Polysialic acid, a unique glycan that is developmentally regulated by two polysialyltransferases, PST and STX, in the central nervous system: From biosynthesis to function

Polysialic acid is a developmentally regulated carbohydrate composed of a linear homopolymer of a-2,a-linked sialic acid residues. This unique glycan is mainly attached to the neural cell adhesion molecule (N-CAM) and implicated in many morphogenic events of the neural cells by modulating the adhesive property of N-CAM. Recently, the cDNA that encodes polysialyltransferase, which is responsible for the polysialylation of N-CAM, was successfully cloned from three mammalian species. This review focuses on the molecular cloning of human polysialyltransferase, designated PST. it then describes the number of enzymes actually required for the polysialylation of N-CAM using an in vitro polysialyltransferase assay. Comparisons between PST and another polysialyltransferase, sialyltransferase X (STX), are made and it Is demonstrated that both enzymes can independently form polysiatic acid In vitro , but that during neural development they coordinately but distinctly synthesize polysialic acid on N-CAM. The role of polysialic acid in the central nervous system is also discussed. Finally, evidence that the two polysialyltransferases, PST and STX, apparently have distinct roles in the development of neural cells is provided by using a neurite outgrowth assay.     

Immunoelectron microscopic study of HTLV-II-producing cells with an anti-envelope gp46 monoclonal antibody

Summary We observed expression of envelope gp46 protein on an HTLV-II-producing T-cell line (Mo) cells by an immunoelectron microscopical method using a monoclonal antibody against HTLV-II gp46. gp46 reactivity was observed on virus-like particles, extracellular vesicles, cell membrane, and partially in nuclear membrane and endoplasmic reticulum.     

Seasonal variation in sweating responses of older and younger men

Eight older (60–65 years) and six younger (20–25 years) men were exposed to a standard heat stress for 60 min in summer, autumn, winter, and spring. The test consisted of placing the lower legs and feet in a 42°C water bath while sitting in constant environmental conditions (30°C and 45% relative humidity). The increase of rectal temperature (T _re) was significantly greater (P < 0.05) in autumn, winter, and spring than in summer for the older group, but significantly greater only in winter than in summer for the younger group (P < 0.05). The T _re was greater for the older group in all seasons, but of significance only in autumn and spring (P < 0.01). There were no significant season-related differences for metabolic heat production (m) and mean skin temperature ( _sk) during the heat test in the respective groups, although the m and _sk were lower for the older group in all seasons (P < 0.01). In the older group total body sweating rate (m_sw) divided by T _re (total m_sw/T _re) decreased from summer to winter (P < 0.02) and did not differ between winter and spring, whereas total m_sw/T _re in the younger group increased in spring after decreasing from autumn to winter (P < 0.03). The variations of the value, local sweating rate on the back and thigh divided by T _re (back m_sw/T _re and thigh m_sw/T _re), were similar to those of the total m_sw/T _re in each group, except for back m_sw/T _re in the younger group, which did not increase from winter to spring. The total m_sw/T _re, back m_sw/T _re and thigh m_sw/T _re were significantly less for the older group in summer, autumn and spring (P < 0.05). The range of seasonal variations was significantly less for the older group (P < 0.001). The results indicated that, compared with younger men in older men, the enhancement of sweating function toward summer occurred later and its reduction toward winter occurred earlier despite a smaller range of seasonal variation and that older men had a somewhat lesser capability to maintainT _re when challenged by heat stress in all seasons.     

Changes in fibre-type composition and myosin heavy-chain IId isoform in rat soleus muscle during recovery period after hindlimb suspension

We examined the changes in myosin heavy-chain (HC) isoforms and fibre-type composition in rat soleus muscle using both myosin adenosine triphosphatase staining and sodium dodecyl sulphate/polyacrylamide gel electrophoresis (SDS-PAGE) analyses during the recovery period after 4 weeks of hindlimb suspension. Although there was no change in type IIc fibres after the suspension, an increase in this type of fibres was observed during the 1- to 4-week recovery period. The increase in type Ilc fibres was considered to be due to a shift from type Ila to IIc fibres. The SDS-PAGE analysis revealed the presence of the HC IId isoform, which was not observed in the control muscle, after a 4-week hindlimb suspension. The HC IId isoform gradually decreased over 3 weeks of recovery and disappeared in the 4th week of recovery after the suspension. These results suggest that the hypogravity conditions induced by hindlimb suspension stimulated the synthesis of the HC IId isoform, whereas an increase in mechanical load to the muscle accelerated the degradation of the HC IId isoform and the synthesis of type Ilc fibres during the recovery period after hindlimb suspension.     

