Cystatin SN Upregulation in Patients with Seasonal Allergic Rhinitis

Seasonal allergic rhinitis (SAR) to the Japanese cedar, Cryptomeria japonica (JC) pollen is an IgE-mediated type I allergy affecting nasal mucosa. However, the molecular events underlying its development remain unclear. We sought to identify SAR-associated altered gene expression in nasal epithelial cells during natural exposure to JC pollen. We recruited study participants in 2009 and 2010 and collected nasal epithelial cells between February and April, which is the period of natural pollen dispersion. Fifteen patients with SAR-JC and 13 control subjects were enrolled in 2009, and 17 SAR-JC patients, 13 sensitized asymptomatic subjects (Sensitized), and 15 control subjects were enrolled in 2010. Total RNA was extracted from nasal epithelial cells and 8 SAR-JC patients and 6 control subjects in 2009 were subjected to microarray analysis with the Illumina HumanRef-8 Expression BeadChip platform. Allergen-stimulated histamine release was examined in the peripheral blood basophils isolated from patients with SAR. We identified 32 genes with significantly altered expression during allergen exposure. One of these, CST1 encodes the cysteine protease inhibitor, cystatin SN. CST1 expression in nasal epithelial cells was significantly upregulated in both the 2009 and 2010 SAR-JC groups compared with the control groups. Immunohistochemical staining confirmed the increased expression of CST1 in the nasal epithelial cells of SAR patients. Addition of exogenous CST1 to basophils inhibited JC allergen-stimulated histamine release in vitro. We propose that CST1 may contribute to inactivation of protease allergens and help re-establish homeostasis of the nasal membranes.     

Relationship between Numerous Mast Cells and Early Follicular Development in Neonatal MRL/MpJ Mouse Ovaries

In the neonatal mouse ovary, clusters of oocytes called nests break into smaller cysts and subsequently form individual follicles. During this period, we found numerous mast cells in the ovary of MRL/MpJ mice and investigated their appearance and morphology with follicular development. The ovarian mast cells, which were already present at postnatal day 0, tended to localize adjacent to the surface epithelium. Among 11 different mouse strains, MRL/MpJ mice possessed the greatest number of ovarian mast cells. Ovarian mast cells were also found in DBA/1, BALB/c, NZW, and DBA/2 mice but rarely in C57BL/6, NZB, AKR, C3H/He, CBA, and ICR mice. The ovarian mast cells expressed connective tissue mast cell markers, although mast cells around the surface epithelium also expressed a mucosal mast cell marker in MRL/MpJ mice. Some ovarian mast cells migrated into the oocyte nests and directly contacted the compressed and degenerated oocytes. In MRL/MpJ mice, the number of oocytes in the nest was significantly lower than in the other strains, and the number of oocytes showed a positive correlation with the number of ovarian mast cells. The gene expression of a mast cell marker also correlated with the expression of an oocyte nest marker, suggesting a link between the appearance of ovarian ? 4mast cells and early follicular development. Furthermore, the expression of follicle developmental markers was significantly higher in MRL/MpJ mice than in C57BL/6 mice. These results indicate that the appearance of ovarian mast cells is a unique phenotype of neonatal MRL/MpJ mice, and that ovarian mast cells participate in early follicular development, especially nest breakdown.     

Antifungal Activity of 2,4-Dihydroxyacylophenones and Related Compounds

The antifungal activity of 2,4-dihydroxyacylophenones and related compounds against Trichophyton spp and other fungi were investigated to determine their structure-activity relationships.

The activity of these compounds was found to be closely related to the length of the acyl and alkyl substituents attached to the 1,3-dihydroxybenzene moiety In addition, differences in activity were observed depending on the position of the alkyl substituents and on the number of substituents attached to the 1,3-dihydroxybenzene moiety. Some compounds tested showed potent antifungal activity against Trichophyton spp. and other fungi that was more active than amphotericin B.     

Serotobenine,a Novel Phenolic Amide from Safflower Seeds (Carthamus tinctorius L.)

The structure of a new indole derivative, serotobenine (1), from sfflower meal (Carthamus tinctorius> L.) is proposed based on ¹H- and ¹³C-NMR spectral data and X-ray crystallography. The known compounds N-feruloyltryptamine (2) and N-(p-coumaroyl)tryptamine (3) were also isolated and identified.     

Binding-site Analysis of the Ether Linkages between Lignin and Hemicelluloses in Lignin–Carbohydrate Complexes by DDQ-Oxidation

Lignin-carbohydrate complexes (LCC; nor-C-1-M, com-C-1-A) isolated from normal and compression woods of Pinus densiflora were hydrolyzed with two types of cellulase preparations, and the hydrolyzates formed were fractionated by adsorption chromatography on polyvinyl gel into water-soluble materials and LCC fragments. To elucidate the binding sites between the lignin and carbohydrate, the cellulase-degraded LCC fragments were subjected to acetylation, and then oxidation with 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ), which was confirmed to oxidatively cleave the benzyl ether linkages between the lignin and carbohydrate. The DDQ-oxidized fraction was then methylated by the method of Prehm, hydrolyzed, reduced and acetylated. A GC-MS analysis of the methylated sugar revealed that alditol acetates from 6-O-methyl mannose, 6-O-methyl galactose, 6-O-methyl glucose and a small amount of their 2-O- or 3-O-methyl isomers existed in both methylated fractions. 2-O-Methyl xylose and 3-O-methyl xylose were also identified in the fraction from the acidic LCC (com-C-1-A). These results led to the conclusion that acetylglucomannan and β-1,4-galactan were preferably bound to the lignin at C-6 position of the hexoses, and that arabinoglucuronoxylan did likewise at the C-2 and C-3 positions of xylose units.     

