GB virus C/hepatitis G virus infection in autoimmune liver diseases.
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Aged↗
Biomedical subjects
M Kanaoka
Publications and source records attributed to M Kanaoka.
Intestinal bacterial hydrolysis is required for the appearance of compound K in rat plasma after oral administration of ginsenoside Rb1 from Panax ginseng.
Ginsenoside Rb1 from Panax ginseng root is transformed into compound K via ginsenosides Rd and F2 by intestinal bacterial flora. Among 31 defined intestinal strains from man, only Eubacterium sp. A-44 transformed ginsenoside Rb1 into compound K via ginsenoside Rd. The ginsenoside Rb1-hydrolysing enzyme isolated from Eubacterium sp. A-44 was identical to a previously purified geniposide-hydrolysing beta-D-glucosidase. When ginsenoside Rb1 (200 mg kg-1) was administered orally to germ-free rats, neither compound K nor any other metabolite was detected in the plasma, intestinal tract or cumulative faeces 7 or 15 h after administration. Most of the ginsenoside Rb1 administered was recovered from the intestinal tract, especially the caeca, and cumulative faeces indicating poor absorption of ginsenoside Rb1. When ginsenoside Rb1 was administered orally to gnotobiote rats mono-associated with Eubacterium sp. A-44, a significant amount of compound K was detected in the plasma and considerable amounts were found in the caecal contents and cumulative faeces 7 and 15 h after administration. A small amount of ginsenoside Rb1 was detected in the caecal contents only 7 h after administration. These results indicate that orally administered ginsenoside Rb1 is poorly absorbed from the gut but that its metabolite compound K, produced by ginsenoside Rb1-hydrolysing bacteria such as Eubacterium sp. A-44 in the lower part of intestine, is absorbed.
Administration, Oral↗
Appearance of compound K, a major metabolite of ginsenoside Rb1 by intestinal bacteria, in rat plasma after oral administration--measurement of compound K by enzyme immunoassay.
Enzyme immunoassay (EIA) for the determination of compound K (C-K), a major metabolite of ginsenoside Rb1 (G-Rb1) from Panax ginseng root by intestinal bacterial flora, was explored. Bovine serum albumin (BSA) was coupled to the C-26 position on the unsaturated side chain of C-K. Beta-D-galactosidase was introduced at the C-26 position of the saturated side chain. Antiserum, obtained by immunization of rabbits with C-K-BSA conjugate, possessed high affinity and specificity toward C-K. The EIA for C-K by the double antibody method was established in the range of 0.1--100 ng/tube. Plasma C-K after the oral administration of C-K and G-Rb1 to rats was determined by the established EIA. C-K was rapidly absorbed from the gastrointestinal tract after the administration, then slowly decreased. On the other hand, C-K appeared late and was retained for a long period of time in the plasma after the administration of G-Rb1, which itself is hardly absorbed.
Administration, Oral↗
D1 cap region involved in the receptor recognition and neural cell survival activity of human ciliary neurotrophic factor.
Human ciliary neurotrophic factor (hCNTF), which promotes the cell survival and differentiation of motor and other neurons, is a protein belonging structurally to the alpha-helical cytokine family. hCNTF was subjected to three-dimensional structure modeling and site-directed mutagenesis to analyze its structure-function relationship. The replacement of Lys-155 with any other amino acid residue resulted in abolishment of neural cell survival activity, and some of the Glu-153 mutant proteins had 5- to 10-fold higher biological activity. The D1 cap region (around the boundary between the CD loop and helix D) of hCNTF, including both Glu-153 and Lys-155, was shown to play a key role in the biological activity of hCNTF as one of the putative receptor-recognition sites. In this article, the D1 cap region of the 4-helix-bundle proteins is proposed to be important in receptor recognition and biological activity common to alpha-helical cytokine proteins reactive with gp130, a component protein of the receptors.
Amino Acids↗
Stereoselective production of (+)-trans-chrysanthemic acid by a microbial esterase: cloning, nucleotide sequence, and overexpression of the esterase gene of Arthrobacter globiformis in Escherichia coli.
