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71.
Yoshihito Shirai Kenji Hashimoto Hideki Yamaji Hiroyuki Kawahara 《Applied microbiology and biotechnology》1988,29(2-3):113-118
Summary The specific oxygen uptake rate of hybridoma cells immobilized in calcium alginate gel particles was measured, and the observed data was compared with those of non-immobilized cells. The uptake rate of the immobilized cells coincided with that of the non-immobilized hybridoma cells just after immobilization, but increased with cell growth. On the other hand, the cellular glucose consumption rate decreased slightly during the experiments. The increased oxygen uptake rate by immobilized cells was closely related to the formation of cell colonies in the gel particles. 相似文献
72.
Morita A Nakahira K Hasegawa T Uchida K Taniguchi Y Takeda S Toyoda A Sakaki Y Shimada A Takeda H Yanagihara I 《Development, growth & differentiation》2012,54(5):588-604
Roberts syndrome and SC phocomelia (RBS/SC) are genetic autosomal recessive syndromes caused by establishment of cohesion 1 homolog 2 ( ESCO 2) mutation. RBS/SC appear to have a variety of clinical features, even with the same mutation of the ESCO2 gene. Here, we established and genetically characterized a medaka model of RBS/SC by reverse genetics. The RBS/SC model was screened from a mutant medaka library produced by the Targeting Induced Local Lesions in Genomes method. The medaka mutant carrying the homozygous mutation at R80S in the conserved region of ESCO2 exhibited clinical variety (i.e. developmental arrest with craniofacial and chromosomal abnormalities and embryonic lethality) as characterized in RBS/SC. Moreover, widespread apoptosis and downregulation of some gene expression, including notch1a, were detected in the R80S mutant. The R80S mutant is the animal model for RBS/SC and a valuable resource that provides the opportunity to extend knowledge of ESCO2. Downregulation of some gene expression in the R80S mutant is an important clue explaining non-correlation between genotype and phenotype in RBS/SC. 相似文献
73.
Takei D Ishihara H Yamaguchi S Yamada T Tamura A Katagiri H Maruyama Y Oka Y 《FEBS letters》2006,580(24):5635-5640
The WFS1 gene, encoding an endoplasmic reticulum (ER) membrane glycoprotein, is mutated in Wolfram syndrome characterized by diabetes mellitus and optic atrophy. Herein, Ca(2+) dynamics were examined in WFS1-knockdown and -overexpressing HEK293 cells. Studies using ER-targeted Ca(2+)-sensitive photoprotein aequorin demonstrated WFS1 protein to positively modulate ER Ca(2+) levels by increasing the rate of Ca(2+) uptake. Furthermore, Ca(2+) imaging with Fura-2 showed the magnitude of the store-operated Ca(2+) entry to parallel WFS1 expression levels. These data indicate that WFS1 protein participates in the regulation of cellular Ca(2+) homeostasis, at least partly, by modulating the filling state of the ER Ca(2+) store. 相似文献
74.
Satoko Uraki Masahiko Tameda Kazushi Sugimoto Katsuya Shiraki Yoshiyuki Takei Tsutomu Nobori Masaaki Ito 《PloS one》2015,10(6)
Background & Aims
It has been suggested that amino acid (aa) substitution at position 70 from arginine (70R) to glutamine (70Q) in the genotype 1b hepatitis C virus (HCV) core protein is associated with insulin resistance and worse prognosis. However, the precise mechanism is still unclear. The aim of this study was to investigate the impact of the substitution at position 70 in HCV core protein on adipokine production by murine and human adipocytes.Methods
The influence of treatment with HCV core protein (70R or 70Q) on adipokine production by both 3T3-L1 and human adipocytes were examined with real-time PCR and enzyme-linked immunosorbent assay (ELISA), and triglyceride content was also analyzed. The effects of toll-like receptor (TLR)2/4 inhibition on IL-6 production by 3T3-L1 induced by HCV core protein were examined.Results
IL-6 production was significantly increased and adiponectin production was reduced without a change in triglyceride content by treatment with 70Q compared to 70R core protein in both murine and human adipocytes. IL-6 induction of 3T3-L1 cells treated by 70Q HCV core protein was significantly inhibited with anti-TLR2 antibody by 42%, and by TLR4 inhibitor by 40%.Conclusions
Our study suggests that extracellular HCV core protein with substitution at position 70 enhanced IL-6 production and reduced adiponectin production from visceral adipose tissue, which can cause insulin resistance, hepatic steatosis, and ultimately development of HCC. 相似文献75.
