Shingo Niimi

Rhodamine 6G, inhibitor of both H+-ejections from mitochondria energized with ATP and with respiratory substrates.

Rhodamine 6G inhibited ATP hydrolysis by oligomycin-sensitive ATPase, purified from rat liver mitochondria, in good accord with the dose-response curve for its inhibition of energy transduction of ATP synthesis in mitochondria, but it did not inhibit ATP hydrolysis by purified F1. Rhodamine 6G also inhibited both H+-ejections from mitochondria energized with respiratory substrates and with ATP. The present findings show that the inhibitory effect of rhodamine 6G on energy transduction is not due to a modification of the transport system for adenine nucleotides, Pi, and respiratory substrates, and that the inhibition sites of rhodamine 6G are on components related with H+-ejection by redox components and also on F0.

Changes in peroxisome proliferator-activated receptor γ-regulated gene expression and inhibin/activin–follistatin system gene expression in rat testis after an administration of di-n-butyl phthalate

Changes in gene expression regulated by peroxisome proliferator-activated receptor gamma (PPARgamma) and in gene expression related to the inhibin/activin-follistatin system in the rat testis induced by a single oral administration of di-n-butyl phthalate (DBP) (8.6 mmol/kg) were examined and compared with those in the control rats using reverse-transcriptase polymerase chain reaction (RT-PCR). The increase in cytochrome P450 4A1 mRNA, which is regulated by PPARalpha, was significant, but not so profound as the increase of P450 4A1 mRNA in the liver. In contrast, a remarkable increase in the mRNA level of plasminogen activator inhibitor-1 (PAI-1) was found in the testis, suggesting the activation of PPARgamma. The substantial increase in PAI-1 may be related to the disruption of spermatogenesis. On the other hand, significant suppression of the mRNA level of inhibin beta(B) and elevation in the mRNA level of follistatin, an activin-binding protein, were observed after the DBP-administration. Activin B, a homodimer of inhibin beta(B), is known to stimulate spermatogonial proliferation. The present results suggest that the suppression of spermatogenesis resulting from the changes in the expression of genes involved in the inhibin/activin-follistatin system is one of the mechanisms of the testicular atrophy induced by DBP.

Hormonal regulation of translatable mRNA of glucose-6-phosphate dehydrogenase in primary cultures of adult rat hepatocytes

The quantity of translatable mRNA of glucose-6-phosphate dehydrogenase (D-glucose-6-phosphate: NADP+ 1-oxidoreductase, EC 1.1.1.49) in primary cultures of adult rat hepatocytes subjected to different hormonal conditions was determined with a reticulocyte-lysate, cell-free system. The level of glucose-6-phosphate dehydrogenase mRNA was about 5-fold higher in the presence of insulin than in its absence. This increase of glucose-6-phosphate dehydrogenase mRNA reached a maximum 12 h after the addition of insulin. The maximum level of induction of glucose-6-phosphate dehydrogenase mRNA required 10(-8) M insulin. Glucagon and triiodothyronine had no effect on the glucose-6-phosphate dehydrogenase mRNA level. The increase of glucose-6-phosphate dehydrogenase activity correlated with the increase in level of mRNA of this enzyme. This suggests that the changes in glucose-6-phosphate dehydrogenase activity in response to the above hormonal changes are primarily due to changes in the amount of mRNA coding for this enzyme.

Effect of cell density on induction of growth hormone receptors by dexamethasone in primary cultured rat hepatocytes

The effect of cell density on the regulation of growth hormone (GH) receptors was studied by measuring specific binding of [125I]hGH to primary cultured hepatocytes with or without dexamethasone, which induces GH receptors. In cell cultures without dexamethasone, the cell density did not affect the level of binding of labeled GH appreciably. On the other hand, in the presence of dexamethasone, which induced an increase in the level of GH receptors on the cells, GH-binding by cultured cells at low cell density (3.3 x 10(4) cells/cm2) was about one-third of that of cells at high cell density (10(5) cells/cm2). Scatchard plot analysis showed that the cell-density dependent change in induction of GH binding, by dexamethasone was due to change in the number of binding sites without significant change in their affinity. The binding capacity of glucocorticoid receptors, measured as specific binding of [3H]dexamethasone to the hepatocytes, was not significantly affected by cell density. These results suggest that cell density modulates GH receptor induction by dexamethasone via events after glucocorticoid receptor binding.

Corrigendum to "Characterization of the cell growth analysis for detection of immortal cellular impurities in human mesenchymal stem cells" [Biologicals 43 (2) (March 2015) 146-149].

Corrigendum to “Characterization of the cell growth analysis for detection of immortal cellular impurities in human mesenchymal stem cells” [Biologicals 43 (2) (March 2015) 146e149] Ken Kono , Nozomi Takada b, , Satoshi Yasuda b, , Rumi Sawada , Shingo Niimi , Akifumi Matsuyama , Yoji Sato b, d, e, f, g, * a Division of Medical Devices, National Institute of Health Sciences, 1-18-1 Kami-yoga, Setagaya, Tokyo, 158-8501, Japan b Division of Cellular & Gene Therapy Products, National Institute of Health Sciences, 1-18-1 Kami-yoga, Setagaya, Tokyo, 158-8501, Japan c Research on Disease Bioresources, Platform of Therapeutics for Rare Disease and Health Policy, National Institute of Biomedical Innovation, Kobe International Business Center Rm#602, 5-5-2 Minatojima-Minami-Machi, Chuo-ku, Kobe, Hyogo, 650-0047, Japan d Foundation for Biomedical Research and Innovation, Hyogo, Japan e Department of Quality Assurance Science for Pharmaceuticals, Graduate School of Pharmaceutical Sciences, Nagoya City University, Aichi, Japan f Department of Cellular & Gene Therapy Products, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka, Japan g Department of Translational Pharmaceutical Sciences, Graduated School of Pharmaceutical Sciences, Kyushu University, Fukuoka, Japan

Study of hepatocytes using RNA interference

RNA interference (RNAi) is the process of sequence-specific gene silencing, initiated by small double-stranded RNA homologous in sequence to the target gene. Various factors involved in the regulation of hepatocyte function have been identified using RNAi, indicating that RNAi is a useful strategy for characterization. There has been some success in treating experimental liver dysfunction using RNAi in several model systems, suggesting a promising new therapeutic strategy. A number of groups have also demonstrated that RNAi can interfere with hepatitis C virus and hepatitis B virus gene expression and replication in several model systems, suggesting a new approach for the treatment of these viral diseases. This review summarizes studies of hepatocytes using RNAi.