The gene coding for a novel esterase which stereoselectively hydrolyzes the (+)-trans (1R,3R) stereoisomer of ethyl chrysanthemate was cloned from Arthrobacter globiformis SC-6-98-28 and overexpressed in Escherichia coli. The cellular content of the active enzyme reached 33% of the total soluble protein in the recombinant E. coli JM105 cells and 5.6 g/liter of culture by high-density cell cultivation. The hydrolytic activity of the recombinant E. coli cells for ethyl chrysanthemate reached 605 mumol of chrysanthemic acid per min per g of dry cells, which is approximately 2,500-fold higher than that of A. globiformis cells. The stereoselective hydrolysis by the recombinant E. coli cells was efficient at substrate concentrations of up to 40% by removing the produced chrysanthemic acid by ultrafiltration. The (+)-trans-chrysanthemic acid produced had 100% optical purity. The amino acid sequence of the esterase was found to be similar to that of several class C beta-lactamases, D,D-carboxypeptidase, D-aminopeptidase, 6-aminohexanoate-dimer hydrolase, and Pseudomonas esterase. The sequence comparison also suggested that the Ser-X-X-Lys motif in the esterase was at the active site of the enzyme.
Amino Acid Sequence↗
3-Monoglucuronyl-glycyrrhetinic acid is a major metabolite that causes licorice-induced pseudoaldosteronism.
18 beta-Glycyrrhetinic acid (GA) has been thought to be one of the major metabolites that causes licorice-induced pseudoaldosteronism. However, we found no difference in the blood level of GA between the patients with and without pseudoaldosteronism. We measured the blood concentration of 3 beta-D-(monoglucuronyl)18 beta-glycyrrhetinic acid (3MGA), another metabolite of 3 beta-D-diglucuronyl-18 beta-glycyrrhetinic acid (glycyrrhizin), by high performance liquid chromatography and found an increased concentration of 3MGA in 10 patients with licorice-induced pseudoaldosteronism, but not in 11 patients without pseudoaldosteronism. To investigate whether 3MGA can inhibit 11 beta-hydroxysteroid dehydrogenase, we incubated rat renal microsome with or without 3MGA and measured the conversion rate of [3H]cortisol to [3H]cortisone. 3MGA was found to be a potent inhibitor of 11 beta-hydroxysteroid dehydrogenase, allowing cortisol to exert its full mineralocorticoid effects. These results suggest that licorice-induced pseudoaldosteronism is due to an increased concentration of 3MGA, but not GA, in the circulating blood of these patients.
11-beta-Hydroxysteroid Dehydrogenases↗
Histopathological evaluation of gastric mucosal environments in peptic ulcer using the endoscopic 5-point gastric biopsy method.
Although a strong association has been established between chronic Helicobacter pylori infection and peptic ulcers, the role of H. pylori is not necessarily causative because there are many patients infected with H. pylori who do not develop peptic ulcer. Therefore, we studied the relationship between the gastric mucosal environment and the development of peptic ulcers. We examined 165 endoscopic biopsy specimens from the gastric mucosa of 33 patients with peptic ulcers using the 5-point gastric biopsy method. The follow-up biopsies done within 3 weeks were well correlated with the first biopsy samples. We also reviewed the clinicohistopathological findings of 2250 endoscopic biopsy specimens from 450 patients with active gastric and/or duodenal ulcers. Over 90% of the patients with duodenal ulcer, with or without gastric ulcer, had no fundic gland atrophy, and a high incidence of intestinal metaplasia and pyloric mucosal atrophy was found in the patients with gastric ulcer. These findings suggest that patients with concomitant active gastric and duodenal ulcers exhibit severe atrophic changes in the antral mucosa but not in the fundic mucosa.
Adult↗
[A case of primary pulmonary B cell lymphoma confirmed by gene analysis].
A 76-year-old man visited our clinic with chest radiographic evidence of a coin lesion in the right middle lung. Chest CT demonstrated an infiltrative shadow mainly in the S4 segment. Lymphoproliferative disorders were suggested by transbronchial lung biopsy. Middle lobectomy was performed. Pathological study showed massive proliferation of mature lymphocytes. Immunoperoxidase studies could not demonstrate a neoplastic monoclonal process. Pseudolymphoma was diagnosed. High molecular weight DNAs were extracted from frozen specimens. Whether genes of immunoglobulins and T-cell receptors were rearranged was investigated using DNA probes against JH and TCR beta-1 receptor gene. One rearranged band was found in the JH chain. No rearranged band appeared in TCR beta-1 chain. B cell lymphoma was diagnosed instead of pseudolymphoma.
