Identification of sialyltransferases of Streptococcus agalactiae

Group B streptococci, Streptococcus agalactiae, produce high-molecular-weight polysaccharides containing N-acetylneuraminic acid. Although the type-specific capsular polysaccharide (CP) synthesis (cps) genes of several S. agalactiae strains have been extensively analyzed, to date, no sialyltransferase activity has been detected from any gene product of the cps gene cluster. Among the cps genes, the cpsK gene products of S. agalactiae types la and Ib showed weak similarity to several bacterial sialyltransferases. In this study, the cpsIaK and cpsIbK gene products were found to show sialyltransferase activity specific for lacto-N-neotetraose and lacto-N-tetraose, respectively. This acceptor specificity seems to reflect the respective CP structure, since the repeating unit of type la CP is sialyllacto-N-neotetraose and that of type Ib CP is sialyllacto-Ntetraose. We also found that the C-terminal regions of CpsKs were almost completely conserved in various S. agalactiae strains.     

Physical and functional association between thymic shared antigen-1/stem cell antigen-2 and the T cell receptor complex

Thymic shared antigen-1 (TSA-1)/stem cell Ag-2 (Sca-2) is a glycosylphosphatidylinositol (GPI)-anchored antigen expressed on lymphocytes. We have previously demonstrated that a signal via TSA-1/Sca-2 inhibits T cell receptor (TCR)-mediated T cell activation and apoptosis. To elucidate a molecular mechanism for TSA-1-mediated modulation of the TCR-signaling pathway, we examined whether TSA-1 is physically coupled to the TCR in the present study. TSA-1 was clearly associated with CD3zeta chains in T cell hybridomas, activated T cells, and COS-7 cells transfected with TSA-1 and CD3zeta cDNA. The physical association was confirmed on the surface of T cells in immunoprecipitation and confocal microscopy. The analysis using stable and transient transfectants expressing a transmembrane form of TSA-1 revealed that the association of CD3zeta did not require the GPI anchor of TSA-1. Finally, tyrosine phosphorylation of CD3zeta chains was induced after stimulation with anti-TSA-1, suggesting that a functional association between these two molecules also exists. These results imply that the physical association to CD3zeta underlies a regulatory role of TSA-1/Sca-2 in the TCR-signaling pathway.     

A stress-sensitive chemokinergic neuronal pathway in the hypothalamo-pituitary system

Recently we found that cytokine-induced neutrophil chemoattractant influenced anterior pituitary hormone release in vitro. These observations prompted us to investigate the possibility of the existence of cytokine-induced neutrophil chemoattractant in the hypothalamus. Immunohistochemistry showed that cytokine-induced neutrophil chemoattractant-like immunoreactivity existed in the paraventricular hypothalamic nucleus, the supraoptic nucleus, both the internal and the external layers of the median eminence and the posterior pituitary. Since the paraventricular hypothalamic nucleus plays a pivotal role in response to stressful stimuli, we examined the effect of a single episode of immobilization stress on cytokine-induced neutrophil chemoattractant messenger RNA expression in the paraventricular hypothalamic nucleus. Immobilization stress induced strong hybridization signals of cytokine-induced neutrophil chemoattractant messenger RNA in the parvocellular and magnocellular subdivision of the paraventricular hypothalamic nucleus within 15 min, and cytokine-induced neutrophil chemoattractant-like immunostaining intensity in the posterior pituitary started to increase around the periphery of the posterior lobe at 30 min after stress and extended to the whole lobe at 1 h after stress. The increase in the serum cytokine-induced neutrophil chemoattractant in response to stress showed a kinetically biphasic pattern. A first phase occurred within 15 min which may be due to an immediate release of stored cytokine-induced neutrophil chemoattractant in the neurohypophysis, since hypophysectomy completely blocked this phase. A second phase may reflect the release of newly synthesized cytokine-induced neutrophil chemoattractant in the paraventricular hypothalamic nucleus and/or peripheral cytokine-induced neutrophil chemoattractant, since hypophysectomy could not reduce this phase. These data suggest that cytokine-induced neutrophil chemoattractant in the paraventricular hypothalamic nucleus was immediately synthesized in response to stress, and then released into the peripheral blood via the hypothalamo-neurohypophysial system, revealing the presence of a stress-sensitive chemokinergic neuronal pathway in the hypothalamo-pituitary system.     

