Yuzu (Citrus junos Tanaka) Peel Attenuates Dextran Sulfate Sodium-induced Murine Experimental Colitis.

Ulcerative colitis is a well-known inflammatory bowel disease. Although there are drugs that are effective against this disease, the prevention and attenuation of ulcerative colitis by food rich in functional ingredients without side effects is desired because some drugs have side effects. In this study, we investigated the effects of yuzu (Citrus junos Tanaka), a citrus fruit native to northeast Asia, on a mouse dextran sulfate sodium (DSS)-induced colitis model. Mice given drinking water containing DSS showed significant weight loss, colon shortening, diarrhea, and visible fecal blood. In contrast, mice fed a diet containing 5% yuzu peel for 14 d before receiving DSS showed significant attenuation of these phenotypes. To clarify the mechanism underlying the attenuation, we investigated the anti-inflammatory and antioxidant effects of yuzu peel. We found that yuzu peel extract suppressed tumor necrosis factor-α (TNF-α) production in lipopolysaccharide (LPS)-stimulated mice and murine macrophage cell line through suppression of nuclear factor-κB (NF-κB) activation. In addition, we confirmed that yuzu peel extract had a moderate antioxidant effect. These results suggest that yuzu peel attenuates the pathologies of DSS-induced colitis by coordinately suppressing inflammation and oxidative stress against lipids in vivo.

Evaluations of lipid peroxidation and inflammation in short-term glycerol-induced acute kidney injury in rats.

Rhabdomyolysis is characterised by acute kidney injury (AKI) resulting from skeletal muscle injury. Lipid peroxidation-mediated oxidant injury and pro-inflammatory cytokine-mediated inflammatory response play critical roles in the pathogenesis of rhabdomyolysis-induced AKI. The present study aimed to investigate the short-term effects of both lipid peroxidation and inflammatory responses on rhabdomyolysis-induced AKI in a rat model of glycerol-induced rhabdomyolysis. Rhabdomyolysis was induced by the intramuscular injection of 50% glycerol in saline (10 mL/kg) into the hind limbs of rats. Rats were killed 1 or 3 hours after glycerol injection. Time-dependent increases in serum biochemical parameters, including blood urea nitrogen, creatinine, lactate dehydrogenase and creatine phosphokinase levels, were observed 1 hour after glycerol injection. In kidneys, glycerol injection resulted in histopathological changes such as renal tubular injury and renal tubular myoglobin deposition. Levels of Nε-(hexanoyl)lysine-modified, 4-hydroxy-2-nonenal-modified, and nitrotyrosine-modified proteins in rat kidneys were unaltered at 1 hour after glycerol injection, but increased significantly at 3 hours. Increases in renal nitric oxide production and the expression levels of inducible nitric oxide synthase, interleukin-6 and tumour necrosis factor-α in the renal parenchyma were observed at 1 hour after glycerol injection and plateaued at 3 hours. Our findings suggest that the pro-inflammatory cytokine-mediated inflammatory response may cause rhabdomyolysis-induced AKI very shortly after glycerol injection, and lipid peroxidation-mediated oxidant injury may promote the development of these pathophysiological processes.

Development of a Valuable Yeast Strain Using a Novel Mutagenesis Technique for the Effective Production of Therapeutic Glycoproteins.

The so-called disparity mutagenesis technique selectively elevates mutation in the lagging strand of DNA by using a mutant form of DNA polymerase δ, encoded on a proofreading-deficient pol3 gene. This novel mutagenesis technique generates a pool of mutants that includes a no-mutant strain together with mutant strains carrying multiple mutations. By using a suitable screening system it is possible to isolate the desired mutant strain from this pool of mutants. Here, we used our novel mutagenesis technique to isolate a yeast strain with good growth characteristics that was glycosylation deficient.

Beneficial effects of vildagliptin combined with miglitol on glucose tolerance and islet morphology in diet-controlled db/db mice.

Dipeptidyl peptidase-4 (DPP-4) inhibitors improve glycemic control in patients with type 2 diabetes primarily by increasing plasma active glucagon-like peptide-1 (GLP-1) levels. While various combination therapies based on DPP-4 inhibitors have been proposed for treatment of type 2 diabetes, the effects of combination therapy of DPP-4 inhibitors and alpha-glucosidase inhibitors on ß-cell function are less characterized. We evaluated the effects of long-term treatment with vildagliptin, a DPP-4 inhibitor, on metabolic parameters and ß-cell function, in combination with miglitol, an alpha-glucosidase inhibitor, in diet-controlled db/db mice. In this study, 6-week-old male db/db mice were provided with standard chow twice a day for 6 weeks. Meal tolerance tests and glucose tolerance tests showed that the combination therapy of vildagliptin with miglitol, but not each alone, suppressed postprandial glycemic excursion, enhanced postprandial active GLP-1 levels and prevented deterioration of glucose tolerance in the db/db mice. The combination treatment did not alter ß-cell mass, but resulted in preserved expression of glucose transporter 2, Zinc transporter 8 and MafA and reduced the number of α cells. These results suggest that the combination of vildagliptin and miglitol prevents the development of overt diabetes in diet-controlled pre-diabetic db/db mice by normalizing postprandial glucose and incretin response, and by preserving ß-cell structure and the expression of factors essential for ß-cell function.

