Peptide Sequence Mapping around Bisecting GlcNAc-Bearing N-Glycans in Mouse Brain

N-glycosylation is essential for many biological processes in mammals. A variety of N-glycan structures exist, of which, the formation of bisecting N-acetylglucosamine (GlcNAc) is catalyzed by N-acetylglucosaminyltransferase-III (GnT-III, encoded by the Mgat3 gene). We previously identified various bisecting GlcNAc-modified proteins involved in Alzheimer’s disease and cancer. However, the mechanisms by which GnT-III acts on the target proteins are unknown. Here, we performed comparative glycoproteomic analyses using brain membranes of wild type (WT) and Mgat3-deficient mice. Target glycoproteins of GnT-III were enriched with E4-phytohemagglutinin (PHA) lectin, which recognizes bisecting GlcNAc, and analyzed by liquid chromatograph-mass spectrometry. We identified 32 N-glycosylation sites (Asn-Xaa-Ser/Thr, Xaa ≠ Pro) that were modified with bisecting GlcNAc. Sequence alignment of identified N-glycosylation sites that displayed bisecting GlcNAc suggested that GnT-III does not recognize a specific primary amino acid sequence. The molecular modeling of GluA1 as one of the good cell surface substrates for GnT-III in the brain, indicated that GnT-III acts on N-glycosylation sites located in a highly flexible and mobile loop of GluA1. These results suggest that the action of GnT-III is partially affected by the tertiary structure of target proteins, which can accommodate bisecting GlcNAc that generates a bulky flipped-back conformation of the modified glycans.     

Long-term stimulation and recording with a penetrating microelectrode array in cat sciatic nerve

We studied the consequences of long-term implantation of a penetrating microelectrode array in peripheral nerve over the time course of 4-6 mo. Electrode arrays without lead wires were implanted to test the ability of different containment systems to protect the array and nerve during contractions of surrounding muscles. Treadmill walking was monitored and the animals showed no functional deficits as a result of implantation. In a different set of experiments, electrodes with lead wires were implanted for up to 7 mo and the animals were tested at 2-4 week intervals at which time stimulation thresholds and recorded sensory activity were monitored for every electrode. It was shown that surgical technique highly affected the long-term stimulation results. Results between measurement sessions were compared, and in the best case, the stimulation properties stabilized in 80% of the electrodes over the course of the experiment (162 days). The recorded sensory signals, however, were not stable over time. A histological analysis performed on all implanted tissues indicated that the morphology and fiber density of the nerve around the electrodes were normal.     

Novel surface modification of cellulose film by heat‐set finishing method using poly(ethylene glycol)‐coated polystyrene nanospheres

The nanosphere having hydrophobic backbone and hydrophilic branches was used as the agent for the surface modification of a cellulose film. They were obtained by dispersion copolymerization of styrene (St) and poly(ethylene glycol) (PEG) macromonomers in an ethanol/water solution at 60°C by using a free‐radical initiator. The PEG‐coated polystyrene (PSt) nanosphere–water dispersions were prepared at concentrations of 0.1, 0.2, 0.5, and 1.0% (w/v). A measure of 1 mL of the dispersion was poured over the cellulose film, cut into a strip of 5 × 5 cm². The film was pressed by plates heated at 200°C with 6.8 g/cm² pressure for 2 min to melt PSt nanospheres and fix them on the cellulose film. The morphology of the film surface was also observed by a scanning electron microscopy (SEM). The resulting modified surface was characterized by X‐ray photoelectron spectroscopy (XPS). The contact angle, the moisture absorption, and the leakage of electrostatic charge from the film were studied. The surface of the film treated with the dispersion had high water‐repellency, although the bulk properties did not change. It was found that the dispersion was effective in making the cellulose surface hydrophobic. The surface modification of cellulose film was successful by using this simple method. © 1999 John Wiley & Sons, Inc. J Appl Polym Sci 74: 1516–1523, 1999     

Surface modification of synthetic fiber nonwoven fabrics with poly(acrylic acid) chains prepared by corona discharge induced grafting