Nanoscale operation of a living cell using an atomic force microscope with a nanoneedle

We have developed a tool for performing surgical operations on living cells at nanoscale resolution using atomic force microscopy (AFM) and a modified AFM tip. The AFM tips are sharpened to ultrathin needles of 200-300 nm in diameter using focused ion beam etching. Force-distance curves obtained by AFM using the needles indicated that the needles penetrated the cell membrane following indentation to a depth of 1-2 microm. The force increase during the indentation process was found to be consistent with application of the Hertz model. A three-dimensional image generated by laser scanning confocal microscopy directly revealed that the needle penetrated both the cellular and nuclear membranes to reach the nucleus. This technique enables the extended application of AFM to analyses and surgery of living cells.     

Growth of the cervical spine with special reference to its lordosis and mobility

STUDY DESIGN: The cervical spine of the healthy Japanese children aged between 1 year and 18 years was radiographically examined. OBJECTIVES: To examine the correlation between growth of the cervical vertebral body and the facet joint and the development of the cervical lordosis and intervertebral motion. SUMMARY OF BACKGROUND DATA: Although the growth of body height and facet angle have been well documented, their correlation with curvature or mobility has not been elucidated. METHODS: We evaluated plain lateral radiographs of 180 boys and 180 girls regarding diameters and central heights of the cervical vertebra, the anterior and posterior vertebral height ratio, body height index, the facet joint angles, and tilting and sliding motions. Cervical length as the summation of the central height from C3 to C7 and the cervical lordosis angle (C3-C7 angle) were also measured. RESULTS: The mean C3-C7 angle and body height index gradually decreased until 9 years of age and then increased. The C3-C7 angle showed a significant correlation with cervical length, body height index, and facet joint angles before 9 years of age, and with cervical length and body height index after 9 years of age but not with facet joint angles. Facet joint angle decreased until 10 years of age and remained almost unchanged thereafter. Total sliding showed a significant age-related decrease and showed a significant correlation with facet joint angle. CONCLUSION: Although the lordosis angle showed a significant correlation with the other values, cervical length, body height index, and facet joint angle, the determinants of the lordosis could not be elucidate in the present study. As for the mobility of the cervical spine, changes of tilting motion were small, whereas changes of sliding motion were restricted by the change of orientation of the facet joints.     

Sub-quarter-micrometer gate-length p-channel MOSFETs with shallowboron counter-doped layer fabricated using channel preamorphization

A technique for forming shallow boron-doped layers for channel doping using preamorphization (channel preamorphization) is described. An extremely shallow boron-doped layer for shallow channel doping has been formed using preamorphization and rapid thermal annealing. Boron peak concentration around the surface is 3.5×10¹⁸ cm ^-3, and the depth at which the boron concentration becomes 10 ¹⁷ cm^-3 is 450 Å. In contrast, the depth is as large as 900 Å for nonpreamorphized samples. It is found that the shallow boron-doped layer formation is made possible because enhanced diffusion arising from ion implantation damage as well as the channeling in boron ion implantation is suppressed by preamorphization. It is also found that preamorphization does not affect MOS capacitor characteristics so long as the amorphous/crystalline interface is sufficiently deep, which allows that channel preamorphization is readily applicable to channel doping in MOSFET fabrication. To substantiate the experimental results, buried-channel p-MOSFETs with a shallow boron counterdoped layer using channel preamorphization have been successfully fabricated. Channel preamorphization did not degrade carrier mobility and improved MOSFET characteristics in the sub-quarter-micrometer-gate-length region suppressing short-channel effects due to the shallower counterdoped boron profile. High-performance 0.2-μm-gate-length p-MOSFETs with good subthreshold characteristics have been fabricated     

A quantitative assessment of serum chylomicron by light scattering intensity: application to the intestinal fat absorption test

We cloned the canine interleukin-12 (IL-12) subunit cDNA. Canine IL-12 exhibited sequence homology to the known sequences of human, mouse, and bovine genes at nucleotide and amino acid levels. Cotransfection of the p35 and p40 subunits of canine IL-12 cDNA clones into COS-1 cells resulted in the secretion of IL-12, which supported proliferation of the stimulated canine lymphocytes, promoted induction of canine interferon-gamma (IFN-gamma) from canine lymphocytes, and showed antitumor effect in vitro. The cloned canine IL-12 will be useful for canine therapeutic applications.