Characterization of an oligomeric carbonyl reductase of dog liver: its identity with peroxisomal tetrameric carbonyl reductase.

Dog liver contains an oligomeric NADPH-dependent carbonyl reductase (CR) with substrate specificity for alkyl phenyl ketones, but its endogenous substrate and primary structure remain unknown. In this study, we examined the molecular weight and substrate specificity of the enzyme purified from dog liver. The enzyme is a ca. 100-kDa tetramer composing of 27-kDa subunit, and reduces all-trans-retinal and alpha-dicarbonyl compounds including isatin, which are substrates for pig peroxisomal tetrameric carbonyl reductase (PTCR). In addition, the dog enzyme resembles pig PTCR in inhibitor sensitivity to flavonoids, myristic acid, lithocholic acid, bromosulfophthalein and flufenamic acid. Furthermore, the amino acid sequence of dog CR determined by protein sequencing and cDNA cloning was 84% identical to that of pig PTCR and had a C-terminal peroxisomal targeting signal type 1, Ser-His-Leu. The immunoprecipitation using the anti-pig PTCR antibody shows that the dog enzyme is a major form of soluble NADPH-dependent all-trans-retinal reductase in dog liver. Thus, dog oligomeric CR is PTCR, and may play a role in retinoid metabolism as a retinal reductase.

Melanogenesis inhibition by an oolong tea extract in b16 mouse melanoma cells and UV-induced skin pigmentation in brownish guinea pigs.

To investigate the new physiological functions of oolong tea, the effects on melanogenesis were studied. An oolong tea extract inhibited melanogenesis without affecting cell growth in B16 mouse melanoma cells. However, the oolong tea extract hardly showed any inhibitory effect on mushroom tyrosinase in a cell-free system. The effects of an oolong tea extract on the intracellular tyrosinase level in B16 cells were therefore studied. All the levels of activity, protein and mRNA were decreased in the oolong tea extract-treated cells. We also investigated the inhibitory effects of oolong tea on the pigmentation induced by ultraviolet B (UVB) by using brownish guinea pigs in vivo. The number of 3,4-dihydroxyphenylalanine (DOPA)-positive melanocytes increased by UVB was repressed by an oral administration of oolong tea. These results imply that oolong tea might be effective in whitening and that its inhibitory effect on melanogenesis was involved in the decrease of intracellular tyrosinase at the mRNA level.

ADAM family protein Mde10 is essential for development of spore envelopes in the fission yeast Schizosaccharomyces pombe.

We report the identification of Schizosaccharomyces pombe mde10+ as a gene possessing a FLEX element, which forms a binding site for the meiosis-specific transcription factor Mei4. In fact, mde10+ is transcribed only in diploid cells that are induced to meiosis in a Mei4-dependent manner. Western blot analysis indicated that the epitope-tagged Mde10 protein accumulates transiently during meiosis and then rapidly decreases. Mde10 is a multidomain protein containing a metalloprotease catalytic domain, a disintegrin domain, a cysteine-rich domain, and membrane-spanning regions, all of which are shared by members of the mammalian ADAM family. A fusion protein of Mde10 and green fluorescent protein localized to the endoplasmic reticulum during meiosis and was located at the peripheral region of spores at the end of meiosis. An mde10Delta deletion mutant showed no apparent defects in meiosis, sporulation, or spore germination. However, the mutant spores exhibited an aberrant surface appearance, in which the ragged outer spore wall was lost to a large extent. Furthermore, mde10Delta spores were found to be less tolerant to ethanol and diethyl ether than were wild-type spores. The mutagenic replacement of the conserved glutamic acid in the putative protease active site with an alanine residue did not affect the surface morphology or the resistance of spores to environmental stress. Our observations indicate that Mde10 is important in the development of the spore envelope, although this function of Mde10 seems to be independent of its metalloprotease activity.

Cloning, expression and tissue distribution of a tetrameric form of pig carbonyl reductase.

In this study, we isolated a cDNA for tetrameric carbonyl reductase (CR) from pig heart. The pig CR showed high amino acid sequence identity (81%) with rabbit NADP(+)-dependent retinol dehydrogenase (NDRD). The purified recombinant pig CR and NDRD were about 100-kDa homotetramers and exhibited high reductase activity towards alkyl phenyl ketones, alpha-dicarbonyl compounds and all-trans-retinal. The identity of NDRD with the tetrameric CR was verified by protein sequencing of CR purified from rabbit heart. Both tetrameric CR and its mRNA were ubiquitously expressed in pig and rabbit tissues. The pig and rabbit enzymes belonged to the short-chain dehydrogenase/reductase family, and their sequences comprise a C-terminal SRL tripeptide, which is a variant of the type 1 peroxisomal targeting signal, SKL. Transfection of HeLa cells with vectors expressing pig CR demonstrated that the enzyme is localized in the peroxisomes. Thus, the tetrameric form of CR represents the first mammalian peroxisomal enzyme that reduces all-trans-retinal as the endogenous substrate.