Immunotherapy for experimental rat autoimmune thyroiditis using a novel immunosuppressant, FTY720

While autoimmune disease needs to be continuously treated via long term, administration of immunosuppressants conventional immunotherapy using drugs such as cyclosporin and tacrolimus may cause adverse effects. Recently, a novel agent, FTY720, which was structurally modified from a natural product, has been shown to have a moderate and different immunosuppressive effect from conventional drugs. In this study, we examined the effect of FTY720 on experimental autoimmune thyroiditis (EAT), which was induced in rats by neonatal thymectomy, followed by subsequent sublethal irradiation. Thyroid stimulating hormone (TSH) was measured before and at the end of drug therapy. Histological and hematological examinations were performed using the samples from sacrificed animals. High TSH levels in the animals returned to the normal range following treatment with FTY720. The severity of the thyroiditis was lower in the FTY720 group than in the control group. FTY720 markedly decreased the number of circulatory lymphocytes, and no infections episodes were observed under this treatment. Thus, FTY720 treatment might be preferred for continuous immunotherapy for autoimmune disease without adverse effects.     

2-Isopropylidenehydrazono-4-oxo-thiazolidin-5-ylacetanilide (OPB-9195) treatment inhibits the development of intimal thickening after balloon injury of rat carotid artery: role of glycoxidation and lipoxidation reactions in vascular tissue damage

We have pursued the hypothesis that the carbonyl modification of proteins by glycoxidation and lipoxidation reactions plays a role in atherogenesis. Human atherosclerotic tissues with fatty streaks and uremic arteriosclerotic tissues were examined, with specific antibodies, to detect protein adducts formed with carbonyl compounds by glycoxidation or lipoxidation reactions, i.e. advanced glycation end products (AGEs) or glycoxidation products, such as carboxymethyllysine (CML) and pentosidine, and lipoxidation products, such as malondialdehyde (MDA)-lysine and 4-hydroxy-nonenal (HNE)-protein adduct. All the four adducts were identified in the proliferative intima and in macrophage-rich fatty streaks. If the carbonyl modification is not a mere result but is a contributor to atherogenesis, inhibition of glycoxidation and lipoxidation reactions might prevent vascular tissue damage. We tested this hypothesis in rats following balloon injury of their carotid arteries, a model exhibiting a remarkable intimal thickening, which are stained positive for all the four adducts. Oral administration of 2-isopropylidenehydrazono-4-oxo-thiazolidin-5-ylacetanili de (OPB-9195), an inhibitor of both glycoxidation and lipoxidation reactions, in rats following balloon injury effectively prevented the intimal thickening. These data suggest a role for the carbonyl modification of proteins by glycoxidation and lipoxidation reactions in most, if not all, types of vascular tissue damage ('carbonyl stress'), and the usefulness of inhibitors of carbonyl reactions for the treatment of vascular tissue damage.     

Chemokine receptor-usage of clinical HIV-1 isolates obtained from patients with HIV-1 infection in late clinical stages using PHA-blast

In the present sudy, chemokine receptor-usage of primary HIV-1 isolates was examined using U87-CD4 cells expressing chemokine receptors CCR3, CCR5 and CXCR4. HIV-1 was isolated from the peripheral blood mononuclear cells (PBMC) and/or plasma of eight HIV-1-infected individuals in late CDC-II and CDC-IV clinical stages using PHA-blast prepared from the PBMC of healthy blood donors. The primary HIV-1 isolates from patients in late CDC-II stage rarely infected monocyte-derived macrophages in the present study, whereas most isolates from patients in the CDC-IV stage infected the macrophages. In the experiments using U87-CD4 cells expressing chemokine receptors, the isolates from patients in the late CDC-II stage infected U87-CD4 cells expressing CXCR4, but not U87-CD4 cells expressing CCR5. In contrast, most isolates from patients in the CDC-IV stage infected both U87-CD4 cells expressing CXCR4 or CCR5. The isolates which infected both U87-CD4 cells were supposed to contain dual tropic HIV-1 or a mixture of CXCR4-tropic and CCR5-tropic HIV-1s. Analysis of the deduced amino acid sequence of the V3 region in proviral env gene showed that the V3 region in U87-CD4 cells infected with CXCR4-tropic HIV-1 isolates was largely different from that in the cells infected with CCR5-tropic isolates, but were highly similar to that in cells infected with dual tropic isolates. These results suggest that PHA-blasts may preferentially support the replication of the CXCR4-tropic and dual tropic HIV-1s, and that CXCR4-tropic and dual tropic HIV-1s are also present in peripheral blood from patients in the late stage of the asymptomatic phase.     

