material was collected after the laboratory experiments and the chemical estimation and the germination test for agricultural reuse were performed. As a result, it was shown that the hydroxyapatite precipitated on the seed material had a 100% fusibility to soil and had characteristics to be a good nutrient source as a fertilizer for plants.

Calcium Phosphates↗

PubMed2001

Highly efficient random mutagenesis in transcription-reverse-transcription cycles by a hydrogen bond ambivalent nucleoside 5'-triphosphate analogue: potential candidates for a selective anti-retroviral therapy.

The nucleoside P can base pair with both A and G. We evaluated the mutation frequency induced by the 5'-triphospbate of the ribonucleoside P (PTP) in an in vitro retroviral replication model. After 4 cycles of replication in the presence of PTP, the mutation frequency was raised to 3.8 x 10(-2) per nucleotide and C-to-U and U-to-C mutations were dominantly observed. These results suggest that ambivalent NTP analogues, like PTP, could induce mutations beyond the error threshold of retroviruses.

Antiviral Agents↗

PubMed2001

Reverse transcriptional mutagenesis induced by ribonucleoside triphosphate analogue, PTP and its availability for anti-HIV-1 therapy.

The bicyclic pyrimidine analogue, 3,4-dihydro-6H,8H-pyrimido[4,5-c][1,2]oxazin-7-one (P) can base pair with both A and G. The riboside 5'-triphsophate of P (PTP) efficiently induces mutation during in vitro transcription and reverse transcription cycles using a phage promoter. In the present study, we have constructed an in vitro transcription system promoted by the human immunodeficiency virus type 1 (HIV-1) 5'-long terminal repeat (LTR) using HeLa nuclear extract supplemented with HIV-1 Tat protein. Using this system, the effects of mutagenic ribonucleotide analogues were studied.

Anti-HIV Agents↗

PubMed2001

Substantial production of ghrelin by a human medullary thyroid carcinoma cell line.

Ghrelin, an endogenous ligand for the GH secretagogue receptor, is a novel acylated peptide produced in the gastrointestinal endocrine cells as well as neuroendocrine cells in the hypothalamus. The Ser(3) residue of ghrelin is modified by n-octanoic acid, a modification necessary for hormonal activity. Human medullary thyroid carcinoma is known to produce a variety of gastrointestinal and neuroendocrine peptides. In the present study we investigated ghrelin production in the thyroid gland, especially in human medullary thyroid carcinoma. PCR amplification demonstrated prepro-ghrelin gene transcripts in normal human thyroid tissue and two medullary thyroid carcinoma cell lines (human TT cells and rat 6-23 cells), but not in a rat thyroid follicular cell line. TT cells showed the expression of prepro-ghrelin mRNA of about 0.6 kb by Northern blot analysis. Furthermore, production of ghrelin in TT cells was demonstrated by RIA and immunocytochemistry. Accumulation of des-n-octanoyl ghrelin in the cultured medium of the cells was confirmed. Finally, human medullary thyroid carcinoma surgical specimens showed significantly higher des-n-octanoyl ghrelin contents than normal thyroid tissues. In conclusion, we revealed that ghrelin was produced by the human thyroid parafollicular carcinoma cell line, TT cells. These findings suggest that ghrelin is produced in the thyroid C cells as well as in medullary thyroid carcinoma and may provide opportunities to investigate its physiological role in the thyroid gland.

Carcinoma, Medullary↗

PubMed2001

A low dose of ghrelin stimulates growth hormone (GH) release synergistically with GH-releasing hormone in humans.

The synergistic relationship between GH-releasing secretagogue (GHS) and GH-releasing hormone (GHRH) with respect to GH secretion is well known. In the present study, we report a similar relationship between GHRH and ghrelin, a recently identified endogenous ligand for the GHS receptor. In normal male adults, various doses of ghrelin were intravenously administered alone or together with 1.0 microg/kg GHRH. At small doses of 0.08 and 0.2 microg/kg ghrelin, combined administration of the two peptides significantly stimulated GH release in a synergistic manner; the mean GH response values of the two peptide combinations were more than the summed mean GH response values of each peptide alone (P < 0.05). In addition, at 1.0 microg/kg ghrelin, the tendency of the synergistic effect was observed, although the comparison was not statistically significant probably due to a submaximal dose ceiling effect. No synergistic effects with respect to ACTH or prolactin secretion were observed. In conclusion, the synergistic interaction between ghrelin and GHRH was clearly shown and might be useful for a provocation test to diagnose GH deficiency.

Adrenocorticotropic Hormone↗

PubMed2001

Dentocraniofacial morphology of 12 Japanese subjects with unilateral cleft lip and palate with a severe Class III malocclusion: a cephalometric study at the pretreatment stage of surgical orthodontic treatment.

OBJECTIVE: The present study clarifies the dentocraniofacial morphology of patients with cleft lip and palate (CLP) with severe Class III malocclusion prior to surgical orthodontic treatment. METHODS: The sample was 12 Japanese male subjects with repaired complete unilateral CLP (surgical CLP group; 21.2 +/- 1.92 years in mean age). Two sets of patients without CLP Class III malocclusion, consisting of 19 male subjects treated by surgical orthodontic treatment (surgical Class III group; 23.4 +/- 6.35 years in mean age) and 14 male subjects treated by nonsurgical orthodontic treatment (nonsurgical Class III group; 18.7 +/- 3.49 years in mean age) were used as controls. Analyses were performed using lateral and posteroanterior (P-A) cephalograms. RESULTS: (1) The surgical CLP group showed significantly smaller values for overjet, SNA angle, and inclination of the maxillary incisor as compared with those of the surgical and nonsurgical Class III controls. The values of SNB, mandibular effective length, and ramus height in the surgical CLP group were significantly smaller than those of the surgical Class III group but were similar to those of the nonsurgical Class III group. (2) The mandible and the upper and lower dental arches deviated laterally toward the cleft side. The displacement of the mandible was correlated with that of the maxilla. These results show that CLP patients who required surgical orthodontic treatment had a characteristic dentocraniofacial morphology, compared to controls without CLP with Class III malocclusion.

Adolescent↗