Isolation and Properties of an α-Amylase Inhibitor (0.53) from Wheat (Triticum aestivum)

A droplet gel-entrapping method used for enzyme immobilization was improved to simplify the procedure and to increase the enzyme stability. This immobilization technique is suitable for coupled enzyme reactions requiring cofactors. Leucine dehydrogenase (LeuDH) and formate dehydrogenase (FDH) were freeze-dried with bovine serum albumin, dextrin and stabilizers. The freeze-dried enzyme powder was suspended in a methylcellosolve solution containing polyethyleneglycol(#4000)diacrylate, N,N′-methylenebisacrylamide and 2-hydroxyethylacrylate, and the suspension was gelled with initiators. The gel was cut up and the pieces were washed in a buffer to remove the methylcellosolve and the dextrin inside. The maximum conversion ratio for a LeuDH-FDH gel column was determined to be 99.8% by means of the recycling reaction. On longterm operation at 30 °C for leucine production, the initial conversion ratio (7.2%) gradually decreased to 6.6% over the first 10 days. However, the conversion ratio remained almost constant after the 10th day. The effects of flow rate, temperature, pH, and the concentrations of formate, α- ketoisocaproate, ammonium and NAD on the leucine productivity with the gel column were also investigated.     

Expression of recombinant alpha and beta tubulins from the yew Taxus cuspidata and analysis of the microtubule assembly in the presence of taxol

Taxol was originally isolated from the yew Taxus brevifolia. Because taxol inhibits the depolymerization of microtubules, the presence of a self-resistance mechanism in Taxus spp. was hypothesized. The cloning of the cDNA for alpha and beta tubulins from Taxus cuspidata and those from the human embryonic kidney cell line HEK293T revealed that the ²⁶Asp, ³⁵⁹Arg, and ³⁶¹Leu residues in the human beta tubulin, which are important for taxol binding, were replaced with Glu, Trp, and Met in the beta tubulin of T. cuspidata, respectively. The microtubule assembly of the recombinant alpha and beta tubulins was monitored turbidimetrically, and the results clearly demonstrated that the microtubule from T. cuspidata is less sensitive to taxol than that from HEK293T cells. The Taxus microtubule composed of the wild-type alpha tubulin and the beta tubulin with the E26D mutation restored the sensitivity to taxol. We thus postulated that the mutation identified in the beta tubulin of T. cuspidata plays a role in the self-resistance of this species against taxol.     

Rate-adjusted cross-linking reaction by photoresponsive α-bromoaldehyde (PBA)-conjugated ODN

We developed a photoresponsive α-bromoaldehyde-conjugated oligonucleotide (PBA-ODN). The PBA-ODN selectively reacted and formed covalent bonds with target oligonucleotides having adenine or cytosine at the frontal position of the aldehyde derivative. Kinetic studies revealed that PBA-ODN has increased kinetic rates for the formation of cross-linked duplexes compared with the corresponding α-chloroaldehyde-conjugated oligonucleotide (PCA-ODN).     

Soluble recombinant human thrombomodulin suppresses inflammation-induced gastrointestinal tumor growth in a murine peritonitis model

Molecular and Cellular Biochemistry - In the original article, Figs. 3b, 4a, c and 5d were published incorrectly. The correct version of the figures are provided in this correction.     

HM1.24/BST-2 is constitutively poly-ubiquitinated at the N-terminal amino acid in the cytoplasmic domain

HM1.24 (also known as BST-2, CD317, and Tetherin) is a type II single-pass transmembrane glycoprotein, which traverses membranes using an N-terminal transmembrane helix and is anchored in membrane lipid rafts via a C-terminal glycosylphosphatidylinositol (GPI). HM1.24 plays a role in diverse cellular functions, including cell signaling, immune modulation, and malignancy. In addition, it also functions as an interferon-induced cellular antiviral restriction factor that inhibits the replication and release of diverse enveloped viruses, and which is counteracted by Vpu, an HIV-1 accessory protein. Vpu induces down-regulation and ubiquitin conjugation to the cytoplasmic domain of HM1.24. However, evidence for ubiquitination site(s) of HM1.24 remains controversial. We demonstrated that HM1.24 is constitutively poly-ubiquitinated at the N-terminal cytoplasmic domain, and that the mutation of all potential ubiquitination sites, including serine, threonine, cysteine, and lysine in the cytoplasmic domain of HM1.24, does not affect the ubiquitination of HM1.24. We further demonstrated that although a GPI anchor is necessary and sufficient for HM1.24 antiviral activities and virion-trapping, the deleted mutant of GPI does not influence the ubiquitination of HM1.24. These results suggest that the lipid raft localization of HM1.24 is not a prerequisite for the ubiquitination. Collectively, our findings demonstrate that the ubiquitination of HM1.24 occurs at the N-terminal amino acid in the cytoplasmic domain and indicate that the constitutive ubiquitination machinery of HM1.24 may differ from the Vpu-induced machinery.