Prevention of gut inflammation by Bifidobacterium in dextran sulfate-treated gnotobiotic mice associated with Bacteroides strains isolated from ulcerative colitis patients

Indigenous Bacteroides strains are closely associated with the occurrence and exacerbation of ulcerative colitis (UC). In this study, we aimed to clarify the effect of Bifidobacterium strains, another major member of colonic bacteria, on the development of gut inflammation using gnotobiotic mouse models associated with Bacteroides strains isolated from UC patients. Dextran sulfate (DSS) administration induced inflammation in the large intestine, in particular of the cecum, in the gnotobiotic mice associated with three strains of Bacteroides vulgatus, judging from the myeloperoxidase activity, occult blood score, and IgG leakage into the intestinal contents. However, the severity of the inflammation was greatly reduced in the gnotobiotic mice associated with both B. vulgatus and Bifidobacterium strains. The severity of the cecal inflammation was well correlated with the concentration of succinic acid in the cecum. Bacteriologically, the density of B. vulgatus strain A (BV-A) greatly decreased and the predominant strain changed from BV-A to BV-B on additional association with Bifidobacterium strains. Among gnotobiotic mice associated with a single B. vulgatus strain, the severity of cecal inflammation in BV-A-associated mice was greater than that in BV-B-associated mice. Each Bifidobacterium strain produced compound(s) more effectively inhibiting the growth of BV-A than BV-B in in vitro culture. Taken together, these results suggest that the severity of DSS-induced gut inflammation is closely associated with a particular B. vulgatus strain, and that Bifidobacterium strains may repress exacerbation of intestinal inflammation through growth inhibition of the B. vulgatus strain.     

Enhancement of albumin expression in bone tissues with healing rat fractures

The characterization of 66 kDa protein molecule, a major protein component which is produced from femoral-diaphyseal tissues with fracture healing (Igarashi and Yamaguchi [2002] Int. J. Mol. Med. 9:503-508), was investigated. Weaning rats were killed at 7 and 14 days after femoral fracture. When the femoral-diaphyseal tissues with fracture healing were cultured for 48 h in a serum-free medium, many proteins in the bone tissues were released into the medium. Analysis with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) showed that a protein molecule of approximately 66 kDa was markedly increased in culture medium from bone tissues with fracture healing. N-terminal sequencing of 66 kDa protein indicated that its N-terminus was identical to that of rat albumin. Western blot analysis of medium 66 kDa protein showed expression of albumin. This expression was significantly enhanced by fracture healing. The expression of albumin was seen in the diaphyseal (cortical bone) and metaphyseal (trabecular bone) tissues of rat femur. When the femoral-diaphyseal tissues obtained at 7 days after femoral fracture were cultured in a serum-free medium containing either vehicle, parathyroid hormone (1-34) (10(-7) M), insulin-like growth factor-I (10(-8) M) or zinc acexamate (10(-4) M), medium albumin was significantly increased in the presence of those bone-stimulating factors. The addition of albumin (0.5 or 1.0 mg/ml of medium) caused a significant increase in calcium and deoxyribonucleic acid contents in the femoral-diaphyseal and -metaphyseal tissues obtained from normal rats in vitro. The present study demonstrates that fracture healing induces a remarkable production of albumin which is a major protein component produced from femoral-diaphyseal tissues of rats, and that albumin has an anabolic effect on bone components.     

Extraction of total RNA from leaves of Eucalyptus and other woody and herbaceous plants using sodium isoascorbate

Rapid extraction of total RNA from Eucalyptus leaves is difficult due to the high content of polyphenolics and polysaccharides. A rapid and simple method was developed by using an extraction buffer containing sodium isoascorbate at a concentration of 500 mM. This method consisted of one or two chloroform extractions, one acid guanidium-phenol-chloroform extraction, and isopropanol precipitation alone. The yields of the RNA fractions were 246-1750 micrograms/g fresh weight when leaves of Eucalyptus, five other woody plants, and four herbaceous plants were used as samples. The contamination of the RNA fractions by proteins and polysaccharides was very limited as judged spectrophotometrically. When the RNA fractions were subjected to agarose gel electrophoresis, intact rRNA bands were detected. The RNA fractions could be used for RT-PCR. These results indicate that our new method achieves a simple and rapid preparation of high-quality RNA from leaves of Eucalyptus and other plant species.     