Aged↗
Humanization of mouse anti-human IL-2 receptor antibody B-B10.
Mouse monoclonal anti-human IL-2 receptor antibody (B-B10) inhibits IL-2-dependent human T-cell proliferation. It has been used in clinical trials in the transplantation field and promising results are being accumulated. Mouse B-B10 antibody was humanized by grafting all CDRs and some framework amino acid residues onto human antibodies, KAS for VH and PAY for V kappa. Nine humanized B-B10s with differently grafted framework residues were constructed and assessed for their biological activities. One of these humanized B-B10, M5, showed nearly the same activity as the mouse B-B10. The 49th residue of V kappa was demonstrated to play a crucial role in the antigen-antibody interaction by 3-D structure analysis with a computer modeling system.
Amino Acid Sequence↗
Intestinal bacterial hydrolysis is indispensable to absorption of 18 beta-glycyrrhetic acid after oral administration of glycyrrhizin in rats.
Gnotobiote rats were prepared by infecting germ-free rats with Eubacterium sp. strain GLH, a human intestinal bacterium capable of hydrolysing glycyrrhizin to 18 beta-glycyrrhetic acid. Their faeces and caecal contents showed glycyrrhizin-hydrolysing activities (31.7 and 31.3 pmol min-1 (mg protein)-1, respectively) similar to those (81.0 and 39.9 pmol min-1 (mg protein)-1, respectively) of conventional rats, although there was no detectable activity in germ-free rats. When glycyrrhizin (100 mg kg-1) was orally administered to conventional, germ-free and gnotobiote rats, no glycyrrhizin could be detected in plasma 4 or 17 h after the administration, using EIA and HPLC assays. Plasma 18 beta-glycyrrhetic acid was not detected 4 or 17 h after the administration of glycyrrhizin to germ-free rats nor could this compound be detected in caecal contents or in the faeces. However, 18 beta-glycyrrhetic acid (0.6-2.6 nmol mL-1) was detected in plasma of the conventional and the gnotobiote rats 4 and 17 h after the administration, and the caecal contents after 4 h and the cumulative faeces up to 17 h of the conventional and the gnotobiote rats contained considerable amounts of 18 beta-glycyrrhetic acid. These findings indicate that orally administered glycyrrhizin is poorly absorbed from the gut, but is hydrolysed to 18 beta-glycyrrhetic acid by intestinal bacteria such as E. sp. strain GLH, and the resulting 18 beta-glycyrrhetic acid is absorbed.
Administration, Oral↗
Modularity of protein function: chimeric interleukin 1 beta s containing specific protease inhibitor loops retain function of both molecules.
Although it is widely recognized that many proteins contain discrete functional domains, it is less certain whether smaller, less obviously discrete, units of structure will retain their specific function when transplanted into a different context. The observation that the potent inflammatory cytokine human interleukin 1 beta has the same overall structure as soybean trypsin inhibitor (STI) (Kunitz) prompted us to replace a tight turn in the cytokine sequence with the large loop in soybean trypsin inhibitor that binds to the active site of trypsin. Wild-type interleukin 1 beta (IL-1 beta) is highly resistant to proteolysis, but the chimeric STI/IL is specifically cleaved by trypsin, apparently in the inserted loop. Other chimeric interleukins have also been constructed, by replacing the same tight turn with inhibitory loops from other protein protease inhibitors: turkey ovomucoid inhibitor (TOI), a chymotrypsin inhibitor, and alpha 1-antitrypsin (AT), an elastase inhibitor. Although these loops come from proteins not related structurally to interleukin 1, they confer specific protease sensitivity or inhibition on the chimeric cytokine. The cytokine properties of these chimeric interleukins have also been evaluated. The chimeras formed from human IL-1 beta and all inhibitory loops tested bind to the interleukin 1 receptor with reasonable affinity. The typical cellular effects of IL-1, however, are not observed with all the recombinant proteins, thus confirming that receptor binding and signal transduction can be uncoupled. When these results are taken together with the results of site-directed mutagenesis of IL-1, reported in this paper and elsewhere, they allow the receptor and intracellular transduction sites on the protein to be mapped in detail.
