Biomedical subjects

T Iimura

Publications and source records attributed to T Iimura.

At least 37 records · Page 2Linked to original sources

PubMed1996

Expression of mRNA encoding intestinal type alkaline phosphatase in rat liver and its increase by fat-feeding.

The presence of types of alkaline phosphatase (ALP) other than the tissue non-specific type enzyme in rat liver and its increase by fat feeding are known. In order to examine expression of intestinal type ALP in liver, specific oligonucleotide primers corresponding to two types of mRNAs of rat intestinal ALP (RTIN-1 and -2) were designed and amplified by means of the reverse transcriptase-polymerase chain reaction (RT-PCR). It was found that RTIN-1 mRNA was expressed only in the intestine but not in the liver, while RTIN-2 mRNA was expressed both in the intestine and in the liver. By fat feeding, expression of RTIN-1 mRNA increased in the intestine and that of RTIN-2 mRNA increased both in the intestine and in the liver. Thus, it was concluded that rat liver expressed one of the intestinal type ALP (RTIN-2) which was enhanced by fat feeding.

Alkaline Phosphatase↗

PubMed1996

Molecular structure of the mouse amelogenin genomic DNA.

A genomic DNA clone (14.5 kbp) for amelogenin, a tooth enamel specific protein, has been isolated from a mouse genomic library. The nucleotide structure of this DNA was determined by the polymerase chain reaction and sequence analysis. The result indicated that the cloned DNA contained all the exons in addition to approximately 5.5 kbp of upstream sequence.

Amelogenin↗

PubMed1995

Expression of prostaglandin E receptor subtypes in bone: expression of EP2 in bone development.

Prostaglandin (PG) E2 displays physiological and pharmacological action in various tissues including bone. It increases intracellular Ca, and stimulates or inhibits cAMP production through the PGE receptor subtypes EP1, EP2, and EP3, respectively. These receptor subtypes have been recently cloned. In the present study, we investigate the expression of these receptor subtypes in bone tissue. RT-PCR revealed that EP1, EP2, and EP3 were expressed in rat calvariae and that osteoblastic cells (MC3T3-E1) expressed EP1 and EP2. In situ hybridization analysis using cryosection of neonatal calvariae revealed that EP2 was expressed by osteoblasts and cells not in contact with bone, probably including preosteoblasts. EP2 expression was observed at an early stage in calvarial development, at 14 days prenatal. EP2 expression was also observed at day 3 in rat bone marrow cell culture in which bone-like mineralized nodules are formed at day 8. It has been established that PGE2 response accompanying cAMP production is one of the characteristics of osteoblasts. The present results indicate that this phenotype appears at an early stage of osteoblastic differentiation and bone development.

3T3 Cells↗

PubMed1995

Identification of bone-type alkaline phosphatase mRNA from human periodontal ligament cells.

Tissue-nonspecific-type alkaline phosphatase is found in the bone, liver, kidney, and other tissues, and its gene consists of 12 exons with the coding sequence beginning in the second exon. Recently, an alternative noncoding first exon was identified in the liver message which differed from that of the previously known osteoblast-derived cDNA sequence. Although these two mRNAs produce an identical protein, they have different promoter regions. The periodontal ligament tissue expresses a high level of alkaline phosphatase activity. To identify its mRNA type, we isolated a full-length cDNA for alkaline phosphatase from a cultured human periodontal ligament cell expression library, using bone-derived tissue-nonspecific alkaline phosphatase cDNA as a hybridization probe. The size of this clone was 2.5 kb, and its 5' and 3' untranslated sequences were identical to those of the human tissue-nonspecific type isolated from osteoblastic cells but not to those of the liver type. In addition, the same fragments as in bone-derived tissue-nonspecific-type cDNA were detected by the treatment of the cDNA clone with restriction enzymes Hinc II and Pst I. The results suggest that expression of the same alkaline phosphatase isozyme in human periodontal ligament cells may be regulated by the same transcriptional mechanism as in bone.

Alkaline Phosphatase↗

PubMed1995

Molecular cloning and sequence of bovine Msx-1 homeobox-containing gene cDNA from a bovine odontoblast library.

Screening of a bovine odontoblast cDNA library from developing incisor with murine Msx-1 and Msx-2 cDNA probes led to the isolation of three positive clones. All of them encoded for a sequence of a protein containing 297 amino acids. The responsible gene was designated as bovine Msx-1 (bMsx-1) due to the high homology with the human MSX-1 and mouse Msx-1 sequences.

Amino Acid Sequence↗

PubMed1995

Cloning and sequence of bone morphogenetic protein 4 (BMP-4) from a human placental cDNA library.

A cDNA clone encoding bone morphogenetic protein-4 (BMP-4) has been isolated from a human placental cDNA library. Sequence analysis of this clone revealed that the nucleotide sequence of 5' region was different from that of human osteosarcoma BMP-4 and the deduced amino acid sequence indicated deletion of N-terminal 6 amino acids. We confirmed the expression of this type of BMP-4 mRNA in one human osteogenic cell line in addition to the placenta by the polymerase chain reaction (PCR).

Amino Acid Sequence↗

PubMed1995

Production of functional human bone morphogenetic protein-2 using a baculovirus/Sf-9 insect cell system.