Thelephantins D-H: five p-terphenyl derivatives from the inedible mushroom Thelephora aurantiotincta

Five p-terphenyl derivatives named thelephantins D-H (1-5) together with nine known compounds, thelephantins A-C (6-8), ganbajunin E (9), p-hydroxylbenzoic acid (10), ganbajunin C (11), thelephorin A (12), 2-O-methylatromentin (13) and atromentin (14), were isolated from the methanolic extract of fruit bodies of the Thelephoraceous Basidiomycete Thelephora aurantiotincta. Their structures were elucidated by high-resolution MS, 2D NMR, IR and UV spectroscopic analysis.     

Sesquiterpene constituents from the liverwort Bazzania japonica

The hydrodistillation products of the liverwort Bazzania japonica were separated by preparative gas chromatography (GC) and investigated by spectroscopic methods. Seven unknown compounds were isolated and identified by GC-MS and NMR. Four of them, the norsesquiterpene hydrocarbons 4-epi-11-nor-aristola-1(10),11-diene (1), 4-epi-11-nor-aristola-1,9,11-triene (2), 4-epi-11-nor-aristola-9,11-diene (3), and one oxygenated sesquiterpene, (-)-aristol-1(10)-en-12-ol (5) are new natural compounds, and one, (+)-himachala-2,4-diene (7), has for the first time been isolated from liverworts. The absolute configurations of 5 and 7 were derived by chemical correlation reactions and/or enantioselective GC using cyclodextrin phases. 1, 2 and 3 have identical absolute configuration.     

A new ecdysteroid, 2-deoxy-5beta,20-dihydroxyecdysone from the fruits of Diploclisia glaucescens

Chemical investigation of ethyl acetate extract of the fruits of Diploclisia glaucescens of the family Menispermaceae furnished a new ecdysteroid 2-deoxy-5beta,20-dihydroxyecdysone, together with 20-hydroxyecdysone, 3-deoxy-1beta,20-dihydroxyecdysone, 2-deoxy-20-hydroxyecdysone, 24-ethyl-20-hydroxyecdysone (makisterone C). Latter two ecdysteroids are reported first time from the family Menispermaceae.     

Structural and immunological characterization of recombinant ovomucoid expressed in Escherichia coli

The expression of recombinant allergens is becoming new insights of an important diagnosis and the therapy of allergies as well as molecular approaches to immunological and structural studies of allergens. Ovomucoid is a major food allergens in the hen's egg white which causes immediate food-hypersensitivity reactions mainly in children. A gene coding for the cDNA representing an entire ovomucoid molecule has been cloned in Escherichia coli under the control of T5 promoter fused with six-Histidine tag at the amino terminal end. Upon induction, the E. coli cells, harbouring this construct, expressed the recombinant protein as a soluble fraction and the recombinant ovomucoid protein was purified to electrophoeretic homogeneity using Ni²⁺ nitrilotriacetic acid agarose affinity chromatography. Immunoblot analysis showed that human IgE and IgG binding activities of the recombinant ovomucoid was identical to that of native analogue. The antigenicity and allergenicity of recombinant ovomucoid were almost same as that of native form when tested with an ELISA using six individual patient's serum. CD spectra indicated that that the recombinant ovomucoid has more -helix and less -structure than native form. These results show that the recombinant ovomucoid constructed in this study could be used for further studies on the immunological and structural studies of ovomucoid.     

Age-related changes of element contents in human tendon of the iliopsoas muscle and the relationships among elements

To elucidate compositional changes of the tendons with aging, the authors investigated age-related changes of element contents in the tendon of the iliopsoas muscle by inductively coupled plasma-atomic emission spectrometry. The subjects consisted of 14 men and 14 women, ranging in age from 4 to 102 yr. The insertion tendons of the iliopsoas were removed from the cadavers after ordinary dissection and also surgically from patients with congenital dislocation of the hip. It was found that both Mg and P in the tendon of the iliopsoas muscle increased significantly with aging, but the other elements, such as S, Ca, Na, Zn, Fe, and Si, hardly changed with aging. With regard to the relationship among element contents, extremely significant correlations were found between P and S contents, between Mg and Na contents, and between S and Zn contents. In addition, very significant correlations were found between Si and either S or Zn contents. However, there was no significant correlation between Ca and P contents. These results revealed that with regard to age-related changes of element contents and the relationships among element contents, the tendon of the iliopsoas muscle was different from the Achilles tendon, in which both Mg and P decreased significantly with aging and there were significant correlations each other among the contents of S, Mg, and P.