Amino Acid Sequence↗
Molecular dynamics simulation of winter flounder antifreeze protein variants in solution: correlation between side chain spacing and ice lattice.
The solution structure of the 38 amino acid C-terminal region of the precursor for the HPLC-6 antifreeze protein from winter flounder has been investigated with molecular dynamics using the AMBER software. The simulation for the peptide in aqueous solution was carried out at a constant temperature of 0 degree C and at atmospheric pressure. The simulation covered 120 ps and the results were analyzed based on data sampled upon reaching a stable equilibrium phase. Information has been obtained on the quality of constant temperature and pressure simulations, the solution structure and dynamics, the hydrogen bonding network, the helix stabilizing role of terminal charges and the interaction with the surrounding water molecules. The Lys18-Glu22 interactions and the terminal charged residues are found to stabilize a helical structure with the side chains of Thr2, Thr13, Thr24 and Thr35 equally spaced on one side of the helix. The spacing between oxygen atoms in the hydroxyl group of the threonine side chains exhibits fluctuations of the order of 2-3 A during the 120 ps of simulation, but values simultaneously close to the repeat distance of 16.6 A between oxygen atoms along the [0112] direction in ice are observed. Furthermore, two engineered variants were studied using the same simulation protocol.
Amino Acid Sequence↗
Intercellular adhesion molecule-1 expression on the hepatocyte membrane of patients with chronic hepatitis B and C.
The expression of intercellular adhesion molecule-1 (ICAM-1) on the hepatocyte membrane was studied in 27 patients with chronic hepatitis B and C (CHB, CHC) by immunostaining using a monoclonal antibody. ICAM-1 was expressed focally in a honeycomb-like pattern by hepatocytes in livers of 26/27 patients. The degree of ICAM-1 expression was closely related to the ALT level and the histological grade of liver damage. Abundant cytotoxic T cells (CD8+, CD11b-) were found in ICAM-1-positive areas of the liver. Zones of focal necrosis contained both ICAM-1-positive hepatocytes and cytotoxic T cells. The expression of ICAM-1 was decreased in 4/6 CHB patients after interferon-alpha therapy. No relationship between the degree of hepatocyte ICAM-1 expression and viral replication markers (DNA polymerase activity and the presence of HBcA in the liver) was observed in patients with CHB. In addition, no positive correlation was found between the distribution of ICAM-1-positive hepatocytes and HBcAg-positive hepatocytes. These results suggest that ICAM-1 may play an important role in the pathogenesis of hepatocellular injury mediated by cytotoxic T cells in CHB and CHC.
Antigens, CD↗
Usual interstitial pneumonia: histologic correlation with high-resolution CT.
The authors reviewed 46 cases of idiopathic pulmonary fibrosis with usual interstitial pneumonia (UIP), correlating findings on high-resolution computed tomographic (HRCT) scans with findings in specimens obtained at open lung biopsy and autopsy. The following HRCT findings were observed: (a) an accumulation of small cystic spaces with thick walls, (b) air bronchiolograms within areas of intense lung attenuation, (c) rugged pleural surfaces, (d) irregularly thickened pulmonary vessels, (e) bronchial wall thickening, and (f) slightly increased lung attenuation. Macroscopic honeycombing correlating with small cystic spaces was demonstrated at HRCT and pathologic examination. Air bronchiolograms in the areas of intense lung attenuation (ie, microscopic honeycombing) corresponded to dilated bronchioles (greater than 1 mm in diameter) with fibrosis. Irregularly thickened vessels and bronchial walls and irregular pleural surfaces were the result of fibrosis in the periphery of the secondary pulmonary lobules. Areas of slightly increased lung attenuation seen on the HRCT scans correlated with patchy alveolar septal fibrosis or inflammation. The authors conclude that microscopic honeycombing and a perilobular distribution in UIP may be clearly identified with HRCT.
