Establishment of a novel human myeloid leukaemia cell line (FKH-1) with t(6;9)(p23;q34) and the expression of dek-can chimaeric transcript

Calcein-labeled B16 melanoma (B16M) cells were injected intraportally, and in vivo video microscopy was used to study the distribution and damage of cancer cells arrested in the liver microvasculature over a period of 4 hours. The contribution of glutathione (GSH)-dependent antioxidant machinery to the possible oxidative stress-resistance mechanism of B16M cell was determined by in vitro incubation with the selective inhibitor of GSH synthesis L-buthionine (S,R)-sulphoximine (BSO) before B16M cell injection in untreated and 0.5-mg/kg lipopolysaccharide (LPS)-treated mice. In addition, untreated and LPS-treated isolated syngeneic hepatic sinusoidal endothelial cells (HSE) were used to determine in vitro their specific contribution to B16M cell damage. Trauma inherent to intrasinusoidal lodgement damaged 35% of B16M cells in both normal and LPS-treated mouse liver. The rest of the arrested B16M cells remained intact in normal liver for at least 4 hours, although their damaged cell percentage significantly (P < .05) increased since the second hour in normal mice injected with BSO-treated cells and since the first hour in LPS-treated mice given untreated cells. Recombinant human interleukin-1 receptor antagonist (rHuIL-1-Ra) given to mice 15 minutes before LPS significantly (P < .05) abrogated B16M cell damage. On the other hand, 40% of the B16M cells co-cultured with unstimulated HSE and 70% of the co-cultured with LPS-treated HSE became sensitive to endothelial cell-mediated damage after BSO treatment. These results demonstrate that a high intracellular level of GSH protects B16M cells from possible in vivo and in vitro sinusoidal cell-mediated oxidative stress, contributing to the mechanism of metastatic cell survival within the hepatic microvasculature.     

Caspase-independent cell killing by Fas-associated protein with death domain

The binding of Fas ligand to Fas recruits caspase 8 to Fas via an adaptor, FADD/MORT1, and activates a caspase cascade leading to apoptosis. Here, we describe a human Jurkat-derived cell line (JB-6) that is deficient in caspase 8. This cell line was resistant to the apoptosis triggered by Fas engagement. However, the multimerization of Fas-associated protein with death domain, through the use of a dimerizing system, killed the JB-6 cells. This killing process was not accompanied by the activation of caspases or DNA fragmentation. The dying cells showed neither condensation nor fragmentation of cells and nuclei, but the cells and nuclei swelled in a manner similar to that seen in necrosis. These results suggested that Fas-associated protein with death domain can kill the cells via two pathways, one mediated by caspases and another that does not involve them.     

Scanning electron microscopy and immunohistochemical observations of the vascular nerve plexuses in the dental pulp of rat incisor

Elimination of auditory nerve activity results in atrophy and death of nucleus magnocellularis (NM) neurons in the chick. One early event in the degeneration of NM neurons is a disruption of their ribosomes. This experiment examines the role of metabotropic glutamate receptors in afferent regulation of ribosomes. The auditory nerve on one side of a chick brainstem slice was stimulated in vitro. Rapid stimulation-dependent changes in ribosomes were visualized by immunolabeling using an antibody, called Y10B, that recognizes ribosomal RNA. In normal media, NM neurons on the stimulated side of the slice show greater Y10B labeling than the unstimulated NM neurons on the opposite side of the same slice. The role of metabotropic glutamate receptors was evaluated by unilaterally stimulating the auditory nerve in media containing the metabotropic glutamate receptor antagonist (RS)-alpha-methyl-4-carboxyphenyl-glycine (MCPG). Addition of MCPG to the bath did not block EPSPs produced by stimulating the auditory nerve. However, MCPG did prevent the stimulation-dependent regulation of ribosomes in NM neurons (as indexed by Y10B labeling). These data suggest that glutamate may play a trophic role in the young auditory system through activation of metabotropic glutamate receptors.     

The hydrogen-bonding network of water molecules and the peptide backbone in the region connecting Asp83, Gly120, and Glu113 in bovine rhodopsin

Difference Fourier transform infrared spectra were recorded between mutants of rhodopsin and their batho products. The pigments studied were single and combined mutants of intramembrane residues of bovine rhodopsin: Asp83, Glu113, Gly120, Gly121, and Glu122. Previous studies [Nagata, T., Terakita, A., Kandori, H., Kojima, D., Shichida, Y., and Maeda, A. (1997) Biochemistry 36, 6164-6170] showed that one of the water molecules which undergoes structural changes in this process forms hydrogen bonds with Glu113 and the Schiff base, and that another water molecule is linked to this structure through the peptide backbone. The present results show that this water molecule is located at the place that is affected by the replacements of Asp83 and Gly120 but only slightly by Gly121 and not at all by Glu122. Asp83 and Gly120 are close to each other, in view of the observations that the carboxylic C=O stretching vibration of Asp83 is affected by the G120A replacement and that each replacement affects the common peptide carbonyl groups. Our results suggest that these residues in the middle of helices B and C are linked-through a hydrogen-bonding network composed of water and the peptide backbone-with the region around Glu113.     

