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271.
Kizawa H Nishi K Ishibashi Y Harada M Asano T Ito Y Suzuki N Hinuma S Fujisawa Y Onda H Nishimura O Fujino M 《Regulatory peptides》2003,113(1-3):79-84
Relaxin 3 has been reported recently as a member of the insulin/IGF/relaxin family. To clarify the function of relaxin 3, we prepared recombinant human relaxin 3 using a mouse adrenocorticotrophic hormone (ACTH)-secreting cell line, AtT20. To detect a mature form of recombinant human relaxin 3, a competitive enzyme immunoassay (EIA) was developed using a monoclonal antibody (mAb; HK4-144-10), which was raised for the N-terminal peptide of human relaxin 3 A-chain. We detected immunoreactive (ir-) relaxin 3 in the culture supernatant of AtT20 cells stably transfected with human relaxin 3 cDNA. After treatment with 5 microM forskolin for 3 days, the concentration of the ir-relaxin 3 in the culture supernatant reached 12 nM. Ir-relaxin 3 was purified from the culture supernatant by a combination of various chromatographies. By analyses of N-terminal amino acid sequence and electrospray ionization mass spectrometry (ESI-MS), we confirmed that the purified material was a mature form of human relaxin 3. The recombinant human relaxin 3 thereby obtained increased intracellular cAMP production in THP-1 cells. Our results demonstrate that the expression of relaxin 3 cDNA in AtT20 cells is a useful tool to produce a bioactive and mature form of relaxin 3. 相似文献
272.
Proton translocation by the vacuolar (H+)-ATPase (or V-ATPase) has been shown by mutagenesis to be dependent upon charged residues present within transmembrane segments of subunit a as well as the three proteolipid subunits (c, c', and c"). Interaction between R735 in TM7 of subunit a and the glutamic acid residue in the middle of TM4 of subunits c and c' or TM2 of subunit c" has been proposed to be essential for proton release to the luminal compartment. In order to determine whether the helical face of TM7 of subunit a containing R735 is capable of interacting with the helical face of TM4 of subunit c' containing the essential glutamic acid residue (Glu-145), cysteine-mediated cross-linking between these subunits in yeast has been performed. Cys-less forms of subunits a and c' as well as forms containing unique cysteine residues were constructed, introduced together into a strain disrupted in both endogenous subunits, and tested for growth at neutral pH, for assembly competence and for cross-linking in the presence of cupric-phenanthroline by SDS-PAGE and Western blot analysis. Four different cysteine mutants of subunit a were each tested pairwise with ten different unique cysteine mutants of subunit c'. Strong cross-linking was observed for the pairs aS728C/c'I142C, aA731C/c'E145C, aA738C/c'F143C, aA738C/c'L147C, and aL739C/c'L147C. Partial cross-linking was observed for an additional 13 of 40 pairs analyzed. When arrayed on a helical wheel diagram, the results suggest that the helical face of TM7 of subunit a containing Arg-735 interacts with the helical face of TM4 of subunit c' centered on Val-146 and bounded by Glu-145 and Leu-147. The results are consistent with a possible rotational flexibility of one or both of these transmembrane segments as well as some flexibility of movement perpendicular to the membrane. 相似文献
273.
Ikeda K Torisawa Y Nishi T Minamikawa J Tanaka K Sato M 《Bioorganic & medicinal chemistry》2003,11(14):3073-3076
A combined system of bismuth triflate [Bi(OTf)(3)] and boron trifluoride etherate (BF(3).OEt(2)) in dichloromethane is an efficient promoter for the glycosylation of N-acetylneuraminic acid derivatives. The co-existence of two acid catalysts such as Bi(OTf)(3)-BF(3).OEt(2) or Bi(OTf)(3)-PPA is confirmed to be essential for obtaining high yields of glycosylation products with p-nitrobenzyl alcohol, which also turned to be superior to those reported previously. 相似文献
274.
Yoshii T Magara S Miyai D Nishimura H Kuroki E Furudoi S Komori T Ohbayashi C 《Cytokine》2002,19(2):59-65
The purpose of this study is to evaluate local levels of interleukin-1 beta (IL-1 beta), -4 (IL-4), -6 (IL-6), and tumour necrosis factor-alpha (TNF-alpha), in a model of murine osteomyelitis due to Staphylococcus aureus.Cytokine levels in supernatants derived from bone homogenates were determined by enzyme-linked immunosorbent assay, for 28 days following the direct implantation of murine tibiae with S.aureus. Levels of IL-1 beta and IL-6 in infected bone were elevated in the early post-infection period and then decreased. In contrast, TNF-alpha levels remained elevated 3 to 28 days post-infection, while IL-4 levels were elevated late in the course of infection. The histopathology of infected bone showed predominant infiltration of inflammatory cells and bone resorption 3 to 7 days after infection, and bone resorption and adjacent areas of formation 14 to 28 days after infection. These results suggest that the elevated IL-1 beta and IL-6 levels induced by infection may be related to bone damage mainly in the early phase of infection, and that TNF-alpha and IL-4 may at least in part be associated with histopathological changes, including both bone resorption and formation in the later phase of this osteomyelitis model. 相似文献
275.