Nobukatsu Horita Kiichiro Tsuchiya Ryohei Hayashi Keita Fukushima Shuji Hibiya Masayoshi Fukuda Yoshihito Kano Tomohiro Mizutani Yasuhiro Nemoto Shiro Yui Ryuichi Okamoto Tetsuya Nakamura Mamoru Watanabe 《Biochemical and biophysical research communications》2014
Background and aims
The dynamics of intestinal stem cells are crucial for regulation of intestinal function and maintenance. Although crypt stem cells have been identified in the intestine by genetic marking methods, identification of plural crypt stem cells has not yet been achieved as they are visualised in the same colour.Methods
Intestinal organoids were transferred into Matrigel® mixed with lentivirus encoding mCherry. The dynamics of mCherry-positive cells was analysed using time-lapse imaging, and the localisation of mCherry-positive cells was analysed using 3D immunofluorescence.Results
We established an original method for the introduction of a transgene into an organoid generated from mouse small intestine that resulted in continuous fluorescence of the mCherry protein in a portion of organoid cells. Three-dimensional analysis using confocal microscopy showed a single mCherry-positive cell in an organoid crypt that had been cultured for >1 year, which suggested the presence of long-lived mCherry-positive and -negative stem cells in the same crypt. Moreover, a single mCherry-positive stem cell in a crypt gave rise to both crypt base columnar cells and transit amplifying cells. Each mCherry-positive and -negative cell contributed to the generation of organoids.Conclusions
The use of our original lentiviral transgene system to mark individual organoid crypt stem cells showed that long-lived plural crypt stem cells might independently serve as intestinal epithelial cells, resulting in the formation of a completely functional villus. 相似文献76.
Osami Shoji Takashi Fujishiro Shingo Nagano Shota Tanaka Takuya Hirose Yoshitsugu Shiro Yoshihito Watanabe 《Journal of biological inorganic chemistry》2010,15(8):1331-1339
Cytochrome P450BSβ, a H2O2-dependent cytochrome P450 catalyzing the hydroxylation of long-alkyl-chain fatty acids, lacks the general acid–base residue
around the heme, which is indispensable for the efficient generation of the active species using H2O2. On the basis of the crystal structure of the palmitic acid bound form of cytochrome P450BSβ, it was suggested that the role of the general acid–base function was provided by the carboxylate group of fatty acids. The
participation of the carboxylate group of the substrate was supported by the fact that cytochrome P450BSβ can catalyze oxidations of nonnatural substrates such as styrene and ethylbenzene in the presence of a series of short-alkyl-chain
carboxylic acids as a dummy molecule of fatty acid. We refer to a series of short-alkyl-chain carboxylic acids as a “decoy
molecule”. As shown here, we have clarified the crystal structure of the decoy-molecule-bound form and elucidated that the
location of its carboxylate group is virtually the same as that of palmitic acid in the heme cavity, indicating that the carboxylate
group of the decoy molecule serves as the general acid–base catalyst. This result further confirms that the role of the acid–base
function is satisfied by the carboxylate group of the substrates. In addition, the structure analysis of the substrate-free
form has clarified that no remarkable structural change is induced by the binding of the decoy molecule as well as fatty acid.
Consequently, whether the carboxylate group is positioned in the active site provides the switching mechanism of the catalytic
cycle of cytochrome P450BSβ. 相似文献
77.
Kumi Nagaoka Yoshiaki Kitamura Yoshihito Ueno Yukio Kitade 《Bioorganic & medicinal chemistry letters》2010,20(3):1186-1188
Human ribonuclease L (RNase L), an interferon-induced endoribonuclease, becomes enzymatically active after binding to 2-5A. The 5′-phosphoryl group of 2-5A is reportedly necessary for the conformational change leading to RNase L activation. However, we found that 5′-O-dephosphorylated 2-5A tetramer analogs with 8-methyladenosine at the 2′-terminus were more effective as an activator of RNase L than the parent 2-5A tetramer. Introduction of 8-methyladenosine is thought to induce a dramatic shift of 2-5A in the binding site of RNase L. 相似文献
78.