Adult↗
Studies on the enzyme immunoassay of bio-active constituents in oriental medicinal drugs. VI. Enzyme immunoassay of ginsenoside Rb1 from Panax ginseng.
Enzyme immunoassay (EIA) of ginsenoside Rb1 (GRb1), one of the glucosides of protopanaxadiol from Panax ginseng, was explored. A carrier protein (bovine serum albumin (BSA)) was coupled to the C-26 position on the unsaturated side chain of the protopanaxadiol moiety to prepare the immunogen. In order to perform bridge heterologous EIA, a label (beta-D-galactosidase) was introduced at C-26 of the saturated side chain to obtain labeled antigen. Anti-GRb1 antisera were elicited in rabbits by immunization with GRb1-BSA conjugate (9). The double antibody method (with goat anti-rabbit IgG antiserum) was used to separate the bound and free GRb1-beta-Gal. A satisfactory standard curve for EIA of GRb1 was obtained in the range of 0.04-10 ng/tube. In a comparison of the assay results obtained by EIA and HPLC, the linear regression equation and correlation coefficient for the two methods were y (EIA) = 9.18x(HPLC)-0.033 and 0.98, respectively. The anti-GRb1 antiserum cross-reacted with GRb2 (21.8%) and GRc (10.6%), which are also constituents of Panax ginseng.
Animals↗
Ion spray mass spectra of native and recombinant streptococcal antitumor proteins.
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Antibiotics, Antineoplastic↗
Insertion of an elastase-binding loop into interleukin-1 beta.
The protease-binding sequence EAIPMSIPPE from alpha 1-antitrypsin has been inserted into the cytokine interleukin-1 beta, replacing residues 50-53. The resulting mutant protein was cleaved specifically at a single site by elastase and chymotrypsin, but not by trypsin. The cleavage by elastase was shown to be between Met and Ser of the inserted loop. In contrast, wild-type interleukin is not susceptible to cleavage by any of these enzymes. The mutant protein acts as an inhibitor of elastase, with a KI of approximately 30 microM. The wild type displays no such inhibitory activity. The overall structure of the mutant, as demonstrated by CD, appears to be indistinguishable from that of the wild type. These results indicate that the protease-binding region of alpha 1-antitrypsin can be recognized and is active even within the context of an entirely different protein structure. Given that interleukin-1 beta binds to, and is internalized by, many types of cells, this hybrid protein also demonstrates the feasibility of using interleukin-1 beta as a delivery system for useful therapeutic agents.
Amino Acid Sequence↗
Effect of oral administration of glycyrrhizin on the pharmacokinetics of prednisolone.
The pharmacokinetics of total and free prednisolone (PSL) in six healthy men, with or without pretreatment with oral glycyrrhizin (GL), was investigated to confirm whether oral administration of GL influences the metabolism of PSL in man. Each subject received an intravenous administration of 0.096 mg/kg of prednisolone hemisuccinate (PSL-HS) with or without pretreatment with 50 mg of oral GL four times. Blood samples were taken from a peripheral vein at 5, 10, 15, 30, 45 min and 1, 1.5, 2, 3, 4, 6, 8, 10, 12 and 24 h after the start of PSL-HS infusion. The concentrations of total PSL in plasma were analyzed by high-performance liquid chromatography, and the free PSL was measured by an isocolloidosmolar equilibrium dialysis method. The pharmacokinetic parameters of PSL were determined by non-compartment analysis. Oral administration of GL was found to significantly increase the concentrations of total PSL at 6, 8 h, and of free PSL at 4, 6 and 8 h after PSL-HS infusion. Moreover, oral administration of GL was also found to modify the pharmacokinetics of both total and free PSL. After oral administration of GL, the area under the curve (AUC) was significantly increased, the total plasma clearance (CL) was significantly decreased, and the mean residence time (MRT) was significantly prolonged. However, the volume of distribution (Vdss) showed no evident change. This suggests that oral administration of GL increases the plasma PSL concentrations and influences its pharmacokinetics by inhibiting its metabolism, but not by affecting its distribution.
Administration, Oral↗