A clone of a human Bone Morphogenetic Protein-2 (hBMP-2) cDNA was obtained from a cDNA library established from human dental pulp cells. After subcloning hBMP-2 cDNA into Autographa californica nuclear polyhedrosis virus, the recombinant baculovirus was transfected to Sf-9 cells. Immuno-reactive recombinant hBMP-2 (rhBMP-2) was detected by a polyclonal antibody against Xenopus BMP-2 in the transfected insect cells but not in the culture media. Three days after treatment with the lysate of the transfected Sf-9 cells, increase in alkaline phosphatase activity of a murine stromal cell line, ST2, was detected. Subcutaneous implantation of rhBMP-2 produced in the insect cells induced formation of cartilage, bone and bone marrow in the rats. The present data indicated that the rhBMP-2 preparation produced in the insect Sf-9 cells had a comparable activity to that produced in mammalian cells.

Alkaline Phosphatase↗

PubMed1995

Mechanism of mutagenesis induced by cytosine analogs bearing N(4)-substitutions.

The mechanism of mutagenesis induced by dihydropyrimido [4,5-c][1,2]oxazin-7-one deoxyriboside, P-nucleoside, was studied. This analog is highly mutagenic toward Escherichia coli and Salmonella typhimurium. In E. coli, it induces GC-to-AT and AT-to-GC transitions specifically. No transversions are inducible. P-nucleoside was highly mutagenic to a wild-type E. coli, but little mutagenic in a strain lacking thymidine kinase. This indicates that P-nucleoside may be phosphorylated by thymidine kinase after its uptake into bacteria. The mutagenesis induced by P-nucleoside was efficiently inhibited by the addition of thymidine. This inhibition further confirmed the involvement of thymidine kinase in the first step of the metabolism of P-nucleoside in the bacterial cells. These findings indicate that P-nucleoside is a mutagen of a nucleoside-analog type, causing mutations by the erroneous incorporation and replication. The experiments to prove its ambiguous nature in DNA synthesis is now under way.

Adenine↗

PubMed1994

Modulation of responses to TGF-beta by 1, 25 dihydroxyvitamin D3 in MG-63 osteoblastic cells: possible involvement of regulation of TGF-beta type II receptor.

The effect of TGF-beta 1 on the expression of alkaline phosphatase (ALPase) activity was examined during osteoblastic cell line (MG-63) differentiation induced by 1,25-dihydroxyvitamin D3 (1,25D3). TGF-beta 1 and 1,25D3 were found to enhance ALPase activity. However, preincubation of the cells with 1,25D3 transiently abolished the effects of TGF- beta 1. Kinetics of the complex responses to TGF- beta 1 and 1,25D3 were found to correlate well with that of the expression level of type II receptor for TGF- beta. These results suggest that 1,25D3 may regulate the cellular responses to TGF- beta 1 in part via regulation of functional receptor for TGF- beta.

Alkaline Phosphatase↗

PubMed1994

Molecular cloning of rat bone morphogenetic protein (BMP) type IA receptor and its expression during ectopic bone formation induced by BMP.

A cDNA for the rat bone morphogenetic protein (BMP) type IA receptor (BMPR-IA) was isolated from a dental pulp cell cDNA library. The rat BMPR-IA cDNA encodes a protein of 532 amino acids with a single transmembrane domain and a putative serine/threonine kinase domain. The overall amino acid sequence identity between the rat and human BMPR-IA was 97%. Reverse transcriptase-polymerase chain reaction analysis revealed that BMPR-IA mRNA was highly expressed in the BMP-induced bone forming tissues throughout the stages tested.

Amino Acid Sequence↗

PubMed1994

Changes in homeobox-containing gene expression during ectopic bone formation induced by bone morphogenetic protein.

Expression of homeobox genes in relation to ectopic bone formation induced by bone morphogenetic protein (BMP) was investigated. Oligonucleotide primers corresponding to highly conserved regions of Hox cluster and Msx genes were designed to detect homeobox sequences by means of the polymerase chain reaction (PCR). Nine rat homologues of Hox cluster genes and two Msx genes were discovered in the BMP-implanted tissue, at earlier stage and later cartilage and bone formation stage, respectively. The PCR study provided evidence of dynamic changes in BMP-induced homeobox gene expression.

Amino Acid Sequence↗

PubMed1994

[Molecular cloning and expression of homeobox-containing genes during hard tissue development].

Although much is known about the hormonal regulation of hard tissue development, much less is known about the nuclear regulatory molecules that affect the process. Homeobox-containing genes are thought to encode DNA-binding transcriptional factors which control the expression of other genes. In this study, molecular cloning of Msx homeobox-containing genes from a bovine odontoblast library and a human dental pulp-derived cells library was performed, and also the expression of mRNAs for Hox and Msx homeobox-containing genes during ectopic bone formation induced by BMP was investigated. Screening of a bovine odontoblast cDNA library and a human dental pulp-derived cells library with murine Msx-1 and Msx-2 cDNA probes led to the isolation of several positive clones. All of the clones from a bovine odontoblast library encoded the bovine counterpart for human MSX-1. All of the clones from a human dental pulp-derived cells library encoded the human counterpart for murine Msx-2. Northern blot analysis probed with a human MSX-2 cDNA indicated the expression of 2.2kb and 1.2kb mRNA in human dental pulp-derived cells. Polymerase chain reaction (PCR)-based analysis in the BMP-implanted tissue revealed that nine rat homologues of Hox homeobox-containing genes were expressed in the early cell migration stage and that two Msx genes were expressed in the cartilage and bone differentiation stages. The PCR study provided evidence of dynamic changes in the BMP-induced homeobox gene expression.

Amino Acid Sequence↗

PubMed1988