Immunomagnetic separation of scum-forming bacteria using polyclonal antibody that recognizes mycolic acids

Mycolic acid-containing bacteria (mycolata) are thought to be involved in scum formation in aeration basins of activated sludge plants due to their ability to produce biosurfactants and their cell surface hydrophobicity. To isolate these bacteria, immunomagnetic separation (IMS) using an anti-mycolic acid polyclonal antibody was investigated. IMS that targeted Gordonia amarae SC1 exhibited a 100% recovery at 5x10(3) CFU ml(-1). At cell concentration of 7.8x10(6) CFU ml(-1), the recovery was lowered, but 80% of cells were still captured. Effect of bead concentrations on the recovery of SC1 at 10(6) CFU ml(-1) was examined. The results showed that addition of more than 6-7x10(6) beads for 1x10(6) CFU reached a maximum recovery (83%). Furthermore, the IMS procedure optimized with SC1 cells was tested with another mycolata. The results suggested that variation of the recovery for each mycolata is dependent on the specificity of the polyclonal antibody and that mycolata which are recognized by the antibody can be recovered by this procedure.     

Acute effects of a single warm-water bath on serum adiponectin and leptin levels in healthy men: A pilot study

To preliminarily assess the acute effects of a single warm-water bath (WWB) on serum adipokine activity, we measured serum adiponectin, leptin and other metabolic profiles before, immediately after and 30 minutes after WWB in seven healthy male volunteers (mean age, 39.7 ± 6.0 years; mean body mass index, 21.6 ± 1.8 kg/m(2)). The subjects were immersed in tap water at 41°C for 10 minutes. Two weeks later, the same subjects underwent a single WWB with a bath additive that included inorganic salts and carbon dioxide (WWB with ISCO(2)) by the same protocol as for the first WWB. Leptin levels significantly increased immediately after WWB with tap water and ISCO(2) (both P < 0.05), and remained significantly higher than those at baseline even 30 minutes after WWB with tap water (P < 0.05). Adiponectin levels showed a slight, but not significant, increase both immediately after and 30 minutes after WWB with tap water or ISCO(2). Some parameters, such as serum total cholesterol, red blood cell count, hemoglobin and hematocrit significantly increased immediately after WWB with tap water or ISCO(2) (all P < 0.05), but they all returned to the baseline levels 30 minutes after bathing under both conditions. The sublingual temperature rose significantly after 10 minutes of WWB with tap water (0.96 ± 0.16°C relative to baseline, P < 0.01) and after the same duration of WWB with ISCO(2) (1.24 ± 0.34°C relative to baseline, P < 0.01). These findings suggest that a single WWB at 41°C for 10 minutes may modulate leptin and adiponectin profiles in healthy men.     

Construction and Systematical Symmetric Studies of a Series of Supramolecular Clusters with Binary or Ternary Ammonium Triphenylacetates

Functions of clusters in nano or sub-nano scale significantly depend on not only kinds of their components but also arrangements, or symmetry, of their components. Therefore, the arrangements in the clusters have been precisely characterized, especially for metal complexes. Contrary to this, characterizations of molecular arrangements in supramolecular clusters composed of organic molecules are limited to a few cases. This is because construction of the supramolecular clusters, especially obtaining a series of the supramolecular clusters, is difficult due to low stability of non-covalent bonds compare to covalent bonds. From this viewpoint, utilization of organic salts is one of the most useful strategies. A series of the supramolecules could be constructed by combinations of a specific organic molecule with various counter ions. Especially, primary ammonium carboxylates are suitable as typical examples of supramolecules because various kinds of carboxylic acids and primary amines are commercially available, and it is easy to change their combinations. Previously, it was demonstrated that primary ammonium triphenylacetates using various kinds of primary amines specifically construct supramolecular clusters, which are composed of four ammoniums and four triphenylacetates assembled by charge-assisted hydrogen bonds, in crystals obtained from non-polar solvents. This study demonstrates an application of the specific construction of the supramolecular clusters as a strategy to conduct systematical symmetric study for clarification of correlations between molecular arrangements in supramolecules and kinds and numbers of their components. In the same way with binary salts composed of triphenylacetates and one kind of primary ammoniums, ternary organic salts composed of triphenylacetates and two kinds of ammoniums construct the supramolecular clusters, affording a series of the supramolecular clusters with various kinds and numbers of the components.     

Differing expression patterns and evolution of the rat kininogen gene family

The present investigation using molecular cloning and sequence analysis concerns the examination of the molecular basis for different expression patterns of two types of the rat kininogen genes. We show that the low molecular weight and high molecular weight forms of K kininogens are produced from a single gene through alternative usage of two 3'-coding regions, whereas only the low molecular weight forms of T kininogens are generated as a result of several mutational changes in the high molecular weight-specifying regions of both T-I and T-II kininogen genes. The mutational changes include a nucleotide substitution at the polyadenylation/processing signal site, nucleotide deletions resulting in the frame-shift mutation, and an insertion of the type 2 Alu-equivalent sequence. Because kininogens represent a multifunctional protein comprising the proteinase-inhibitory activity, the kinin moiety, and the clotting activity, these results present evidence indicating the molecular basis for the disappearance of a part of the gene functions. We also show that the K and T kininogen genes as well as the two T kininogen genes are extremely homologous, excluding and including the above mutational changes, respectively. These structural relationships allow us to envisage evolutionary processes for the generation of the rat kininogen gene family, particularly for the disappearance of a part of the gene functions.