Association of reduced heme oxygenase-1 with excessive Toll-like receptor 4 expression in peripheral blood mononuclear cells in Behçet's disease

Introduction

Toll-like receptors (TLRs) mediate signaling that triggers activation of the innate immune system, whereas heme oxygenase (HO)-1 (an inducible heme-degrading enzyme that is induced by various stresses) suppresses inflammatory responses. We investigated the interaction between TLR and HO-1 in an inflammatory disorder, namely Behçet's disease.

Methods

Thirty-three patients with Behçet's disease and 30 healthy control individuals were included in the study. Expression levels of HO-1, TLR2 and TLR4 mRNA were semiquantitatively analyzed using a real-time PCR technique, and HO-1 protein level was determined by immunoblotting in peripheral blood mononuclear cells (PBMCs) and polymorphonuclear leukocytes. In some experiments, cells were stimulated with lipopolysaccharide or heat shock protein-60; these proteins are known to be ligands for TLR2 and 4.

Results

Levels of expression of HO-1 mRNA were significantly reduced in PBMCs from patients with active Behçet's disease, whereas those of TLR4, but not TLR2, were increased in PBMCs, regardless of disease activity. Moreover, HO-1 expression in PBMCs from patients with Behçet's disease was repressed in the presence of either lipopolysaccharide or heat shock protein-60.

Conclusion

The results suggest that upregulated TLR4 is associated with HO-1 reduction in PBMCs from patients with Behçet's disease, leading to augmented inflammatory responses.     

Genomewide analysis of PRC1 and PRC2 occupancy identifies two classes of bivalent domains

In embryonic stem (ES) cells, bivalent chromatin domains with overlapping repressive (H3 lysine 27 tri-methylation) and activating (H3 lysine 4 tri-methylation) histone modifications mark the promoters of more than 2,000 genes. To gain insight into the structure and function of bivalent domains, we mapped key histone modifications and subunits of Polycomb-repressive complexes 1 and 2 (PRC1 and PRC2) genomewide in human and mouse ES cells by chromatin immunoprecipitation, followed by ultra high-throughput sequencing. We find that bivalent domains can be segregated into two classes -- the first occupied by both PRC2 and PRC1 (PRC1-positive) and the second specifically bound by PRC2 (PRC2-only). PRC1-positive bivalent domains appear functionally distinct as they more efficiently retain lysine 27 tri-methylation upon differentiation, show stringent conservation of chromatin state, and associate with an overwhelming number of developmental regulator gene promoters. We also used computational genomics to search for sequence determinants of Polycomb binding. This analysis revealed that the genomewide locations of PRC2 and PRC1 can be largely predicted from the locations, sizes, and underlying motif contents of CpG islands. We propose that large CpG islands depleted of activating motifs confer epigenetic memory by recruiting the full repertoire of Polycomb complexes in pluripotent cells.     

Characterization of the carbohydrase productions of an ectomycorrhizal fungus, Tricholoma matsutake

To evaluate the potential of the production of the ectomycorrhizal fungus Tricholoma matsutake to produce carbohydrases, (1) the distribution of carbohydrase activities among the different strains (18 strains) was investigated and (2) the abilities of T. matsutake and saprophytic fungi to produce β-glucosidase were compared. The results showed that the carbohydrase productions patterns of T. matsutake still resemble one another. Moreover, this fungus exhibited markedly higher β-glucosidase than did the saprophytic mushrooms. Tricholoma matsutake showed weak production of α-amylase and α-glucosidase in a static cultur filtrate. On the other hand, glucoamylase activity was not observed. Surprisingly, we discovered that β-glucosidase demonstrated strong activity. This finding suggests that this fungus has saprotrophic abilities. The carbohydrase production systems in T. matsutake were characterized from our experimental results. Also, we point out some weak points in the carbohydrase production systems of T. matsutake.     

Restriction on conjugational transfer of pLS20 in Bacillus subtilis 168

Conjugational transfer of pLS20 in Bacillus subtilis Marburg 168 is restricted by the BsuM restriction-modification system. Restriction efficiency was measured using pLS20 derivatives possessing various numbers of XhoI sites, which are known to be recognized by BsuM. An increase in XhoI sites clearly reduced the conjugational efficiency of pLS20 as compared with that of pUB110 plasmid lacking XhoI.     