The surface modification of synthetic fiber fabrics via corona discharge treatment and subsequent graft polymerization was investigated. Polyethylene (PE) nonwoven fabric and polyamide-6 (PA-6) nonwoven fabric were used as base fabrics. Acrylic acid (AAc) was graft polymerized onto the fabrics via corona discharge pre-treatment. The grafted amounts of PAAc were dependent on the grafting time, that of PA-6 being higher than that of PE. It was confirmed that the surface of the fibers constructing the fabric was fully covered with PAAc after the 20 min reaction. The surface of the PAAc grafted fabrics was characterized by X-ray photoelectron spectroscopy. The leakage of electrostatic charge from the fabric was determined and the dyeability was studied with methylene blue. The period of time in which the charge potential attenuated to 1/2 of the initial potential decreased drastically by grafting with PAAc. The grafted amount was enough for dyeing the entire fabrics.     

Ezrin Regulates Ca2+ Ionophore-Induced Plasma Membrane Translocation of Aquaporin-5

Aquaporin-5 (AQP5) is selectively expressed in the apical membrane of exocrine glands, such as salivary, sweat, and submucosal airway glands, and plays important roles in maintaining their secretory functions. Because AQP5 is not regulated by gating, localization on the plasma membrane is important for its water-permeable function. Ezrin is an ezrin–radixin–moesin family protein that serves as a crosslinker between the plasma membrane and actin cytoskeleton network. It plays important roles in translocation of various membrane proteins to mediate vesicle trafficking to the plasma membrane. In this study, we examined the effects of ezrin inhibition on membrane trafficking of AQP5. Ezrin inhibition selectively suppressed an ionomycin-induced increase in AQP5 translocation to the plasma membrane of mouse lung epithelial cells (MLE-12) without affecting the steady-state level of plasma membrane AQP5. Taken together, our data suggest that AQP5 translocates to the plasma membrane through at least two pathways and that ezrin is selectively involved in a stimulation-dependent pathway.     

Synthesis and Sintering of Cerium(III) Sulfide Powders

Cerium(III) sulfide (Ce₂S₃) powder was synthesized via the sulfurization of ceria (CeO₂) powder using carbon disulfide gas. Single-phase α-Ce₂S₃ could be formed via sulfurization at 973 K for 28.8 ks. The preparation of α-Ce₂S₃ became feasible at low temperature, in comparison to sulfurization using hydrogen sulfide gas. According to the fact that the formation of α-Ce₂S₃ was accelerated by the addition of carbon black to the CeO₂ powder, carbothermic reduction was considered to become a dominant reaction, as the temperature increased. To obtain the activation energy for the densification of β-Ce₂S₃ powder, which was prepared by vacuum heating α-Ce₂S₃, the data of densification by hot pressing was analyzed by a kinetic equation that was proposed by other researchers. As a result, the sintering behavior could be best explained by a grain-boundary-diffusion mechanism that had an apparent activation energy of 382 kJ/mol.     

IL-17A Is the Critical Cytokine for Liver and Spleen Amyloidosis in Inflammatory Skin Disease

Systemic amyloidosis is recognized as a serious complication of rheumatoid arthritis or inflammatory bowel disease, but also of inflammatory skin disease. However, the detailed molecular mechanism of amyloidosis associated with cutaneous inflammation remains unclear, and therapeutic approaches are limited. Here, we investigated the pathophysiology of amyloidosis secondary to cutaneous inflammation and the therapeutic effects of Janus kinase (JAK) inhibitors by examining a mouse model of spontaneous dermatitis (KCASP1Tg mice). Moreover, KCASP1Tg mice were crossed with interleukin-17A (IL-17A) knockout mice to generate IL-17A-/KCASP1Tg and examine the role of IL-17A in amyloidosis under cutaneous inflammation. KCASP1Tg mice showed severe amyloid deposition in the liver and spleen. Increased serum-neutral fat levels and decreased lymphocyte production were observed in the spleen. Overproduction of amyloidosis was partially ameliorated by the administration of JAK inhibitors and was further improved in IL-17A-/KCASP1Tg mice. IL-17A-producing cells included CD4, gamma delta, and CD8 T cells. In summary, our results from the analysis of a mouse model of dermatitis revealed that skin-derived inflammatory cytokines can induce amyloid deposition in the liver and spleen, and that the administration of JAK inhibitors and, even more, IL-17A ablation, reduced amyloidosis. This study demonstrates that active control of skin inflammation is essential to prevent internal organ amyloidosis.