Yoshito Yamashiro Kimiko Takei Masato Umikawa Minoru Oshiro Takahiro Ishikawa Hiroshi Uezato 《Biochemical and biophysical research communications》2010,399(3):365-2106
Cutaneous squamous cell carcinoma (cSCC) results from transformation of epidermal keratinocytes. Invasion of transformed keratinocytes through the basement membrane into the dermis results in invasive cSCC with substantial metastatic potential. To better understand the mechanisms for invasion and metastasis, we compared the protein expression profiles of a non-metastatic transformed mouse keratinocyte line and its metastatic derivative. Keratin 8 (Krt8) and Krt18, not seen in normal keratinocytes, were coexpressed and formed Krt8/18 filaments in the metastatic line. The metastatic line efficiently invaded an artificial basement membrane in vitro owing to the Krt8/18-coexpression, since coexpression of exogenous Krt8/18 in the non-invasive parental line conferred invasiveness. To test whether the Krt8/18-coexpression is induced and is involved in cSCC invasion, we examined specimens from 21 pre-invasive and 24 invasive cSCC patients by immunohistochemistry, and the ectopic Krt8/18-coexpression was almost exclusively found in invasive cSCC. Further studies are needed to examine the clinical significance of ectopic Krt8/18-coexpression in cSCC. 相似文献
79.
Reduced pain hypersensitivity and inflammation in mice lacking microsomal prostaglandin e synthase-1 总被引:8,自引:0,他引:8
Kamei D Yamakawa K Takegoshi Y Mikami-Nakanishi M Nakatani Y Oh-Ishi S Yasui H Azuma Y Hirasawa N Ohuchi K Kawaguchi H Ishikawa Y Ishii T Uematsu S Akira S Murakami M Kudo I 《The Journal of biological chemistry》2004,279(32):33684-33695
We examined the in vivo role of membrane-bound prostaglandin E synthase (mPGES)-1, a terminal enzyme in the PGE2-biosynthetic pathway, using mPGES-1 knockout (KO) mice. Comparison of PGES activity in the membrane fraction of tissues from mPGES-1 KO and wild-type (WT) mice indicated that mPGES-1 accounted for the majority of lipopolysaccharide (LPS)-inducible PGES in WT mice. LPS-stimulated production of PGE2, but not other PGs, was impaired markedly in mPGES-1-null macrophages, although a low level of cyclooxygenase-2-dependent PGE2 production still remained. Pain nociception, as assessed by the acetic acid writhing response, was reduced significantly in KO mice relative to WT mice. This phenotype was particularly evident when these mice were primed with LPS, where the stretching behavior and the peritoneal PGE2 level of KO mice were far less than those of WT mice. Formation of inflammatory granulation tissue and attendant angiogenesis in the dorsum induced by subcutaneous implantation of a cotton thread were reduced significantly in KO mice compared with WT mice. Moreover, collagen antibody-induced arthritis, a model for human rheumatoid arthritis, was milder in KO mice than in WT mice. Collectively, our present results provide unequivocal evidence that mPGES-1 contributes to the formation of PGE2 involved in pain hypersensitivity and inflammation. 相似文献
80.
Junichi Nemoto Chihiro Harada Yoshihito Takei Naoto Katakura Masao Kaneko 《Photochemical & photobiological sciences》2007,6(1):77-82
Visible light decomposition of aqueous NH3 to N2 was investigated using a photocatalyst aqueous solution based on molecular photoelectron relay systems of either sensitizer (tris(2,2'-bipyridine)ruthenium(II), (Ru(bpy)3(2+))/potassium peroxodisulfate(K(2)S(2)O(8)) or Ru(bpy)3(2+)/methylviologen dichloride(MV2+)/O2, capable of using visible light instead of UV-driven semiconductors such as TiO2. It was confirmed by using an in situ visible absorption spectral change under irradiation that the Ru(II) complex is oxidized to the Ru(III) complex by K(2)S(2)O(8), and that the Ru(III) complex formed is stable without NH3, while the added NH3 was oxidized by the Ru(III) complex to produce the Ru(II) complex. In the presence of 1 mM NH3 aqueous solution, the Ru(III) complex was the predominant species under the photostationary state, but in the presence of 100 mM NH3, Ru(II) predominated. Gas-chromatographic analysis of the gaseous phase in the presence of 8.1 M NH3 showed that the photochemical oxidation of ammonia yielded N2. It was also demonstrated by using the in situ visible absorption spectrum under irradiation of the NH3 (1 M)/Ru(bpy)3(2+) (0.1 mM)/MV2+ (10 mM) system under Ar that MV+* is accumulated, showing that NH3 works as an electron donor for MV+* accumulation with simultaneous formation of the oxidized product of ammonia ((NH3)ox) without producing N2. It was suggested that the reduced product (MV+*) and the oxidized product ((NH3)ox) are in a kind of dynamic equilibrium prohibiting further oxidation of (NH3)ox by Ru(bpy)3(3+) to N2. In the O2 atmosphere, the oxidation of MV+* to MV2+ takes place to accumulate Ru(III) complex, so that (NH3)ox was further oxidized to N2. The high activity of IrO2 as a cocatalyst in this system was demonstrated. 相似文献