Correlation Between Superconducting and Magnetic Transition Temperatures of R1−x Nd x Ni2B2C (R=Y, Ho, Er)

The variations of the superconducting transition temperature T _c and the magnetic transition temperature T _N as a function of the Nd concentration x in R _1–x Nd _x Ni ₂ B ₂ C (R=Y, Ho, and Er) systems is systematically studied by the measurements of electrical resistivity and magnetization. It is found that the Nd substitution in R _1–x Nd _x Ni ₂ B ₂ C show an effect to break Cooper pairs. The de Gennes scaling on the variation of T _c is also discussed.     

Gel biomachine based on muscle proteins

Summary We have created an ATP-fueled soft gel machine constructed from muscle proteins. Chemically cross-linked gels of the polymer-actin complex of the length several decades times the length of native actin filament (F-actin) move on myosin-coated surface with a velocity as high as that of native F-actin, by coupling to ATP hydrolysis. The motility observed in muscle protein-gels suggests that one might construct a soft machine fueled by chemical energy using actin and myosin molecules as elements. We have investigated the growth process of polymer-actin complexes and the correlation between the polarity and the motility of polymer-actin complex gels.     

Transforming growth factor-beta1 increases mRNA levels of osteoclastogenesis inhibitory factor in osteoblastic/stromal cells and inhibits the survival of murine osteoclast-like cells

Osteoclastogenesis inhibitory factor (OCIF), also termed osteoprotegerin (OPG), is a secreted member of the tumor necrosis factor (TNF) receptor family. It inhibits bone resorption in vivo and osteoclast-like cell (OCL) formation in vitro. To better understand the biological role of OCIF, we first examined the effects of various osteotropic agents on OCIF mRNA levels in mouse calvarial osteoblasts. Northern blot analysis showed that stimulators of OCL formation such as 1,25-(OH)2D3, prostaglandin E2 (PGE2), parathyroid hormone (PTH), and interleukin 1 (IL-1) decreased OCIF mRNA levels. In contrast, transforming growth factor (TGF)-beta1 increased OCIF mRNA levels in primary osteoblasts as well as in osteoblastic/stromal cell lines. Since it was reported that both TGF-beta1 and OCIF not only inhibited OCL formation but also impaired the survival of OCL by inducing apoptosis in vitro, we next examined the possible involvement of OCIF in TGF-beta1-induced impairment of OCL survival. In a mouse bone marrow culture, we confirmed that addition of OCIF or TGF-beta1 decreased the number of surviving OCL. Anti-OCIF IgG, which completely neutralized the effect of OCIF, partially prevented the TGF-beta1-induced decrease in the number of OCL. Our results suggest that (i) downregulation of OCIF expression is one of the mechanisms for the stimulatory effects of 1,25(OH)2D3, PGE2, PTH, and IL-1 on osteoclastogenesis; and (ii) the TGF-beta1-induced apoptosis of OCL is mediated, at least in part, by upregulation of OCIF expression.     

Preparation of an Organic-Inorganic Hybrid Ionic Conductive Material with Thermal and Chemical Stability

An organic–inorganic hybrid ionic conductive material was prepared through oxidation of thiol groups in a surface-modified porous glass. Surface-modified pores in the hybrid materials acted as permeation paths for protons. Proton conductivity of the hybrid material was 4.2 × 10⁻² S/cm at 120°C under 100% relative humidity. This hybrid material was thermally stable at high temperature (above 100°C) due to the temperature-tolerant inorganic frameworks.     

HSP47, a collagen-specific molecular chaperone, delays the secretion of type III procollagen transfected in human embryonic kidney cell line 293: a possible role for HSP47 in collagen modification

HSP47 is a stress protein (heat shock protein) which resides in the endoplasmic reticulum, and is postulated to function as a collagen-specific molecular chaperone. To elucidate the role of HSP47 in procollagen biosynthesis, we have established human embryonic kidney 293 cell lines, which were stably transfected with alpha1(III) procollagen chains with or without HSP47. 293 cells do not produce any extracellular matrix proteins including collagens, and the level of HSP47 expression is almost undetectable in this cell line. Recombinant type III procollagens in 293 cells form trypsin-resistant homotrimers, which are secreted into the medium as trimers in the presence or absence of recombinant mouse HSP47. The secretion of procollagen III was delayed in 293 cells stably transfected with proalpha1(III) collagen chains [293+proalpha1(III) cells] in comparison with human rhabdomyosarcoma cell line RD, which normally produces type III procollagens. In this study, we examined the rate of type III procollagen secretion in detail. In cells cotransfected with mouse HSP47 [293+proalpha1(III)+HSP47 cells], the rate of type III procollagen secretion was slower than in 293+proalpha1(III) cells. The binding of HSP47 with proalpha1(III) collagen chains was confirmed by immunoprecipitation using the chemical cross-linker, DSP. The electrophoretic mobility of proalpha1(III) collagen chains in 293+proalpha1(III) cells was slightly slower than that in RD cells, whereas the recombinant proalpha1(III) chains of 293+proalpha1(III)+HSP47 cells showed almost the same electrophoretic mobility as those of RD cells. The melting temperature (Tm) of type III procollagen in 293+proalpha1(III)+HSP47 cells was almost the same as that in RD cells, and the Tm in 293+proalpha1(III) cells was slightly higher than that in RD cells. These data suggest that the recombinant proalpha1(III) collagen chain is overmodified in 293+proalpha1(III) cells, but not in 293+proalpha1(III)+HSP47 cells.