Therapeutic effect of topical administration of SN50, an inhibitor of nuclear factor-kappaB, in treatment of corneal alkali burns in mice

We evaluated the therapeutic efficacy of topical administration of SN50, an inhibitor of nuclear factor-kappaB, in a corneal alkali burn model in mice. An alkali burn was produced with 1 N NaOH in the cornea of C57BL/6 mice under general anesthesia. SN50 (10 microg/microl) or vehicle was topically administered daily for up to 12 days. The eyes were processed for histological or immunohistochemical examination after bromodeoxyuridine labeling or for semi-quantification of cytokine mRNA. Topical SN50 suppressed nuclear factor-kappaB activation in local cells and reduced the incidence of epithelial defects/ulceration in healing corneas. Myofibroblast generation, macrophage invasion, activity of matrix metalloproteinases, basement membrane destruction, and expression of cytokines were all decreased in treated corneas compared with controls. To elucidate the role of tumor necrosis factor (TNF)-alpha in epithelial cell proliferation, we performed organ culture of mouse eyes with TNF-alpha, SN50, or an inhibitor of c-Jun N-terminal kinase (JNK) and examined cell proliferation in healing corneal epithelium in TNF-alpha-/- mice treated with SN50. An acceleration of epithelial cell proliferation by SN50 treatment was found to depend on TNF-alpha/JNK signaling. In conclusion, topical application of SN50 is effective in treating corneal alkali burns in mice.     

Biological activities of Bacteroides forsythus lipoproteins and their possible pathological roles in periodontal disease

Bacteroides forsythus is a gram-negative, anaerobic, fusiform bacterium and is considered to be an etiological agent in periodontal disease. A lipoprotein fraction prepared from B. forsythus cells by Triton X-114 phase separation (BfLP) activated human gingival fibroblasts and a human monocytic cell line, THP-1, to induce interleukin-6 production and tumor necrosis factor alpha production. BfLP was found to be capable of inducing nuclear factor-kappaB translocation in human gingival fibroblasts and THP-1 cells. By using Chinese hamster ovary K1 cells transfected with Toll-like receptor genes together with a nuclear factor-kappaB-dependent CD25 reporter plasmid, it was found that signaling by BfLP was mediated by Toll-like receptor 2 but not by CD14 or Toll-like receptor 4. BfLP induced apoptotic cell death in human gingival fibroblasts, KB cells (an oral epithelial cell line), HL-60 cells (a human myeloid leukemia cell line), and THP-1 cells but not in MOLT4 cells (a T-cell leukemia cell line). Caspase-8, an initiator caspase in apoptosis, was found to be activated in these cells in response to BfLP stimulation. Thus, this study suggested that BfLP plays some etiological roles in oral infections, especially periodontal disease, by induction of cell activation or apoptosis.     

Co-expression in CHO cells of two muscle proteins involved in excitation-contraction coupling

Summary Ryanodine receptors and dihydropyridine receptors are located opposite each other at the junctions between sarcoplasmic reticulum and either the surface membrane or the transverse tubules in skeletal muscle. Ryanodine receptors are the calcium release channels of the sarcoplasmic reticulum and their cytoplasmic domains form the feet, connecting sarcoplasmic reticulum to transverse tubules. Dihydropyridine receptors are L-type calcium channels that act as the voltage sensors of excitation-contraction coupling: they sense surface membrane and tranverse tubule depolarization and induce opening of the sarcoplasmic reticulum release channels. In skeletal muscle, ryanodine receptors are arranged in extensive arrays and dihydropyridine receptors are grouped into tetrads, which in turn are associated with the four subunits of ryanodine receptors. The disposition allows for a direct interaction between the two sets of molecules.CHO cells were stably transformed with plasmids for skeletal muscle ryanodine receptors and either the skeletal dihydropyridine receptor, or a skeletal-cardiac dihydropyridine receptor chimera (CSk3) which can functionally substitute for the skeletal dihydropyridine receptor, in addition to plasmids for the ₂, and subunits. RNA blot hybridization gave positive results for all components. Immunoblots, ryanodine binding, electron microscopy and exposure to caffeine show that the expressed ryanodine receptors forms functional tetrameric channels, which are correctly inserted into the endoplasmic reticulum membrane, and form extensive arrays with the same spacings as in skeletal muscle. Since formation of arrays does not require coexpression of dihydropyridine receptors, we conclude that self-aggregation is an independent property of ryanodine receptors. All dihydropyridine receptor-expressing clones show high affinity binding for dihydropyridine and immunolabelling with antibodies against dihydropyridine receptor. The presence of calcium currents with fast kinetics and immunolabelling for dihydropyridine receptors in the surface membrane of CSk3 clones indicate that CSk3-dihydropyridine receptors are appropriately targeted to the cell's plasmalemma. The expressed skeletal-type dihydropyridine receptors, however, remain mostly located within perinuclear membranes. In cells coexpressing functional dihydropyridine receptors and ryanodine receptors, no junctions between feet-bearing endoplasmic reticulum elements and surface membrane are formed, and dihydropyridine receptors do not assemble into tetrads. A separation between dihydropyridine receptors and ryanodine receptors is not unique to CHO cells, but is found also in cardiac muscle, in muscles of invertebrates and, under certain conditions, in skeletal muscle. We suggest that failure to form junctions in co-transfected CHO cell may be due to lack of an essential protein necessary either for the initial docking of the endoplasmic reticulum to the surface membrane or for maintaining the interaction between dihydropyridine receptors and ryanodine receptors. We also conclude that formation of tetrads requires a close interaction between dihydropyridine receptors and ryanodine receptors.