A tightly bound quinone functions in the ubiquinone reaction sites of quinoprotein alcohol dehydrogenase of an acetic acid bacterium, Gluconobacter suboxydans

Quinoprotein alcohol dehydrogenase (ADH) of acetic acid bacteria is a membrane-bound enzyme that functions as the primary dehydrogenase in the ethanol oxidase respiratory chain. It consists of three subunits and has a pyrroloquinoline quinone (PQQ) in the active site and four heme c moieties as electron transfer mediators. Of these, three heme c sites and a further site have been found to be involved in ubiquinone (Q) reduction and ubiquinol (QH2) oxidation respectively (Matsushita et al., Biochim. Biophys. Acta, 1409, 154-164 (1999)). In this study, it was found that ADH solubilized and purified with dodecyl maltoside, but not with Triton X-100, had a tightly bound Q, and thus two different ADHs, one having the tightly bound Q (Q-bound ADH) and Q-free ADH, could be obtained. The Q-binding sites of both the ADHs were characterized using specific inhibitors, a substituted phenol PC16 (a Q analog inhibitor) and antimycin A. Based on the inhibition kinetics of Q2 reductase and ubiquinol-2 (Q2H2) oxidase activities, it was suggested that there are one and two PC16-binding sites in Q-bound ADH and Q-free ADH respectively. On the other hand, with antimycin A, only one binding site was found for Q2 reductase and Q2H2 oxidase activities, irrespective of the presence of bound Q. These results suggest that ADH has a high-affinity Q binding site (QH) besides low-affinity Q reduction and QH2 oxidation sites, and that the bound Q in the QH site is involved in the electron transfer between heme c moieties and bulk Q or QH2 in the low-affinity sites.     

Synthesis of 5'-branched derivatives of cyclic ADP-carbocyclic-ribose, a potent Ca2+-mobilizing agent: The first antagonists modified at the N1-ribose moiety

The 5'-branched cyclic ADP-carbocyclic-ribose derivatives were designed and synthesized. These target compounds were identified as the first antagonists of cADPR without a substituent at the adenine 8-position, and were shown to be stable due to the N1-carbocyclic-ribosyl structure.     

Single molecule measurements and molecular motors

Single molecule imaging and manipulation are powerful tools in describing the operations of molecular machines like molecular motors. The single molecule measurements allow a dynamic behaviour of individual biomolecules to be measured. In this paper, we describe how we have developed single molecule measurements to understand the mechanism of molecular motors. The step movement of molecular motors associated with a single cycle of ATP hydrolysis has been identified. The single molecule measurements that have sensitivity to monitor thermal fluctuation have revealed that thermal Brownian motion is involved in the step movement of molecular motors. Several mechanisms have been suggested in different motors to bias random thermal motion to directional movement.     

Excretion of urinary cadmium, copper, and zinc in cadmium-exposed and nonexposed subjects, with special reference to urinary excretion of β2-microglobulin and metallothioneinand metallothionein

The objectives of this study were to examine the association between urinary excretion of cadmium (U-Cd), copper (U-Cu), and zinc (U-Zn) and the severity of two different indicators of renal toxicity (urinary excretion of beta2-microglobulin [U-beta2-MG] and metallothionein [U-MT]) in Cd-exposed subjects compared to controls, and to assess the physiologic mechanisms by which the exposure to environmental Cd affects U-Cd, U-Cu, and U-Zn. The target population included 3508 Cd-exposed and 294 nonexposed participants who received a health survey conducted among the population of the Kakehashi River basin. Increases of U-Cd, U-beta2-MG, and U-MT in the Cd-exposed population were observed relative to excretion of these substances in controls. Regression analysis using a general linear model revealed that the correlations between U-Cd or U-Cu, and U-beta2-MG and between U-Cd, U-Cu or U-Zn, and U-MT were statistically significant in both sexes, but the correlation between U-Zn and U-beta2-MG excretion was significant only in men. These results suggest U-Cd and U-Cu is affected by dysfunction in renal tubular absorption (indicated by U-beta2-MG), whereas not only U-Cd and U-Cu but also U-Zn appear to be a function of renal cellular desquamation (indicated by U-MT).