276.
Atsubo Kushi Koji Edamura Masato Noguchi Kiyotaka Akiyama Yoshisuke Nishi Hitoshi Sasai 《Mammalian genome》1998,9(4):269-273
A method of generating mice from embryonic stem (ES) cells with a large chromosomal deletion produced by X-ray irradiation
has been developed. Fifty-two mutant ES clones were made that carried a nested set of chromosomal deletions up to approximately
10 cM in length around the hprt locus on the X Chromosome (Chr). Germline chimeras were generated from three ES clones with deletions ranging from 200 to
700 kb. In germline male mice from two independent clones, deletions around the hprt locus yielded a runty phenotype or caused death at birth. The runty mice had approximately ⅓ the body weight and size of
wild littermates and did not survive more than 3 weeks after birth. The most plausible cause of these phenotypes is defects
in regions flanking the hprt locus. This method of creating mutant mice with a large chromosomal deletion is very useful for the identification and understanding
of gene functions.
Received: 31 October 1996 / Accepted: 16 December 1997 相似文献
277.
Nobuhiko Sugano Kaoru Koide Yoshinori Ogawa Yoshiko Moriya Arasuke Nishi 《Phytochemistry》1978,17(8):1235-1237
The levels of glutamic acid dehydrogenase (GDH), phenylalanine ammonia-lyase (PAL), cinnamic acid 4-hydroxylase (CAH) and O-methyltransferase (OMT) were measured during the formation of phenolic acids in carrot cells in suspension culture. Caffeic, ferulic and p-hydroxybenzoic acids were always present as the culture proceded. Total content of these acids increased at the early logarithmic and linear phases. GDH showed high activity at the early logarithmic and stationary phases. PAL activity was much enhanced at the linear and stationary phases. CAH activity was found in actively growing cells, especially at the early and late logarithmic phases OMT behaved similarly to PAL. The increases in GDH and CAH might be responsible for the rapid synthesis of phenolic acid at the early logarithmic phase. The increase in phenolic acid at the linear phase would certainly be due to enhancements of both PAL and OMT. On the other hand, the accumulation of vanillic acid was observed in cells which were transferred and cultured on an agar medium, but not in cells in suspension culture. This accumulation is related to increases in OMT levels and also to changes in the degree of β-oxidation. 相似文献
278.
A method of direct measurement of the heat and vapor transfer properties of clothing is described when worn in situ by human subjects during rest and exercise. Additional experimental observations were used to verify that a model for predicting Fpcl is reasonably accurate. From the thermal efficiency factor (Fcl), derived from
s and Tcl measurements, an effective value for Iclo can be determined and thus vapor permeation efficiency (Fpcl) can be predicted. A single coefficient () that characterizes the boundary environment over the skin surface is the ratio (hFcl)/(heFpcl) and is determined for any environment in terms of directly observed Fcl factor or clo value and relative air movement concerned. Ultimately,-factor may be used as a key biometeorological parameter to describe any heat balance condition graphically on a psychrometric chart. Its usefulness is also evident in prediction of the probable combination of operative temperature and humidity necessary for thermal acceptable zones, the limits for evaporative regulation, and the zone in which some degree of thermal regulatory failure can be expected.Presented at the Seventh International Biometeorological Congress, 17–23 August 1975, College Park, Maryland, USA. 相似文献
279.
280.
The spatial development of the cultivation of a bone marrow stromal cell line (SR-4987) in porous carriers was investigated
in order to construct a three-dimensional hematopoietic culture system. Low-rate continuous agitation, 20 rpm, was an appropriate
method to achieve initial adhesion of cells onto a cellulose porous beads (CPB, 100 μm pore diameter) in a spinner bottle,
compared with other methods such as centrifugation and intermittent agitation. Cell growth with continuous agitation at 70
rpm after initial cell adhesion was not inferior to that at 20 rpm. A 2- and 10-fold increase in the inoculum cell concentration
for CPB and another type of porous cellulose beads (Micro-cube (MC), 500 μm pore diameter) resulted in a 1.2- and 2-fold increase
in final cell concentrationm, respectively. Cells attached to the MC beads and a polyester nonwoven dic (Fibra-cell (FC))
could grow and spread well on the carriers and a fibroblast-like shape was observed under scanning electron microscopy while
the cells on CPB were globular. The flatness and inner surface area of these carriers may be the reason for the differences
in cell morphology.
This revised version was published online in July 2006 with corrections to the Cover Date. 相似文献