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10. Expression and activity of Runx2 mediated by hyaluronan during chondrocyte differentiation

Author

Tanne, Yuki, Tanimoto, Kotaro, Tanaka, Nobuaki, Ueki, Masashi, Lin, Y. Y., Ohkuma, S, Kamiya, Takashi, Tanaka, Eiji, Tanne, Kazuo, Tanne, Yuki, Tanimoto, Kotaro, Tanaka, Nobuaki, Ueki, Masashi, Lin, Y. Y., Ohkuma, S, Kamiya, Takashi, Tanaka, Eiji, and Tanne, Kazuo

Abstract

type:text, During endochondral ossification, the production of hyaluronan (HA) is strictly and selectively regulated by chondrocytes, with a temporal peak at the hypertrophic stage. This study was conducted to clarify the effects of HA on expression and activity of runt-related gene 2 (Runx2), a potent transcription factor for chondrocyte differentiation in hypertrophic chondrocytes. Immature chondrocytes from an ATDC5 cell line were cultured and differentiated in DMEM/Ham's F12 with pre-defined supplements. Using real-time PCR, the gene expressions of type II collagen, MMP-13, HAS2, and Runx2 in cultured chondrocytes were analysed from days 0 to 18 of cell differentiation. The activity and expression of Runx2 in hypertrophic chondrocytes were analysed after the treatment with HA oligosaccharide (HAoligo) using AML-3/Runx2 binding, real-time PCR and Western blot analysis. The effects of pre-incubation of anti-CD44 antibody on Runx2 expression were also examined. Expression of type X collagen and Runx2 mRNAs reached a maximum at the terminal differentiation of chondrocytes. The activity and expression of Runx2 was significantly inhibited in hypertrophic chondrocytes treated with HAoligo compared to the untreated controls. High molecular weight-HA did not affect the expression or activity of Runx2. The expression of Runx2 mRNA was significantly decreased in hypertrophic chondrocytes treated with anti-CD44 antibody. These results suggest that HAoligo may affect the terminal differentiation of chondrocytes during the endochondral ossification by inhibiting the expression and activity of Runx2.

Published
2008

16. Chemotherapeutic choice of ranimustine or nimustine on the basis of regional polyamine levels in rat brain

Author

Satoru Watanabe, Nagase S, Ohkuma S, and Shoichi Sato

Subjects
Male, endocrine system, medicine.medical_specialty, Central nervous system, Spermine, Antineoplastic Agents, Striatum, Biology, Nitrosourea Compounds, Rats, Sprague-Dawley, chemistry.chemical_compound, Diencephalon, Internal medicine, medicine, Animals, Brain Chemistry, Nimustine, Biogenic Polyamines, Rats, Spermidine, Endocrinology, medicine.anatomical_structure, nervous system, chemistry, Putrescine, Polyamine
Abstract

The aim of the present study was to improve the chemotherapeutic efficacy of anticancer drugs by choosing on the basis of the polyamine level induced by the drug in each host cancer-bearing tissue. We propose an "organ-specific therapy" in the article. The polyamines, putrescine, spermidine and spermine are strongly associated with tumor cell growth. The effects of ranimustine (MCNU) and nimustine (ACNU) on body weight, regional brain weights and concentrations of putrescine, spermidine and spermine in the cerebellum, hippocampus, corpus striatum, cortex, combined thalamus and hypothalamus and diencephalon of the brain were examined in rats. MCNU and ACNU reduced spermidine and spermine in the corpus striatum, and spermine in the diencephalon, but increased putrescine in the corpus striatum and combined thalamus and hypothalamus. These results indicate that both MCNU and ACNU are suitable for the treatment of cancers of the corpus striatum, but ACNU is not suitable for cancers of the corpus striatum, thalamus and hypothalamus.

Published
2008
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44. V-Type H+-ATPase/synthase from a thermophilic eubacterium, Thermus thermophilus. Subunit structure and operon.

Author

Yokoyama, K, Ohkuma, S, Taguchi, H, Yasunaga, T, Wakabayashi, T, and Yoshida, M

Abstract

V-type ATPase (V(o)V(1)) capable of ATP-driven H(+) pumping and of H(+) gradient driven ATP synthesis was isolated from a thermophilic eubacterium, Thermus thermophilus. When the enzyme was analyzed by gel electrophoresis in the presence of sodium dodecyl sulfate, it showed eight polypeptide bands of which four were subunits of V(1). We also isolated the V(o)V(1) operon, containing nine genes in the order of atpG-I-L-E-X-F-A-B-D, which encoded proteins with molecular sizes of 13, 43, 10, 20, 35, 11, 64, 53, and 25 kDa, respectively. The last four genes were identified as those for V(1) subunits; atpA, B, D, and F encoded the A, B, gamma, and delta subunits, respectively. The first five genes, atpG-atpX, were identified as genes for the V(o) subunits. The product of atpL, the proteolipid subunit, lacked a 19-amino acid presequence and, unlike V-type ATPases, contained two membrane-spanning domains rather than four. The hydrophobic 43-kDa product of atpI is the smallest member so far found of the eukaryotic 100-kDa subunit family. Its electrophoretic band overlapped with the band of the A subunit. Therefore, all the gene products were found in our purified V(o)V(1). We isolated the A(3)B(3) subcomplex reconstituted from the isolated subunits and the A(3)B(3)gamma subcomplex from subunit-expressing Escherichia coli. Electron microscopic observation of these subcomplexes revealed that the gamma subunit of V(1) filled the central cavity of A(3)B(3) and might be central subunit, similar to the gamma subunit of F(1)-ATPase.

Published
2000

48. Increase of diazepam binding inhibitor mRNA levels in the brains of chronically ethanol-treated and -withdrawn mice.

Author

Katsura, M, Ohkuma, S, Tsujimura, A, and Kuriyama, K

Abstract

Although the molecular mechanisms underlying behavioral responses observed in ethanol (EtOH)-dependent and -withdrawn animals are poorly understood, several lines of evidence have suggested that some of the pharmacological actions of EtOH are mediated via the potentiation of gamma-aminobutyric acid (GABA)ergic transmission. In the present study, the effects of acute and chronic EtOH treatment on the expression of diazepam binding inhibitor (DBI) mRNA in the mouse brain and the functional involvement of the benzodiazepine (BDZ) receptor in EtOH-induced alteration of the DBI mRNA expression were examined. The expression of cerebral DBI mRNA significantly increased in EtOH-treated (130% of control) and EtOH-withdrawn (220% of control) mice in comparison with untreated mice, whereas the DBI mRNA level was not altered after a single administration of EtOH (3 g/kg i.p.). The increase in the DBI mRNA expression in the mouse cerebral cortex after EtOH withdrawal diminished over 14 days despite the disappearance of withdrawal signs within 2 days after the withdrawal of EtOH. Simultaneous administration of flunitrazepam (10 mg/kg i.p.) with EtOH completely abolished the EtOH-induced increase in DBI mRNA expression. On the other hand, the level of beta-actin mRNA was not affected by similar EtOH treatment. These results indicate that changes in the expression of cerebral DBI mRNA induced by long-term EtOH treatment may be involved in the establishment of alcohol dependence, and such changes may be also regulated by BDZ receptors.

Published
1995

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